Hepatic stellate cells and liver fibrosis.

Hepatic Stellate Cells and Liver Fibrosis Juan E. Puche,1,2 Yedidya Saiman,1 and Scott L. Friedman*1 ABSTRACT Hepatic stellate cells are resident perisinusoidal cells distributed throughout the liver, with a remarkable range of functions in normal and injured liver. Derived embryologically from septum transversum mesenchyme, their precursors include submesothelial cells that invade the liver parenchyma from the hepatic capsule. In normal adult liver, their most characteristic feature is the presence of cytoplasmic perinuclear droplets that are laden with retinyl (vitamin A) esters. Normal stellate cells display several patterns of intermediate filaments expression (e.g., desmin, vimentin, and/or glial fibrillary acidic protein) suggesting that there are subpopulations within this parental cell type. In the normal liver, stellate cells participate in retinoid storage, vasoregulation through endothelial cell interactions, extracellular matrix homeostasis, drug detoxification, immunotolerance, and possibly the preservation of hepatocyte mass through secretion of mitogens including hepatocyte growth factor. During liver injury, stellate cells activate into alpha smooth muscle actin-expressing contractile myofibroblasts, which contribute to vascular distortion and increased vascular resistance, thereby promoting portal hypertension. Other features of stellate cell activation include mitogen-mediated proliferation, increased fibrogenesis driven by connective tissue growth factor, and transforming growth factor beta 1, amplified inflammation and immunoregulation, and altered matrix degradation. Evolving areas of interest in stellate cell biology seek to understand mechanisms of their clearance during fibrosis resolution by either apoptosis, senescence, or reversion, and their contribution to hepatic stem cell amplification, reC 2013 American Physiological Society. Compr Physiol generation, and hepatocellular cancer. 3:1473-1492, 2013.

Introduction Liver fibrosis is a common outcome of virtually all chronic hepatic insults including viral hepatitis (i.e., hepatitis B and C), alcoholic or obesity-associated steatohepatitis (i.e., nonalcoholic steatohepatitis (NASH)), parasitic disease (i.e., schistosomiasis), metabolic disorders (i.e., Wilson’s), hemochromatosis and other storage diseases, congenital abnormalities, autoimmune and chronic inflammatory conditions (i.e., sarcoidosis), and drug toxicity, among others (102). Fibrosis was described in 1951 as a passive process with “no direct evidence of fibrous tissue proliferation but the suggestion of connective tissue appearance just by stromal condensation due to hepatocyte cell collapse.” However, in 1978, a growing body of evidence prompted the World Health Organization to update its definition of fibrosis as “the presence of excess collagen due to new fiber formation (3).” It is virtually axiomatic now that liver fibrosis results from enhanced production of extracellular matrix (ECM) due to accumulation and activation of myofibroblasts in the context of ongoing or repetitive liver damage. Early studies focused on methods to isolate and grow primary hepatic stellate cells (HSCs) in culture establishing them as one of the main sources of myofibroblasts in liver parenchymal disease resulting from hepatocyte as oppsed to biliary injury (49, 71). Immediately thereafter, the concept of stelllate cell “activation” emerged, which represents a transdifferentiation of the cell during liver injury from a quiescent state that is rich in vitamin A droplets,

Volume 3, October 2013

to one that is proliferative fibrogenic and contractile (62, 82, 83, 234, 241, 289). While HSCs are a major source of myofibroblasts, mounting evidence also implicates portal fibroblasts as a source when the injury is to bile ducts rather than hepatocytes (53, 162). From early studies focusing on the role of the stellate cells solely as a source of ECM during liver injury, a sustained effort has subsequently uncovered broadening roles of the cell type as a source of regenerative cytokines, an immunomodulatory cell with a range of activities and one that serves roles far beyond those envisioned at the time of its isolation 35 years ago (65). From these studies realistic hopes have emerged for exploiting features of stellate cell activation and hepatic inflammation in devising effective antifibrotic and regenerative therapies for patients with chronic liver disease and fibrosis. This article builds upon a comprehensive review of stellate cell biology in 2008 by one of the authors (S.L.F.) (65),

* Correspondence

to [email protected] of Liver Diseases, Icahn School of Medicine at Mount Sinai Hospital, New York, New York 2 University CEU-San Pablo, School of Medicine, Institute of Applied Molecular Medicine (IMMA), Madrid, Spain Published online, October 2013 (comprehensivephysiology.com) DOI: 10.1002/cphy.c120035 C American Physiological Society. Copyright 1 Division

1473

Hepatic Stellate Cells and Liver Fibrosis

Comprehensive Physiology

Figure 1

Appearance of hepatic stellate cells and the sinusoidal microenvironment in normal and injured liver. In normal liver, stellate cells (shown in blue) are laden with perinuclear retinoid droplets and preserve the differentiated function of surrounding cells, including hepatocytes and sinusoidal endothelial cells. In liver injury, the cells multiply, lose vitamin A and become embedded within dense extracellular matrix. This leads to deterioration of hepatocyte function manifested as loss of microvilli, and decreased size and number of endothelial fenestrations. Reprinted, with permission, from (68).

by highlighting the accelerating pace of progress and increasingly nuanced understanding of a cell type that is unique not only in liver, but throughout mammalian biology. The fascination with stellate cells may explain why the published literature contains more than 2800 articles about this cell type only in the last ten years (Pubmed search using the keyword “hepatic stellate cell” from 2002-2012).

Hepatic Stellate Cell Biology, Origin and Ultrastructure HSCs are resident nonparenchymal cells located in the subendothelial space of Disse, between the basolateral surface of hepatocytes and the antiluminal side of sinusoidal endothelial cells (Fig. 1). This privileged location, together with their dendritic cytoplasmic processes, facilitates their direct contact with hepatocytes, endothelial cells, other stellate cells, and Kupffer cells up to 140 μm away (86, 126). This intimate contact between stellate cells and their neighboring cell types may facilitate intercellular transport of soluble mediators and cytokines. In addition, stellate cells are directly adjacent to nerve endings (19, 299), which is consistent with reports identifying neurotrophin receptors (36, 137, 284, 333), and with functional studies confirming neurohumoral responsiveness of stellate cells (158, 203, 249). Interestingly, apart from the different patterns of distribution (pericentral vs. periportal predominances) among species (28, 65, 305), stellate cells only represent ∼10% of the total number of resident cells in normal liver (86) and ∼1.5% of total liver volume, a low proportion of the total cell number in contrast to their remarkably divergent functions in normal and diseased liver. Prominent dendritic cytoplasmic processes from stellate cells contact hepatocytes and endothelial cells (65, 266, 302, 304). These subendothelial processes have three cell surfaces: inner, outer, and lateral. The inner one is smooth and adheres to the adluminal (basal) surface of the liver sinusoidal

1474

endothelial cells (LSECs) while the outer surface, facing the space of Disse, has numerous micro-projections which contact with hepatocytes and may function in detecting chemotactic signals, and transmitting them to the cell’s mechanical apparatus to generate a contractile force that regulates blood flow (180). Actin filaments and microtubules are distributed in both the periphery and the core of the cell’s processes, respectively, and could be responsible for their extension and retraction (115, 148, 204, 246, 252). While no electron dense basement membrane can be identified within the perisinusoidal space of Disse, basement membrane proteins, including type IV collagen, nidogen, and laminin, have been identified, which are thought be functionally important in preserving differentiated hepatocellular function and stellate cell quiescence (20, 29, 70). HSCs were first described by Kupffer in 1876 (303), using a gold chloride method for detection of neuronal components in the liver. Their star-shaped characteristics led Kupffer to call them “sternzellen” (“star cell,” in German). However, in 1898, a misinterpretation based on India ink staining led him to conclude that they were actually “special endothelial cells of the sinusoids with phagocytic capacity,” thereby confusing them with macrophages (now often called “Kupffer cells” (304)). Ironically, recent studies have indeed established that stellate cells have phagocytic properties (35, 129, 190, 301). Since Kupffer first identified stellate cells nearly 150 years ago (303), a number of new techniques for their detection and isolation have been developed based on their lipid droplet content, cytoskeletal features, and cell surface markers (288). These approaches have aided in further defining their ultrastructure and enhancing their purity during isolation, for example, by using vitamin A fluorescence to isolate the cells using flow cytometry (45). In normal liver, stellate cells have spindle-shaped cell bodies with perikaryons within recesses between neighboring parenchymal cells, with oval or elongated nuclei. Vitamin A lipid droplets are conspicuous features of the cytoplasm. Stellate cells also have



well-developed juxtanuclear small Golgi complex and rough endoplasmic reticulum spaces indicating active biosynthesis of secreted polypeptides or proteins (56). The presence of active lysosomes in stellate cell cytoplasm described for decades (33, 129) is now recognized to indicate a high capacity for the cells to undergo autophagy during cellular activation (104, 105). Endosomes and multivesicular bodies are also present in HSCs (267) and contribute to the generation of vitamin A-containing lipid droplets. In contrast to the well-established origins of other liver populations (endoderm-derived hepatocytes and cholangiocytes, mesoderm-derived endothelial cells and fibroblasts, and ectoderm-derived neurons), the ontogeny of this enigmatic cell has been controversial for decades, because the cells express a pattern of cytoskeletal markers reflecting a range of origins including ectoderm (e.g., glial fibrillary acidic protein (GFAP), nestin, neurotrophins and their receptors, nerve growth factor (NGF), brain-derived neurotrophic factor, synaptophysin, and N-CAM) and mesoderm (e.g., vimentin, desmin, alpha smooth muscle actin, hematopoietic markers) (80, 186). More recently, elegant developmental studies have established that stellate cells are traced to mesothelial cells (likely derived from septum transversum mesenchyme), which appear to give rise to cells that invade the hepatic bud (8, 11, 165). However, due to the limited labeling efficiency of the mesothelium, the proportion of stellate cells derived from the mesothelium is not known and other cellular sources may also be possible. There is ample evidence that HSCs are remarkably heterogeneous in their content of retinoid, cytoskeletal phenotype, potential for activation, and even their capacity to revert to a quiescent state after liver injury resolves (45, 80, 143, 170). For example, there is a subpopulation of stellate cells that may lack typical cytoskeletal markers (16, 235, 257) depending on the lobular location. Pericentral areas are rich in stellate cells with longer cytoplasmic processes (i.e., more astrocytic morphology), predominant GFAP expression (instead of desmin), decreased number and size of lipid droplets, and more differentiated. Periportal stellate cells are typically desmin positive with shorter cytoplasmic processes (with a more contractile phenotype), contain more and larger lipid droplets and may be less differentiated (82, 195, 304, 305). While the origin of stellate cells is becoming less controversial, still uncertain is the possibility that stellate cells are pluripotent and can give rise to multiple cell lineages. This phenomenon has been reported in at least two studies (154, 324), but based only on cell culture findings, where modulation of cellular phenotype is notoriously promiscuous and may not reflect events in vivo. On the other hand, it is increasingly clear that HSCs are intimately associated with the progenitor cell niche and typically surround cells that have stem cell-like properties (39, 40, 89, 219, 311). These findings indicate a strong likelihood that stellate cells support stem cell expansion, although underlying mechanisms and mediators that drive this interaction are not known.



Extrahepatic stellate cells have also been described throughout many organs in particular pancreas, where they retain their characteristic shape and markers (267). Stellate cells in other tissues typically store vitamin A and synthesize and secrete ECM components. Best characterized among these are pancreatic stellate cells, which clearly contribute to pancreatic fibrosis and tumor stroma (4, 5, 57). Pancreatic stellate cells reportedly generate stem cells that may also directly contribute to liver regeneration through differentiation into hepatocytes and duct-forming cholangiocytes across tissue boundaries, but this observation requires confirmation (153).

Role of Hepatic Stellate Cells in Normal Liver Because of their recognized role in hepatic fibrosis, most studies of HSCs have focused primarily on their behavior during liver injury, but have neglected their contribution to normal liver homeostasis. With an increased availability of tools to selectively express transgenes in stellate cells, their role in normal liver development and function is now being illuminated (Fig. 2).

Contribution of stellate cells to liver development As noted above, stellate cells can be identified within the progenitor cell niche (near the canals of Hering) in normal, developing, and regenerating liver (248, 326). Additionally, murine fetal liver-derived Thy1+ cells, which express classical markers of HSCs (α-SMA, desmin, and vimentin), promote maturation of hepatic progenitors through cell-cell contact in culture (12). Pleiotrophin, a morphogen secreted by stellate cell precursors (ALCAM+ submesothelial cells) during liver development, may contribute to liver organogenesis and regeneration (9, 10). Quiescent stellate cells also express epimorphin, a mesenchymal morphogenic protein involved

Extracellular matrix homeostasis Vasoregulation

Preservation of hepatocyte mass

Normal liver development

Quiescent stellate cell

Retinoid metabolism

Drug metabolism and detoxification

Figure 2

Roles of hepatic stellate cells in normal liver.

1475


in differentiation of rat hepatic stem-like cells by a putative epithelial-mesenchymal contact that promotes bile duct epithelial morphogenesis (184), which involves the RhoA and C/EBPβ pathways. These findings complement evidence of paracrine interactions between bile duct epithelium and either stellate cells or portal fibroblasts both in culture (156, 166) and in vivo (141, 156, 285). While not limited to stellate cellbile duct crosstalk, components of the Notch (254) and Wnt pathways (178, 330), purinergic signaling (52), chemokines (156) and the Dlk1 protein (298, 330) are also important in hepatic development. Stellate cell precursors, isolated from fetal liver based on UV fluorescence in flow cytometry (157), display extensive proliferative activity, and a high capacity to express hepatocyte growth factor (HGF), CXCL12, and homeobox transcription factors, supporting their potential contributions to both hepatic development and hematopoiesis (157). A recent study has identified stellate cells in zebrafish using a reporter gene driven by the Hand2 promoter (325). Use of this model will further clarify the stellate cell’s contributions to hepatic development and homeostasis.

Retinoid metabolism (see section on Perpetuation, Section V, below) Vitamin A (retinoid) is primarily stored in the liver in mammals, and among liver cell types stellate cells are the primary cellular depot (303). Normally, dietary retinoids are absorbed by the gut and transported in chylomicron remnants as retinyl esters to hepatocytes, where they are hydrolyzed into free retinol. Retinol is then transferred to stellate cells, where they are reesterified (101). Importantly, these droplets contain not only retinoids, but also triglycerides, phospholipids, cholesterol, and free fatty acids, among others (188, 320). Recent studies have identified a family of proteins that coat lipid droplets known as perilipins (27, 28, 253). One perilipin, adipose-differentiation-related protein, is expressed by stellate cells and its levels are reduced as the cells activate and lose retinoid droplets (161). The contributions of these lipid droplets go beyond the simple storage of Vitamin A, and extend to the regulation of stellate cell activation (206, 207), possibly through the impact of lipids in fueling autophagy (103, 104). The importance of these retinoid mechanisms in fibrosis however has been challenged, however, as mice deficient in lecithin retinol acetyl transferase (LRAT), the enzyme that catalyzes the esterification of retinol into retinyl esters nonetheless undergo fibrosis (144) following toxic liver injury, perhaps indicating an alternate or more complex role for retinoid metabolism in hepatic and stellate cell homeostasis.

Extracellular matrix homeostasis In normal liver, the ECM comprises ∼0.5% of the total liver weight (245) and is distributed between portal triads, central veins and Glisson’s capsule, with only a small portion present in the space of Disse (81). Normal ECM components include

1476


collagens (type I, III, IV, V, VI, XIV, and XVIII), elastin, structural glycoproteins (laminin, fibronectin, nidogen/ entactin, tenascin, osteopontin, and secreted acidic proteins rich in cysteine), proteoglycans (heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate, and dermatan sulfate, syndecan, biglycan, and decorin), and the free glycosaminoglycan hyaluronan (74, 136, 259). While quantitatively modest in the latter location, ECM in the space of Disse has an important role in preserving liver homeostasis and has a unique spatial expression pattern. Type IV collagen is primarily located between LSECs and stellate cells while type I and III fibrillar collagens are located between HCSs and hepatocytes in normal liver (20, 70). Primarily, the ECM composition can affect the behavior of surrounding liver cells through cell surface receptors, especially integrins, of which stellate cells express α1β1, α1β2 (227), αvβ6 (222), α5 β1 (110), αvβ6 (216, 222), as well as integrin linked kinase (269). The three cell types surrounding the space of Disse (hepatocytes, endothelial cells, and stellate cells) each produce ECM components in normal liver. While all of them express collagen type I, hepatocytes mainly produce fibronectin (233), endothelial cells express collagen IV, and quiescent stellate cells secrete laminin and collagen types III and IV (83, 171), among several other ECM proteins. The maintenance of ECM homeostasis requires turnover in which production of new components is offset by parallel rates of degradation. Matrix metalloproteinases (MMPs) are the primary effectors of ECM degradation, whose activity is regulated in turn by tissue inhibitors of metalloproteinases (TIMPs) (6, 145). Several liver cell populations (i.e., Kupffer cells, myofibroblast, and hepatocytes) can produce both MMPs and TIMPs (6, 145), however a subgroup of this family, the A Disintegrin and Metalloproteinase-domain proteins (ADAMs) may elude TIMP action and contribute to transforming growth factor beta (TGF-β) activation (26), the most potent stimulus for collagen I production by stellate cells (91, 116, 296). While most ADAMs are expressed by more than one liver cell type, at least two (ADAM 13 and 28) are produced solely by stellate cells (194, 261, 273, 314).

Secretion of mediators While stellate cell-derived molecules are a major driving force in hepatic fibrosis they may also play an important role in preserving liver homeostasis and promoting regeneration, although the data do not fully support such a role yet. The specific spatial and temporal expression patterns of these molecules may therefore be important to promoting proper hepatic development and regeneration after injury. In steady state conditions, stellate cells are reported to secrete a range of molecules detailed in the following sections.

