Liraglutide, a glucagon-like peptide-1 receptor agonist, facilitates osteogenic proliferation and differentiation in MC3T3-E1 cells through phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT), extracellular signal-related kinase (ERK)1/2, and cAMP/protein kinase A (PKA) signaling pathways involving β-catenin.
Wu Xuelun,Li Shilun,Xue Peng,Li Yukun
Experimental cell research
Previous studies have proven that glucagon-like peptide-1 (GLP-1) and its receptor agonist exert favorable anabolic effects on skeletal metabolism. However, whether GLP-1 could directly impact osteoblast-mediated bone formation is still controversial, and the underlying molecular mechanism remains to be elucidated. Thus in this paper, we investigated the effects of liraglutide, a glucagon-like peptide-1 (GLP-1) receptor agonist, on murine MC3T3-E1 preosteoblasts proliferation and differentiation and explored the potential cellular basis. Our study confirmed the presence of GLP-1R in MC3T3-E1, and demonstrated that liraglutide promotes osteoblasts proliferation at an intermediate concentration (100nM) and time (48h), upregulated the expression of osteoblastogenic biomarkers at various stages, and stimulated osteoblastic mineralization. Liraglutide also elevated the intracellular cAMP level and phosphorylation of AKT, ERK and β-catenin simultaneously with increased nuclear β-catenin content and transcriptional activity. Pretreatment of cells with the inhibitors LY294002, PD98059, H89 and GLP-1R and β-catenin siRNA partially blocked the liraglutide-induced signaling activation and attenuated the facilitating effect of liraglutide on MC3T3-E1 cells. Collectively, liraglutide was capable of acting upon osteoblasts directly through GLP-1R by activating PI3K/AKT, ERK1/2, cAMP/PKA/β-cat-Ser675 signaling to promote bone formation via GLP-1R. Thus, GLP-1 analogues may be potential therapeutic strategy for the treatment of osteoporosis in diabetics.
The Glucagon-Like Peptide-1 Receptor Agonist Exendin-4 Inhibits Lipopolysaccharide-Induced Osteoclast Formation and Bone Resorption via Inhibition of TNF- Expression in Macrophages.
Shen Wei-Ren,Kimura Keisuke,Ishida Masahiko,Sugisawa Haruki,Kishikawa Akiko,Shima Kazuhiro,Ogawa Saika,Qi Jiawei,Kitaura Hideki
Journal of immunology research
Glucagon-like peptide-1 (GLP-1) receptor agonists are an effective treatment approach for type 2 diabetes. Recently, anti-inflammatory effects of GLP-1 receptor agonists have also been reported. Lipopolysaccharide (LPS) induces inflammation and osteoclast formation. In this study, we investigated the effect of exendin-4, a widely used GLP-1 receptor agonist, in LPS-induced osteoclast formation and bone resorption. LPS with or without exendin-4 was administered on mouse calvariae by daily subcutaneous injection. The number of osteoclasts, the ratio of bone resorption pits, and the level of C-terminal cross-linked telopeptide of type I collagen (CTX) were significantly lower in LPS- and exendin-4-coadministered mice than in mice administered with LPS alone. RANKL and TNF- mRNA expression levels were lower in the exendin-4- and LPS-coadministered group than in the LPS-administered group. Our results showed no direct effects of exendin-4 on RANKL-induced osteoclast formation, TNF--induced osteoclast formation, or LPS-induced RANKL expression in stromal cells. Conversely, TNF- mRNA expression was inhibited in the exendin-4- and LPS-cotreated macrophages compared with cells treated with LPS alone. These results indicate that the GLP-1 receptor agonist exendin-4 may inhibit LPS-induced osteoclast formation and bone resorption by inhibiting LPS-induced TNF- production in macrophages.
Glucagon-like peptide-1(GLP-1) receptor agonists: potential to reduce fracture risk in diabetic patients?
