LIVER TRANSPLANTATION 20:376–385, 2014

ORIGINAL ARTICLE

Heat Shock Protein 70 Is Required for Optimal Liver Regeneration After Partial Hepatectomy in Mice Joshua H. Wolf,1 Tricia R. Bhatti,2 Suomi Fouraschen,1 Shourjo Chakravorty,1 Liqing Wang,2 Sunil Kurian,3 Daniel Salomon,3 Kim M. Olthoff,1 Wayne W. Hancock,2 and Matthew H. Levine1 1 Department of Surgery, University of Pennsylvania School of Medicine, Philadelphia, PA; 2Division of Transplant Immunology, Department of Pathology and Laboratory Medicine, Children’s Hospital of Philadelphia/University of Pennsylvania School of Medicine, Philadelphia, PA; and 3Scripps Research Institute, La Jolla, CA

Liver regeneration is a complex process that restores functional tissue after resection or injury, and it is accompanied by transient adenosine triphosphate depletion and metabolic stress in hepatic parenchymal cells. Heat shock protein 70 (Hsp70) functions as a chaperone during periods of cellular stress and induces the expression of several inflammatory cytokines identified as key players during early liver regeneration. We, therefore, hypothesized that Hsp70 is required for the initiation of regeneration. Investigations were carried out in a 70% partial hepatectomy mouse model with mice lacking inducible Hsp70 (Hsp702/2). Liver regeneration was assessed postoperatively with the liver weight/body weight (LW/BW) ratio, and sera and tissues were collected for analysis. In addition, the expression of Hsp-related genes was assessed in a cohort of 23 human living donor liver transplantation donors. In mice, the absence of Hsp70 was associated with a reduced postoperative LW/BW ratio, Ki-67 staining, and tumor necrosis factor a (TNF-a) expression in comparison with wild-type mice. TNF-a expression was also reduced in livers from Hsp702/2 mice after induction with lipopolysaccharide (1 mg/kg). Clinically, the transcription of multiple Hsp genes (especially Hsp70 family members) was up-regulated after donor hepatectomy. Together, these results suggest that the early phase of successful liver regeneration requires the presence of Hsp70 to induce TNF-a. Further studies are required to determine whether Hsp70 contributes to liver regeneration as a chaperone by stabilizing specific interactions required for growth signaling or as a paracrine inflammatory signal, as can occur in modC 2013 AASLD. els of shock. Liver Transpl 20:376-385, 2014. V Received June 10, 2013; accepted November 21, 2013. Liver regeneration is a biological response to hepatocellular injury or loss involving a complex network of inflammatory, proliferative, and metabolic signals.1 After partial hepatectomy (PH), 75% to 95% of hepato-

cytes in a regenerating rodent liver undergo multiple rounds of mitosis, and this results in complete parenchymal restoration within a period of approximately 7 days.2 This leads to a large energy demand in the

Abbreviations: 17-DMAG, 17-dimethylaminoethylamino-17-demethoxygeldanamycin; ALT, alanine aminotransferase; ATP, adenosine triphosphate; AUC, area under the curve; CXCL, chemokine (C-X-C motif) ligand; DC, dendritic cell; DNAJ, DnaJ (heat shock protein 40) homolog; GRO-(a), growth regulated protein (a); HGF, hepatocyte growth factor; Hsp, heat shock protein; Hsp70-KO, heat shock protein 70–knockout; IL, interleukin; KC, keratinocyte chemoattractant; LDLT, living donor liver transplantation; LPS, lipopolysaccharide; LW/BW, liver weight/body weight; Ly6C, lymphocyte antigen 6c; mRNA, messenger RNA; NS, not significant; PH, partial hepatectomy; POD, postoperative day; POST biopsy sample, left lobe sample after hepatectomy and just before abdominal closure; PRE biopsy sample, right lobe sample before hepatectomy; qPCR, quantitative polymerase chain reaction; RT-PCR, reverse-transcription polymerase chain reaction; TNF-a, tumor necrosis factor a; WT, wild type. This study was supported by funds from the Biesecker Center of the Children’s Hospital of Philadelphia (to Wayne W. Hancock and Kim M. Olthoff) and by the National Institute of Diabetes and Digestive and Kidney Diseases (grant 5-U01-AI-063589-05 to Kim M. Olthoff). Address reprint requests to Wayne W. Hancock, M.B.B.S., Ph.D., Division of Transplant Immunology, Department of Pathology and Laboratory Medicine, Children’s Hospital of Philadelphia, 3615 Civic Center Boulevard, Philadelphia PA 19104. Telephone: 215-590-8709; FAX: 215-5907384; E-mail: [email protected] DOI 10.1002/lt.23813 View this article online at wileyonlinelibrary.com. LIVER TRANSPLANTATION.DOI 10.1002/lt. Published on behalf of the American Association for the Study of Liver Diseases