Growth factors HGF is the most potent mitogen for hepatocytes (256). Quiescent stellate cells can produce HGF, but interestingly, during



liver damage the expression of this growth factor is downregulated in stellate cells by the action of transforming growth factor β (TGF-β) (232). This temporal expression of HGF may, therefore, explain the decreased rate of hepatic regeneration in a fibrotic/injured liver. TGF-β is among most potent cytokines that regulate stellate cell phenotype (21, 51). In normal liver TGF-β isoform expression (TGF-β1,2,3 ) is shared between hepatocytes, Kupffer cells and stellate cells (21). While TGF-β1 is more highly expressed by Kupffer cells than HSCs, TGF-β3 is only expressed by stellate cells (48). Regardless, TGF-β is secreted in its latent form, and requires a further activation of the latent molecule to exert its action. The predominant pathways of TGF-β activation diverge among different tissues. In liver, integrins, fibrinogen, and urokinase-type plasminogen activator, among others, can activate latent TGF-β during liver injury, which eventually induces stellate cell activation (25, 227, 278). On the other hand, it can be inactivated by binding to the proteoglycan decorin (15). The recent elucidation of the latent TGF-β structure (271) could yield important new approaches to selectively blocking its activation in vivo. Vascular endothelial growth factor (VEGF) is also expressed by quiescent stellate cells (316). Its potent mitogenic effect toward sinusoidal and endothelial cells underscores the key role that stellate cells play in communication and control of liver homeostasis. This important paracrine signaling pathway between stellate cells and sinusoidal endothelium, mediated by VEGF and soluble guanylate cyclase, may be critical for sinusoidal homeostasis in normal liver and regeneration (309, 310, 319). Other growth factors synthesized by stellate cells are insulin-like growth factors (IGF-I and IGF-II), transforming growth factor α (TGF-α), EGF, stem cell factor, and fibroblast growth factors (both acidic and basic FGF), although their contributions may be more critical during liver development and regeneration (39, 40, 75, 177, 182, 191, 247, 298, 332).

Neurotrophins and their receptors NGF, brain-derived neurotrophin, neurotrophin 3, neurotrophin 4/5, the low-affinity NGF receptor p75 and the highaffinity tyrosine kinase receptors B and C are all expressed by HSCs (36), and/or their precursors (284). A number of potential functions of these pathways are suggested by their activities in other tissues; however, to date, their best known function in liver is in contributing to stellate cell activation and tissue repair (36, 137, 215).

Other mediators Endothelin-1 (ET-1) is a potent vasoconstrictor produced primarily by endothelial cells in normal liver (322), but also by stellate cells. Interestingly, during liver injury, endothelial cells decrease their production and stellate cells become the dominant source of ET-1, which correlates with stellate cell activation (138, 221, 242, 270) highlighting the complex interplay of ET-1 between stellate and endothelial



cells. A carefully regulated pathway exists for activation of latent endothelin in stellate cells (138, 164). Human stellate cells have also functional receptors for adrenomedullin (ADM), a peptide produced by most contractile cells, which modulates the contractile effect of ET-1 (88). Moreover, cultured human stellate cells secrete ADM in baseline conditions, and its production is markedly increased by cytokines (88), a surprising finding given that stellate cell activation promotes cellular contraction. ADM can also attenuate activation of stellate cells by inhibiting TGF-β1 production and TGF-β-induced MMP-2 expression partially through the ERK pathway (312). These results suggest that ADM regulates stellate cell activation and contractility in an autocrine manner. A more comprehensive assessment of protein production by stellate cells was generated by an unbiased proteomic analysis of the cells and their surrounding ECM (13, 127, 236). In initial work by Kristensen, the patterns of protein expression were compared between quiescent, in vivo activated and in vitro activated stellate cells, by two-dimensional-gel electrophoresis. From the 300 identified proteins, 83 were found to be secreted, including collagen α1 (I), α1 (III), and α2 (I); α1antitrypsin; calcyclin, calgizzarin, and galectin-1; proteases including plasminogen activator inhibitor-1 and cathepsin A, B, and D; ganglioside GM2; among others. A more recent analysis by Ji et al. has emphasized the importance of stellate cells in also generating immunoregulatory molecules, consistent with their function in conferring immune tolerance in liver (130, 290, 318).

Drug metabolism and detoxification HSCs express both alcohol- and acetaldehydedehydrogenases, but not cytochrome P450-2E1 (34) and it is likely that their contribution to ethanol detoxification is minimal compared to hepatocytes. Apart from P450-2E1, other isoforms of cytochrome p450 are expressed by stellate cells, and are downregulated during cellular activation (321); however, their roles in cellular quiescence and activation are unknown. Some cytochrome p450 isoforms have been identified in stellate cells (160), implicating their participation in xenobiotic detoxification and oxidant stress response.

Role of Hepatic Stellate Cells in Liver Injury and Fibrosis The framework for understanding stellate cell activation was established several years ago (63), and remains a practical and relevant template for characterizing the cell’s response to injury. A common consequence of liver injury is parenchymal damage with an increase in apoptotic bodies, Kupffer cell activation, production of oxidative species, and ECM remodeling (65), which all function as triggers for cellular activation. Activation of stellate cells comprises two well-established phases: initiation (also called “preinflammatory stage”) and perpetuation, which can be followed by a

1477


potential third phase, resolution, if the liver injury resolves (58, 64, 69). During resolution of fibrosis, loss of activated stellate cells occurs through numerous pathways and there is now evidence that stellate cells can not only undergo apoptosis, but are also able to either become senescent or revert to a quiescent phenotype (67, 143, 294). The elucidation of molecular mechanisms underlying these events may accelerate the discovery of potential antifibrotic drug targets.

Initiation of stellate cell activation Stellate cell initiation promotes changes in gene expression and phenotype that render the cells susceptible to the changing environment and stimuli in the injured liver, thereby promoting the transition to the perpetuation phase. The earliest signals triggering the initiation of stellate cell activation result from paracrine stimulation by neighboring cell populations (endothelial cells, platelets, immune cells, and hepatocytes) and changes in its surrounding ECM (Fig. 3). As endothelial cells comprise the vascular lining of the liver’s sinusoids, they play a vital role in these early stages. They induce the activation of stellate cells by secreting fibronectin (125) and by activating latent transforming TGF-β (149), as well as through secretion of a range of mediators that modulate inflammation and participate in cellular crosstalk (275, 319). Fibronectin’s effects are largely promigratory and not fibrogenic, suggesting that stellate cell migration is an important first step in responding to injury (210). Platelets also contribute by secreting TGF-β, as well as EGF and plateletderived growth factor (PDGF), the most potent stellate cell mitogen identified (14, 24). Overall however, the resident hepatic macrophage population may be the main source of PDGF, as well as other paracrine mediators that drive stellate cell activation (106, 287, 308). There is an increasingly nuanced understanding of how inflammatory and immune cells regulate stellate cell responses and activation. In particular, T cells, dendritic cells (DC), and macrophage subsets all have well defined interactions with stellate cells [see (76, 131, 152, 179, 313) for reviews]. Among these, recent studies have characterized a specific macrophage subset in rodents, Ly-6Clo , that are vital for regression of hepatic fibrosis (231). On the other hand, different macrophages can drive stellate cell function including stimulation of matrix synthesis, cell proliferation, and retinoid release by secreting TGF-β, TNF-α, and MMP-9, and production of reactive oxygen species (ROS) and lipid peroxides (230). Moreover, ROS produced by hepatic macrophages can initiate downstream signals that include osteopontin, an ECM protein that can induce collagen (300) and perpetuate the activated stellate cell phenotype. Recent studies further implicate an inhibitory role of platelets in blocking stellate cell activation based on the following: (i) transgenic thrombocytopenic mice develop exacerbated liver fibrosis, with increased expression of type I collagen α1 and α2, during cholestasis (147); (ii) in vitro experiments reveal that, upon exposure to stellate cells, platelets became activated, released

1478


HGF, and then inhibited stellate cell expression of the type I collagen gene in a Met signal-dependent manner (147); and, (iii) activation of human stellate cells in culture is suppressed by human platelets or platelet-derived ATP via the adenosinecAMP signaling pathway (114). Hepatocytes are the main target for most forms of liver injury including viral infection, alcohol, and obesity, among others (198). Following injury, damaged hepatocytes become a major source of lipid peroxides and apoptotic bodies that initiate stellate activation through a process mediated by Fas and TRAIL (32). The contribution of hepatocytederived apoptotic bodies to stellate cell activation is independent of the inflammatory response, since in cultured stellate cells, addition of hepatocyte apoptotic fragments are directly fibrogenic (33), and can also activate Kupffer cells (31). Hepatocytes also express P450-2E1, an important enzyme involved in the metabolism of xenobiotics as ethanol, and a potent source of ROS (193) that can stimulate stellate cell fibrogenesis (193). Changes in the composition and stiffness of ECM also impact on stellate cell responses (84, 121, 315) suggesting a feed-forward loop where stellate cell mediated changes in ECM further drive stellate cell activation. Early changes in transcription factor activity in response to ECM, as well as soluble signals set the stage for a broad phenotypic transition of the cells, and a large number of nuclear factors have been implicated [see (173) for review]. Additionally, pathways of translational, transcriptional and posttranscriptional regulatory control (including epigenetic pathways and miRNAs) contribute to this process (18, 41, 99, 163, 172, 174, 218, 243, 297). While the initial presumption that transcription factors largely stimulate stellate cell activation, it appears equally true that other factors repress activation, and their activity is downregulated during cellular activation. Three key examples include Lhx2 (306), KLF6 (85), and Foxf1 (133), in which each contribute to preservation of a quiescent phenotype, such that their loss or downregulation derepresses the activation program.

Perpetuation of stellate cell activation—mechanisms and implications After the initial liver injury, stellate cells initiate activation followed by a process of perpetuation, leading to accumulation of ECM and culminating in the formation of scar tissue. Perpetuation of stellate cell activation is a tightly orchestrated process that includes a number of functional responses including proliferation, fibrogenesis, chemotaxis, contractility, matrix degradation, retinoid loss, and cytokine/chemokine expression (Fig. 4).

Proliferation PDGF is most potent stellate cell mitogen during liver injury. An increase in available PDGF and stellate cell responsiveness due to increased expression of PDGF receptor




(A) Normal liver Portal triad Bile duct

Hepatocytes

HSC Sinusoidal space of Disse Portal vein

Sinusoidal endothelial cells

KC

Terminal hepatic vein

Hepatic arteriole

(B) Fibrotic liver HSC activation and proliferation

Loss of endothelial fenestrations

Loss of hepatocyte microvilli

Distortion of veins

Increase in fibril-forming collagen in space of Disse Fibril-forming collagens (types I, III, and V) Basement membrane collagens (type IV and VI) Glycoconjugates (laminin, fibronectin, glycosaminoglycans, and tensacin)

Hernandez-Gea V, Friedman SL. 2011. Annu. Rev. Pathol. Mech. Dis. 6:425–56 Figure 3

Matrix and cellular alteration in hepatic fibrosis. Normal liver parenchyma contains epithelial cells (hepatocytes) and nonparenchymal cells: fenestrated sinusoidal endothelium, hepatic stellate cells (HSCs), and Kupffer cells (KCs). (A) Sinusoids are separated from hepatocytes by a low-density basement membrane-like matrix confined to the space of Disse, which ensures metabolic exchange. Upon injury, the stellate cells become activated and secrete large amounts of extracellular matrix (ECM), resulting in progressive thickening of the septa. (B) Deposition of ECM in the space of Disse leads to the loss of both endothelial fenestrations and hepatocyte microvilli, which results in both the impairment of normal bidirectional metabolic exchange between portal venous flow and hepatocytes and the development of portal hypertension.


1479



Perpetuation

Initiation

Injury

Proliferation Oxidative stress Apoptotic bodies LPS Paracrine stimuli

PDGF VEGF FGF

Contractility ET–1

NO

Fibrogenesis

TGF-β1/ CTGF MMP-2&9;

MT-1-MMP

TIMP-1,2

Reversion

Altered matrix degradation

PDGF Chemokines

Resolution Chemokines TLR ligands

Adenosine

HSC chemotaxis

TIMP-1,2 TRAIL Fas

Apoptosis

T cells B cells NK cells NK-T cells

Inflammatory signaling

Figure 4

Pathways of hepatic stellate cell activation and loss during liver injury and resolution. Features of stellate cell activation can be distinguished between those that stimulate initiation and those that contribute to perpetuation. Initiation is provoked by soluble stimuli that include oxidant stress signals (reactive oxygen intermediates), apoptotic bodies, lipopolysaccharide (LPS), and paracrine stimuli from neighboring cell types including hepatic macrophages (Kupffer cells), sinusoidal endothelium, and hepatocytes. Perpetuation follows, characterized by a number of specific phenotypic changes including proliferation, contractility, fibrogenesis, altered matrix degradation, chemotaxis, and inflammatory signaling. During resolution of hepatic fibrosis, there is both programmed cell death (apoptosis) to clear fibrogenic cells, as well as reversion to a more quiescient phenotype. FGF, fibroblast growth factor; ET-1, endothelin-1; NK, natural killer; NO, nitric oxide; MT, membrane type. Reprinted, with permission, from (66).

results in rapid proliferation and an overall increase in the absolute number of stellate cells with a profibrogeneic phenotype (24, 220, 317). Tumor necrosis factor (TNF) alpha signaling also contributes to PDGF-mediated stellate cell proliferation primarily through the TNF receptor 1 (291). Stellate cells are also responsive to a wide array of factors including, VEGF (327), thrombin, EGF, keratinocytre growth factor (282), and bFGF (328). Among those, VEGF is one of the major cytokines secreted by activated HSCs, which drives both angiogenesis and fibrogenesis, as described above (44, 135, 159). VEGF production is dependent on the overexpression of COX-2 protein via phospho-p42/44 MAP kinase activation (329). Overall, HSCs contribute to both wound healing and tumor growth, a conclusion underscored by several studies implicating this cell type in the development and growth of both primary and metastatic tumors (47, 134, 209). Tissue inhibitors of matrix metalloproteinases (TIMPs) are profibrogenic by inhibiting matrix degradation, and promoting stellate cell survival. Increased TIMP-1 expression

1480

by stellate cells also drives stellate cell proliferation in an AKT-dependent manner (61). This mechanism might contribute to the antiproliferative effects of activated Vitamin D, 1,25(OH)(2)D(3) (1). Vitamin D receptor is expressed by quiescent stellate cells (79), and its expression is downregulated with activation. Consequently, treatment of stellate cells with 1,25(OH)(2)D(3) dampens proliferation via cyclin D1 suppression and decreases expression of type I collagen and TIMP-1 while simultaneously increasing MMP-9 expression. Patients with liver disease exhibit vitamin D deficiency, and appropriate supplementation might prove to be a useful therapeutic intervention (225). MicroRNAs are small 19-24 noncoding RNA sequences that can regulate posttransciptional gene expression by sequence-specific binding to the 3 -UTR of mRNAs to promote their degradation. The role of miRNAs in fibrosis progression is being clarified, as miRNA levels change in livers of patients with fibrotic disease and in stellate cells during activation (93, 99, 112, 196, 197, 205, 243, 307). In stellate



cells, miRNAs control proliferation and fibrogenesis by regulating protein expression of proproliferative and profibrogenic signaling pathways. In particular, miR-27a and -27b and miR29b are upregulated in activated stellate cells, whereas their suppression leads to decreased proliferation and an increase in lipid droplets indicative of the quiescent phenotype (205, 244). miR-27a -27b directly target the 3 -UTR of retinoid X receptor α (RXRα) to inhibit its expression. RXRα can decrease DNA synthesis, leading to growth arrest in stellate cells (100). Furthermore, RXRα regulates adipogenesis by activation of peroxisome proliferator-activated receptor γ (PPARγ) which is a master regulator of stellate cell activation (297). RXRα expression is decreased in stellate cell activation and its expression in activated stellate cells increases with inactivation of miR-27a -27b, indicating a direct interaction between RXRα and miR27a -27b (128). In contrast, miR195 is downregulated during stellate cell proliferation and its expression is induced upon treatment with IFN-β, which exhibits antifibrotic effects independent of its antiviral activity. Treatment with IFN-β downregulates cyclin E1 and upregulates p21 in a miR-195 specific manner thereby promoting cell cycle arrest and decreased stellate cell proliferation (265).

Fibrogenesis Production of ECM, in particular collagen type I, is a hallmark of activated stellate cells. Production of collagen type I by stellate cells is regulated both transcriptionally and posttransciptionally (2, 37, 73, 117-119, 167, 175, 214, 238, 279,280, 295). TGF-β1 is major driver of this process through autocrine and paracrine stimulation of ECM production (see above). The other well-characterized fibrogenic cytokine towards stellate cells is connective tissue growth factor (CTGF/CCN2). Levels of CTGF are increased in liver injury and the cytokine promotes a range of profibrotic activities toward stellate cells, mediated by a G-coupled protein receptor (78, 92, 111). CTGF represents a very appealing target for antifibrotic therapy, since unlike antagonism of TGF-β1, CTGF inhibition should have no impact on hepatocyte growth or confer a risk of carcinogenesis. There is a growing list of other factors that contribute to fibrogenesis, including signaling molecules, chemokines, and cellular stressors (250). For example, osteopontin, an ECM cytokine expressed by stellate cells, activates collagen I expression via integrin α(V)β(3) engagement and activation of the PI3K/pAkT/NFκB signaling pathways (300). Furthermore, the recent identification of receptors to profibrogenic chemokines on stellate cells including CXCR4 (108), CCR1, CCR5 (262), CXCR2 (281), and CCR2 (263), add to the repertoire of signals promoting stellate cell activation. The targetability of chemokine receptors with small molecule inhibitors makes them ideal candidates for antifibrotic therapies. Recently, blockade of IL-17A has been proposed as a potential strategy for cirrhosis treatment due to its induction (together with its receptor) in response to liver injury. IL-17A may promote fibrosis by activating inflammatory and



liver resident cells and inducing collagen type I production in HSCs through engagement of the signal transducer and activator of transcription 3 signaling pathway (181). As discussed above, (see “Proliferation”) miRNAs play a significant role in stellate cell biology and can modulate collagen synthesis. MiR-29b binds directly to the 3 -UTR of collagen-IαI and -IV, thereby inhibiting its translation. miR29b expression is repressed by TGF-β, and its overexpression inhibits TGF-β induced collagen expression via a SMADindependent mechanism, while HGF, which exhibits antifibrotic effects in stellate cells, induces miR-29b expression (244, 266). MiR-21, whose expression is enhanced during fibrotic disease, is also controlled by TGF-β signaling. TGFβ functions via 2 distinct mechanisms to increase miR-21 production. Smad3 induces miR-21 transcription, while both Smad2 and Smad3 enhance miR-21 maturation.