Luo Guojing,Liu Hong,Lu Hongyun
British journal of clinical pharmacology
This review summarizes current knowledge about glucagon-like peptide 1 receptor agonists (GLP-1 RA) and their effects on bone metabolism and fracture risk. Recent in vivo and in vitro experiments indicated that GLP-1 RA could improve bone metabolism. GLP-1 could affect the fat-bone axis by promoting osteogenic differentiation and inhibiting adipogenic differentiation of bone mesenchymal precursor cells (BMSCs), which express the GLP-1 receptor. GLP-1 RA may also influence the balance between osteoclasts and osteoblasts, thus leading to more bone formation and less bone resorption. Wnt/β-catenin signalling is involved in this process. Mature osteocytes, which also express the GLP-1 receptor, produce sclerostin which inhibits Wnt/β-catenin signalling by binding to low density lipoprotein receptor-related protein (LRP) 5 and preventing the binding of Wnt. GLP-1 RA also decreases the expression of sclerostin (SOST) and circulating levels of SOST. In addition, GLP-1 receptors are expressed in thyroid C cells, where GLP-1 induces calcitonin release and thus indirectly inhibits bone resorption. Furthermore, GLP-1 RA influences the osteoprotegerin(OPG)/receptor activator of nuclear factor-κB ligand (RANKL)/receptor activator of nuclear factor-κB (RANK) system by increasing OPG gene expression, and thus reverses the decreased bone mass in rats models. However, a recent meta-analysis and a cohort study did not show a significant relationship between GLP-RA use and fracture risk. Future clinical trials will be necessary to investigate thoroughly the relationship between GLP-1 RA use and fracture risk in diabetic patients.
Activation of GLP-1 Receptor Promotes Bone Marrow Stromal Cell Osteogenic Differentiation through β-Catenin.
Meng Jingru,Ma Xue,Wang Ning,Jia Min,Bi Long,Wang Yunying,Li Mingkai,Zhang Huinan,Xue Xiaoyan,Hou Zheng,Zhou Ying,Yu Zhibin,He Gonghao,Luo Xiaoxing
Stem cell reports
Glucagon-like peptide 1 (GLP-1) plays an important role in regulating bone remodeling, and GLP-1 receptor agonist shows a positive relationship with osteoblast activity. However, GLP-1 receptor is not found in osteoblast, and the mechanism of GLP-1 receptor agonist on regulating bone remodeling is unclear. Here, we show that the GLP-1 receptor agonist exendin-4 (Ex-4) promoted bone formation and increased bone mass and quality in a rat unloading-induced bone loss model. These functions were accompanied by an increase in osteoblast number and serum bone formation markers, while the adipocyte number was decreased. Furthermore, GLP-1 receptor was detected in bone marrow stromal cells (BMSCs), but not in osteoblast. Activation of GLP-1 receptor by Ex-4 promoted the osteogenic differentiation and inhibited BMSC adipogenic differentiation through regulating PKA/β-catenin and PKA/PI3K/AKT/GSK3β signaling. These findings reveal that GLP-1 receptor regulates BMSC osteogenic differentiation and provide a molecular basis for therapeutic potential of GLP-1 against osteoporosis.
Liraglutide Attenuates Preestablished Atherosclerosis in Apolipoprotein E-Deficient Mice via Regulation of Immune Cell Phenotypes and Proinflammatory Mediators.