C 2013 American Association for the Study of Liver Diseases. V

LIVER TRANSPLANTATION, Vol. 20, No. 3, 2014

growing liver tissue because hepatocytes must suddenly balance available resources between synthesis/ proliferation and the maintenance of metabolic homeostasis. Transient signs of metabolic stress after PH have been observed in animal studies; these include the depletion of available adenosine triphosphate (ATP; a decline in the ATP to adenosine diphosphate ratio), mitochondrial swelling, increased mitochondrial membrane permeability, and a decreased adenylate energy charge.3-7 These changes can be extremely rapid (as early as 30 seconds after PH) and can persist for several days after surgery. The heat shock protein 70 (Hsp70) family consists of a group of related molecular chaperones that respond to cellular stress, but its link to the metabolic changes during early liver regeneration is relatively unexplored. The final common pathway for the various sources of cellular stress that activate Hsp70 (including temperature, hypoxia, acidosis, and starvation) involves ATP depletion, which leads to the denaturing of folded intracellular proteins, the formation of disordered protein aggregates, and compromised cell viability. Inducible Hsp70 chaperone activity enables the cell to cope with the increased burden of misfolded proteins by a direct bind-and-release mechanism that promotes refolding. This mechanism relies on an intrinsic adenosine triphosphatase at the Hsp70 N-terminus, whose activity is catalyzed by other heat shock–responsive components, including members of the Hsp40 family and nuclear exchange factors. Combinatorial interactions with various Hsp40 proteins and nuclear exchange factors provide for a large range of Hsp70 substrate specificities.8,9 Mounting evidence suggests that Hsp70 is also released from stressed cells as an extracellular protein that can serve as a paracrine signal. Extracellular Hsp70 can stimulate innate immune mechanisms by promoting the expression of tumor necrosis factor a (TNF-a) and interleukin-6 (IL-6) and downstream nuclear factor kappa B signaling, all of which are biologically important components of early liver regeneration.1,10,11 We, therefore, hypothesized that the Hsp70 stress response is activated immediately after hepatectomy by transient metabolic stress, and this induction leads to the production of inflammatory cytokines critical to liver regeneration, such as TNF-a and IL-6. Hence, the Hsp70 stress response serves as an essential early trigger for liver regeneration. This study tested whether inducible Hsp70 is required for liver regeneration in a PH mouse model and whether there is a clinical correlation with HSP gene expression in humans after the related procedure of donor right hepatic lobectomy.

MATERIALS AND METHODS Animals Animals housed in the Laboratory Animal Facility of the Children’s Hospital of Philadelphia were studied according to a protocol approved by the Institutional Animal Care and Use Committee. Hsp702/2 mice were originally developed on a 129 background, and they

WOLF ET AL. 377

lacked both inducible forms of Hsp70 (Hsp70.1 and Hsp70.3).12 All control wild-type (WT) 129 mice and Hsp702/2 mice used in our studies were males that were 6 to 8 weeks of age and weighed 20 to 25 g. Hsp702/2 mice were maintained on site as a stable inbred colony, and WT mice were purchased (Jackson Laboratory, Bar Harbor, ME). Both groups were housed in groups of 6 mice per cage and were fed normal ad libitum diets preoperatively and postoperatively.