Chemotaxis Stellate cell chemotaxis is an important event in the generation of fibrotic septae by allowing activated cells to align within regions of injury. Stellate cells migrate primarily towards chemoattractant cytokines (chemokines), and they express a range of chemokine receptors and their cognate chemokine ligands. Notably, stellate cells migrate towards PDGF (113, 142), VEGF, Ang-1 (199), TGF-β1 , EGF (323), b-FGF (59), CCL2 (176), and CXCR4 (254), and CXCR3 specific ligands (23). The mechanism of chemotaxis includes a cytoskeletal remodeling with cell spreading at the tip, movement of the cell body towards the stimulus, and retraction of trailing protrusions (180). Oxidant signaling contributes to these responses. Specifically, numerous chemoattractant signals (PDGF, VEGF, and CCL2) increase NADPH oxidasedependent intracellular ROS and activation of the ERK1/2 and JNK1/2 pathways (50, 212). Furthermore, generation of intracellular superoxide anion or H2 O2 by treatment with menadione promotes cell migration even in the absence of specific chemoattractants (198). Hypoxia is another broad activator of stellate cell migration, which functions via two distinct mechanisms. After induction of hypoxic conditions, mitochondrial-generated ROS activate the ERK1/2 and JNK1/2 pathways, driving migration. Sustained hypoxia leads to a HIF-1α-dependent increased production and secretion of VEGF by stellate cells, promoting their mobility (200). Since stellate cell mobilization is also required for tissue wound healing, it has been reported that the space of Disse microenvironment, per se, is another key factor in regulating the migratory behavior of stellate cells. ECM components including MMP-2 and type I collagen are able to induce stellate cell migration (208, 323). Cellular fibronectin containing an alternatively spliced domain A (EIIA) is upregulated during liver injury. Signaling specifically by the EIIA fibronectin variant though integrin receptor α(9)β(1) on stellate cells promotes formation of lamellipodia and cellular motility, further implicating ECM signaling in stellate cell

1481


biology (210). The hyaluronic acid receptor (CD44) is also increased in liver injury and repair, promoting both activation and migration of stellate cells (140). Interestingly, a specific splice variant (CD44v6) is responsible for up to 50% of this migration, confirming the idea that activated stellate cells may depend, to some degree, on CD44v6 and hyaluronic acid for migration.

Contraction Stellate cell contraction is thought to be a primary determinant of portal hypertension in patients with end-stage liver disease. The factors leading to portal hypertension include increased blood flow, increased intrahepatic resistance, and disrupted liver architecture. During injury, the hepatic sinusoids undergo both morphological and functional changes mediated by HSCs. Dramatic remodeling occurs, characterized by deposition of collagen matrix, loss of fenestrations and increased density of contractile HSCs (293). Additionally, there is an imbalance of vasoactive forces characterized by deficient nitric oxide production and an increase in vasoconstrictive substances including ET-1, angiotensinogen II, eicosanoids, atrial natriuretic peptide, somatostatin, and carbon monoxide (237, 239, 240, 292). Together, these factors lead to an increase in sinusoidal resistance and portal hypertenstion. The final step in the induction of cellular contraction is phosphorylation of MLC-2, resulting from a calciumdependent and a calcium-sensitive pathway. In the calciumdependent pathway, typically in skeletal and cardiac muscle, release of Ca2+ from the endoplasmic/sarcoplasmic reticulum leads to activation of myosin light-chain kinase (MLCK) by calmodulin and subsequent phosphorylation of MLC. Alternatively, in the Ca2+ -sensitive pathway, the dominant pathway in stellate cell mediated contraction and smooth muscle cells, Rho-kinase inactivates the myosin binding subunit by phosphorylation, thereby preventing it from dephosphorylating MLC and inhibiting contraction. The net effect of Rho-kinase activation is increased phosphorylated-MLC and contraction (168). Adenosine plays an important role in stellate cell differentiation, proliferation, and type I collagen production. Despite these profibrotic effects, however, adenosine inhibits stellate cell contraction (as well as chemotaxis) via loss of actin stress fibers. Engagement of the A2a receptor by adenosine promotes PKA activity and Rho A inhibition (276), which establishes a rationale for Rho antagonism as a strategy for treatment of portal hypertension (132). Adenosine therefore promotes both injurious and protective effects on stellate cells and understanding these different functions will be important in the development of antifibrotic agents. Interestingly, the antifibrotic activities of caffeine, now validated in several large cohorts (43, 187, 211), may reflect the compound’s effect in reducing adenosine signaling in stellate cells (38).

1482


Retinoid loss Recently, there has been a renewed focus on the role of retinoid loss in stellate cell activation and collagen production. Stellate cell activation is characterized by the loss of perinuclear retinoid droplets (65, 70); however, their function in activation and fibrogenesis is only now being revealed. Stellate cells are the largest reserve of retinoids in the body (∼60%) and conversion of retinol into retinyl ester is a hallmark of stellate cell activation. The abundance of vitamin A in stellate cells is heterogenous depending on the intralobular position of the cell (80) and may be indicative of alternate activation states. Quiescent stellate cells that are isolated based on their collagen 1 expression display an increase in CYP251 retinoid catabolizing cytochrome, a decrease in retinyl esters, and a more activated phenotype compared to cells isolated based on their buoyancy in gradient centrifugation (45). LRAT, which catalyzes the esterification of retinol into retinyl ester, is the sole acyltransferase found in the liver. With stellate cell activation, LRAT expression is lost. Additionally, treatment with IL-1 promotes decreased LRAT expression (139). Despite its apparent role in stellate cell activation, mice deficient in LRAT neither display spontaneous fibrogenesis, nor do they exhibit increased fibrogenesis in liver injury models, indicating that perhaps retinoid loss is a marker of activation, but is not crucial for stellate cell activation (144). However, treatment with retinoic acid can decrease stellate cell activation as reflected in reduced collagen I, MMP-9, and α-SMA by inhibiting expression of TGF-β (98). In contrast to the lack of dependence on LRAT for fibrogenesis by stellate cells, mice deficient in LRAT are protected from chemical hepatocarcinogenesis. An increase in active retinoids due to their lack of conversion to retinol storage form leads to an overall antiproliferative effect, increased p21 levels, and inhibition of tumor progression (144, 272). An additional link between retinoid metabolism to stellate cell activation has emerged through the recognition that this process requires cellular autophagy (104). Specifically, hydrolysis of retinyl esters liberates fatty acids that are metabolized by β-oxidation, generating the substrates that are essential for fueling the energy-intensive pathways of cellular activation. The free retinol can be detected in the extracellular milieu under these conditions (72), but pathways enabling its cellular egress are not known. Similar to stellate cells, autophagy contributes to the intracellular catabolism of lipids in hepatocytes, fibroblasts (274), and neurons (150, 151). Moreover, inhibition of autophagy in hepatocytes leads to reduced rates of β-oxidation and marked lipid accumulation in cytosolic lipid droplets (274).

Matrix degradation Fibrosis is a dynamic process of matrix production and degradation. Fibrotic progression is characterized by the replacement of normal basement membrane, collagen type IV, with scar forming collagen type I. Early matrix degradation is an important step in fibrosis and may be essential for stellate cell



migration to sites of injury. Stellate cells are the prominent source for MMP-2, MMP-9, and MMP-13 (7, 96, 183, 302), which function as either interstitial collagenases (MMP-13) or gelatinases (MMP-2 and -9). Despite the dependence of fibrosis progression on MMPs, these molecules paradoxically exhibit antifibrotic properties, and their expression is suppressed with fibrotic progression. MMP-2 inhibits collagen type I production by stellate cells and mice lacking MMP-2 therefore exhibit increased fibrosis (229). Furthermore, MMP-2 is able to regulate stellate cell apoptosis by cleavage of the extracellular domain of Ncadherin, further supporting its antifibrotic role (97). Some of the paradoxical activity of MMPs in vivo may be explained by their simultaneous activation of macrophages (87) a feature often overlooked in interpreting their contribution to fibrosis. Regulation of MMP expression occurs through several mechanisms. TIMPs, which are prominently expressed by stellate cells, bind MMPs, inactivating them. Additionally, with progressive fibrosis, MMP-9 and MMP-13 are repressed at the chromatin level and access by the transcription factors NF-κB and AP-1 is inhibited. Impaired histone acetylation is associated with permanently silenced genes, and activated stellate cells have a global increase in histone deacetlyase-4 leading to decreases in acetylation in the MMP-9 and -13 promoter regions and gene repression (226). While the source of enzymes that degrade ECM in liver has been elusive, findings increasingly point to subsets of macrophages that have fibrolytic potential. Indeed, a recent study has characterized a specific subtype of macrophages that express the surface marker Ly-6C as a recruited cellular subset that secretes a range of proteases that have the capacity to degrade ECM (230, 231).

Immunoregulation In recent years stellate cells have emerged as a prominent determinant of hepatic immunoregulation during injury. They express of a battery of chemokines including CCL2 (277), CCL5 (260), CXCL2, CCL21 (22), CXCL8, CXCL9, CXCL10, CXCL12, and CX3CL3 which have been shown to recruit neutrophils, macrophages/monocytes, NK/NKT cells, DCs, and T-cells, thereby establishing their role in immune cell infiltration. Many of these chemokines also function independent of immune cells and modulate cellular differentiation, survival, proliferation, and apoptosis. Stellate cells can control immune cell function through three interrelated mechanisms (283, 288): (i) cell surface expression of chemokines and/or delivery of chemokines to endothelial cells promotes lymphocyte adhesion and subsequent migration and activated stellate cells specifically promote ICAM-1 and VCAM-1 dependent adhesion and migration (107); (ii) increased expression of stellate cell derived chemokines establishes a chemoattractant gradient between the peripheral blood and the liver, thus driving immune cell migration into the liver; and (iii) stellate cell interaction with immune cells has a direct role in promoting/inhibiting



their maturation within the liver. For example, stellate cells can inhibit the priming of na¨ıve T cells in a cell contactdependent CD54 mechanism. Incubation of stellate cells with DCs and OT-1 T-cells inhibits T cell proliferation and reduces CD25 and CD44 activation markers (255). The ability of stellate cells to inhibit T cell proliferation is dependent on their activation state, since quiescent stellate cells do not possess this inhibitory function. Proteomics analysis reinforces this immune suppressive role (127). In the transplantation setting, endotoxin-stimulated stellate cells are important immune regulators by inducing selective expansion of tolerance-promoting Tregs to reduce inflammation and alloimmunity (46). Pattern recognition receptors, particularly TLR4, contribute significantly to stellate cell responses. Stimulation of stellate cells with the TLR4 ligands enhances TGF-β signaling and production of inflammatory and chemotactic cytokines (CCL2, IL-6), leading to a more profibrogenic phenotype (94, 185, 213, 264). Additionally, stellate cell TLR3 activation confers antifibrotic activities by stimulating interleukin-10 production (30). During late stage fibrosis when portal hypertension is present there is an increased bacterial load delivery from the gut to the liver due to increased intestinal permeability, leading to an increase in LPS and TLR4 activation by stellate cells. Treatment of mice with the intestinal decontaminant rifaximin, decreases portal pressure, fibrosis, and angiogenesis in a TLR4-specific manner (331). Specifically, rifaximin treatment inhibits production of stellate cell derived fibronectin. LSECs are the key mediators during angiogenesis. Stellate cell derived fibronectin is able to induce LSEC migration, and tubulogenesis, thereby contributing to the pathogenic effects of LPS (331). The stellate cell’s capabilities now extend to their role in DC development (109). DCs exposed to stellate cells or their supernatants express low CD11c, CD86, and major histocompatibility complex class II, which elicits an inferior allostimulatory function compared with conventional DC. From the complementary point of view, the inflammatory response can also modulate stellate cell activation and fibrogenesis. Therefore, a positive feedback loop exists in which inflammatory and fibrogenic cells stimulate each other in amplifying fibrosis. Thus, cell types regulating progression and resolution of fibrosis include macrophages (a pivotal cell with the potential for both a pro- and antifibrotic capacity by secreting/regulating TGF-β1, PDGF, MMPs, and TIMPs) (230, 231), natural killer cells (antifibrotic activity by inhibiting and/or killing activated stellate cells) (77, 189), T-cells (responsible for initiating/maintaining the adaptive immune response and may induce liver injury upon CCL3/MIP1α recruitment of CCR1 expressing CD4+ T cells) (286), B-cells (fibrosis promotion in an antibody- and T cell-independent manner) (201), DCs (involved in both proinflammatory and immunogenic responses) (42, 131), as well as endothelial cells (antigen-presenting cells for CD4+ and CD8+ T populations with antigen clearance activity) (146).

1483


Stellate Cells and Resolution of Liver Fibrosis Despite the historical conception of liver fibrosis as a passive and irreversible process due to hepatocyte collapse (223,224), the idea of fibrosis regression was proposed in the 1970s (217, 224) and demonstrated in the 1990s (17, 95). At a certain stage of disease, fibrosis may become irreversible, likely due to significant collagen cross-linking and development of an insoluble and relatively hypocellular matrix, which may coincide with the appearance of clinical cirrhosis manifestations (124). Since stellate cells are an important contributor to ECM remodeling in liver fibrosis, three possibilities may account for regression of fibrosis, either their apoptosis, senescence, or reversion to their quiescent stage. Apoptosis of stellate cells during liver fibrosis recovery contributes to a decrement in the number of activated stellate cells (120, 123, 251). This situation coincides with a decrease in TIMP-1 expression (122), which protects the cells from apoptosis and inhibits the MMPs functions (192), favoring the partial degradation of ECM (55). Of interest, the NF-κB transcription factor plays a role in the stellate cell’s protection from apoptosis during liver fibrosis resolution, since its inhibition can accelerate recovery from liver fibrosis (202). Moreover, apoptosis can be spontaneously initiated in activated stellate cells via CD95L (Fas ligand) Bcl-2 and p53 (90). Finally, other cell populations can contribute to stellate cell apoptosis. Hepatocytes can secrete NGF, which is apoptotic towards stellate cells (228) (203). Natural killer cells can also provoke stellate cell apoptosis through TRAIL and NKG2D, a stellate cell membrane receptor that is specifically expressed by activated stellate cells, rendering them susceptible to NK action (228). Kupffer cells can also induce stellate cell apoptosis by a caspase-9-mediated mechanism (60). Recent studies have now established that HSCs can undero p53-mediated senescence in culture (258) and in vivo (155). Initial studies in cultured HSCs suggested that as the cells reached their replicative limit, they adopt a more inflammatory and less fibrogenic phenotype (258). Based on these observations, elegant in vivo approaches confirmed the development of senescence in HSCs in vivo (155, 169). Moreover, this response is p53-dependent, because animals in which p53 is specifically deleted in stellate cells have more fibrosis after toxic liver injury (169). Remarkably, this senescence program regulates the polarization of macrophages towards a phenotype that is protective against experimental carcinogenesis, unearthing a noncell autonomous pathway of tumor suppression by p53 through its impact on the surrounding stromal biology (169). Until recently, reversion from activated to quiescent HSC was only demonstrated in cultured cells. However, elegant studies have now used genetic methods to document reversion of activated cells to a quiescent phenotype in vivo (67, 143, 294). Reverted cells, however, have a heightened capacity to reactivate, compared to those that have never activated. Quantitatively, reversion of activated stellate cells to quiescence is

1484


likely to be a significant pathway in fibrosis regression that involves approximately 50% of the stellate cell population, and may in part be regulated by changes in PPARγ activity.

Conclusion HSCs are among the most fascinating and versatile of cell types in mammalian biology. While their contributions to hepatic injury and fibrosis has been well established for at least two decades, clarifying this role further continues to benefit from remarkable discoveries uncovering the signals that control cell plasticity, survival, and intracellular signaling. A growing range of genes, epigenetic changes, mediators, and secretory proteins of stellate cells has been uncovered using complementary methods of gene profiling and proteomics, which has led to a more integrated understanding of the cell’s regulation. Concurrently, genetic models including new methods to delete genes specifically in stellate cells further clarify their roles in normal and injured liver. In aggregate, these discoveries have enhanced the perceived importance of HSCs not only to the fibrotic response in liver, but also to normal liver homeostasis that includes blood flow regulation, regeneration, and stem cell responses. A key issue in studying HSCs is the choice of model and species. Studies over the past 25 years have utilized both rodent and human cells, as well as several methods to immortalize cultured cells, including SV40 T antigen gene transfection, spontaneous immortalization in low serum, or ectopic telomerase gene expression. Whereas human cells have the advantage of direct human disease relevance, they lack the capacity for in vivo gene deletion using knockout technology, which is now widely used for mouse models and isolated stellate cells. This raises an important issue, as yet unanswered, whether the genotypes and phenotypes of rodent and human stellate cells in normal and diseased liver are sufficiently similar to allow investigators to rely on rodent studies to understand and predict human disease targets. This concern has become more pertinent based on a recent study indicating that rodent models of inflammation bear little resemblance to human disease (268). While one approach might be to “humanize” mice so that they contain human stellate cells, methods to achieve this goal would be tedious and would likely require use of immunodeficient animals. An alternative approach would be to perform comprehensive comparative analysis of mRNAs, microRNAs, proteins, and/or epigenetics between human and rodent cells from normal and injured liver to determine how faithful rodent cells are to their human counterparts. Based on studies to date, there is no evidence that fibrogenic responses by stellate cells between species are widely divergent, but the question has not yet been rigorously addressed. Despite progress in the field, several other key issues also remain unresolved. The contribution of stellate cells to regeneration has long been assumed, but recent reports suggest that HSC depletion has no effect on regeneration (54, 144). Furthermore, use of transgenic animals has begun to trace



stellate cell fate during fibrosis regression (143, 230, 294) but underlying mechanisms are still incomplete. Similarly, it is increasingly clear that stellate cells and the fibrotic stroma they generate in injured liver accelerate the risk of hepatocellular carcinoma, but methods to clarify pathways linking fibrosis to cancer are only now emerging. Also unclear is the full range of paracrine interactions in stellate cell crosstalk with sinusoidal endothelial cells, biliary epithelial cells, and immune cell subsets. Finally, although activation of HSCs into contractile myofibroblasts is well recognized as the central event in hepatic fibrosis, this insight has not yet translated into an effective antifibrotic therapy for patients with chronic fibrosing liver diseases. Future studies in animal models will further clarify these vital roles, while their translation into effective antifibrotic drugs for fibrotic liver disease is sure to emerge in the coming years.