Bruen Robyn,Curley Seán,Kajani Sarina,Lynch Gina,O'Reilly Marcella E,Dillon Eugène T,Brennan Eoin P,Barry Mary,Sheehan Stephen,McGillicuddy Fiona C,Belton Orina
The Journal of pharmacology and experimental therapeutics
We have shown that the glucagon-like peptide-1 receptor agonist (GLP-1RA) liraglutide (Lir) inhibits development of early atherosclerosis in vivo by modulating immune cell function. We hypothesized that Lir could attenuate pre-established disease by modulating monocyte or macrophage phenotype to induce atheroprotective responses. Human atherosclerotic plaques obtained postendarterectomy and human peripheral blood macrophages were treated ex vivo with Lir. In parallel, apolipoprotein E-deficient (ApoE) mice received a high-fat, high-cholesterol diet to induce atherosclerosis for 8 weeks, after which ApoE mice received 300 g/kg of Lir daily or vehicle control for a further 4 weeks to investigate the attenuation of atherosclerosis. Lir inhibited proinflammatory monocyte chemoattractant protein-1 secretion from human endarterectomy samples and monocyte chemoattractant protein-1, tumor necrosis factor-, and interleukin (IL)-1 secretion from human macrophages after ex vivo treatment. An increase in CD206 mRNA and IL-10 secretion was also detected, which implies resolution of inflammation. Importantly, Lir significantly attenuated pre-established atherosclerosis in ApoE mice in the whole aorta and aortic root. Proteomic analysis of ApoE bone marrow cells showed that Lir upregulated the proinflammatory cathepsin protein family, which was abolished in differentiated macrophages. In addition, flow cytometry analysis of bone marrow cells induced a shift toward reduced proinflammatory and increased anti-inflammatory macrophages. We concluded that Lir attenuates pre-established atherosclerosis in vivo by altering proinflammatory mediators. This is the first study to describe a mechanism through which Lir attenuates atherosclerosis by increasing bone marrow proinflammatory protein expression, which is lost in differentiated bone marrow-derived macrophages. This study contributes to our understanding of the anti-inflammatory and cardioprotective role of GLP-1RAs. SIGNIFICANCE STATEMENT: It is critical to understand the mechanisms through which liraglutide (Lir) mediates a cardioprotective effect as many type 2 diabetic medications increase the risk of myocardial infarction and stroke. We have identified that Lir reduces proinflammatory immune cell populations and mediators from plaque-burdened murine aortas in vivo and augments proresolving bone marrow-derived macrophages in attenuation of atherosclerotic disease, which provides further insight into the atheroprotective effect of Lir.
Liraglutide treatment reduced interleukin-6 in adults with type 1 diabetes but did not improve established autonomic or polyneuropathy.
Brock Christina,Hansen Christian Stevns,Karmisholt Jesper,Møller Holger Jon,Juhl Anne,Farmer Adam Donald,Drewes Asbjørn Mohr,Riahi Sam,Lervang Hans Henrik,Jakobsen Poul Erik,Brock Birgitte
British journal of clinical pharmacology
AIMS:Type 1 diabetes can be complicated with neuropathy that involves immune-mediated and inflammatory pathways. Glucagon-like peptide-1 receptor agonists such as liraglutide, have shown anti-inflammatory properties, and thus we hypothesized that long-term treatment with liraglutide induced diminished inflammation and thus improved neuronal function. METHODS:The study was a randomized, double-blinded, placebo-controlled trial of adults with type 1 diabetes and confirmed symmetrical polyneuropathy. They were randomly assigned (1:1) to receive either liraglutide or placebo. Titration was 6 weeks to 1.2-1.8 mg/d, continuing for 26 weeks. The primary endpoint was change in latency of early brain evoked potentials. Secondary endpoints were changes in proinflammatory cytokines, cortical evoked potential, autonomic function and peripheral neurophysiological testing. RESULTS:Thirty-nine patients completed the study, of whom 19 received liraglutide. In comparison to placebo, liraglutide reduced interleukin-6 (-22.6%; 95% confidence interval [CI]: -38.1, -3.2; P = .025) with concomitant numerical reductions in other proinflammatory cytokines. However neuronal function was unaltered at the central, autonomic or peripheral level. Treatment was associated with -3.38 kg (95% CI: -5.29, -1.48; P < .001] weight loss and a decrease in urine albumin/creatinine ratio (-40.2%; 95% CI: -60.6, -9.5; P = .02). CONCLUSION:Hitherto, diabetic neuropathy has no cure. Speculations can be raised whether mechanism targeted treatment, e.g. lowering the systemic level of proinflammatory cytokines may lead to prevention or treatment of the neuroinflammatory component in early stages of diabetic neuropathy. If ever successful, this would serve as an example of how fundamental mechanistic principles are translated into clinical practice similar to those applied in the cardiovascular and nephrological clinic.