PH Model To test the role of Hsp70 in liver regeneration, 70% murine PH was performed as described.13 Each mouse was anesthetized with a charcoal-filtered induction chamber and flow meter with a mixture of isoflurane (3%-5%) and supplementary oxygen (2 L/minute). The abdomen was accessed via midline laparotomy, and the bowel and the liver were gently retracted to expose the hila of the left and median lobes, which together accounted for approximately 70% of the murine liver mass. Each hilum was ligated separately with a single 3-0 silk suture and resected. The abdomen was closed with a full-thickness running silk suture, and the mouse was placed in an incubator (37 C) for a brief period of recovery (5-10 minutes). At the designated postoperative endpoints (4, 24, 48, or 96 hours), each mouse was given a lethal dose of pentobarbital, and tissues were harvested. The surgical incision was reopened, and blood was drawn directly from the inferior vena cava with a 28-gauge syringe. The right and caudate lobes were dissected free from surrounding structures, placed into ice-cold phosphate-buffered saline, and weighed on a scale. Once a weight was obtained, the right lobe tissue was divided into 8 to 10 specimens for analysis and partitioned into preservation tubes with RNAlater (Ambion, Austin, TX) or formalin. Plasma was collected for the analysis of alanine aminotransferase (ALT) and total bilirubin through the core pathology facilities (Children’s Hospital of Philadelphia), and plasma cytokines/chemokines were measured with Luminex (Invitrogen, Carlsbad, CA). Frozen samples of liver tissue and plasma were kept at 280 C.

Quantitative Polymerase Chain Reaction (qPCR) RNA purification was performed by mechanical disruption with a rotor-blade homogenizer and lysis/ extraction solutions from the RNeasy kit (Qiagen, Inc., Valencia, CA). Purified RNA was reverse-transcribed with TaqMan reagents (Applied Biosystems, Carlsbad, CA). Complementary DNAs for each specimen were amplified with primers for each gene of interest and normalized against expression levels of 18S. Results were analyzed with a StepOnePlus 96-well plate reader (Life Technologies, Grand Island, NY).

Endotoxin Model Male WT mice and Hsp702/2 mice, 6 to 8 weeks old, were evaluated at the baseline and after the injection

378 WOLF ET AL.

of lipopolysaccharide (LPS; 1 mg/kg intraperitoneally) with 3 mice per group. Readouts included qPCR detection of cytokine messenger RNA (mRNA) and flow cytometry analysis of liver and spleen samples, and the flow cytometry of macrophage surface markers and TNF-a was performed with monoclonal antibodies purchased from BD Biosciences.

Immunohistochemistry and Image Analysis Formalin-fixed, paraffin-embedded liver sections were stained with hematoxylin and eosin. Additional sections were treated in a pressure cooker with a citrate buffer and incubated with an antibody to Ki-67 (AB1667, Abcam; 1:400) overnight at 4 C; this was followed by an avidin-biotin complex (PK-6100, Vector Laboratories). Immunostained slides were scanned with the Aperio ScanScope CS slide scanner (Aperio Technologies, Vista, CA). Digitized images were analyzed with Aperio ImageScope software (version 10.0.1346.1807, Aperio Technologies) for the determination of the percentage of cells with nuclear positivity among the total number of cells present on the slide. Cells with 21 or 31 intensity of staining were considered positive.

Acquisition of Patient Samples Twenty-three living donor liver transplantation (LDLT) donors underwent right lobe hepatectomy at 3 US transplant centers between 2006 and 2009 as part of a study ancillary to the Adult-to-Adult Living Donor Liver Transplantation Cohort Study (Genomics and Regeneration in the Transplant Setting).14 Institutional review board approval was obtained at each participating institution (University of Pennsylvania, Columbia University, and Northwestern University) before the investigation. Two biopsy samples were obtained: (1) a right lobe sample before hepatectomy (PRE biopsy sample) and (2) a left lobe sample after hepatectomy and just before abdominal closure (POST biopsy sample). The time between the acquisition of the 2 samples was approximately 2 to 3 hours.

Luminex Analysis Sera were assayed with a Luminex 100 array reader (Luminex Corp., Austin, TX) at 5 different times (0, 4, 24, 48, and 96 hours). The 20 analytes were IL-1a, IL1b, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, IL-13, IL-17, TNF-a, granulocyte-macrophage colony stimulating factor, interferon-g, interferon-inducible protein 10 [also known as chemokine (C-X-C motif) ligand (CXCL10)], KC (also known as GRO-a and CXCL1), monokine induced by interferon-g (also known as CXCL9), monocyte chemoattractant protein 1 [chemokine (C-C motif) ligand 2], macrophage inflammatory protein 1a [also known as chemokine (C-C motif) ligand 3], fibroblast growth factor, and vascular endothelial growth factor.

