PERSPECTIVE
JBMR
Energy Excess, Glucose Utilization, and Skeletal Remodeling: New Insights Beata Lecka-Czernik,1,2 and Clifford J Rosen3,4 1
Department of Orthopaedic Surgery, University of Toledo Health Sciences Campus, Toledo, OH, USA Department of Physiology and Pharmacology, Center for Diabetes and Endocrine Research, University of Toledo Health Sciences Campus, Toledo, OH, USA 3 Center for Clinical & Translational Research, Maine Medical Center Research Institute, Scarborough, ME, USA 4 Tufts University School of Medicine, Boston, MA, USA 2
ABSTRACT Skeletal complications have recently been recognized as another of the several comorbidities associated with diabetes. Clinical studies suggest that disordered glucose and lipid metabolism have a profound effect on bone. Diabetes-related changes in skeletal homeostasis result in a significant increased risk of fractures, although the pathophysiology may differ from postmenopausal osteoporosis. Efforts to understand the underlying mechanisms of diabetic bone disease have focused on the direct interaction of adipose tissue with skeletal remodeling and the potential influence of glucose utilization and energy uptake on these processes. One aspect that has emerged recently is the major role of the central nervous system in whole-body metabolism, bone turnover, adipose tissue remodeling, and beta cell secretion of insulin. Importantly, the skeleton contributes to the metabolic balance inherent in physiologic states. New animal models have provided the insights necessary to begin to dissect the effects of obesity and insulin resistance on the acquisition and maintenance of bone mass. In this Perspective, we focus on potential mechanisms that underlie the complex interactions between adipose tissue and skeletal turnover by focusing on the clinical evidence and on preclinical studies indicating that glucose intolerance may have a significant impact on the skeleton. In addition, we raise fundamental questions that need to be addressed in future studies to resolve the conundrum associated with glucose intolerance, obesity, and osteoporosis. © 2015 American Society for Bone and Mineral Research. KEY WORDS: BONE-FAT INTERACTIONS; SYSTEMS BIOLOGY; BONE INTERACTORS; CELL/TISSUE SIGNALING; TRANSCRIPTION FACTORS; DISEASES AND DISORDERS OF/RELATED TO BONE
Clinical Insight Into the Effect of Disordered Glucose and Lipids Homeostasis on Bone
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iabetes mellitus (DM) can be considered a disease of intracellular glucose starvation in muscle and fat, due to either lack of insulin (Type 1 [T1D]) or impaired insulin action (Type 2 [T2D]). Both types of DM are associated with an increase in fractures; however, the skeletal phenotype is very different. T1D patients have reduced bone mineral density (BMD), due to insulin and amylin deficiency, whereas T2D patients have either normal or high BMD. T2D is a disease of disordered energy storage as well as glucose intolerance and altered bone strength. Its prevalence is high and not abating. In the United States, 29.1 million people— 9.3% of the population—have diabetes; among them, nearly 60% are characterized as obese (http://www.cdc.gov). Insulin resistance is a characteristic feature of T2D in liver, muscle, and fat, leading to glucose intolerance, elevated cardiometabolic risk factors (eg, low-density lipoprotein [LDL] cholesterol), and excess hepatic glucose output. Visceral adipocytes, in particular, are large and surrounded by fibrous and inflammatory elements.
In clinical medicine a major paradox is the relationship of obesity to the skeleton. Obese individuals tend to have higher areal bone mineral density (aBMD), but in some studies they have a greater risk of fractures.(1) A meta-analysis of several large studies confirmed that obesity was generally associated with a lower fracture risk.(2,3) But, in a cohort study of older men (Osteoporotic Fractures in Men [MrOS]) that was controlled for BMD, as well as in a registry study of postmenopausal women from England, higher BMI was found to be a significant risk factor for certain fractures.(4) Several studies in children have shown that obesity increases the risk for radial fractures.(5) Similarly, patients with T2D have high or normal BMD, but a greater risk of fracture.(6–8) On the other hand, some obese older adults may have low BMD and an increased fracture risk.(9,10) This association is confounded by the location of the adipose depot and its relationship to bone. Bredella and colleagues(11) showed that visceral adiposity, which relates to increased inflammation and metabolic syndrome, was negatively associated with lumbar volumetric BMD measured by QCT, but positively related to marrow adipose volume measured by MRI. Similarly,
Received in original form March 3, 2015; revised form June 7, 2015; accepted June 8, 2015. Accepted manuscript online June 10, 2015. Address correspondence to: Beata Lecka-Czernik, PhD, University of Toledo Health Sciences Campus, 3000 Arlington Avenue, Toledo, OH 43614, USA. E-mail: [email protected] Journal of Bone and Mineral Research, Vol. 30, No. 8, August 2015, pp 1356–1361 DOI: 10.1002/jbmr.2574 © 2015 American Society for Bone and Mineral Research
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the Mayo group reported an inverse relationship between visceral adipose tissue and CT volumetric bone density.(12) Moreover, Cohen and colleagues(13) described a negative association between visceral fat volume and reduced bone volume fractions in iliac crest biopsies of premenopausal women with normal areal and spine volumetric BMD but low bone formation. In contrast, Dimitri and colleagues,(14) using high-resolution mCT, showed that obese individuals, both young and old, have higher trabecular bone volume fractions in the radius and tibia than healthy controls. One possible mechanism may be that increased body weight has a beneficial effect because of mechanical loading.(15) These conflicting results may be explained, at least in part, by the type of obesity involving either subcutaneous “good” fat or visceral “bad” fat. Thus, the effect on skeleton of peripheral and central obesity may differ, and needs to be considered in future studies. The effect of chronic hyperglycemia on bone mass is another area that remains to be clarified. In most patients with T2D, aBMD is either normal or high; yet skeletal fragility, particularly in the appendicular skeleton, is a characteristic feature.(8,16) Newer techniques for skeletal microarchitecture have allowed investigators to identify several unique aspects of the appendicular and axial skeleton from T2D patients. These include greater cortical porosity, smaller cortical area, decreased bone material strength measured by microindentation, and high bone marrow adiposity.