Acknowledgements Work in the authors’ laboratory was supported by NIH Grants DK56621, AA020709 (to S.L.F.), NIDDK F30-DK090986, and T32-GM007280 (to Y.S.), and funds from the Alfonso Martin Escudero Fundation (to J.E.P.).

References 1.

2. 3.

4. 5. 6. 7.

8. 9.

10.

11.

12. 13.

Abramovitch S, Dahan-Bachar L, Sharvit E, Weisman Y, Ben Tov A, Brazowski E, Reif S. Vitamin D inhibits proliferation and profibrotic marker expression in hepatic stellate cells and decreases thioacetamideinduced liver fibrosis in rats. Gut 60: 1728-1737, 2011. Anania FA, Womack L, Jiang M, Saxena NK. Aldehydes potentiate alpha(2)(I) collagen gene activity by JNK in hepatic stellate cells. Free Radic Biol Med 30: 846-857, 2001. Anthony PP, Ishak KG, Nayak NC, Poulsen HE, Scheuer PJ, Sobin LH. The morphology of cirrhosis. Recommendations on definition, nomenclature, and classification by a working group sponsored by the World Health Organization. J Clin Pathol 31: 395-414, 1978. Apte MV, Pirola RC, Wilson JS. Pancreatic stellate cells: A starring role in normal and diseased pancreas. Front Physiol 3: 344, 2012. Apte MV, Wilson JS. Dangerous liaisons: Pancreatic stellate cells and pancreatic cancer cells. J Gastroenterol Hepatol 27(Suppl 2): 69-74, 2012. Arthur MJ, Friedman SL, Roll FJ, Bissell DM. Lipocytes from normal rat liver release a neutral metalloproteinase that degrades basement membrane (type IV) collagen. J Clin Invest 84: 1076-1085, 1989. Arthur MJ, Stanley A, Iredale JP, Rafferty JA, Hembry RM, Friedman SL. Secretion of 72 kDa type IV collagenase/gelatinase by cultured human lipocytes. Analysis of gene expression, protein synthesis and proteinase activity. Biochem J 287: 701-707, 1992. Asahina K. Hepatic stellate cell progenitor cells. J Gastroenterol Hepatol 27(Suppl 2): 80-84, 2012. Asahina K, Sato H, Yamasaki C, Kataoka M, Shiokawa M, Katayama S, Tateno C, Yoshizato K. Pleiotrophin/heparin-binding growthassociated molecule as a mitogen of rat hepatocytes and its role in regeneration and development of liver. Am J Pathol 160: 2191-2205, 2002. Asahina K, Tsai SY, Li P, Ishii M, Maxson RE, Jr., Sucov HM, Tsukamoto H. Mesenchymal origin of hepatic stellate cells, submesothelial cells, and perivascular mesenchymal cells during mouse liver development. Hepatology 49: 998-1011, 2009. Asahina K, Zhou B, Pu WT, Tsukamoto H. Septum transversumderived mesothelium gives rise to hepatic stellate cells and perivascular mesenchymal cells in developing mouse liver. Hepatology 53: 983-995, 2011. Baba S, Fujii H, Hirose T, Yasuchika K, Azuma H, Hoppo T, Naito M, Machimoto T, Ikai I. Commitment of bone marrow cells to hepatic stellate cells in mouse. J Hepatol 40: 255-260, 2004. Bach Kristensen D, Kawada N, Imamura K, Miyamoto Y, Tateno C, Seki S, Kuroki T, Yoshizato K. Proteome analysis of rat hepatic stellate cells. Hepatology 32: 268-277, 2000.



14. Bachem MG, Melchior R, Gressner AM. The role of thrombocytes in liver fibrogenesis: Effects of platelet lysate and thrombocyte-derived growth factors on the mitogenic activity and glycosaminoglycan synthesis of cultured rat liver fat storing cells. J Clin Chem Clin Biochem 27: 555-565, 1989. 15. Baghy K, Iozzo RV, Kovalszky I. Decorin-TGFbeta axis in hepatic fibrosis and cirrhosis. J Histochem Cytochem 60: 262-268, 2012. 16. Ballardini G, Groff P, Badiali dGL, Schuppan D, Bianchi FB. Ito cell heterogeneity: Desmin-negative Ito cells in normal rat liver. Hepatology 19: 440-446, 1994. 17. Bataller R, Brenner DA. Liver fibrosis. J Clin Invest 115: 209-218, 2005. 18. Bian EB, Huang C, Ma TT, Tao H, Zhang H, Cheng C, Lv XW, Li J. DNMT1-mediated PTEN hypermethylation confers hepatic stellate cell activation and liver fibrogenesis in rats. Toxicol Appl Pharmacol 264: 13-22, 2012. 19. Bioulac-Sage P, Lafon ME, Saric J, Balabaud C. Nerves and perisinusoidal cells in human liver. J Hepatol 10: 105-112, 1990. 20. Bissell DM, Arenson DM, Maher JJ, Roll FJ. Support of cultured hepatocytes by a laminin-rich gel. Evidence for a functionally significant subendothelial matrix in normal rat liver. J Clin Invest 79: 801-812, 1987. 21. Bissell DM, Wang SS, Jarnagin WR, Roll FJ. Cell-specific expression of transforming growth factor-beta in rat liver. Evidence for autocrine regulation of hepatocyte proliferation. J Clin Invest 96: 447-455, 1995. 22. Bonacchi A, Petrai I, Defranco RM, Lazzeri E, Annunziato F, Efsen E, Cosmi L, Romagnani P, Milani S, Failli P, Batignani G, Liotta F, Laffi G, Pinzani M, Gentilini P, Marra F. The chemokine CCL21 modulates lymphocyte recruitment and fibrosis in chronic hepatitis C. Gastroenterology 125: 1060-1076, 2003. 23. Bonacchi A, Romagnani P, Romanelli RG, Efsen E, Annunziato F, Lasagni L, Francalanci M, Serio M, Laffi G, Pinzani M, Gentilini P, Marra F. Signal transduction by the chemokine receptor CXCR3: Activation of Ras/ERK, Src, and phosphatidylinositol 3-kinase/Akt controls cell migration and proliferation in human vascular pericytes. J Biol Chem 276: 9945-9954, 2001. 24. Borkham-Kamphorst E, van Roeyen CR, Ostendorf T, Floege J, Gressner AM, Weiskirchen R. Pro-fibrogenic potential of PDGF-D in liver fibrosis. J Hepatol 46: 1064-1074, 2007. 25. Botella LM, Sanchez-Elsner T, Sanz-Rodriguez F, Kojima S, Shimada J, Guerrero-Esteo M, Cooreman MP, Ratziu V, Langa C, Vary CP, Ramirez JR, Friedman S, Bernabeu C. Transcriptional activation of endoglin and transforming growth factor-beta signaling components by cooperative interaction between Sp1 and KLF6: Their potential role in the response to vascular injury. Blood 100: 4001-4010, 2002. 26. Bourd-Boittin K, Bonnier D, Leyme A, Mari B, Tuffery P, Samson M, Ezan F, Baffet G, Theret N. Protease profiling of liver fibrosis reveals the ADAM metallopeptidase with thrombospondin type 1 motif, 1 as a central activator of transforming growth factor beta. Hepatology 54: 2173-2184, 2011. 27. Brasaemle DL, Wolins NE. Packaging of fat: An evolving model of lipid droplet assembly and expansion. J Biol Chem 287: 2273-2279, 2012. 28. Buers I, Hofnagel O, Ruebel A, Severs NJ, Robenek H. Lipid droplet associated proteins: An emerging role in atherogenesis. Histol Histopathol 26: 631-642, 2011. 29. Burt AD, Griffiths MR, Schuppan D, Voss B, MacSween RN. Ultrastructural localization of extracellular matrix proteins in liver biopsies using ultracryomicrotomy and immuno-gold labelling. Histopathology 16: 53-58, 1990. 30. Byun JS, Suh YG, Yi HS, Lee YS, Jeong WI. Activation of toll-like receptor 3 attenuates alcoholic liver injury by stimulating Kupffer cells and stellate cells to produce interleukin-10 in mice. J Hepatol 58: 342349, 2013. 31. Canbay A, Feldstein AE, Higuchi H, Werneburg N, Grambihler A, Bronk SF, Gores GJ. Kupffer cell engulfment of apoptotic bodies stimulates death ligand and cytokine expression. Hepatology 38: 1188-1198, 2003. 32. Canbay A, Higuchi H, Bronk SF, Taniai M, Sebo TJ, Gores GJ. Fas enhances fibrogenesis in the bile duct ligated mouse: A link between apoptosis and fibrosis. Gastroenterology 123: 1323-1330, 2002. 33. Canbay A, Taimr P, Torok N, Higuchi H, Friedman S, Gores GJ. Apoptotic body engulfment by a human stellate cell line is profibrogenic. Lab Invest 83: 655-663, 2003. 34. Casini A, Pellegrini G, Ceni E, Salzano R, Parola M, Robino G, Milani S, Dianzani MU, Surrenti C. Human hepatic stellate cells express class I alcohol dehydrogenase and aldehyde dehydrogenase but not cytochrome P4502E1. J Hepatol 28: 40-45, 1998. 35. Cassiman D, Barlow A, Vander Borght S, Libbrecht L, Pachnis V. Hepatic stellate cells do not derive from the neural crest. J Hepatol 44: 1098-1104, 2006.

1485


36. Cassiman D, Denef C, Desmet VJ, Roskams T. Human and rat hepatic stellate cells express neurotrophins and neurotrophin receptors. Hepatology 33: 148-158, 2001. 37. Challa AA, Vukmirovic M, Blackmon J, Stefanovic B. Withaferin-A reduces type I collagen expression in vitro and inhibits development of myocardial fibrosis in vivo. PLoS One 7: e42989, 2012. 38. Chan ES, Montesinos MC, Fernandez P, Desai A, Delano DL, Yee H, Reiss AB, Pillinger MH, Chen JF, Schwarzschild MA, Friedman SL, Cronstein BN. Adenosine A(2A) receptors play a role in the pathogenesis of hepatic cirrhosis. Br J Pharmacol 148: 1144-1155, 2006. 39. Chan KM, Fu YH, Wu TJ, Hsu PY, Lee WC. Hepatic stellate cells promote the differentiation of embryonic stem cell-derived definitive endodermal cells into hepatic progenitor cells. Hepatol Res 43: 648657, 2012. 40. Chen L, Zhang W, Zhou QD, Yang HQ, Liang HF, Zhang BX, Long X, Chen XP. HSCs play a distinct role in different phases of oval cellmediated liver regeneration. Cell Biochem Funct 30: 588-596, 2012. 41. Chen SL, Zheng MH, Yang T, Song M, Chen YP. Disparate profiles of dys-regulated miRNAs in activated hepatic stellate cells. Hepatology 57: 1285-1286, 2013. 42. Connolly MK, Bedrosian AS, Mallen-St Clair J, Mitchell AP, Ibrahim J, Stroud A, Pachter HL, Bar-Sagi D, Frey AB, Miller G. In liver fibrosis, dendritic cells govern hepatic inflammation in mice via TNF-alpha. J Clin Invest 119: 3213-3225, 2009. 43. Costentin CE, Roudot-Thoraval F, Zafrani ES, Medkour F, Pawlotsky JM, Mallat A, Hezode C. Association of caffeine intake and histological features of chronic hepatitis C. J Hepatol 54: 1123-1129, 2011. 44. Coulouarn C, Corlu A, Glaise D, Guenon I, Thorgeirsson SS, Clement B. Hepatocyte-stellate cell cross-talk in the liver engenders a permissive inflammatory microenvironment that drives progression in hepatocellular carcinoma. Cancer Res 72: 2533-2542, 2012. 45. D’Ambrosio DN, Walewski JL, Clugston RD, Berk PD, Rippe RA, Blaner WS. Distinct populations of hepatic stellate cells in the mouse liver have different capacities for retinoid and lipid storage. PLoS One 6: e24993, 2011. 46. Dangi A, Sumpter TL, Kimura S, Stolz DB, Murase N, Raimondi G, Vodovotz Y, Huang C, Thomson AW, Gandhi CR. Selective expansion of allogeneic regulatory T cells by hepatic stellate cells: Role of endotoxin and implications for allograft tolerance. J Immunol 188: 3667-3677, 2012. 47. Dapito DH, Mencin A, Gwak GY, Pradere JP, Jang MK, Mederacke I, Caviglia JM, Khiabanian H, Adeyemi A, Bataller R, Lefkowitch JH, Bower M, Friedman R, Sartor RB, Rabadan R, Schwabe RF. Promotion of Hepatocellular Carcinoma by the Intestinal Microbiota and TLR4. Cancer Cell 21: 504-516, 2012. 48. De Bleser PJ, Niki T, Rogiers V, Geerts A. Transforming growth factorbeta gene expression in normal and fibrotic rat liver. J Hepatol 26: 886-893, 1997. 49. de Leeuw AM, McCarthy SP, Geerts A, Knook DL. Purified rat liver fat-storing cells in culture divide and contain collagen. Hepatology 4: 392-403, 1984. 50. De Minicis S, Brenner DA. NOX in liver fibrosis. Arch Biochem Biophys 462: 266-272, 2007. 51. Dooley S, Delvoux B, Lahme B, Mangasser-Stephan K, Gressner AM. Modulation of transforming growth factor beta response and signaling during transdifferentiation of rat hepatic stellate cells to myofibroblasts. Hepatology 31: 1094-1106, 2000. 52. Dranoff JA, Ogawa M, Kruglov EA, Gaca MD, Sevigny J, Robson SC, Wells RG. Expression of P2Y nucleotide receptors and ectonucleotidases in quiescent and activated rat hepatic stellate cells. Am J Physiol Gastrointest Liver Physiol 287: G417-G424, 2004. 53. Dranoff JA, Wells RG. Portal fibroblasts: Underappreciated mediators of biliary fibrosis. Hepatology 51: 1438-1444, 2010. 54. Ebrahimkhani MR, Oakley F, Murphy LB, Mann J, Moles A, Perugorria MJ, Ellis E, Lakey AF, Burt AD, Douglass A, Wright MC, White SA, Jaffre F, Maroteaux L, Mann DA. Stimulating healthy tissue regeneration by targeting the 5-HT(2B) receptor in chronic liver disease. Nat Med 17: 1668-1673, 2011. 55. Elsharkawy AM, Oakley F, Mann DA. The role and regulation of hepatic stellate cell apoptosis in reversal of liver fibrosis. Apoptosis 10: 927-939, 2005. 56. Enzan H, Hayashi Y, Miyazaki E, Naruse K, Tao R, Kuroda N, Nakayama H, Kiyoku H, Hiroi M, Saibara T. Morphological aspects of hepatic fibrosis and Ito cells (hepatic stellate cells), with special reference to their myofibroblastic transformation. In: Tanikawa K and Ueno T, editors. Liver Diseases and Hepatic Sinusoidal Cells. Tokyo: Springer-Verlag, 1999, p. 219-231. 57. Erkan M, Adler G, Apte MV, Bachem MG, Buchholz M, Detlefsen S, Esposito I, Friess H, Gress TM, Habisch HJ, Hwang RF, Jaster R, Kleeff J, Kloppel G, Kordes C, Logsdon CD, Masamune A, Michalski CW, Oh J, Phillips PA, Pinzani M, Reiser-Erkan C, Tsukamoto H, Wilson J. StellaTUM: Current consensus and discussion on pancreatic stellate cell research. Gut 61: 172-178, 2012.