LIVER TRANSPLANTATION, March 2014

RNA Purification and Microarray Analysis of Human Liver Biopsies Total RNA was extracted from the PRE and POST biopsy samples with TRIzol (Invitrogen); afterward, the RNA was further purified with the RNeasy kit (Qiagen) according to the manufacturer’s instructions. Biotinylated complementary RNA was prepared with the Ambion MessageAmp Biotin II kit; afterward, labeled complementary RNA was hybridized to Affymetrix Human Gene 1.0 ST Array GeneChips (Affymetrix, Santa Clara, CA) with standard Affymetrix protocols.

Statistics Gene expression data from Affymetrix GeneChips were analyzed with BRB-ArrayTools software (developed by Dr. Richard Simon and the BRB-ArrayTools Development Team). Normalized signals were generated with RMA; afterward, class comparisons were performed with a paired t test with random variance and with a cutoff of P < 0.001 for significance.15,16 Differentially expressed genes were identified with a false discovery rate < 10% and a P value < 0.001. Liver regeneration in the PH model was assessed with the liver weight/body weight (LW/BW) ratio, and means were compared with the Student t test. The liver remnant in post-operative mice consisted of only caudate and right lobes; we therefore used the weights of these lobes to calculate LW/BW in pre-operative mice. Preoperative results for LW/BW ratios, reverse-transcription polymerase chain reaction (RT-PCR), and immunohistochemistry reflected means from 3 mice. After PH, the sample sizes for LW/BW ratios, RT-PCR, and immunohistochemistry were 6, 8, and 3 mice per time point, respectively, and a P value of 0.05 was deemed significant.

RESULTS Liver Regeneration Is Impaired in Hsp702/2 Mice We studied whether the 2 stress-inducible Hsp70 genes (HSPA1A and HSPA1B) were required for liver recovery and regeneration in a murine PH model. In comparison with WT mice, Hsp702/2 mice did not exhibit any obvious developmental defects, anatomical variations, or differences in lifespan at the baseline. The 2 groups were also similar with respect to their preoperative liver weights, body weights, and LW/BW ratios (Table 1). However, after PH, Hsp702/2 mice had significantly lower LW/BW ratios in comparison with WT mice, and this indicated a reduced capacity for regenerative growth [postoperative day 1 (POD1), 1.5% versus 1.9%, P < 0.001; POD2, 1.9% versus 2.8%, P < 0.001; POD4, 2.6% versus 3.2%, P 5 0.02; Fig. 1). This did not appear to be related to differences in hepatocellular injury or synthetic function because Hsp702/2 mice and WT mice had comparable plasma levels of ALT and total bilirubin at preoperative and postoperative time points (Fig. 2).

Attenuated Liver Growth in Hsp702/2 Mice Is Associated With Reduced Cellular Proliferation To test whether lower LW/BW ratios in Hsp702/2 mice reflected decreased cellular proliferation, sections

LIVER TRANSPLANTATION, Vol. 20, No. 3, 2014

WOLF ET AL. 379

TABLE 1. Baseline Weights for WT and Hsp70-KO Mice WT Total liver (g) Caudate + right lobe (g) Body weight (g) LW/BW ratio (%)

Hsp70-KO P Value

1.10 6 0.04 1.10 6 0.03 0.35 6 0.01 0.33 6 0.01

0.93 0.14

26.2 6 0.6 26.6 6 1.2 1.35 6 0.06 1.27 6 0.04

0.43 0.19

NOTE: The data are presented as means and standard deviations. LW/BW 5 caudate 1 right lobes / body weight.

Figure 1. Reduced liver regeneration after PH in Hsp702/2 mice. PH was performed in WT mice and Hsp702/2 mice, and regeneration was assessed with LW/BW measurements at various postoperative time points. The boxes represent the mean LW/BW ratios at each time point (6 mice per group), and the bars indicate the standard errors. Preoperative LW/BW was derived using weights from caudate and right hepatic lobes only, in order to maintain a consistent comparison to post-operative mice. P values were obtained with unpaired Student t tests for WT mice versus Hsp702/2 mice at each time point.