(17–19) In addition, the composition of the skeletal matrix may be significantly altered by higher concentrations of advanced glycation end products (AGEs). (20) All may contribute to the higher frequency of fractures documented in epidemiologic studies. There remains significant debate about the importance of insulin signaling in the skeletal remodeling of humans and its contribution to the hypothesized “fast-forward” system, which proposes positive and reciprocal association between insulin signaling and skeletal remodeling, and point to the bonespecific proteins (eg, osteocalcin) defining this association.(21) For example, insulin-resistant obese and T2D patients generally exhibit lower bone turnover than nondiabetic normal subjects, as measured by histomorphometry and serum bone turnover markers.(22–24) In particular, those with previous vertebral fractures have been shown to have higher sclerostin and lower serum IGF-I and PTH levels consistent with reduced bone formation and lower bone turnover.(25,26) Whether this reflects the insulin-resistant state of the individual and hence impairment in the fast-forward cycle, or another process, remains to be determined. On the other hand, patients treated with antiresorptive agents have not been shown to have a greater frequency of impaired insulin sensitivity, glucose intolerance, or T2D.(27) There are likely other skeletal factors that are important in regulating glucose metabolism. For example, with high bone turnover, matrix-bound IGF-I is released locally to recruit new osteoblasts; it is also conceivable that this could impact glucose tolerance because IGF-I can signal through insulin receptors. Others have postulated another, yet to be identified, bone factor that regulates insulin sensitivity.(28,29) One candidate may be FGF-23, a skeletal factor produced by osteocytes that regulates phosphate homeostasis.(30) It belongs to a subset of the FGF family (FGF-19, FGF-21, and FGF-23) that has systemic glucoselowering properties. Recent studies showed that the C-terminal portion of FGF-23 is positively associated with increased BMI and insulin resistance as measured by Homeostasis Model Assessment (HOMA), but negatively associated with ferritin and serum
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iron.(31–34) Weight loss has been shown to reduce the serum levels of FGF-23.(35) Another study showed that the response of FGF-23 and PTH to a phosphate load is impaired in T2D.(36) This is a new area of investigation, and more studies are needed to define the potential impact of skeletal factors besides osteocalcin on glucose tolerance in both animals and humans. Finally, there is preliminary evidence that RANKL may have a negative impact on glucose homeostasis by its effects on hepatic glucose output. High serum concentrations of RANKL are independent risk predictors for development of T2D in humans and, as shown in animals, blockage of RANKL signaling improves hepatic insulin sensitivity and normalizes blood glucose levels.(37)
Preclinical Insights Central and peripheral controls of metabolic homeostasis and bone turnover It has been recognized for over the decade that the central nervous system (CNS) regulates both energy and bone homeostasis in a highly coordinated manner. Hypothalamic axes that regulate food intake and satiety also control bone remodeling. Several mechanisms have been identified, some relying on integrating signals within the brain(38) and others on integrating signals from peripheral tissues. The latter are exemplified by the adipocyte-derived hormone leptin, which signals to the hypothalamus via induction of serotonergic neurons in the brain stem to regulate bone mass, food intake, and peripheral fat activity. As fat cells expand and release leptin, circulating concentrations increase to suppress orexigenic factors, alter hypothalamic reproductive factors, and enhance sympathetic nervous system (SNS) tone, which increases adrenergic activity.(39) Activation of b2 adrenergic receptors in osteoblasts from increased SNS tone results in suppressed bone formation and enhanced bone resorption. On the other hand, there is little evidence that leptin has a direct effect on osteoblasts or osteoclasts, although the leptin receptor is expressed early in osteoblast progenitors. In contrast, adiponectin, an adipokine with insulin sensitizing activity, has both direct and indirect effects on bone, first by directly suppressing bone formation, and later by blocking SNS outflow from the CNS.(40) This “yin and yang” model, proposed and validated by the Karsenty laboratory, provides strong evidence that metabolic status, integrated in the brain but originating from the adipocyte, has a major impact on the state of the skeleton.(40) Another aspect of common regulatory circuits in energy and bone metabolism via the SNS is fat metabolism. Beige or brownlike adipocytes develop in subcutaneous white adipose depots (WAT) in response to different hormonal and environmental stimuli including chronic cold exposure, adrenergic signaling, and pharmacologic and nutritional factors, which increase mitochondrial number and enhance expression of brown fat (BAT)-specific transcription factors to uncouple respiration.(41) Conditions that stimulate beige fat development usually involve the SNS. However, the triad of SNS, beige fat, and bone mass is very complex. In states in which SNS tone is high (eg, cold exposure, FGF-21 excess), beige cells are found in the subcutaneous or inguinal depot.(42) On the other hand, bone mass is significantly reduced during high SNS tone, principally due to activation of the b2 receptor on osteoblasts. Activation of this receptor suppresses the key transcription factor, ATF4, while it enhances RANKL production.(43–45)
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Fig. 1. Efferent and afferent signaling regulating energy metabolism and bone turnover. In conditions of balanced energy metabolism, signaling from fat, bone, liver, pancreas, and brain maintain insulin sensitivity and normal bone turnover. Conditions of unbalanced energy metabolism lead to increased SNS/adrenergic signaling, which attenuates positive axes regulating glucose metabolism and bone turnover. Fat tissue is depicted as a group of white-type adipocytes (shapes in gray), which in normal conditions may be infiltrated by beige adipocytes (shapes in yellow, left panel); however, in obesity and diabetes it is infiltrated by macrophages (red shapes, right panel). SNS ¼ sympathetic nervous system.