1486


58. Fallowfield JA, Iredale JP. Reversal of liver fibrosis and cirrhosis–an emerging reality. Scott Med J 49: 3-6, 2004. 59. Fibbi G, Pucci M, D’Alessio S, Grappone C, Pellegrini G, Salzano R, Casini A, Milani S, Del Rosso M. Transforming growth factor beta-1 stimulates invasivity of hepatic stellate cells by engagement of the cell-associated fibrinolytic system. Growth Factors 19: 87-100, 2001. 60. Fischer R, Cariers A, Reinehr R, Haussinger D. Caspase 9-dependent killing of hepatic stellate cells by activated Kupffer cells. Gastroenterology 123: 845-861, 2002. 61. Fowell AJ, Collins JE, Duncombe DR, Pickering JA, Rosenberg WM, Benyon RC. Silencing tissue inhibitors of metalloproteinases (TIMPs) with short interfering RNA reveals a role for TIMP-1 in hepatic stellate cell proliferation. Biochem Biophys Res Commun 407: 277-282, 2011. 62. Friedman SL. Seminars in medicine of the Beth Israel Hospital, Boston. The cellular basis of hepatic fibrosis. Mechanisms and treatment strategies. N Engl J Med 328: 1828-1835, 1993. 63. Friedman SL. Molecular regulation of hepatic fibrosis, an integrated cellular response to tissue injury. J Biol Chem 275: 2247-2250, 2000. 64. Friedman SL. Hepatic fibrosis-Overview. Toxicology 254: 120-129, 2008. 65. Friedman SL. Hepatic stellate cells – protean, multifunctional, and enigmatic cells of the liver. Physiol Rev 88: 125-172, 2008. 66. Friedman SL. Mechanisms of hepatic fibrogenesis. Gastroenterology 134: 1655-1669, 2008. 67. Friedman SL. Fibrogenic cell reversion underlies fibrosis regression in liver. Proc Natl Acad Sci U S A 109: 9230-9231, 2012. 68. Friedman SL, Arthur MJ. Reversing hepatic fibrosis. Sci Med 8: 194205, 2002. 69. Friedman SL, Bansal MB. Reversal of hepatic fibrosis – fact or fantasy? Hepatology 43: S82-S88, 2006. 70. Friedman SL, Roll FJ, Boyles J, Arenson DM, Bissell DM. Maintenance of differentiated phenotype of cultured rat hepatic lipocytes by basement membrane matrix. J Biol Chem 264: 10756-10762, 1989. 71. Friedman SL, Roll FJ, Boyles J, Bissell DM. Hepatic lipocytes: The principal collagen-producing cells of normal rat liver. Proc Natl Acad Sci U S A 82: 8681-8685, 1985. 72. Friedman SL, Wei S, Blaner WS. Retinol release by activated rat hepatic lipocytes: Regulation by Kupffer cell-conditioned medium and PDGF. Am J Physiol 264: G947-G952, 1993. 73. Fritz D, Stefanovic B. RNA-binding protein RBMS3 is expressed in activated hepatic stellate cells and liver fibrosis and increases expression of transcription factor Prx1. J Mol Biol 371: 585-595, 2007. 74. Frizell E, Liu SL, Abraham A, Ozaki I, Eghbali M, Sage EH, Zern MA. Expression of SPARC in normal and fibrotic livers. Hepatology 21: 847-854, 1995. 75. Fujio K, Evarts RP, Hu Z, Marsden ER, Thorgeirsson SS. Expression of stem cell factor and its receptor, c kit, during liver regeneration from putative stem cells in adult rat. Lab Invest 70: 511-516, 1994. 76. Gao B, Radaeva S. Natural killer and natural killer T cells in liver fibrosis. Biochim Biophys Acta 1832: 1061-1069, 2012. 77. Gao B, Radaeva S, Park O. Liver natural killer and natural killer T cells: Immunobiology and emerging roles in liver diseases. J Leukoc Biol 86: 513-528, 2009. 78. Gao R, Brigstock DR. Connective tissue growth factor (CCN2) induces adhesion of rat activated hepatic stellate cells by binding of its Cterminal domain to integrin alpha(v)beta(3) and heparan sulfate proteoglycan. J Biol Chem 279: 8848-8855, 2004. 79. Gascon-Barre M, Demers C, Mirshahi A, Neron S, Zalzal S, Nanci A. The normal liver harbors the vitamin D nuclear receptor in nonparenchymal and biliary epithelial cells. Hepatology 37: 1034-1042, 2003. 80. Geerts A. History, heterogeneity, developmental biology, and functions of quiescent hepatic stellate cells. Semin Liver Dis 21: 311-335, 2001. 81. Geerts A, Bouwens L, Wisse E. Ultrastructure and function of hepatic fat-storing and pit cells. J Electr Micr Tech 14: 247-256, 1990. 82. Geerts A, Lazou JM, De Bleser P, Wisse E. Tissue distribution, quantitation and proliferation kinetics of fat-storing cells in carbon tetrachloride-injured rat liver. Hepatology 13: 1193-1202, 1991. 83. Geerts A, Vrijsen R, Rauterberg J, Burt A, Schellinck P, Wisse E. In vitro differentiation of fat-storing cells parallels marked increase of collagen synthesis and secretion. J Hepatol 9: 59-68, 1989. 84. Georges PC, Hui JJ, Gombos Z, McCormick ME, Wang AY, Uemura M, Mick R, Janmey PA, Furth EE, Wells RG. Increased stiffness of the rat liver precedes matrix deposition: Implications for fibrosis. Am J Physiol Gastrointest Liver Physiol 293: G1147-G1154, 2007. 85. Ghiassi-Nejad Z, Hernandez-Gea V, Woodrell C, Lang UE, Dumic K, Kwong A, Friedman SL. Reduced hepatic stellate cell expression of Kruppel-like factor 6 tumor suppressor isoforms amplifies fibrosis during acute and chronic rodent liver injury. Hepatology 57: 786-796, 2013. 86. Giampieri MP, Jezequel AM, Orlandi F. The lipocytes in normal human liver. Digestion 22: 165-169, 1981.



87. Gong Y, Hart E, Shchurin A, Hoover-Plow J. Inflammatory macrophage migration requires MMP-9 activation by plasminogen in mice. J Clin Invest 118: 3012-3024, 2008. 88. Gorbig MN, Gines P, Bataller R, Nicolas JM, Garcia-Ramallo E, Cejudo P, Sancho-Bru P, Jimenez W, Arroyo V, Rodes J. Human hepatic stellate cells secrete adrenomedullin: Potential autocrine factor in the regulation of cell contractility. J Hepatol 34: 222-229, 2001. 89. Greenbaum LE, Wells RG. The role of stem cells in liver repair and fibrosis. Int J Biochem Cell Biol 43: 222-229, 2011. 90. Gressner AM. The cell biology of liver fibrogenesis - an imbalance of proliferation, growth arrest and apoptosis of myofibroblasts. Cell Tissue Res 292: 447-452, 1998. 91. Gressner AM, Weiskirchen R, Breitkopf K, Dooley S. Roles of tgf-Beta in hepatic fibrosis. Front Biosci 7: D793-D807, 2002. 92. Gressner OA, Gressner AM. Connective tissue growth factor: A fibrogenic master switch in fibrotic liver diseases. Liver Int 28: 1065-1079, 2008. 93. Guo CJ, Pan Q, Li DG, Sun H, Liu BW. miR-15b and miR-16 are implicated in activation of the rat hepatic stellate cell: An essential role for apoptosis. J Hepatol 50: 766-778, 2009. 94. Guo J, Loke J, Zheng F, Hong F, Yea S, Fukata M, Tarocchi M, Abar OT, Huang H, Sninsky JJ, Friedman SL. Functional linkage of cirrhosispredictive single nucleotide polymorphisms of Toll-like receptor 4 to hepatic stellate cell responses. Hepatology 49: 960-968, 2009. 95. Hammel P, Couvelard A, O’Toole D, Ratouis A, Sauvanet A, Flejou JF, Degott C, Belghiti J, Bernades P, Valla D, Ruszniewski P, Levy P. Regression of liver fibrosis after biliary drainage in patients with chronic pancreatitis and stenosis of the common bile duct. N Engl J Med 344: 418-423, 2001. 96. Han YP, Yan C, Zhou L, Qin L, Tsukamoto H. A matrix metalloproteinase-9 activation cascade by hepatic stellate cells in transdifferentiation in the three-dimensional extracellular matrix. J Biol Chem 282: 12928-12939, 2007. 97. Hartland SN, Murphy F, Aucott RL, Abergel A, Zhou X, Waung J, Patel N, Bradshaw C, Collins J, Mann D, Benyon RC, Iredale JP. Active matrix metalloproteinase-2 promotes apoptosis of hepatic stellate cells via the cleavage of cellular N-cadherin. Liver Int 29: 966-978, 2009. 98. He H, Mennone A, Boyer JL, Cai SY. Combination of retinoic acid and ursodeoxycholic acid attenuates liver injury in bile duct-ligated rats and human hepatic cells. Hepatology 53: 548-557, 2011. 99. He Y, Huang C, Zhang SP, Sun X, Long XR, Li J. The potential of microRNAs in liver fibrosis. Cell Signal 24: 2268-2272, 2012. 100. Hellemans K, Verbuyst P, Quartier E, Schuit F, Rombouts K, Chandraratna RA, Schuppan D, Geerts A. Differential modulation of rat hepatic stellate phenotype by natural and synthetic retinoids. Hepatology 39: 97-108, 2004. 101. Hendriks HF, Verhoofstad WA, Brouwer A, de Leeuw AM, Knook DL. Perisinusoidal fat-storing cells are the main vitamin A storage sites in rat liver. Exp Cell Res 160: 138-149, 1985. 102. Hernandez-Gea V, Friedman SL. Pathogenesis of liver fibrosis. Annu Rev Pathol 6: 425-456, 2011. 103. Hernandez-Gea V, Friedman SL. Autophagy fuels tissue fibrogenesis. Autophagy 8: 849-850, 2012. 104. Hernandez-Gea V, Ghiassi-Nejad Z, Rozenfeld R, Gordon R, Fiel MI, Yue Z, Czaja MJ, Friedman SL. Autophagy releases lipid that promotes fibrogenesis by activated hepatic stellate cells in mice and in human tissues. Gastroenterology 142: 938-946, 2012. 105. Hernandez-Gea V, Hilscher M, Rozenfeld R, Lim MP, Nieto N, Werner S, Devi LA, Friedman SL. Endoplasmic reticulum stress induces fibrogenic activity in hepatic stellate cells through autophagy. J Hepatol 59: 98-104, 2013. 106. Heymann F, Trautwein C, Tacke F. Monocytes and macrophages as cellular targets in liver fibrosis. Inflamm Allergy Drug Targets 8: 307318, 2009. 107. Holt AP, Haughton EL, Lalor PF, Filer A, Buckley CD, Adams DH. Liver myofibroblasts regulate infiltration and positioning of lymphocytes in human liver. Gastroenterology 136: 705-714, 2009. 108. Hong F, Tuyama A, Lee TF, Loke J, Agarwal R, Cheng X, Garg A, Fiel MI, Schwartz M, Walewski J, Branch A, Schecter AD, Bansal MB. Hepatic stellate cells express functional CXCR4: Role in stromal cellderived factor-1alpha-mediated stellate cell activation. Hepatology 49: 2055-2067, 2009. 109. Hsieh CC, Chou HS, Fung JJ, Qian S, Lu L. The role of liver stromal cells in dendritic cells development in mice. Transplant Proc 42: 42794281, 2010. 110. Huang G, Brigstock DR. Integrin expression and function in the response of primary culture hepatic stellate cells to connective tissue growth factor (CCN2). J Cell Mol Med 15: 1087-1095, 2011. 111. Huang G, Brigstock DR. Regulation of hepatic stellate cells by connective tissue growth factor. Front Biosci 17: 2495-2507, 2012. 112. Iizuka M, Ogawa T, Enomoto M, Motoyama H, Yoshizato K, Ikeda K, Kawada N. Induction of microRNA-214-5p in human and rodent liver fibrosis. Fibrogenesis Tissue Repair 5: 12, 2012.



113. Ikeda K, Wakahara T, Wang YQ, Kadoya H, Kawada N, Kaneda K. In vitro migratory potential of rat quiescent hepatic stellate cells and its augmentation by cell activation. Hepatology 29: 1760-1767, 1999. 114. Ikeda N, Murata S, Maruyama T, Tamura T, Nozaki R, Kawasaki T, Fukunaga K, Oda T, Sasaki R, Homma M, Ohkohchi N. Platelet-derived adenosine 5 -triphosphate suppresses activation of human hepatic stellate cell: In vitro study. Hepatol Res 42: 91-102, 2012. 115. Imai K, Senoo H. Morphology of sites of adhesion between hepatic stellate cells (vitamin A-storing cells) and a three-dimensional extracellular matrix. Anat Rec 250: 430-437, 1998. 116. Inagaki Y, Kushida M, Higashi K, Itoh J, Higashiyama R, Hong YY, Kawada N, Namikawa K, Kiyama H, Bou-Gharios G, Watanabe T, Okazaki I, Ikeda K. Cell type-specific intervention of transforming growth factor beta/Smad signaling suppresses collagen gene expression and hepatic fibrosis in mice. Gastroenterology 129: 259-268, 2005. 117. Inagaki Y, Mamura M, Kanamaru Y, Greenwel P, Nemoto T, Takehara K, Ten Dijke P, Nakao A. Constitutive phosphorylation and nuclear localization of Smad3 are correlated with increased collagen gene transcription in activated hepatic stellate cells. J Cell Physiol 187: 117-123, 2001. 118. Inagaki Y, Nemoto T, Kushida M, Sheng Y, Higashi K, Ikeda K, Kawada N, Shirasaki F, Takehara K, Sugiyama K, Fujii M, Yamauchi H, Nakao A, De Crombrugghe B, Watanabe T, Okazaki I. Interferon alfa down-regulates collagen gene transcription and suppresses experimental hepatic fibrosis in mice. Hepatology 38: 890-899, 2003. 119. Inagaki Y, Truter S, Greenwel P, Rojkind M, Unoura M, Kobayashi K, Ramirez F. Regulation of the alpha 2(I) collagen gene transcription in fat-storing cells derived from a cirrhotic liver. Hepatology 22: 573-579, 1995. 120. Iredale JP. Hepatic stellate cell behavior during resolution of liver injury. Semin Liver Dis 21: 427-436, 2001. 121. Iredale JP. Models of liver fibrosis: Exploring the dynamic nature of inflammation and repair in a solid organ. J Clin Invest 117: 539-548, 2007. 122. Iredale JP, Benyon RC, Pickering J, McCullen M, Northrop M, Pawley S, Hovell C, Arthur MJ. Mechanisms of spontaneous resolution of rat liver fibrosis. Hepatic stellate cell apoptosis and reduced hepatic expression of metalloproteinase inhibitors. J Clin Invest 102: 538-549, 1998. 123. Issa R, Williams E, Trim N, Kendall T, Arthur MJ, Reichen J, Benyon RC, Iredale JP. Apoptosis of hepatic stellate cells: Involvement in resolution of biliary fibrosis and regulation by soluble growth factors. Gut 48: 548-557, 2001. 124. Issa R, Zhou X, Constandinou CM, Fallowfield J, Millward-Sadler H, Gaca MD, Sands E, Suliman I, Trim N, Knorr A, Arthur MJ, Benyon RC, Iredale JP. Spontaneous recovery from micronodular cirrhosis: Evidence for incomplete resolution associated with matrix cross-linking. Gastroenterology 126: 1795-1808, 2004. 125. Jarnagin WR, Rockey DC, Koteliansky VE, Wang SS, Bissell DM. Expression of variant fibronectins in wound healing: Cellular source and biological activity of the EIIIA segment in rat hepatic fibrogenesis. J Cell Biol 127: 2037-2048, 1994. 126. Jezequel AM, Novelli G, Venturini C, Orlandi F. Quantitative analysis of the perisinusoidal cells in human liver; the lipocytes. Front Gastrointestinal Res 8: 85-90, 1984. 127. Ji J, Yu F, Ji Q, Li Z, Wang K, Zhang J, Lu J, Chen L, Qun E, Zeng Y, Ji Y. Comparative proteomic analysis of rat hepatic stellate cell activation: A comprehensive view and suppressed immune response. Hepatology 56: 332-349, 2012. 128. Ji J, Zhang J, Huang G, Qian J, Wang X, Mei S. Over-expressed microRNA-27a and 27b influence fat accumulation and cell proliferation during rat hepatic stellate cell activation. FEBS Lett 583: 759-766, 2009. 129. Jiang JX, Chen X, Hsu DK, Baghy K, Serizawa N, Scott F, Takada Y, Fukada H, Chen J, Devaraj S, Adamson R, Liu FT, Torok NJ. Galectin-3 modulates phagocytosis-induced stellate cell activation and liver fibrosis in vivo. Am J Physiol Gastrointest Liver Physiol 302: G439-G446, 2012. 130. Jiang Z, Chen Y, Feng X, Jiang J, Chen T, Xie H, Zhou L, Zheng S. Hepatic stellate cells promote immunotolerance following orthotopic liver transplantation in rats via induction of T cell apoptosis and regulation of Th2/Th3-like cell cytokine production. Exp Ther Med 5: 165-169, 2013. 131. Jiao J, Sastre D, Fiel MI, Lee UE, Ghiassi-Nejad Z, Ginhoux F, Vivier E, Friedman SL, Merad M, Aloman C. Dendritic cell regulation of carbon tetrachloride-induced murine liver fibrosis regression. Hepatology 55: 244-255, 2012. 132. Kageyama Y, Ikeda H, Watanabe N, Nagamine M, Kusumoto Y, Yashiro M, Satoh Y, Shimosawa T, Shinozaki K, Tomiya T, Inoue Y, Nishikawa T, Ohtomo N, Tanoue Y, Yokota H, Koyama T, Ishimaru K, Okamoto Y, Takuwa Y, Koike K, Yatomi Y. Antagonism of sphingosine 1-phosphate receptor 2 causes a selective reduction of portal

1487


133.

134. 135.

136.

137.

138. 139.

140. 141.

142.

143.

144.

145.

146. 147.

148.

149.

150.

151.