of livers from Hsp702/2 mice and WT mice were stained for Ki-67, a nuclear marker for cellular proliferation, and were evaluated by quantitative image analysis. Preoperative nuclear Ki-67 expression was the same in both Hsp702/2 mice and WT mice (positive nuclei: 1.7% 6 0.3% versus 1.2% 6 0.1%, P 5 0.17). Peak Ki-67 expression occurred at 48 hours, but it was 20% lower in Hsp702/2 mice versus WT mice (positive nuclei: 48% 6 3% versus 69% 6 5%, P < 0.01). The area under the curve (AUC) across all measured time points was lower for Hsp702/2 mice versus WT mice (60.4 versus 90.5; Fig. 3A). Lower levels of Ki-67 expression in Hsp702/2 mice versus WT mice were also observed at 24 hours in a genotype-blinded review by a pathologist (Fig. 3B), although this difference did not reach statistical significance with automated image analysis (positive nuclei: 1.5% 6 0.4% versus 5.8% 6 2.3%, P 5 0.09). Ki67 expression remained elevated to the same degree in both groups at 96 hours (positive nuclei: 10.6% 6 2.7% versus 11.5% 6 1.5%, P 5 0.69). Parallel sections stained with hematoxylin and eosin 24 and 48 hours after PH showed normal parenchymal architecture with no signs of hepatocellular damage or necrosis, and there were no differences between WT mice and Hsp702/2 mice (Fig. 3C). These studies suggest that

Figure 2. No difference between WT mice and Hsp702/2 mice in the degree of postoperative liver injury or synthetic function. Plasma was collected from WT mice and Hsp702/2 mice and was sent to the institutional core laboratory facility for ALT and total bilirubin testing.

cellular proliferation in mice after PH peaks on POD2 and remains increased through POD4, but this response is blunted when inducible Hsp70 is absent.

Reduced TNF-a Expression in Hsp702/2 Mice After PH Inflammatory cytokines and chemokines play critical roles in the early stages of murine liver regeneration.2 Several such proteins, especially TNF-a and IL-6, are released in response to Hsp70 paracrine activity in other biological contexts.10 We, therefore, questioned whether Hsp70 contributes to the release of cytokines/chemokines during early liver regeneration and whether the lower regeneration seen in Hsp702/2 mice was due to a loss of this function. We undertook Luminex screening of sera for cytokine/chemokine expression in WT mice and Hsp702/2 mice during the immediate period after PH. Only 4 of the 20 proteins assayed were detectable: IL-6, IL-10, IL-12, and CXCL1 (also known as KC). IL-10 and IL-12 were not significantly elevated at 4 hours in either genotype. IL-6 and CXCL1 levels were significantly elevated, but no differences were found in the Hsp702/2 group versus the WT group (IL-6: 450.2 6 223.5 versus 330.0 6 129.3 pg/mL, P 5 0.67; CXCL1: 843.7 6 353.1 versus 1376 6 395.2 pg/mL, P 5 0.37; Fig. 4A). To explore whether a lack of Hsp70 affects intrahepatic cytokine/chemokine production, we next evaluated gene expression levels of TNF-a, IL-6, CXCL1, and hepatocyte growth factor (HGF) in Hsp702/2 and

380 WOLF ET AL.

Figure 3. Hsp702/2 mice exhibited reduced cellular proliferation after PH. (A) Quantitative image analysis showed significantly less nuclear staining for Ki-67 in Hsp702/2 mice 48 hours after hepatectomy. (B) Differences are visualized in representative sections of Ki-67–stained liver tissue at 24 hours. (C) No significant histological differences in adjacent sections were noted with hematoxylin and eosin staining.

WT liver samples. Expression was measured both before surgery and 4 hours after PH with qPCR (Fig. 4B). Preoperative levels were not significantly different in Hsp702/2 mice versus WT mice, although they appeared slightly higher in Hsp702/2 mice for TNFa (7.6 6 3.8 versus 1.7 6 0.6 fold, P 5 0.20) and CXCL1 (9.0 6 3.7 versus 1.1 6 0.05 fold, P 5 0.10). In both groups, all measured factors were up-regulated at 4 hours in comparison with preoperative levels. At 4 hours, Hsp702/2 and WT mice had comparable expression levels for IL-6 (15.1 6 4.6 versus 11.9 6 3.2 fold, P 5 0.60), CXCL1 (104.0 6 21.0 versus 135.7 6 19.2 fold, P 5 0.32), and HGF (1.9 6 0.1 versus 2.1 6 0.3 fold, P 5 0.43), whereas TNF-a expres-