Liraglutide Improves the Kidney Function in a Murine Model of Chronic Kidney Disease.
Ougaard Maria E,Sembach Frederikke E,Jensen Henrik E,Pyke Charles,Knudsen Lotte B,Kvist Peter H
BACKGROUND:Chronic kidney disease (CKD) is a global health burden, and the current treatment options only slow down the disease progression. GLP-1 receptor agonists (GLP-1 RA) have shown a renal protective effect in models of CKD; however, the mechanism behind the beneficial effect is not understood. In this study, we investigate the effect of the GLP-1 RA liraglutide in the nephrotoxic serum nephritis (NTN) CKD model. Moreover, we compare the gene expression pattern of liraglutide-treated mice to the gene expression pattern of mice treated with the angiotensin converting enzyme inhibitor, enalapril. METHODS:The effect of liraglutide was tested in the NTN model by evaluating the glomerular filtration rate (GFR), albuminuria, mesangial expansion, renal fibrosis, and renal inflammation. Furthermore, the regulation of selected genes involved in CKD and in glomerular, cortical tubulointerstitial, and whole kidney structures was analyzed using a gene expression array on samples following laser capture microdissection. RESULTS:Treatment with liraglutide improved CKD hallmarks including GFR, albuminuria, mesangial expansion, renal inflammation, and renal fibrosis. The gene expression revealed that both liraglutide and enalapril reversed the regulation of several fibrosis and inflammation associated genes, which are also regulated in human CKD patients. Furthermore, liraglutide and enalapril both regulated genes in the kidney involved in blood pressure control. CONCLUSIONS:Treatment with liraglutide improved the kidney function and diminished renal lesions in NTN-induced mice. Both liraglutide and enalapril reversed the regulation of genes involved in CKD and regulated genes involved in blood pressure control.
Liraglutide treatment improves endothelial function in the Ldlr-/- mouse model of atherosclerosis and affects genes involved in vascular remodelling and inflammation.
Bjørnholm Katrine Dahl,Skovsted Gry Freja,Mitgaard-Thomsen Anne,Rakipovski Günaj,Tveden-Nyborg Pernille,Lykkesfeldt Jens,Povlsen Gro Klitgaard
Basic & clinical pharmacology & toxicology
Recent clinical intervention studies have shown that the GLP1 analogue liraglutide lowers cardiovascular risk, but the underlying mechanism has not yet been fully elucidated. This study investigated the effects of liraglutide on endothelial function in the Ldlr-/- mouse model. Mice (n = 12/group) were fed Western diet (WD) or chow for 12 weeks followed by 4 weeks of treatment with liraglutide (1 mg/kg/day) or vehicle subcutaneously. Weight loss, blood lipid content, plaque burden, vasomotor function of the aorta and gene expression pattern in aorta and brachiocephalic artery were monitored. Liraglutide treatment significantly induced weight loss (P < .0001), decreased blood triglycerides (P < .0001) and total cholesterol (P < .0001) in WD-fed mice but did not decrease plaque burden. Liraglutide also improved endothelium-mediated dilation of the distal thoracis aorta (P = .0067), but it did not affect phenylephrine or sodium nitroprusside responses. Fluidigm analyses of 96 genes showed significantly altered expression of seven genes related to inflammation, vascular smooth muscle cells and extracellular matrix composition in liraglutide-treated animals. We conclude that treatment with liraglutide decreased endothelial dysfunction and that this could be linked to decreased inflammation or regulation of vascular remodelling.