In contrast, genetically manipulated mice with adipocytespecific expression of FoxC2 transcription factor, which increases sensitivity to the beta-adrenergic-cAMP-protein kinase (PKA) signaling pathway,(46) are characterized by high bone mass and conversion of epididymal WAT to beige fat, and beiging of marrow adipocytes. A secretome of beige adipocytes overexpressing FoxC2 includes bone anabolic Wnt10b, IGFBP2, and IGF-1 proteins. Conditioned media collected from cultures of beige adipocytes induce expression of osteoblast-specific gene markers in marrow mesenchymal stem cells (MSCs) and suppress SOST expression in osteocytes.(47) Thus, it is possible that systemic negative effects on bone of SNS are counterbalanced by either endocrine or local bone anabolic effects of beige fat. There is no doubt that the interrelationship among bone, fat, and brain signaling constitutes a powerful network regulating simultaneous bone and energy metabolism.
Skeletal control of metabolic balance The integration of bone remodeling with energy metabolism links the anabolic effect of insulin signaling in osteoblasts with bone turnover and regulation of insulin sensitivity in peripheral organs.(48,49) Thus, in osteoblasts, insulin signaling regulates expression of Runx2 and production of osteocalcin. In addition, insulin increases support for osteoclastogenesis by decreasing expression of osteoprotegerin (OPG), a decoy receptor for RANKL. As a result, insulin increases bone turnover and production of undercarboxylated osteocalcin, which, in endocrine fashion, regulates insulin release from b-cells in the pancreas and production of adiponectin in adipose tissue.(21,50) The net result is an increase in insulin sensitivity and secretion that ultimately leads to greater bone resorption. Further proof of that tenet comes from studies of Esp1, a phosphatase that is responsible for the degradation of osteocalcin in murine bone cells. The genetic absence of Esp1 in mice results in significant hypoglycemia, due to excess undercarboxylated osteocalcin.(48,51) On the other hand,
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FoxO1 upregulates Esp1 expression, thereby impairing glucose tolerance.(52) Thus, in mice there is a complex interaction between glucose homeostasis and skeletal remodeling. However, moving from the preclinical to the clinical setting has been more difficult in establishing the relationship between this system and metabolic homeostasis.(23,53,54)
Bioenergetics of the skeleton More recently, attention has focused on other mechanisms related to stem cell fate; in particular, the metabolic program of MSC progenitors. There is now emerging evidence that adipocytes and osteoblasts have distinct energy utilization pathways as they move down their respective lineages during differentiation. Early progenitors and pluripotent stem cells utilize glucose as their primary fuel, even in aerobic states, in a manner analogous to cancer cells (ie, the Warburg effect). But preadipocytes, in particular, use fatty acids to supply energy for their metabolic needs. That comes at a cost because mitochondrial fatty acid oxidation leads to the generation of reactive oxygen species (ROS). Such compounds (eg, H2O2, superoxides) can suppress mitochondrial respiration and promote an adipogenic program that is associated with more insulin resistance and less lipolysis.(55,56) In contrast, preosteoblasts utilize a distinct metabolic program. These cells differentiate under the influence of various ligands, particularly the Wnts. Esen and colleagues(57) showed that glycolysis is a major feature of Wnt3a-induced osteoblast differentiation. Remarkably, much older ex vivo studies from Nichols and Neuman(58) showed that PTH produces lactic acid in osteoblasts, supporting the tenet that osteoblasts utilize glucose to fuel collagen synthesis and mineralization. Guntur and colleagues(59) also showed that glycolysis was essential for terminal differentiation of osteoblasts and that oxidative phosphorylation was more important early in the differentiation scheme. More recently, Regan and colleagues,(60) of the Long
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group, reported that HIF1a is a critical transcriptional regulator of glycolysis, triggered in part by relative hypoxia in the niche. In that same vein, Karner and colleagues(61) showed that glutaminolysis is also essential for osteoblast differentiation through the Wnt signaling system. In sum, the bioenergetics of the niche must be fully defined in order to understand lineage allocation and the role transcription factors play in defining the ultimate fate of MSCs in the marrow. It is remarkable, however, that a picture is emerging of a metabolic program in osteoblasts that resembles tumor cell bioenergetics. The relative hypoxic environment, the additive effects of glutaminolysis with glycolysis on ATP generation, and the evolutionary importance of substrate availability that is intimately tied to glycolysis in single-cell organisms provide us with potential clues about progenitor cells and their ultimate fate either to store energy (ie, adipocytes) or to use sufficient energy to build and mineralize matrix (ie, osteoblasts).