LIVER TRANSPLANTATION, March 2014

Figure 4. Serum and liver cytokine/chemokine expression in Hsp702/2 mice after PH. (A) A Luminex-based assay was used to screen sera from Hsp702/2 mice and WT mice (3 mice per group) for cytokine/chemokine expression after PH. Sixteen of the 20 measured factors were below the detection thresholds. Levels at 4 hours for IL-6, IL-10, IL-12, and CXCL1 are shown. (B) Gene expression was assayed from liver biopsy samples taken at the baseline and during early liver regeneration with RT-PCR. Levels of TNF-a were significantly lower in Hsp702/2 mice versus WT mice at 4 hours. RT-PCR comparisons were performed with Student t tests and with a sample size of 8 mice per group (4 hours) or 3 mice per group (0 hours).

sion was significantly lower in Hsp702/2 (8.9 6 1.6 versus 26.2 6 5.4 fold, P 5 0.01).

mice

Reduced Endotoxin Responses in Hsp702/2 Mice Although the phenotypes of Hsp702/2 mice and WT mice appeared similar before PH, we were interested in assessing whether the macrophage populations of these mice were comparable in terms of basal and LPS-induced levels of expression of TNF-a and related cytokines. We found that the basal levels of TNF-a, IL1b, and IL-6 mRNA were similarly very low in the

LIVER TRANSPLANTATION, Vol. 20, No. 3, 2014

WOLF ET AL. 381

Figure 5. Impaired cytokine mRNA production in Hsp702/2 mice after LPS injection. Cytokine mRNA expression levels in Hsp702/2 mice and WT mice (3 mice per group) were analyzed with qPCR before and 3 hours after the injection of LPS (1 mg/kg). Levels were normalized to 18S and are shown as the relative expression (means and standard deviations; **P < 0.01).

livers and spleens of Hsp702/2 mice and WT mice (Fig. 5). However, although considerable induction of all 3 cytokines in both tissues was observed 3 hours after LPS, the level of cytokine mRNA in each case was significantly decreased (P < 0.01) in Hsp702/2 mice versus WT mice. Likewise, although a flow cytometry analysis of splenic macrophage and dendritic cell (DC) populations in Hsp702/2 mice and WT mice showed only modest differences in the proportions of Ly6C1/CD11b1 cells, LPS administration resulted in decreased TNF-a production by splenic CD11b1 macrophages and CD11c1 DCs in Hsp702/2 mice versus WT mice (Fig. 6).

HSP70 Expression Is Up-Regulated During Early Liver Regeneration in Humans We tested the expression of HSP70-related genes in a human PH setting by assessing RNA transcripts in human LDLT donor livers with Affymetrix Human Gene 1.0 ST Array GeneChips. In a cohort of 23 human LDLT donors, HSP expression was significantly induced during the first hours of liver regeneration (Table 2). Transcripts from 39 molecules from 4 different HSP families were up-regulated in POST biopsy samples versus PRE biopsy samples (18 HSP40s, 9 HSP70s, 9 HSP90s, and 3 others). The 2 stress-inducible members of the HSP70 family, HSPA1A and HSPA1B, were both found to be up-

regulated after hepatectomy (1.50- and 1.68-fold increases, respectively). Two other HSP70 family members were notable for having more pronounced inductions than any other HSP genes: HSPA13 (fold increase 5 3.55) and HSPA5 (fold increase 5 2.68). HSPA13 is a constitutively expressed, noninducible member of the HSP70 family with ubiquitin-related functions, and it has been implicated in Alzheimer’s disease and epilepsy.17,18 HSPA5 is a form of HSP70 specific to the endoplasmic reticulum with critical housekeeping functions at the baseline.19 With respect to clinical outcomes in the living donor cohort, all patients had full recovery of liver function, and there were no cases of perioperative morbidity or mortality.

DISCUSSION More than 80 years of investigation have passed since Higgins and Anderson first described liver regeneration in a rodent hepatectomy model, yet the most proximal components initiating this process remain unclear.20 Identifying these molecular triggers has important significance for patients recovering from liver resection (whether due to cancer, transplant donation, ischemic injury, or trauma) because impairments in liver regeneration can lead to organ failure and because no clinical process to augment liver regeneration currently exists. Several unique features

382 WOLF ET AL.

LIVER TRANSPLANTATION, March 2014

Figure 6. Impaired macrophage and DC production of TNF-a by Hsp702/2 mice after LPS injection. A flow cytometry analysis of macrophage and DC populations in spleens of Hsp702/2 mice and WT mice (3 mice per group) was performed before and 3 hours after the injection of LPS (1 mg/kg). Representative flow data are shown on the left, with the proportion of labeled cells indicated in each panel; histograms on the right show pooled percentage data (means and standard deviations; *P < 0.05).