152. 153.

vein pressure in bile duct-ligated rodents. Hepatology 56: 1427-1438, 2012. Kalinichenko VV, Bhattacharyya D, Zhou Y, Gusarova GA, Kim W, Shin B, Costa RH. Foxf1+/− mice exhibit defective stellate cell activation and abnormal liver regeneration following CCl4 injury. Hepatology 37: 107-117, 2003. Kang N, Gores GJ, Shah VH. Hepatic stellate cells: Partners in crime for liver metastases? Hepatology 54: 707-713, 2011. Kaur S, Tripathi D, Dongre K, Garg V, Rooge S, Mukopadhyay A, Sakhuja P, Sarin SK. Increased number and function of endothelial progenitor cells stimulate angiogenesis by resident liver sinusoidal endothelial cells (SECs) in cirrhosis through paracrine factors. J Hepatol 57: 1193-1198, 2012. Kawashima R, Mochida S, Matsui A, YouLuTu ZY, Ishikawa K, Toshima K, Yamanobe F, Inao M, Ikeda H, Ohno A, Nagoshi S, Uede T, Fujiwara K. Expression of osteopontin in Kupffer cells and hepatic macrophages and Stellate cells in rat liver after carbon tetrachloride intoxication: A possible factor for macrophage migration into hepatic necrotic areas. Biochem Biophys Res Commun 256: 527-531, 1999. Kendall TJ, Hennedige S, Aucott RL, Hartland SN, Vernon MA, Benyon RC, Iredale JP. p75 Neurotrophin receptor signaling regulates hepatic myofibroblast proliferation and apoptosis in recovery from rodent liver fibrosis. Hepatology 49: 901-910, 2009. Khimji AK, Rockey DC. Endothelin and hepatic wound healing. Pharmacol Res 63: 512-518, 2011. Kida Y, Xia Z, Zheng S, Mordwinkin NM, Louie SG, Zheng SG, Feng M, Shi H, Duan Z, Han YP. Interleukin-1 as an injury signal mobilizes retinyl esters in hepatic stellate cells through down regulation of lecithin retinol acyltransferase. PLoS One 6: e26644, 2011. Kikuchi S, Griffin CT, Wang SS, Bissell DM. Role of CD44 in epithelial wound repair: Migration of rat hepatic stellate cells utilizes hyaluronic acid and CD44v6. J Biol Chem 280: 15398-15404, 2005. Kinnman N, Francoz C, Barbu V, Wendum D, Rey C, Hultcrantz R, Poupon R, Housset C. The myofibroblastic conversion of peribiliary fibrogenic cells distinct from hepatic stellate cells is stimulated by platelet-derived growth factor during liver fibrogenesis. Lab Invest 83: 163-173, 2003. Kinnman N, Hultcrantz R, Barbu V, Rey C, Wendum D, Poupon R, Housset C. PDGF-mediated chemoattraction of hepatic stellate cells by bile duct segments in cholestatic liver injury. Lab Invest 80: 697-707, 2000. Kisseleva T, Cong M, Paik Y, Scholten D, Jiang C, Benner C, Iwaisako K, Moore-Morris T, Scott B, Tsukamoto H, Evans SM, Dillmann W, Glass CK, Brenner DA. Myofibroblasts revert to an inactive phenotype during regression of liver fibrosis. Proc Natl Acad Sci U S A 109: 9448-9453, 2012. Kluwe J, Wongsiriroj N, Troeger JS, Gwak GY, Dapito DH, Pradere JP, Jiang H, Siddiqi M, Piantedosi R, O’Byrne SM, Blaner WS, Schwabe RF. Absence of hepatic stellate cell retinoid lipid droplets does not enhance hepatic fibrosis but decreases hepatic carcinogenesis. Gut 60: 1260-1268, 2011. Knittel T, Mehde M, Kobold D, Saile B, Dinter C, Ramadori G. Expression patterns of matrix metalloproteinases and their inhibitors in parenchymal and non-parenchymal cells of rat liver: Regulation by TNF-alpha and TGF-beta1. J Hepatol 30: 48-60, 1999. Knolle PA, Gerken G. Local control of the immune response in the liver. Immunol Rev 174: 21-34, 2000. Kodama T, Takehara T, Hikita H, Shimizu S, Li W, Miyagi T, Hosui A, Tatsumi T, Ishida H, Tadokoro S, Ido A, Tsubouchi H, Hayashi N. Thrombocytopenia exacerbates cholestasis-induced liver fibrosis in mice. Gastroenterology 138: 2487-2498, 2498 e2481-2487, 2010. Kojima N, Sato M, Imai K, Miura M, Matano Y, Senoo H. Hepatic stellate cells (vitamin A-storing cells) change their cytoskeleton structure by extracellular matrix components through a signal transduction system. Histochem Cell Biol 110: 121-128, 1998. Kojima S, Hayashi S, Shimokado K, Suzuki Y, Shimada J, Crippa MP, Friedman SL. Transcriptional activation of urokinase by the Kruppellike factor Zf9/COPEB activates latent TGF-beta1 in vascular endothelial cells. Blood 95: 1309-1316, 2000. Komatsu M, Waguri S, Chiba T, Murata S, Iwata J, Tanida I, Ueno T, Koike M, Uchiyama Y, Kominami E, Tanaka K. Loss of autophagy in the central nervous system causes neurodegeneration in mice. Nature 441: 880-884, 2006. Komatsu M, Wang QJ, Holstein GR, Friedrich VL, Jr., Iwata J, Kominami E, Chait BT, Tanaka K, Yue Z. Essential role for autophagy protein Atg7 in the maintenance of axonal homeostasis and the prevention of axonal degeneration. Proc Natl Acad Sci U S A 104: 14489-14494, 2007. Kong X, Horiguchi N, Mori M, Gao B. Cytokines and STATs in liver fibrosis. Front Physiol 3: 69, 2012. Kordes C, Sawitza I, Gotze S, Haussinger D. Stellate cells from rat pancreas are stem cells and can contribute to liver regeneration. PLoS One 7: e51878, 2012.

1488


154. Kordes C, Sawitza I, Muller-Marbach A, Ale-Agha N, Keitel V, Klonowski-Stumpe H, Haussinger D. CD133+ hepatic stellate cells are progenitor cells. Biochem Biophys Res Commun 352: 410-417, 2007. 155. Krizhanovsky V, Yon M, Dickins RA, Hearn S, Simon J, Miething C, Yee H, Zender L, Lowe SW. Senescence of activated stellate cells limits liver fibrosis. Cell 134: 657-667, 2008. 156. Kruglov EA, Nathanson RA, Nguyen T, Dranoff JA. Secretion of MCP1/CCL2 by bile duct epithelia induces myofibroblastic transdifferentiation of portal fibroblasts. Am J Physiol Gastrointest Liver Physiol 290: G765-G771, 2006. 157. Kubota H, Yao HL, Reid LM. Identification and characterization of vitamin A-storing cells in fetal liver: Implications for functional importance of hepatic stellate cells in liver development and hematopoiesis. Stem Cells 25: 2339-2349, 2007. 158. Laleman W, Van Landeghem L, Severi T, Vander Elst I, Zeegers M, Bisschops R, Van Pelt J, Roskams T, Cassiman D, Fevery J, Nevens F. Both Ca2+ -dependent and -independent pathways are involved in rat hepatic stellate cell contraction and intrahepatic hyperresponsiveness to methoxamine. Am J Physiol Gastrointest Liver Physiol 292: G556G564, 2007. 159. Lee JS, Semela D, Iredale J, Shah VH. Sinusoidal remodeling and angiogenesis: A new function for the liver-specific pericyte? Hepatology 45: 817-825, 2007. 160. Lee SJ, Friedman SL, Whalen R, Boyer TD. Cellular sources of glutathione S-transferase P in primary cultured rat hepatocytes: Localization by in situ hybridization. Biochem J 299: 79-83, 1994. 161. Lee TF, Mak KM, Rackovsky O, Lin YL, Kwong AJ, Loke JC, Friedman SL. Downregulation of hepatic stellate cell activation by retinol and palmitate mediated by adipose differentiation-related protein (ADRP). J Cell Physiol 223: 648-657, 2010. 162. Lemoinne S, Cadoret A, Mourabit HE, Thabut D, Housset C. Origins and functions of liver myofibroblasts. Biochim Biophys Acta 1832: 948-954, 2013. 163. Li J, Ghazwani M, Zhang Y, Lu J, Fan J, Gandhi CR, Li S. miR-122 regulates collagen production via targeting hepatic stellate cells and suppressing P4HA1 expression. J Hepatol 58: 522-528, 2013. 164. Li T, Shi Z, Rockey DC. Preproendothelin-1 expression is negatively regulated by IFNgamma during hepatic stellate cell activation. Am J Physiol Gastrointest Liver Physiol 302: G948-G957, 2012. 165. Li Y, Wang J, Asahina K. Mesothelial cells give rise to hepatic stellate cells and myofibroblasts via mesothelial-mesenchymal transition in liver injury. Proc Natl Acad Sci U S A 110: 2324-2329, 2013. 166. Libbrecht L, Cassiman D, Desmet V, Roskams T. The correlation between portal myofibroblasts and development of intrahepatic bile ducts and arterial branches in human liver. Liver 22: 252-258, 2002. 167. Lindquist J, Stefanovic B, Brenner D. Regulation of collagen alpha1(I) expression in hepatic stellate cells. J Gastroenterol 35(Suppl 12): 80-83, 2000. 168. Liu Z, Rossen EV, Timmermans JP, Geerts A, van Grunsven LA, Reynaert H. Distinct roles for non-muscle myosin II isoforms in mouse hepatic stellate cells. J Hepatol 54: 132-141, 2011. 169. Lujambio A, Akkari L, Simon J, Grace D, Tschaharganeh DF, Bolden JE, Zhao Z, Thapar V, Joyce JA, Krizhanovsky V, Lowe SW. Non-CellAutonomous Tumor Suppression by p53. Cell 153: 449-460, 2013. 170. Magness ST, Bataller R, Yang L, Brenner DA. A dual reporter gene transgenic mouse demonstrates heterogeneity in hepatic fibrogenic cell populations. Hepatology 40: 1151-1159, 2004. 171. Maher JJ, McGuire RF. Extracellular matrix gene expression increases preferentially in rat lipocytes and sinusoidal endothelial cells during hepatic fibrosis in vivo. J Clin Invest 86: 1641-1648, 1990. 172. Mann J, Chu DC, Maxwell A, Oakley F, Zhu NL, Tsukamoto H, Mann DA. MeCP2 controls an epigenetic pathway that promotes myofibroblast transdifferentiation and fibrosis. Gastroenterology 138: 705-714, 714 e701-704, 2010. 173. Mann J, Mann DA. Transcriptional regulation of hepatic stellate cells. Adv Drug Deliv Rev 61: 497-512, 2009. 174. Mann J, Oakley F, Akiboye F, Elsharkawy A, Thorne AW, Mann DA. Regulation of myofibroblast transdifferentiation by DNA methylation and MeCP2: Implications for wound healing and fibrogenesis. Cell Death Differ 14: 275-285, 2007. 175. Manojlovic Z, Stefanovic B. A novel role of RNA helicase A in regulation of translation of type I collagen mRNAs. RNA 18: 321-334, 2012. 176. Marra F, Romanelli RG, Giannini C, Failli P, Pastacaldi S, Arrighi MC, Pinzani M, Laffi G, Montalto P, Gentilini P. Monocyte chemotactic protein-1 as a chemoattractant for human hepatic stellate cells. Hepatology 29: 140-148, 1999. 177. Marsden ER, Hu Z, Fujio K, Nakatsukasa H, Thorgeirsson SS, Evarts RP. Expression of acidic fibroblast growth factor in regenerating liver and during hepatic differentiation. Lab Invest 67: 427-433, 1992. 178. Matsumoto K, Miki R, Nakayama M, Tatsumi N, Yokouchi Y. Wnt9a secreted from the walls of hepatic sinusoids is essential for



179.

180. 181.

182.

183.

184.

185.

186.

187. 188. 189. 190.

191. 192.

193.

194.

195.

196.

197. 198.

morphogenesis, proliferation, and glycogen accumulation of chick hepatic epithelium. Dev Biol 319: 234-247, 2008. Mehal WZ, Friedman SL. The role of inflammation and immunity in the pathogenesis of liver fibrosis. In: Gershwin ME, Veirling JM, Manns MP, editors. Liver Immunology. Totowa, New Jersey: Humana Press, 2007, p. 99-109. Melton AC, Yee HF. Hepatic stellate cell protrusions couple plateletderived growth factor-BB to chemotaxis. Hepatology 45: 1446-1453, 2007. Meng F, Wang K, Aoyama T, Grivennikov SI, Paik Y, Scholten D, Cong M, Iwaisako K, Liu X, Zhang M, Osterreicher CH, Stickel F, Ley K, Brenner DA, Kisseleva T. Interleukin-17 signaling in inflammatory, Kupffer cells, and hepatic stellate cells exacerbates liver fibrosis in mice. Gastroenterology 143: 765-776 e761-763, 2012. Meyer DH, Bachem MG, Gressner AM. Modulation of hepatic lipocyte proteoglycan synthesis and proliferation by Kupffer cell-derived transforming growth factors type beta 1 and type alpha. Biochem Biophys Res Commun 171: 1122-1129, 1990. Milani S, Herbst H, Schuppan D, Grappone C, Pellegrini G, Pinzani M, Casini A, Calabr´o A, Ciancio G, Stefanini F, Ciancio AK, Surrenti C. Differential expression of matrix-metalloproteinase-1 and -2 genes in normal and fibrotic human liver. Am J Pathol 144: 528-537, 1994. Miura K, Nagai H, Ueno Y, Goto T, Mikami K, Nakane K, Yoneyama K, Watanabe D, Terada K, Sugiyama T, Imai K, Senoo H, Watanabe S. Epimorphin is involved in differentiation of rat hepatic stem-like cells through cell-cell contact. Biochem Biophys Res Commun 311: 415-423, 2003. Miura K, Yang L, van Rooijen N, Ohnishi H, Seki E. Hepatic recruitment of macrophages promotes nonalcoholic steatohepatitis through CCR2. Am J Physiol Gastrointest Liver Physiol 302: G1310-G1321, 2012. Miyata E, Masuya M, Yoshida S, Nakamura S, Kato K, Sugimoto Y, Shibasaki T, Yamamura K, Ohishi K, Nishii K, Ishikawa F, Shiku H, Katayama N. Hematopoietic origin of hepatic stellate cells in the adult liver. Blood 111: 2427-2435, 2008. Modi AA, Feld JJ, Park Y, Kleiner DE, Everhart JE, Liang TJ, Hoofnagle JH. Increased caffeine consumption is associated with reduced hepatic fibrosis. Hepatology 51: 201-209, 2010. Moriwaki H, Blaner WS, Piantedosi R, Goodman DS. Effects of dietary retinoid and triglyceride on the lipid composition of rat liver stellate cells and stellate cell lipid droplets. J Lipid Res 29: 1523-1534, 1988. Muhanna N, Abu Tair L, Doron S, Amer J, Azzeh M, Mahamid M, Friedman S, Safadi R. Amelioration of hepatic fibrosis by NK cell activation. Gut 60: 90-98, 2011. Muhanna N, Doron S, Wald O, Horani A, Eid A, Pappo O, Friedman SL, Safadi R. Activation of hepatic stellate cells after phagocytosis of lymphocytes: A novel pathway of fibrogenesis. Hepatology 48: 963977, 2008. Mullhaupt B, Feren A, Fodor E, Jones A. Liver expression of epidermal growth factor RNA. Rapid increases in immediate-early phase of liver regeneration. J Biol Chem 269: 19667-19670, 1994. Murphy FR, Issa R, Zhou X, Ratnarajah S, Nagase H, Arthur MJ, Benyon C, Iredale JP. Inhibition of apoptosis of activated hepatic stellate cells by tissue inhibitor of metalloproteinase-1 is mediated via effects on matrix metalloproteinase inhibition: Implications for reversibility of liver fibrosis. J Biol Chem 277: 11069-11076, 2002. Nieto N, Friedman SL, Cederbaum AI. Cytochrome P450 2E1-derived reactive oxygen species mediate paracrine stimulation of collagen I protein synthesis by hepatic stellate cells. J Biol Chem 277: 9853-9864, 2002. Niiya M, Uemura M, Zheng XW, Pollak ES, Dockal M, Scheiflinger F, Wells RG, Zheng XL. Increased ADAMTS-13 proteolytic activity in rat hepatic stellate cells upon activation in vitro and in vivo. J Thromb Haemost 4: 1063-1070, 2006. Niki T, De Bleser PJ, Xu G, Van Den Berg K, Wisse E, Geerts A. Comparison of glial fibrillary acidic protein and desmin staining in normal and CCl4-induced fibrotic rat livers. Hepatology 23: 1538-1545, 1996. Noetel A, Elfimova N, Altmuller J, Becker C, Becker D, Lahr W, Nurnberg P, Wasmuth H, Teufel A, Buttner R, Dienes HP, Odenthal M. Next generation sequencing of the Ago2 interacting transcriptome identified chemokine family members as novel targets of neuronal microRNAs in hepatic stellate cells. J Hepatol 58: 335-341, 2013. Noetel A, Kwiecinski M, Elfimova N, Huang J, Odenthal M. microRNA are central players in anti- and profibrotic gene regulation during liver fibrosis. Front Physiol 3: 49, 2012. Novo E, Busletta C, Bonzo LV, Povero D, Paternostro C, Mareschi K, Ferrero I, David E, Bertolani C, Caligiuri A, Cannito S, Tamagno E, Compagnone A, Colombatto S, Marra F, Fagioli F, Pinzani M, Parola M. Intracellular reactive oxygen species are required for directional migration of resident and bone marrow-derived hepatic pro-fibrogenic cells. J Hepatol 54: 964-974, 2011.