Animal models to dissect the effects of obesity, insulin resistance, and hyperglycemia on the acquisition and maintenance of bone mass Animal models provide important insights into the interactions between adipose tissue and bone, but a similar paradox as in humans has been noted in obese mice. C57BL/6J, the most commonly used inbred strain, has a genetic predisposition to obesity and glucose intolerance, particularly when fed a high-fat diet (HFD). This weight gain, seen in both young and aging mice, is associated with visceral adiposity, insulin resistance, and, in some instances, bone loss.(62,63) But other studies have shown that C57BL/6 mice on an HFD have higher marrow adipose tissue yet either normal or increased aBMD.(64,65) Moreover, rates of bone formation in mice fed an HFD are either normal(64) or are temporally increased followed by a significant decrease in bone turnover concurrent with development of obesity and glucose intolerance.(65) Similarly, animals with induced insulin resistance in osteoblasts by ablation of insulin receptor respond to HFD with a relative increase in bone mass due to attenuated bone turnover.(66) For the laboratory mouse, confounders such as strain, age, gender, composition of the diet, duration of feeding, and the gut microbiota, play important roles in determining the effects of an HFD on the magnitude of obesity, metabolic impairment, bone marrow adipocyte infiltration, and ultimately on the skeletal response. Indeed, even in genetically identical C57BL/6J mice there are subsets of high- and low-weight gaining mice during high-fat feeding, suggesting there have to be strong epigenetic effects.(67) Similarly, human studies are confounded by nutrient and environmental factors as well as genetic heterogeneity, which may explain the different impact of obesity on skeleton, as discussed in “Clinical Insight” section. Animal models provide insights into the molecular and cellular mechanisms of bone remodeling related to obesity and T2D, and have served to illuminate this complex interaction of tissues. L-SACC mice, with impaired insulin clearance in the liver, serve as a model of hyperinsulinemia with normal nonfasting glucose levels. These mice are characterized by high bone mass due to attenuated bone resorption as a result, at least in part, of insulin’s negative effect on osteoclasts.(68) However, there are diabesity models associated with both pronounced hyperglycemia and obesity, including New Zealand Obese (NZO/HlLt and NZO/HlBomDife), yellow agouti Avy/a, and the TALLYHO/JngJ mouse. The yellow agouti and TALLYHO/JngJ mice may be particularly useful in future studies because in these models the
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onset of T2D occurs early, thereby paralleling the rising epidemic of T2D in adolescence.(69,70) Although yellow agouti mice have normal bone mass, TALLYHO/JngJ mice have severely impaired peak bone mass and reduced bone formation, a characteristic feature of bone from adult and juvenile patients with T2D, respectively.
Summary: Future Challenges The relationship between the skeleton and adipose tissue is tied directly to fuel (glucose and lipids) utilization and energy generation at both the bone and whole-body level. The skeletal remodeling unit expends tremendous energy during both resorption and formation. Because it has now been established that there is central control over this process, and that the adipocyte is a major sensor as well as modulator of energy status, it seems certain that the CNS permits the integration of fuel homeostasis in the individual, balancing the demands of multiple tissues including bone. Glycolysis appears to be the preferred substrate for skeletal remodeling, hence glucose intolerance and T2D present a major challenge for the organism. Yet we still know little about the precise sequence of events that leads to insulin resistance in osteoblasts, nor do we know for sure that the changes in bone cell function and matrix constituents associated with T2D are related to insulin resistance or to other processes such as locally high concentrations of reactive oxygen species or other cell non-autonomous processes. The hypoxic environment of the bone marrow niche presents an additional challenge for developing experimental paradigms to study bone-fat interactions at the local level. Yet, ironically, HIF1a is a major determinant of lineage allocation and osteogenesis. Similarly, there are major challenges for defining the origin of the marrow adipocyte and its function in normal and impaired glucose homeostasis. At the patient level, there are no new drugs that specifically target diabetic bone disease, in part because the syndrome is heterogeneous, particularly for T2D. Moreover, some antidiabetic drugs cause bone loss. In addition, the molecular mechanisms related to glucose intolerance at the osteoblastic, osteocytic, and osteoclastic level are only just beginning to be elucidated. It is complicated further by changes in glycosylated proteins in the matrix that alter the structural aspects of the skeleton without directly affecting BMD. Although animal models have been helpful, they often do not fully reflect the complexity of the clinical disorder. Notwithstanding these many limitations, it is clear that we are entering a new era in bone biology—one that encompasses whole-body physiology. Hence, changes in energy metabolism have profound impacts on the skeleton and certainly alterations in the skeletal remodeling unit can affect whole body metabolism.
Disclosures The authors state that they have no conflicts of interest.
Acknowledgments This work was supported by funding from the American Diabetes Association (7-13-BS-089 to BLC) and the NIDDK (R24DK092759-05 to CJR). Authors’ role: BLC and CJR contributed equally to the literature search, discussion, and writing of this manuscript.
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52. Rached MT, Kode A, Silva BC, et al. FoxO1 expression in osteoblasts regulates glucose homeostasis through regulation of osteocalcin in mice. J Clin Invest. 2010;120(1):357–68. pez A, Bullo M, Juanola-Falgarona M, et al. Reduced serum 53. Dıaz-Lo concentrations of carboxylated and undercarboxylated osteocalcin are associated with risk of developing type 2 diabetes mellitus in a high cardiovascular risk population: a nested case-control study. J Clin Endocrinol Metab. 2013;98(11):4524–31. 54. Yeap BB, Alfonso H, Chubb SA, et al. Higher serum undercarboxylated osteocalcin and other bone turnover markers are associated with reduced diabetes risk and lower estradiol concentrations in older men. J Clin Endocrinol Metab. 2015;100(1):63–71. 55. Tormos KV, Anso E, Hamanaka RB, et al. Mitochondrial complex III ROS regulate adipocyte differentiation. Cell Metab. 2011;14(4): 537–44. 56. Wang T, Si Y, Shirihai OS, et al. Respiration in adipocytes is inhibited by reactive oxygen species. Obesity (Silver Spring). 2010;18(8):1493– 502. 57. Esen E, Chen J, Karner CM, Okunade AL, Patterson BW, Long F. WNTLRP5 signaling induces Warburg effect through mTORC2 activation during osteoblast differentiation. Cell Metab. 2013;17(5):745–55. 58. Nichols FC, Neuman WF. Lactic acid production in mouse calvaria in vitro with and without parathyroid hormone stimulation: lack of acetazolamide effects. Bone. 1987;8(2):105–9.