Liraglutide Attenuates Non-Alcoholic Fatty Liver Disease in Mice by Regulating the Local Renin-Angiotensin System.
Yang Mengying,Ma Xiaoyi,Xuan Xiuping,Deng Hongjun,Chen Qi,Yuan Li
Frontiers in pharmacology
The renin-angiotensin system (RAS) is involved in the pathogenesis of non-alcoholic fatty liver disease (NAFLD) and represents a potential therapeutic target for NAFLD. Glucagon-like peptide-1 (GLP-1) signaling has been shown to regulate the RAS within various local tissues. In this study, we aimed to investigate the functional relationship between GLP-1 and the local RAS in the liver during NAFLD. Wild-type and ACE2 knockout mice were used to establish a high-fat-induced NAFLD model. After the mice were treated with liraglutide (a GLP-1 analogue) for 4 weeks, the key RAS component genes were up-regulated in the liver of NAFLD mice. Liraglutide treatment regulated the RAS balance, preventing a reduction in fatty acid oxidation gene expression and increasing gluconeogenesis and the expression of inflammation-related genes caused by NAFLD, which were impaired in ACE2 knockout mice. Liraglutide-treated HepG2 cells exhibited activation of the ACE2/Ang1-7/Mas axis, increased fatty acid oxidation gene expression, and decreased inflammation, which could be reversed by A779 and AngII. These results indicate that the local RAS in the liver becomes overactivated in response to NAFLD. Moreover, ACE2 knockout increases the severity of liver steatosis. Liraglutide has a negative and antagonistic effect on the ACE/AngII/AT1R axis, a positive impact on the ACE2/Ang1-7/Mas axis, and is mediated through the PI3K/AKT pathway. This may represent a potential new mechanism by which liraglutide improves NAFLD.
GLP‑1R agonists ameliorate peripheral nerve dysfunction and inflammation via p38 MAPK/NF‑κB signaling pathways in streptozotocin‑induced diabetic rats.
Ma Jingjing,Shi Min,Zhang Xiangcheng,Liu Xiaoning,Chen Juan,Zhang Ridong,Wang Xingzhou,Zhang Hong
International journal of molecular medicine
The present study aimed to investigate the mechanism of glucagon‑like peptide‑1 receptor (GLP‑1R) agonists in the progression of diabetic peripheral neuropathy (DPN) in streptozotocin (STZ)‑induced diabetic rats, through inflammatory signaling pathways. The DPN rat model was generated by intraperitoneal injection of STZ and then treated with the GLP‑1R agonist liraglutide or saline for 8 weeks. These animals were randomly divided into 4 groups (10 rats in each): The normal control + saline group, the normal control + liraglutide group, the diabetic + saline (DM) group and the diabetic + liraglutide (DML) group. The nerve conduction velocity (NCV) in the sciatic nerves of the rats was monitored over a period of 8 weeks. Peripheral serum was obtained for the measurement of blood glucose, tumor necrosis factor‑α (TNF‑α), interleukin‑6 (IL‑6) and IL‑1β level. The protein levels of phosphorylated (p‑) and total extracellular signal‑regulated kinase, c‑Jun NH2‑terminal kinases, p38 mitogen‑activated protein kinases (MAPK), and nuclear and cytoplasmic nuclear factor‑κB (NF‑κB) were measured through western blot analysis. Sciatic nerve mRNA expression levels of proinflammatory chemokines (TNF‑α, IL‑6 and IL‑1β), chemokines [monocyte chemoattractant protein‑1 (MCP‑1)], adhesion molecules [intercellular adhesion molecule 1 (ICAM‑1)], neurotrophic factors [neuritin, nerve growth factor (NGF) and neuron‑specific enolase (NSE)] and NADPH oxidase 4 (NOX4) were evaluated by reverse transcription-quantitative polymerase chain reaction. Subsequent to 8 weeks of treatment with liraglutide, the density of myelin nerve fibers was partially restored in the DML group. The delayed motor NCV and sensory NCV in the DML group were improved. The IOD value of NOX4 