LIVER TRANSPLANTATION, Vol. 20, No. 3, 2014

WOLF ET AL. 383

TABLE 2. Up-Regulated Gene Expression for HSPs in Human Liver Regeneration Fold Family

Symbol

Gene Name

Change*

HSP40

DNAJB11 DNAJB9 DNAJC2 DNAJB1 DNAJC3 DNAJA1 DNAJC10 DNAJA3 DNAJB4 DNAJA2 DNAJB6 DNAJC11 DNAJC1 DNAJC16 DNAJC21 DNAJC7 DNAJC5 DNAJB6 HSPA13 HSPA5 HSPA8 HSPA4 HSPA1B HSPA14 HSPA1A HSPA4L HSPA9 HSP90AB1 HSP90AB3P HSP90AA6P HSP90AA1 HSP90AA2 HSP90B1

DnaJ (heat shock protein 40) homolog, subfamily B, member 11 DnaJ (heat shock protein 40) homolog, subfamily B, member 9 DnaJ (heat shock protein 40) homolog, subfamily C, member 2 DnaJ (heat shock protein 40) homolog, subfamily B, member 1 DnaJ (heat shock protein 40) homolog, subfamily C, member 3 DnaJ (heat shock protein 40) homolog, subfamily A, member 1 DnaJ (heat shock protein 40) homolog, subfamily C, member 10 DnaJ (heat shock protein 40) homolog, subfamily A, member 3 DnaJ (heat shock protein 40) homolog, subfamily B, member 4 DnaJ (heat shock protein 40) homolog, subfamily A, member 2 DnaJ (heat shock protein 40) homolog, subfamily B, member 6 DnaJ (heat shock protein 40) homolog, subfamily C, member 11 DnaJ (heat shock protein 40) homolog, subfamily C, member 1 DnaJ (heat shock protein 40) homolog, subfamily C, member 16 DnaJ (heat shock protein 40) homolog, subfamily C, member 21 DnaJ (heat shock protein 40) homolog, subfamily C, member 7 DnaJ (heat shock protein 40) homolog, subfamily C, member 5 DnaJ (heat shock protein 40) homolog, subfamily B, member 6 Heat shock protein 70-kDa family, member 13 Heat shock 70-kDa protein 5 (glucose-regulated protein, 78 kDa) Heat shock 70-kDa protein 8 Heat shock 70-kDa protein 4 Heat shock 70-kDa protein 1B Heat shock 70-kDa protein 14 Heat shock 70-kDa protein 1A Heat shock 70-kDa protein 4-like Heat shock 70-kDa protein 9 (mortalin) Heat shock protein 90-kDa alpha (cytosolic), class B member 1 Heat shock protein 90-kDa alpha (cytosolic), class B member 3 Heat shock protein 90-kDa alpha (cytosolic), class A member 6 Heat shock protein 90-kDa alpha (cytosolic), class A member 1 Heat shock protein 90-kDa alpha (cytosolic), class A member 2 Heat shock protein 90-kDa beta (94-kDa glucose-regulated protein), member 1 Heat shock protein 90-kDa alpha (cytosolic), class B member 4 Heat shock protein 90-kDa alpha (cytosolic), class B member 2 Heat shock protein 90-kDa beta (94-kDa glucose-regulated protein), member 3 Heat shock 105-kDa/110-kDa protein 1 Heat shock 60-kDa protein 1 (chaperonin) Heat shock 10-kDa protein 1 (chaperonin 10)

2.59 2.58 2.33 2.22 1.89 1.80 1.67 1.42 1.42 1.41 1.39 1.38 1.29 1.25 1.22 1.19 1.18 1.17 3.55 2.68 1.73 1.71 1.68 1.52 1.50 1.44 1.35 1.76 1.69 1.56 1.43 1.43 1.42

Heat shock protein 70 is required for optimal liver regeneration after partial hepatectomy in mice.

Liver regeneration is a complex process that restores functional tissue after resection or injury, and it is accompanied by transient adenosine tripho...
541KB Sizes 0 Downloads 0 Views