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66. Wei J, Ferron M, Clarke CJ, et al. Bone-specific insulin resistance disrupts whole-body glucose homeostasis via decreased osteocalcin activation. J Clin Invest. 2014;124(4):1–13. 67. Koza RA, Nikonova L, Hogan J, et al. Changes in gene expression foreshadow diet-induced obesity in genetically identical mice. PLoS Genet. 2006;2(5):e81. 68. Huang S, Kaw M, Harris MT, et al. Decreased osteoclastogenesis and high bone mass in mice with impaired insulin clearance due to liverspecific inactivation to CEAC AM1. Bone. 2010;46(4):1138–45. 69. Devlin MJ, Van Vliet M, Motyl K, et al. Early-onset type 2 diabetes impairs skeletal acquisition in the male TALLYHO/JngJ mouse. Endocrinology. 2014;155(10):3806–16. 70. Kolli V, Stechschulte LA, Dowling AR, Rahman S, Czernik PJ, LeckaCzernik B. Partial agonist, telmisartan, maintains PPARg serine 112 phosphorylation, and does not affect osteoblast differentiation and bone mass. PLoS One. 2014;9(5):e96323.
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Energy Excess, Glucose Utilization, and Skeletal Remodeling: New Insights Beata Lecka-Czernik,1,2 and Clifford J Rosen3,4 1
Department of Orthopaedic Surgery, University of Toledo Health Sciences Campus, Toledo, OH, USA Department of Physiology and Pharmacology, Center for Diabetes and Endocrine Research, University of Toledo Health Sciences Campus, Toledo, OH, USA 3 Center for Clinical & Translational Research, Maine Medical Center Research Institute, Scarborough, ME, USA 4 Tufts University School of Medicine, Boston, MA, USA 2
ABSTRACT Skeletal complications have recently been recognized as another of the several comorbidities associated with diabetes. Clinical studies suggest that disordered glucose and lipid metabolism have a profound effect on bone. Diabetes-related changes in skeletal homeostasis result in a significant increased risk of fractures, although the pathophysiology may differ from postmenopausal osteoporosis. Efforts to understand the underlying mechanisms of diabetic bone disease have focused on the direct interaction of adipose tissue with skeletal remodeling and the potential influence of glucose utilization and energy uptake on these processes. One aspect that has emerged recently is the major role of the central nervous system in whole-body metabolism, bone turnover, adipose tissue remodeling, and beta cell secretion of insulin. Importantly, the skeleton contributes to the metabolic balance inherent in physiologic states. New animal models have provided the insights necessary to begin to dissect the effects of obesity and insulin resistance on the acquisition and maintenance of bone mass. In this Perspective, we focus on potential mechanisms that underlie the complex interactions between adipose tissue and skeletal turnover by focusing on the clinical evidence and on preclinical studies indicating that glucose intolerance may have a significant impact on the skeleton. In addition, we raise fundamental questions that need to be addressed in future studies to resolve the conundrum associated with glucose intolerance, obesity, and osteoporosis. © 2015 American Society for Bone and Mineral Research. KEY WORDS: BONE-FAT INTERACTIONS; SYSTEMS BIOLOGY; BONE INTERACTORS; CELL/TISSUE SIGNALING; TRANSCRIPTION FACTORS; DISEASES AND DISORDERS OF/RELATED TO BONE
Clinical Insight Into the Effect of Disordered Glucose and Lipids Homeostasis on Bone
D
iabetes mellitus (DM) can be considered a disease of intracellular glucose starvation in muscle and fat, due to either lack of insulin (Type 1 [T1D]) or impaired insulin action (Type 2 [T2D]). Both types of DM are associated with an increase in fractures; however, the skeletal phenotype is very different. T1D patients have reduced bone mineral density (BMD), due to insulin and amylin deficiency, whereas T2D patients have either normal or high BMD. T2D is a disease of disordered energy storage as well as glucose intolerance and altered bone strength. Its prevalence is high and not abating. In the United States, 29.1 million people— 9.3% of the population—have diabetes; among them, nearly 60% are characterized as obese (http://www.cdc.gov). Insulin resistance is a characteristic feature of T2D in liver, muscle, and fat, leading to glucose intolerance, elevated cardiometabolic risk factors (eg, low-density lipoprotein [LDL] cholesterol), and excess hepatic glucose output. Visceral adipocytes, in particular, are large and surrounded by fibrous and inflammatory elements.