199. Novo E, Cannito S, Zamara E, Valfre di Bonzo L, Caligiuri A, Cravanzola C, Compagnone A, Colombatto S, Marra F, Pinzani M, Parola M. Proangiogenic cytokines as hypoxia-dependent factors stimulating migration of human hepatic stellate cells. Am J Pathol 170: 1942-1953, 2007. 200. Novo E, Povero D, Busletta C, Paternostro C, di Bonzo LV, Cannito S, Compagnone A, Bandino A, Marra F, Colombatto S, David E, Pinzani M, Parola M. The biphasic nature of hypoxia-induced directional migration of activated human hepatic stellate cells. J Pathol 226: 588-597, 2012. 201. Novobrantseva TI, Majeau GR, Amatucci A, Kogan S, Brenner I, Casola S, Shlomchik MJ, Koteliansky V, Hochman PS, Ibraghimov A. Attenuated liver fibrosis in the absence of B cells. J Clin Invest 115: 3072-3082, 2005. 202. Oakley F, Meso M, Iredale JP, Green K, Marek CJ, Zhou X, May MJ, Millward-Sadler H, Wright MC, Mann DA. Inhibition of inhibitor of kappaB kinases stimulates hepatic stellate cell apoptosis and accelerated recovery from rat liver fibrosis. Gastroenterology 128: 108-120, 2005. 203. Oakley F, Trim N, Constandinou CM, Ye W, Gray AM, Frantz G, Hillan K, Kendall T, Benyon RC, Mann DA, Iredale JP. Hepatocytes express nerve growth factor during liver injury: Evidence for paracrine regulation of hepatic stellate cell apoptosis. Am J Pathol 163: 18491858, 2003. 204. Odena G, Bataller R. Actin-binding proteins as molecular targets to modulate hepatic stellate cell proliferation. Focus on “MARCKS actinbinding capacity mediates actin filament assembly during mitosis in human hepatic stellate cells”. Am J Physiol Cell Physiol 303: C355C356, 2012. 205. Ogawa T, Enomoto M, Fujii H, Sekiya Y, Yoshizato K, Ikeda K, Kawada N. MicroRNA-221/222 upregulation indicates the activation of stellate cells and the progression of liver fibrosis. Gut 61: 1600-1609, 2012. 206. Ohata M, Lin M, Satre M, Tsukamoto H. Diminished retinoic acid signaling in hepatic stellate cells in cholestatic liver fibrosis. Am J Physiol 272: G589-G596, 1997. 207. Okuno M, Moriwaki H, Imai S, Muto Y, Kawada N, Suzuki Y, Kojima S. Retinoids exacerbate rat liver fibrosis by inducing the activation of latent TGF-beta in liver stellate cells. Hepatology 26: 913-921, 1997. 208. Olaso E, Arteta B, Benedicto A, Crende O, Friedman SL. Loss of discoidin domain receptor 2 promotes hepatic fibrosis after chronic carbon tetrachloride through altered paracrine interactions between hepatic stellate cells and liver-associated macrophages. Am J Pathol 179: 2894-2904, 2011. 209. Olaso E, Salado C, Egilegor E, Gutierrez V, Santisteban A, Sancho-Bru P, Friedman SL, Vidal-Vanaclocha F. Proangiogenic role of tumoractivated hepatic stellate cells in experimental melanoma metastasis. Hepatology 37: 674-685, 2003. 210. Olsen AL, Sackey BK, Marcinkiewicz C, Boettiger D, Wells RG. Fibronectin extra domain-A promotes hepatic stellate cell motility but not differentiation into myofibroblasts. Gastroenterology 142: 928-937 e923, 2012. 211. Ong A, Wong VW, Wong GL, Chan HL. The effect of caffeine and alcohol consumption on liver fibrosis - a study of 1045 Asian hepatitis B patients using transient elastography. Liver Int 31: 1047-1053, 2011. 212. Paik YH, Iwaisako K, Seki E, Inokuchi S, Schnabl B, Osterreicher CH, Kisseleva T, Brenner DA. The nicotinamide adenine dinucleotide phosphate oxidase (NOX) homologues NOX1 and NOX2/gp91(phox) mediate hepatic fibrosis in mice. Hepatology 53: 1730-1741, 2011. 213. Paik YH, Schwabe RF, Bataller R, Russo MP, Jobin C, Brenner DA. Toll-like receptor 4 mediates inflammatory signaling by bacterial lipopolysaccharide in human hepatic stellate cells. Hepatology 37: 1043-1055, 2003. 214. Parsons CJ, Stefanovic B, Seki E, Aoyama T, Latour AM, Marzluff WF, Rippe RA, Brenner DA. Mutation of the 5 -untranslated region stemloop structure inhibits alpha1(I) collagen expression in vivo. J Biol Chem 286: 8609-8619, 2011. 215. Passino MA, Adams RA, Sikorski SL, Akassoglou K. Regulation of hepatic stellate cell differentiation by the neurotrophin receptor p75NTR. Science 315: 1853-1856, 2007. 216. Patsenker E, Stickel F. Role of integrins in fibrosing liver diseases. Am J Physiol Gastrointest Liver Physiol 301: G425-G434, 2011. 217. Perez-Tamayo R. Cirrhosis of the liver: A reversible disease? Pathol Annu 14(Pt 2): 183-213, 1979. 218. Perugorria MJ, Wilson CL, Zeybel M, Walsh M, Amin S, Robinson S, White SA, Burt AD, Oakley F, Tsukamoto H, Mann DA, Mann J. Histone methyltransferase ASH1 orchestrates fibrogenic gene transcription during myofibroblast transdifferentiation. Hepatology 56: 1129-1139, 2012. 219. Pintilie DG, Shupe TD, Oh SH, Salganik SV, Darwiche H, Petersen BE. Hepatic stellate cells’ involvement in progenitor-mediated liver regeneration. Lab Invest 90: 1199-1208, 2010.

1489


220. Pinzani M. PDGF and signal transduction in hepatic stellate cells. Front Biosci 7: d1720-d1726, 2002. 221. Pinzani M, Milani S, De FR, Grappone C, Caligiuri A, Gentilini A, Tosti GC, Maggi M, Failli P, Ruocco C, Gentilini P. Endothelin 1 is overexpressed in human cirrhotic liver and exerts multiple effects on activated hepatic stellate cells. Gastroenterology 110: 534-548, 1996. 222. Popov Y, Patsenker E, Stickel F, Zaks J, Bhaskar KR, Niedobitek G, Kolb A, Friess H, Schuppan D. Integrin alphavbeta6 is a marker of the progression of biliary and portal liver fibrosis and a novel target for antifibrotic therapies. J Hepatol 48: 453-464, 2008. 223. Popper H, Schaffner F. Hepatic cirrhosis. A problem in communication. Isr J Med Sci 4: 1-7, 1968. 224. Popper H, Udenfriend S. Hepatic fibrosis. Correlation of biologic and morphologic investigations. Am J Med 49: 707-721, 1970. 225. Putz-Bankuti C, Pilz S, Stojakovic T, Scharnagl H, Pieber TR, Trauner M, Obermayer-Pietsch B, Stauber RE. Association of 25hydroxyvitamin D levels with liver dysfunction and mortality in chronic liver disease. Liver Int 32: 845-851, 2012. 226. Qin L, Han YP. Epigenetic repression of matrix metalloproteinases in myofibroblastic hepatic stellate cells through histone deacetylases 4: Implication in tissue fibrosis. Am J Pathol 177: 1915-1928, 2010. 227. Racine-Samson L, Rockey DC, Bissell DM. Expression of the collagenbinding integrins alpha1 beta1 and alpha 2 beta1 during activation of rat hepatic stellate cells in vivo. J Biol Chem 272(49): 30911-30917, 1997. 228. Radaeva S, Sun R, Jaruga B, Nguyen VT, Tian Z, Gao B. Natural killer cells ameliorate liver fibrosis by killing activated stellate cells in NKG2D-dependent and tumor necrosis factor-related apoptosisinducing ligand-dependent manners. Gastroenterology 130: 435-452, 2006. 229. Radbill BD, Gupta R, Ramirez MC, DiFeo A, Martignetti JA, Alvarez CE, Friedman SL, Narla G, Vrabie R, Bowles R, Saiman Y, Bansal MB. Loss of matrix metalloproteinase-2 amplifies murine toxin-induced liver fibrosis by upregulating collagen I expression. Dig Dis Sci 56: 406-416, 2011. 230. Ramachandran P, Iredale JP. Macrophages: Central regulators of hepatic fibrogenesis and fibrosis resolution. J Hepatol 56: 1417-1419, 2012. 231. Ramachandran P, Pellicoro A, Vernon MA, Boulter L, Aucott RL, Ali A, Hartland SN, Snowdon VK, Cappon A, Gordon-Walker TT, Williams MJ, Dunbar DR, Manning JR, van Rooijen N, Fallowfield JA, Forbes SJ, Iredale JP. Differential Ly-6C expression identifies the recruited macrophage phenotype, which orchestrates the regression of murine liver fibrosis. Proc Natl Acad Sci U S A 109: E3186-E3195, 2012. 232. Ramadori G, Neubauer K, Odenthal M, Nakamura T, Knittel T, Schwogler S, Meyer zum Buschenfelde KH. The gene of hepatocyte growth factor is expressed in fat-storing cells of rat liver and is downregulated during cell growth and by transforming growth factor-beta. BBRC 183: 739-742, 1992. 233. Ramadori G, Rieder H, Theiss F, Meyer zum Buschenfelde KH. Fatstoring (Ito) cells of rat liver synthesize and secrete apolipoproteins: Comparison with hepatocytes. Gastroenterology 97: 163-172, 1989. 234. Ramadori G, Veit T, Schwogler S, Dienes HP, Knittel T, Rieder H, Meyer zum Buschenfelde KH. Expression of the gene of the alpha smooth muscle alpha actin isoform in rat liver and in rat fat-storing (ITO) cells. Virchows Arch B Cell Pathol Incl Mol Pathol 59: 349-357, 1990. 235. Ramm GA, Britton RS, O’Neill R, Blaner WS, Bacon BR. Vitamin Apoor lipocytes: A novel desmin-negative lipocyte subpopulation, which can be activated to myofibroblasts. Am J Physiol 269: G532-G541, 1995. 236. Rashid ST, Humphries JD, Byron A, Dhar A, Askari JA, Selley JN, Knight D, Goldin RD, Thursz M, Humphries MJ. Proteomic analysis of extracellular matrix from the hepatic stellate cell line LX-2 identifies CYR61 and Wnt-5a as novel constituents of fibrotic liver. J Proteome Res 11: 4052-4064, 2012. 237. Reynaert H, Thompson MG, Thomas T, Geerts A. Hepatic stellate cells: Role in microcirculation and pathophysiology of portal hypertension. Gut 50: 571-581, 2002. 238. Rippe RA. Role of transcriptional factors in stellate cell activation. Alcohol Clin Exp Res 23: 926-929, 1999. 239. Rockey DC. Hepatic blood flow regulation by stellate cells in normal and injured liver. Semin Liver Dis 21: 337-350, 2001. 240. Rockey DC. Vascular mediators in the injured liver. Hepatology 37: 4-12, 2003. 241. Rockey DC, Boyles JK, Gabbiani G, Friedman SL. Rat hepatic lipocytes express smooth muscle actin upon activation in vivo and in culture. J Submicrosc Cytol Pathol 24: 193-203, 1992. 242. Rockey DC, Fouassier L, Chung JJ, Carayon A, Vallee P, Rey C, Housset C. Cellular localization of endothelin-1 and increased production in liver injury in the rat: Potential for autocrine and paracrine effects on stellate cells. Hepatology 27: 472-480, 1998.

1490


243. Roderburg C, Luedde M, Cardenas DV, Vucur M, Mollnow T, Zimmermann HW, Koch A, Hellerbrand C, Weiskirchen R, Frey N, Tacke F, Trautwein C, Luedde T. miR-133a mediates TGF-beta-dependent de-repression of collagen-synthesis in hepatic stellate cells during liver fibrosis. J Hepatol 58: 736-742, 2012. 244. Roderburg C, Urban GW, Bettermann K, Vucur M, Zimmermann H, Schmidt S, Janssen J, Koppe C, Knolle P, Castoldi M, Tacke F, Trautwein C, Luedde T. Micro-RNA profiling reveals a role for miR29 in human and murine liver fibrosis. Hepatology 53: 209-218, 2011. 245. Rojkind M, Giambrone MA, Biempica L. Collagen types in normal and cirrhotic liver. Gastroenterology 76: 710-719, 1979. 246. Rombouts K, Mello T, Liotta F, Galli A, Caligiuri A, Annunziato F, Pinzani M. MARCKS actin-binding capacity mediates actin filament assembly during mitosis in human hepatic stellate cells. Am J Physiol Cell Physiol 303: C357-C367, 2012. 247. Rosenbaum J, Blazejewski S, Preaux AM, Mallat A, Dhumeaux D, Mavier P. Fibroblast growth factor 2 and transforming growth factor beta 1 interactions in human liver myofibroblasts. Gastroenterology 109: 1986-1996, 1995. 248. Roskams T. Different types of liver progenitor cells and their niches. J Hepatol 45: 1-4, 2006. 249. Roskams T, Cassiman D, De Vos R, Libbrecht L. Neuroregulation of the neuroendocrine compartment of the liver. Anat Rec A Discov Mol Cell Evol Biol 280: 910-923, 2004. 250. Sahin H, Trautwein C, Wasmuth HE. Functional role of chemokines in liver disease models. Nat Rev Gastroenterol Hepatol 7: 682-690, 2010. 251. Saile B, Matthes N, Knittel T, Ramadori G. Transforming growth factor beta and tumor necrosis factor alpha inhibit both apoptosis and proliferation of activated rat hepatic stellate cells. Hepatology 30: 196-202, 1999. 252. Sato M, Kojima N, Miura M, Imai K, Senoo H. Induction of cellular processes containing collagenase and retinoid by integrin-binding to interstitial collagen in hepatic stellate cell culture. Cell Biol Int 22: 115-125, 1998. 253. Sauvant P, Cansell M, Atgie C. Vitamin A and lipid metabolism: Relationship between hepatic stellate cells (HSCs) and adipocytes. J Physiol Biochem 67: 487-496, 2011. 254. Sawitza I, Kordes C, Reister S, Haussinger D. The niche of stellate cells within rat liver. Hepatology 50: 1617-1624, 2009. 255. Schildberg FA, Wojtalla A, Siegmund SV, Endl E, Diehl L, Abdullah Z, Kurts C, Knolle PA. Murine hepatic stellate cells veto CD8 T cell activation by a CD54-dependent mechanism. Hepatology 54: 262-272, 2011. 256. Schirmacher P, Geerts A, Pietrangelo A, Dienes HP, Rogler CE. Hepatocyte growth factor/hepatopoietin A is expressed in fat-storing cells from rat liver but not myofibroblast-like cells derived from fat-storing cells. Hepatology 15: 5-11, 1992. 257. Schmitt-Graff A, Desmouliere A, Gabbiani G. Heterogeneity of myofibroblast phenotypic features: An example of fibroblastic cell plasticity. Virchows Archiv 425: 3-24, 1994. 258. Schnabl B, Purbeck CA, Choi YH, Hagedorn CH, Brenner D. Replicative senescence of activated human hepatic stellate cells is accompanied by a pronounced inflammatory but less fibrogenic phenotype. Hepatology 37: 653-664, 2003. 259. Schuppan D. Structure of the extracellular matrix in normal and fibrotic liver: Collagens and glycoproteins. Semin Liver Dis 10: 1-10, 1990. 260. Schwabe RF, Bataller R, Brenner DA. Human hepatic stellate cells express CCR5 and RANTES to induce proliferation and migration. Am J Physiol Gastrointest Liver Physiol 285: G949-G958, 2003. 261. Schwettmann L, Wehmeier M, Jokovic D, Aleksandrova K, Brand K, Manns MP, Lichtinghagen R, Bahr MJ. Hepatic expression of A disintegrin and metalloproteinase (ADAM) and ADAMs with thrombospondin motives (ADAM-TS) enzymes in patients with chronic liver diseases. J Hepatol 49: 243-250, 2008. 262. Seki E, De Minicis S, Gwak GY, Kluwe J, Inokuchi S, Bursill CA, Llovet JM, Brenner DA, Schwabe RF. CCR1 and CCR5 promote hepatic fibrosis in mice. J Clin Invest 119: 1858-1870, 2009. 263. Seki E, de Minicis S, Inokuchi S, Taura K, Miyai K, van Rooijen N, Schwabe RF, Brenner DA. CCR2 promotes hepatic fibrosis in mice. Hepatology 50: 185-197, 2009. 264. Seki E, De Minicis S, Osterreicher CH, Kluwe J, Osawa Y, Brenner DA, Schwabe RF. TLR4 enhances TGF-beta signaling and hepatic fibrosis. Nat Med 13: 1324-1332, 2007. 265. Sekiya Y, Ogawa T, Iizuka M, Yoshizato K, Ikeda K, Kawada N. Downregulation of cyclin E1 expression by microRNA-195 accounts for interferon-beta-induced inhibition of hepatic stellate cell proliferation. J Cell Physiol 226: 2535-2542, 2011. 266. Sekiya Y, Ogawa T, Yoshizato K, Ikeda K, Kawada N. Suppression of hepatic stellate cell activation by microRNA-29b. Biochem Biophys Res Commun 412: 74-79, 2011. 267. Senoo H, Yoshikawa K, Morii M, Miura M, Imai K, Mezaki Y. Hepatic stellate cell (vitamin A-storing cell) and its relative – past, present and future. Cell Biol Int 34: 1247-1272, 2010.