In clinical medicine a major paradox is the relationship of obesity to the skeleton. Obese individuals tend to have higher areal bone mineral density (aBMD), but in some studies they have a greater risk of fractures.(1) A meta-analysis of several large studies confirmed that obesity was generally associated with a lower fracture risk.(2,3) But, in a cohort study of older men (Osteoporotic Fractures in Men [MrOS]) that was controlled for BMD, as well as in a registry study of postmenopausal women from England, higher BMI was found to be a significant risk factor for certain fractures.(4) Several studies in children have shown that obesity increases the risk for radial fractures.(5) Similarly, patients with T2D have high or normal BMD, but a greater risk of fracture.(6–8) On the other hand, some obese older adults may have low BMD and an increased fracture risk.(9,10) This association is confounded by the location of the adipose depot and its relationship to bone. Bredella and colleagues(11) showed that visceral adiposity, which relates to increased inflammation and metabolic syndrome, was negatively associated with lumbar volumetric BMD measured by QCT, but positively related to marrow adipose volume measured by MRI. Similarly,
Received in original form March 3, 2015; revised form June 7, 2015; accepted June 8, 2015. Accepted manuscript online June 10, 2015. Address correspondence to: Beata Lecka-Czernik, PhD, University of Toledo Health Sciences Campus, 3000 Arlington Avenue, Toledo, OH 43614, USA. E-mail: [email protected] Journal of Bone and Mineral Research, Vol. 30, No. 8, August 2015, pp 1356–1361 DOI: 10.1002/jbmr.2574 © 2015 American Society for Bone and Mineral Research
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the Mayo group reported an inverse relationship between visceral adipose tissue and CT volumetric bone density.(12) Moreover, Cohen and colleagues(13) described a negative association between visceral fat volume and reduced bone volume fractions in iliac crest biopsies of premenopausal women with normal areal and spine volumetric BMD but low bone formation. In contrast, Dimitri and colleagues,(14) using high-resolution mCT, showed that obese individuals, both young and old, have higher trabecular bone volume fractions in the radius and tibia than healthy controls. One possible mechanism may be that increased body weight has a beneficial effect because of mechanical loading.(15) These conflicting results may be explained, at least in part, by the type of obesity involving either subcutaneous “good” fat or visceral “bad” fat. Thus, the effect on skeleton of peripheral and central obesity may differ, and needs to be considered in future studies. The effect of chronic hyperglycemia on bone mass is another area that remains to be clarified. In most patients with T2D, aBMD is either normal or high; yet skeletal fragility, particularly in the appendicular skeleton, is a characteristic feature.(8,16) Newer techniques for skeletal microarchitecture have allowed investigators to identify several unique aspects of the appendicular and axial skeleton from T2D patients. These include greater cortical porosity, smaller cortical area, decreased bone material strength measured by microindentation, and high bone marrow adiposity.(17–19) In addition, the composition of the skeletal matrix may be significantly altered by higher concentrations of advanced glycation end products (AGEs). (20) All may contribute to the higher frequency of fractures documented in epidemiologic studies. There remains significant debate about the importance of insulin signaling in the skeletal remodeling of humans and its contribution to the hypothesized “fast-forward” system, which proposes positive and reciprocal association between insulin signaling and skeletal remodeling, and point to the bonespecific proteins (eg, osteocalcin) defining this association.(21) For example, insulin-resistant obese and T2D patients generally exhibit lower bone turnover than nondiabetic normal subjects, as measured by histomorphometry and serum bone turnover markers.(22–24) In particular, those with previous vertebral fractures have been shown to have higher sclerostin and lower serum IGF-I and PTH levels consistent with reduced bone formation and lower bone turnover.(25,26) Whether this reflects the insulin-resistant state of the individual and hence impairment in the fast-forward cycle, or another process, remains to be determined. On the other hand, patients treated with antiresorptive agents have not been shown to have a greater frequency of impaired insulin sensitivity, glucose intolerance, or T2D.(27) There are likely other skeletal factors that are important in regulating glucose metabolism. For example, with high bone turnover, matrix-bound IGF-I is released locally to recruit new osteoblasts; it is also conceivable that this could impact glucose tolerance because IGF-I can signal through insulin receptors. Others have postulated another, yet to be identified, bone factor that regulates insulin sensitivity.(28,29) One candidate may be FGF-23, a skeletal factor produced by osteocytes that regulates phosphate homeostasis.(30) It belongs to a subset of the FGF family (FGF-19, FGF-21, and FGF-23) that has systemic glucoselowering properties. Recent studies showed that the C-terminal portion of FGF-23 is positively associated with increased BMI and insulin resistance as measured by Homeostasis Model Assessment (HOMA), but negatively associated with ferritin and serum
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iron.(31–34) Weight loss has been shown to reduce the serum levels of FGF-23.(35) Another study showed that the response of FGF-23 and PTH to a phosphate load is impaired in T2D.(36) This is a new area of investigation, and more studies are needed to define the potential impact of skeletal factors besides osteocalcin on glucose tolerance in both animals and humans. Finally, there is preliminary evidence that RANKL may have a negative impact on glucose homeostasis by its effects on hepatic glucose output. High serum concentrations of RANKL are independent risk predictors for development of T2D in humans and, as shown in animals, blockage of RANKL signaling improves hepatic insulin sensitivity and normalizes blood glucose levels.(37)
Preclinical Insights Central and peripheral controls of metabolic homeostasis and bone turnover It has been recognized for over the decade that the central nervous system (CNS) regulates both energy and bone homeostasis in a highly coordinated manner. Hypothalamic axes that regulate food intake and satiety also control bone remodeling. Several mechanisms have been identified, some relying on integrating signals within the brain(38) and others on integrating signals from peripheral tissues. The latter are exemplified by the adipocyte-derived hormone leptin, which signals to the hypothalamus via induction of serotonergic neurons in the brain stem to regulate bone mass, food intake, and peripheral fat activity. As fat cells expand and release leptin, circulating concentrations increase to suppress orexigenic factors, alter hypothalamic reproductive factors, and enhance sympathetic nervous system (SNS) tone, which increases adrenergic activity.(39) Activation of b2 adrenergic receptors in osteoblasts from increased SNS tone results in suppressed bone formation and enhanced bone resorption. On the other hand, there is little evidence that leptin has a direct effect on osteoblasts or osteoclasts, although the leptin receptor is expressed early in osteoblast progenitors. In contrast, adiponectin, an adipokine with insulin sensitizing activity, has both direct and indirect effects on bone, first by directly suppressing bone formation, and later by blocking SNS outflow from the CNS.(40) This “yin and yang” model, proposed and validated by the Karsenty laboratory, provides strong evidence that metabolic status, integrated in the brain but originating from the adipocyte, has a major impact on the state of the skeleton.(40) Another aspect of common regulatory circuits in energy and bone metabolism via the SNS is fat metabolism. Beige or brownlike adipocytes develop in subcutaneous white adipose depots (WAT) in response to different hormonal and environmental stimuli including chronic cold exposure, adrenergic signaling, and pharmacologic and nutritional factors, which increase mitochondrial number and enhance expression of brown fat (BAT)-specific transcription factors to uncouple respiration.(41) Conditions that stimulate beige fat development usually involve the SNS. However, the triad of SNS, beige fat, and bone mass is very complex. In states in which SNS tone is high (eg, cold exposure, FGF-21 excess), beige cells are found in the subcutaneous or inguinal depot.(42) On the other hand, bone mass is significantly reduced during high SNS tone, principally due to activation of the b2 receptor on osteoblasts. Activation of this receptor suppresses the key transcription factor, ATF4, while it enhances RANKL production.(43–45)
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Fig. 1. Efferent and afferent signaling regulating energy metabolism and bone turnover. In conditions of balanced energy metabolism, signaling from fat, bone, liver, pancreas, and brain maintain insulin sensitivity and normal bone turnover. Conditions of unbalanced energy metabolism lead to increased SNS/adrenergic signaling, which attenuates positive axes regulating glucose metabolism and bone turnover. Fat tissue is depicted as a group of white-type adipocytes (shapes in gray), which in normal conditions may be infiltrated by beige adipocytes (shapes in yellow, left panel); however, in obesity and diabetes it is infiltrated by macrophages (red shapes, right panel). SNS ¼ sympathetic nervous system.
In contrast, genetically manipulated mice with adipocytespecific expression of FoxC2 transcription factor, which increases sensitivity to the beta-adrenergic-cAMP-protein kinase (PKA) signaling pathway,(46) are characterized by high bone mass and conversion of epididymal WAT to beige fat, and beiging of marrow adipocytes. A secretome of beige adipocytes overexpressing FoxC2 includes bone anabolic Wnt10b, IGFBP2, and IGF-1 proteins. Conditioned media collected from cultures of beige adipocytes induce expression of osteoblast-specific gene markers in marrow mesenchymal stem cells (MSCs) and suppress SOST expression in osteocytes.(47) Thus, it is possible that systemic negative effects on bone of SNS are counterbalanced by either endocrine or local bone anabolic effects of beige fat. There is no doubt that the interrelationship among bone, fat, and brain signaling constitutes a powerful network regulating simultaneous bone and energy metabolism.
Skeletal control of metabolic balance The integration of bone remodeling with energy metabolism links the anabolic effect of insulin signaling in osteoblasts with bone turnover and regulation of insulin sensitivity in peripheral organs.(48,49) Thus, in osteoblasts, insulin signaling regulates expression of Runx2 and production of osteocalcin. In addition, insulin increases support for osteoclastogenesis by decreasing expression of osteoprotegerin (OPG), a decoy receptor for RANKL. As a result, insulin increases bone turnover and production of undercarboxylated osteocalcin, which, in endocrine fashion, regulates insulin release from b-cells in the pancreas and production of adiponectin in adipose tissue.(21,50) The net result is an increase in insulin sensitivity and secretion that ultimately leads to greater bone resorption. Further proof of that tenet comes from studies of Esp1, a phosphatase that is responsible for the degradation of osteocalcin in murine bone cells. The genetic absence of Esp1 in mice results in significant hypoglycemia, due to excess undercarboxylated osteocalcin.(48,51) On the other hand,
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FoxO1 upregulates Esp1 expression, thereby impairing glucose tolerance.(52) Thus, in mice there is a complex interaction between glucose homeostasis and skeletal remodeling. However, moving from the preclinical to the clinical setting has been more difficult in establishing the relationship between this system and metabolic homeostasis.(23,53,54)
Bioenergetics of the skeleton More recently, attention has focused on other mechanisms related to stem cell fate; in particular, the metabolic program of MSC progenitors. There is now emerging evidence that adipocytes and osteoblasts have distinct energy utilization pathways as they move down their respective lineages during differentiation. Early progenitors and pluripotent stem cells utilize glucose as their primary fuel, even in aerobic states, in a manner analogous to cancer cells (ie, the Warburg effect). But preadipocytes, in particular, use fatty acids to supply energy for their metabolic needs. That comes at a cost because mitochondrial fatty acid oxidation leads to the generation of reactive oxygen species (ROS). Such compounds (eg, H2O2, superoxides) can suppress mitochondrial respiration and promote an adipogenic program that is associated with more insulin resistance and less lipolysis.(55,56) In contrast, preosteoblasts utilize a distinct metabolic program. These cells differentiate under the influence of various ligands, particularly the Wnts. Esen and colleagues(57) showed that glycolysis is a major feature of Wnt3a-induced osteoblast differentiation. Remarkably, much older ex vivo studies from Nichols and Neuman(58) showed that PTH produces lactic acid in osteoblasts, supporting the tenet that osteoblasts utilize glucose to fuel collagen synthesis and mineralization. Guntur and colleagues(59) also showed that glycolysis was essential for terminal differentiation of osteoblasts and that oxidative phosphorylation was more important early in the differentiation scheme. More recently, Regan and colleagues,(60) of the Long
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group, reported that HIF1a is a critical transcriptional regulator of glycolysis, triggered in part by relative hypoxia in the niche. In that same vein, Karner and colleagues(61) showed that glutaminolysis is also essential for osteoblast differentiation through the Wnt signaling system. In sum, the bioenergetics of the niche must be fully defined in order to understand lineage allocation and the role transcription factors play in defining the ultimate fate of MSCs in the marrow. It is remarkable, however, that a picture is emerging of a metabolic program in osteoblasts that resembles tumor cell bioenergetics. The relative hypoxic environment, the additive effects of glutaminolysis with glycolysis on ATP generation, and the evolutionary importance of substrate availability that is intimately tied to glycolysis in single-cell organisms provide us with potential clues about progenitor cells and their ultimate fate either to store energy (ie, adipocytes) or to use sufficient energy to build and mineralize matrix (ie, osteoblasts).