268. Seok J, Warren HS, Cuenca AG, Mindrinos MN, Baker HV, Xu W, Richards DR, McDonald-Smith GP, Gao H, Hennessy L, Finnerty CC, Lopez CM, Honari S, Moore EE, Minei JP, Cuschieri J, Bankey PE, Johnson JL, Sperry J, Nathens AB, Billiar TR, West MA, Jeschke MG, Klein MB, Gamelli RL, Gibran NS, Brownstein BH, Miller-Graziano C, Calvano SE, Mason PH, Cobb JP, Rahme LG, Lowry SF, Maier RV, Moldawer LL, Herndon DN, Davis RW, Xiao W, Tompkins RG. Genomic responses in mouse models poorly mimic human inflammatory diseases. Proc Natl Acad Sci U S A 110: 3507-3512, 2013. 269. Shafiei MS, Rockey DC. The function of integrin-linked kinase in normal and activated stellate cells: Implications for fibrogenesis in wound healing. Lab Invest 92: 305-316, 2012. 270. Shao R, Yan W, Rockey DC. Regulation of endothelin-1 synthesis by endothelin-converting enzyme-1 during wound healing. J Biol Chem 274: 3228-3234, 1999. 271. Shi M, Zhu J, Wang R, Chen X, Mi L, Walz T, Springer TA. Latent TGF-beta structure and activation. Nature 474: 343-349, 2011. 272. Shirakami Y, Gottesman ME, Blaner WS. Diethylnitrosamine-induced hepatocarcinogenesis is suppressed in lecithin:retinol acyltransferasedeficient mice primarily through retinoid actions immediately after carcinogen administration. Carcinogenesis 33: 268-274, 2012. 273. Shomron N, Hamasaki-Katagiri N, Hunt R, Hershko K, Pommier E, Geetha S, Blaisdell A, Dobkin A, Marple A, Roma I, Newell J, Allen C, Friedman S, Kimchi-Sarfaty C. A splice variant of ADAMTS13 is expressed in human hepatic stellate cells and cancerous tissues. Thromb Haemost 104: 531-535, 2010. 274. Singh R, Kaushik S, Wang Y, Xiang Y, Novak I, Komatsu M, Tanaka K, Cuervo AM, Czaja MJ. Autophagy regulates lipid metabolism. Nature 458: 1131-1135, 2009. 275. Smedsrod B, Le Couteur D, Ikejima K, Jaeschke H, Kawada N, Naito M, Knolle P, Nagy L, Senoo H, Vidal-Vanaclocha F, Yamaguchi N. Hepatic sinusoidal cells in health and disease: Update from the 14th International Symposium. Liver Int 29: 490-501, 2009. 276. Sohail MA, Hashmi AZ, Hakim W, Watanabe A, Zipprich A, Groszmann RJ, Dranoff JA, Torok NJ, Mehal WZ. Adenosine induces loss of actin stress fibers and inhibits contraction in hepatic stellate cells via Rho inhibition. Hepatology 49: 185-194, 2009. 277. Sprenger H, Kaufmann A, Garn H, Lahme B, Gemsa D, Gressner AM. Differential expression of monocyte chemotactic protein-1 (MCP-1) in transforming rat hepatic stellate cells. J Hepatol 30: 88-94, 1999. 278. Starkel P, Sempoux C, Leclercq I, Herin M, Deby C, Desager JP, Horsmans Y. Oxidative stress, KLF6 and transforming growth factor-beta up-regulation differentiate non-alcoholic steatohepatitis progressing to fibrosis from uncomplicated steatosis in rats. J Hepatol 39: 538-546, 2003. 279. Stefanovic B, Hellerbrand C, Brenner DA. Regulatory role of the conserved stem-loop structure at the 5 end of collagen alpha1(I) mRNA. Mol Cell Biol 19: 4334-4342, 1999. 280. Stefanovic B, Hellerbrand C, Holcik M, Briendl M, Aliebhaber S, Brenner DA. Posttranscriptional regulation of collagen alpha1(I) mRNA in hepatic stellate cells. Mol Cell Biol 17: 5201-5209, 1997. 281. Stefanovic L, Brenner DA, Stefanovic B. Direct hepatotoxic effect of KC chemokine in the liver without infiltration of neutrophils. Exp Biol Med (Maywood) 230: 573-586, 2005. 282. Steiling H, Muhlbauer M, Bataille F, Scholmerich J, Werner S, Hellerbrand C. Activated hepatic stellate cells express keratinocyte growth factor in chronic liver disease. Am J Pathol 165: 1233-1241, 2004. 283. Su YH, Shu KH, Hu C, Cheng CH, Wu MJ, Yu TM, Chuang YW, Huang ST, Chen CH. Hepatic stellate cells attenuate the immune response in renal transplant recipients with chronic hepatitis. Transplant Proc 44: 725-729, 2012. 284. Suzuki K, Tanaka M, Watanabe N, Saito S, Nonaka H, Miyajima A. p75 Neurotrophin receptor is a marker for precursors of stellate cells and portal fibroblasts in mouse fetal liver. Gastroenterology 135: 270-281 e273, 2008. 285. Syal G, Fausther M, Dranoff JA. Advances in cholangiocyte immunobiology. Am J Physiol Gastrointest Liver Physiol 303: G1077-G1086, 2012. 286. Szabo G, Mandrekar P, Dolganiuc A. Innate immune response and hepatic inflammation. Semin Liver Dis 27: 339-350, 2007. 287. Tacke F. Monocytes and Macrophages as Cellular Targets in Liver Fibrosis. Inflamm Allergy Drug Targets 8: 307-318, 2009. 288. Tacke F, Weiskirchen R. Update on hepatic stellate cells: Pathogenic role in liver fibrosis and novel isolation techniques. Expert Rev Gastroenterol Hepatol 6: 67-80, 2012. 289. Takahara T, Kojima T, Miyabayashi C, Inoue K, Sasaki H, Muragaki Y, Ooshima A. Collagen production in fat-storing cells after carbon tetrachloride intoxication in the rat. Immunoelectron microscopic observation of type I, type III collagens, and prolyl hydroxylase. Lab Invest 59: 509-521, 1988. 290. Tanaka H, Leung PS, Kenny TP, Gershwin ME, Bowlus CL. Immunological orchestration of liver fibrosis. Clin Rev Allergy Immunol 43: 220-229, 2012.



291. Tarrats N, Moles A, Morales A, Garcia-Ruiz C, Fernandez-Checa JC, Mari M. Critical role of tumor necrosis factor receptor 1, but not 2, in hepatic stellate cell proliferation, extracellular matrix remodeling, and liver fibrogenesis. Hepatology 54: 319-327, 2011. 292. Taura K, De Minicis S, Seki E, Hatano E, Iwaisako K, Osterreicher CH, Kodama Y, Miura K, Ikai I, Uemoto S, Brenner DA. Hepatic stellate cells secrete angiopoietin 1 that induces angiogenesis in liver fibrosis. Gastroenterology 135: 1729-1738, 2008. 293. Thabut D, Routray C, Lomberk G, Shergill U, Glaser K, Huebert R, Patel L, Masyuk T, Blechacz B, Vercnocke A, Ritman E, Ehman R, Urrutia R, Shah V. Complementary vascular and matrix regulatory pathways underlie the beneficial mechanism of action of sorafenib in liver fibrosis. Hepatology 54: 573-585, 2011. 294. Troeger JS, Mederacke I, Gwak GY, Dapito DH, Mu X, Hsu CC, Pradere JP, Friedman RA, Schwabe RF. Deactivation of hepatic stellate cells during liver fibrosis resolution in mice. Gastroenterology 143: 1073-1083 e1022, 2012. 295. Tsukada S, Parsons CJ, Rippe RA. Mechanisms of liver fibrosis. Clin Chim Acta 364: 33-60, 2006. 296. Tsukada S, Westwick JK, Ikejima K, Sato N, Rippe RA. SMAD and p38 MAPK signaling pathways independently regulate alpha1(I) collagen gene expression in unstimulated and transforming growth factor-betastimulated hepatic stellate cells. J Biol Chem 280: 10055-10064, 2005. 297. Tsukamoto H. Epigenetic mechanism of stellate cell transdifferentiation. J Hepatol 46: 352-353, 2007. 298. Tsukamoto H, Zhu NL, Wang J, Asahina K, Machida K. Morphogens and hepatic stellate cell fate regulation in chronic liver disease. J Gastroenterol Hepatol 27 Suppl 2: 94-98, 2012. 299. Ueno T, Sata M, Sakata R, Torimura T, Sakamoto M, Sugawara H, Tanikawa K. Hepatic stellate cells and intralobular innervation in human liver cirrhosis. Hum Pathol 28: 953-959, 1997. 300. Urtasun R, Lopategi A, George J, Leung TM, Lu Y, Wang X, Ge X, Fiel MI, Nieto N. Osteopontin, an oxidant stress sensitive cytokine, up-regulates collagen-I via integrin alpha(V)beta(3) engagement and PI3K/pAkt/NFkappaB signaling. Hepatology 55: 594-608, 2012. 301. Vinas O, Bataller R, Sancho-Bru P, Gines P, Berenguer C, Enrich C, Nicolas JM, Ercilla G, Gallart T, Vives J, Arroyo V, Rodes J. Human hepatic stellate cells show features of antigen-presenting cells and stimulate lymphocyte proliferation. Hepatology 38: 919-929, 2003. 302. Vyas SK, Leyland H, Gentry J, Arthur MJ. Rat hepatic lipocytes synthesize and secrete transin (stromelysin) in early primary culture. Gastroenterology 109: 889-898, 1995. 303. Wake K. “Sternzellen” in the liver: Perisinuosoidal cells with special reference to storage of vitamin A. Am J Anat 132: 429-462, 1971. 304. Wake K. Perisinusoidal stellate cells (Fat-storing cells, interstitial cells, lipocytes) their related structure in and around liver sinusoids, and vitamin A storing cells in extrahepatic organs. Int Rev Cytology 66: 303-353, 1980. 305. Wake K, Sato T. Intralobular heterogeneity of perisinusoidal stellate cells in porcine liver. Cell Tissue Res 273: 227-237, 1993. 306. Wandzioch E, Kolterud A, Jacobsson M, Friedman SL, Carlsson L. Lhx2-/- mice develop liver fibrosis. Proc Natl Acad Sci U S A 101: 16549-16554, 2004. 307. Wang B, Li W, Guo K, Xiao Y, Wang Y, Fan J. miR-181b Promotes hepatic stellate cells proliferation by targeting p27 and is elevated in the serum of cirrhosis patients. Biochem Biophys Res Commun 421: 4-8, 2012. 308. Wang J, Leclercq I, Brymora JM, Xu N, Ramezani-Moghadam M, London RM, Brigstock D, George J. Kupffer cells mediate leptininduced liver fibrosis. Gastroenterology 137: 713-723, 2009. 309. Wang L, Wang X, Chiu JD, van de Ven G, Gaarde WA, Deleve LD. Hepatic vascular endothelial growth factor regulates recruitment of rat liver sinusoidal endothelial cell progenitor cells. Gastroenterology 143: 1555-1563 e1552, 2012. 310. Wang L, Wang X, Xie G, Hill CK, DeLeve LD. Liver sinusoidal endothelial cell progenitor cells promote liver regeneration in rats. J Clin Invest 122: 1567-1573, 2012. 311. Wang Y, Yao HL, Cui CB, Wauthier E, Barbier C, Costello MJ, Moss N, Yamauchi M, Sricholpech M, Gerber D, Loboa EG, Reid LM. Paracrine signals from mesenchymal cell populations govern the expansion and differentiation of human hepatic stem cells to adult liver fates. Hepatology 52: 1443-1454, 2010. 312. Wang Y, Zhang JS, Qian J, Huang GC, Chen Q. Adrenomedullin regulates expressions of transforming growth factor-beta1 and beta1induced matrix metalloproteinase-2 in hepatic stellate cells. Int J Exp Pathol 87: 177-184, 2006. 313. Watanabe A, Sohail MA, Gomes DA, Hashmi A, Nagata J, Sutterwala FS, Mahmood S, Jhandier MN, Shi Y, Flavell RA, Mehal WZ. Inflammasome-mediated regulation of hepatic stellate cells. Am J Physiol Gastrointest Liver Physiol 296: G1248-G1257, 2009. 314. Watanabe N, Ikeda H, Kume Y, Satoh Y, Kaneko M, Takai D, Tejima K, Nagamine M, Mashima H, Tomiya T, Noiri E, Omata M, Matsumoto M, Fujimura Y, Yatomi Y. Increased production of ADAMTS13 in hepatic

1491


315. 316.

317.

318.

319.

320. 321. 322. 323.

324.

325.

stellate cells contributes to enhanced plasma ADAMTS13 activity in rat models of cholestasis and steatohepatitis. Thromb Haemost 102: 389-396, 2009. Wells RG. The role of matrix stiffness in hepatic stellate cell activation and liver fibrosis. J Clin Gastroenterol 39: S158-S161, 2005. wIshikawa K, Mochida S, Mashiba S, Inao M, Matsui A, Ikeda H, Ohno A, Shibuya M, Fujiwara K. Expressions of vascular endothelial growth factor in nonparenchymal as well as parenchymal cells in rat liver after necrosis. Biochem Biophys Res Commun 254: 587-593, 1999. Wong L, Yamasaki G, Johnson RJ, Friedman SL. Induction of betaplatelet-derived growth factor receptor in rat hepatic lipocytes during cellular activation in vivo and in culture. J Clin Invest 94: 1563-1569, 1994. Wu TJ, Wang YC, Wu TH, Lee CF, Chan KM, Lee WC. Inhibition of allogenic T-cell cytotoxicity by hepatic stellate cell via CD4(+) CD25(+) Foxp3(+) regulatory T cells in vitro. Transplant Proc 44: 1055-1059, 2012. Xie G, Wang X, Wang L, Atkinson RD, Kanel GC, Gaarde WA, Deleve LD. Role of differentiation of liver sinusoidal endothelial cells in progression and regression of hepatic fibrosis in rats. Gastroenterology 142: 918-927 e916, 2012. Yamada M, Blaner WS, Soprano DR, Dixon JL, Kjeldbye HM, Goodman DS. Biochemical characteristics of isolated rat liver stellate cells. Hepatology 7: 1224-1229, 1987. Yamada T, Imaoka S, Kawada N, Seki S, Kuroki T, Kobayashi K, Monna T, Funae Y. Expression of cytochrome P450 isoforms in rat hepatic stellate cells. Life Sci 61: 171-179, 1997. Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayashi M, Mitsui Y, Yazaki Y, Goto K, Masaki T. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature 332: 411-415, 1988. Yang C, Zeisberg M, Mosterman B, Sudhakar A, Yerramalla U, Holthaus K, Xu L, Eng F, Afdhal N, Kalluri R. Liver fibrosis: Insights into migration of hepatic stellate cells in response to extracellular matrix and growth factors. Gastroenterology 124: 147-159, 2003. Yang L, Jung Y, Omenetti A, Witek RP, Choi S, Vandongen HM, Huang J, Alpini GD, Diehl AM. Fate-mapping evidence that hepatic stellate cells are epithelial progenitors in adult mouse livers. Stem Cells 26: 2104-2113, 2008. Yin C, Evason KJ, Maher JJ, Stainier DY. The basic helix-loop-helix transcription factor, heart and neural crest derivatives expressed transcript 2, marks hepatic stellate cells in zebrafish: Analysis of stellate cell entry into the developing liver. Hepatology 56: 1958-1970, 2012.

1492


326. Yin L, Lynch D, Ilic Z, Sell S. Proliferation and differentiation of ductular progenitor cells and littoral cells during the regeneration of the rat liver to CCl4/2-AAF injury. Histol Histopathol 17: 65-81, 2002. 327. Yoshiji H, Kuriyama S, Yoshii J, Ikenaka Y, Noguchi R, Hicklin DJ, Wu Y, Yanase K, Namisaki T, Yamazaki M, Tsujinoue H, Imazu H, Masaki T, Fukui H. Vascular endothelial growth factor and receptor interaction is a prerequisite for murine hepatic fibrogenesis. Gut 52: 1347-1354, 2003. 328. Yu C, Wang F, Jin C, Huang X, Miller DL, Basilico C, McKeehan WL. Role of fibroblast growth factor type 1 and 2 in carbon tetrachlorideinduced hepatic injury and fibrogenesis. Am J Pathol 163: 1653-1662, 2003. 329. Zhao Y, Wang Y, Wang Q, Liu Z, Liu Q, Deng X. Hepatic stellate cells produce vascular endothelial growth factor via phospho-p44/42 mitogen-activated protein kinase/cyclooxygenase-2 pathway. Mol Cell Biochem 359: 217-223, 2012. 330. Zhu NL, Asahina K, Wang J, Ueno A, Lazaro R, Miyaoka Y, Miyajima A, Tsukamoto H. Hepatic stellate cell-derived delta-like homolog 1 (DLK1) protein in liver regeneration. J Biol Chem 287: 10355-10367, 2012. 331. Zhu Q, Zou L, Jagavelu K, Simonetto DA, Huebert RC, Jiang ZD, DuPont HL, Shah VH. Intestinal decontamination inhibits TLR4 dependent fibronectin-mediated cross-talk between stellate cells and endothelial cells in liver fibrosis in mice. J Hepatol 56: 893-899, 2012. 332. Zindy F, Lamas E, Schmidt S, Kirn A, Brechot C. Expression of insulinlike growth factor II (IGF-II) and IGF-II, IGF-I and insulin receptors mRNAs in isolated non-parenchymal rat liver cells. J Hepatol 14: 3034, 1992. 333. Zvibel I, Atias D, Phillips A, Halpern Z, Oren R. Thyroid hormones induce activation of rat hepatic stellate cells through increased expression of p75 neurotrophin receptor and direct activation of Rho. Lab Invest 90: 674-684, 2010.

Further Reading Friedman SL, Sheppard D, Duffield J and Violette S. Therapy for fibrotic diseases: Nearing the starting line. Sci Transl Med, 5:167sr1, 2013. Friedman SL, guest editor. Special issue on fibrosis. Biochim Biophys Acta. 2013 Mar 16. pii: S0925-4439(13)00077-X. doi: 10.1016/j.bbadis.


R-spondin2 activates hepatic stellate cells and promotes liver fibrosis.

Targeting activated hepatic stellate cells (aHSCs) for liver fibrosis imaging.

Levels of hepatic Th17 cells and regulatory T cells upregulated by hepatic stellate cells in advanced HBV-related liver fibrosis.

Apamin suppresses biliary fibrosis and activation of hepatic stellate cells.

Berberine Inhibition of Fibrogenesis in a Rat Model of Liver Fibrosis and in Hepatic Stellate Cells.

Curcumin attenuates angiogenesis in liver fibrosis and inhibits angiogenic properties of hepatic stellate cells.

Inhibitory effects of capsaicin on hepatic stellate cells and liver fibrosis.

The role of miRNAs in stress-responsive hepatic stellate cells during liver fibrosis.

PTP1B confers liver fibrosis by regulating the activation of hepatic stellate cells.

NF-κB inhibition alleviates carbon tetrachloride-induced liver fibrosis via suppression of activated hepatic stellate cells.

Acyl-CoA:cholesterol acyltransferase 1 mediates liver fibrosis by regulating free cholesterol accumulation in hepatic stellate cells.

Isolation and culture of hepatic stellate cells from mouse liver.

Osthole ameliorates hepatic fibrosis and inhibits hepatic stellate cell activation.

Bone Marrow Stem Cells Anti-liver Fibrosis Potency: Inhibition of Hepatic Stellate Cells Activity and Extracellular Matrix Deposition.

Human Menstrual Blood-Derived Stem Cells Ameliorate Liver Fibrosis in Mice by Targeting Hepatic Stellate Cells via Paracrine Mediators.

Inhibition of ASICs reduces rat hepatic stellate cells activity and liver fibrosis: an in vitro and in vivo study.

Fibronectin peptides as potential regulators of hepatic fibrosis through apoptosis of hepatic stellate cells.

TAZ degradation.

Liver fibrosis and protection mechanisms action of medicinal plants targeting apoptosis of hepatocytes and hepatic stellate cells.

Novel matrine derivative MD-1 attenuates hepatic fibrosis by inhibiting EGFR activation of hepatic stellate cells.

Lyn kinase enhanced hepatic fibrosis by modulating the activation of hepatic stellate cells.

Focal Adhesion Kinase Regulates Hepatic Stellate Cell Activation and Liver Fibrosis.

Suppressive effect of SATB1 on hepatic stellate cell activation and liver fibrosis in rats.

Secreted frizzled-related protein 5 (Sfrp5) decreases hepatic stellate cell activation and liver fibrosis.