Animal models to dissect the effects of obesity, insulin resistance, and hyperglycemia on the acquisition and maintenance of bone mass Animal models provide important insights into the interactions between adipose tissue and bone, but a similar paradox as in humans has been noted in obese mice. C57BL/6J, the most commonly used inbred strain, has a genetic predisposition to obesity and glucose intolerance, particularly when fed a high-fat diet (HFD). This weight gain, seen in both young and aging mice, is associated with visceral adiposity, insulin resistance, and, in some instances, bone loss.(62,63) But other studies have shown that C57BL/6 mice on an HFD have higher marrow adipose tissue yet either normal or increased aBMD.(64,65) Moreover, rates of bone formation in mice fed an HFD are either normal(64) or are temporally increased followed by a significant decrease in bone turnover concurrent with development of obesity and glucose intolerance.(65) Similarly, animals with induced insulin resistance in osteoblasts by ablation of insulin receptor respond to HFD with a relative increase in bone mass due to attenuated bone turnover.(66) For the laboratory mouse, confounders such as strain, age, gender, composition of the diet, duration of feeding, and the gut microbiota, play important roles in determining the effects of an HFD on the magnitude of obesity, metabolic impairment, bone marrow adipocyte infiltration, and ultimately on the skeletal response. Indeed, even in genetically identical C57BL/6J mice there are subsets of high- and low-weight gaining mice during high-fat feeding, suggesting there have to be strong epigenetic effects.(67) Similarly, human studies are confounded by nutrient and environmental factors as well as genetic heterogeneity, which may explain the different impact of obesity on skeleton, as discussed in “Clinical Insight” section. Animal models provide insights into the molecular and cellular mechanisms of bone remodeling related to obesity and T2D, and have served to illuminate this complex interaction of tissues. L-SACC mice, with impaired insulin clearance in the liver, serve as a model of hyperinsulinemia with normal nonfasting glucose levels. These mice are characterized by high bone mass due to attenuated bone resorption as a result, at least in part, of insulin’s negative effect on osteoclasts.(68) However, there are diabesity models associated with both pronounced hyperglycemia and obesity, including New Zealand Obese (NZO/HlLt and NZO/HlBomDife), yellow agouti Avy/a, and the TALLYHO/JngJ mouse. The yellow agouti and TALLYHO/JngJ mice may be particularly useful in future studies because in these models the
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onset of T2D occurs early, thereby paralleling the rising epidemic of T2D in adolescence.(69,70) Although yellow agouti mice have normal bone mass, TALLYHO/JngJ mice have severely impaired peak bone mass and reduced bone formation, a characteristic feature of bone from adult and juvenile patients with T2D, respectively.
Summary: Future Challenges The relationship between the skeleton and adipose tissue is tied directly to fuel (glucose and lipids) utilization and energy generation at both the bone and whole-body level. The skeletal remodeling unit expends tremendous energy during both resorption and formation. Because it has now been established that there is central control over this process, and that the adipocyte is a major sensor as well as modulator of energy status, it seems certain that the CNS permits the integration of fuel homeostasis in the individual, balancing the demands of multiple tissues including bone. Glycolysis appears to be the preferred substrate for skeletal remodeling, hence glucose intolerance and T2D present a major challenge for the organism. Yet we still know little about the precise sequence of events that leads to insulin resistance in osteoblasts, nor do we know for sure that the changes in bone cell function and matrix constituents associated with T2D are related to insulin resistance or to other processes such as locally high concentrations of reactive oxygen species or other cell non-autonomous processes. The hypoxic environment of the bone marrow niche presents an additional challenge for developing experimental paradigms to study bone-fat interactions at the local level. Yet, ironically, HIF1a is a major determinant of lineage allocation and osteogenesis. Similarly, there are major challenges for defining the origin of the marrow adipocyte and its function in normal and impaired glucose homeostasis. At the patient level, there are no new drugs that specifically target diabetic bone disease, in part because the syndrome is heterogeneous, particularly for T2D. Moreover, some antidiabetic drugs cause bone loss. In addition, the molecular mechanisms related to glucose intolerance at the osteoblastic, osteocytic, and osteoclastic level are only just beginning to be elucidated. It is complicated further by changes in glycosylated proteins in the matrix that alter the structural aspects of the skeleton without directly affecting BMD. Although animal models have been helpful, they often do not fully reflect the complexity of the clinical disorder. Notwithstanding these many limitations, it is clear that we are entering a new era in bone biology—one that encompasses whole-body physiology. Hence, changes in energy metabolism have profound impacts on the skeleton and certainly alterations in the skeletal remodeling unit can affect whole body metabolism.
Disclosures The authors state that they have no conflicts of interest.
Acknowledgments This work was supported by funding from the American Diabetes Association (7-13-BS-089 to BLC) and the NIDDK (R24DK092759-05 to CJR). Authors’ role: BLC and CJR contributed equally to the literature search, discussion, and writing of this manuscript.
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