staining in the DML group (24.43±9.01) was reduced compared with that in the DM group (56.60±6.91). The levels of TNF‑α, IL‑1β and IL‑6 in the peripheral serum of the DML group were significantly suppressed compared with those of the DM group. It was also observed that the mRNA expression levels of TNF‑α, IL‑6, IL‑1β, MCP‑1, ICAM‑1 and NOX4 in the sciatic nerve were attenuated in the DML group. The mRNA expression of neuritin and NGF was significantly increased in the DML group compared with that of the DM group; NSE was reduced in the sciatic nerves of the DML group compared with that of the DM group. Additionally, the protein expression of p‑p38 MAPK and NF‑κB in the DML group was significantly suppressed. These data demonstrated that GLP‑1R agonists may prevent nerve dysfunction in the sciatic nerves of diabetic rats via p38 MAPK/NF‑κB signaling pathways independent of glycemic control. GLP‑1R agonists may be a useful therapeutic strategy for slowing the progression of DPN.
After the LEADER trial and SUSTAIN-6, how do we explain the cardiovascular benefits of some GLP-1 receptor agonists?
Vergès B,Charbonnel B
Diabetes & metabolism
Recent cardiovascular outcome trials - the LEADER with liragutide and SUSTAIN-6 with semaglutide - have shown significant reductions of major cardiovascular (CV) events with these glucagon-like peptide (GLP)-1 receptor agonists. Progressive separation of the treatment and placebo curves, starting clearly between 12 and 18 months of the trial period, and significant reductions in the risk of myocardial infarction and stroke, indicate that the beneficial CV effects observed with GLP-1 receptor agonists could be due to an antiatherogenic effect. So far, the reasons for such an effect of GLP-1 receptor agonists have not been entirely clear, although several hypotheses may be proposed. As the reductions in glycated haemoglobin and systolic blood pressure (SBP) in these trials were modest, and both trials lasted only a short period of time, reductions in hyperglycaemia and SBP are unlikely to be involved in the beneficial CV effects of GLP-1 receptor agonists. On the other hand, their effect on lipids and, in particular, the dramatic decrease in postprandial hypertriglyceridaemia may explain their beneficial CV actions. Reduction of body weight, including a significant decrease in visceral fat in patients using GLP-1 receptor agonists, may also have beneficial CV effects by reducing chronic proatherogenic inflammation. In addition, there are in-vitro data showing a direct anti-inflammatory effect with these agents that could also be involved in their beneficial CV effects. Moreover, studies in humans have shown significant beneficial effects on ischaemic myocardium after a very short treatment period, suggesting a direct effect of GLP-1 receptor agonists on myocardium, although the precise mechanism remains unclear. Finally, as a reduction in insulin resistance has been associated with a decrease in CV risk, it cannot be ruled out that the lowering of insulin resistance induced by GLP-1 receptor agonists might also be involved in their beneficial CV actions.
Exenatide mitigated diet-induced vascular aging and atherosclerotic plaque growth in ApoE-deficient mice under chronic stress.
Yang Guang,Lei Yanna,Inoue Aiko,Piao Limei,Hu Lina,Jiang Haiying,Sasaki Takeshi,Wu Hongxian,Xu Wenhu,Yu Chenglin,Zhao Guangxian,Ogasawara Shinyu,Okumura Kenji,Kuzuya Masafumi,Cheng Xian-Wu
BACKGROUND AND AIMS:Exposure to psychosocial stress is a risk factor for cardiovascular disorders. Because the glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) agonist prevents cardiovascular injury, we investigated the beneficial effects and mechanism of the GLP-1 analogue exenatide on stress-related vascular senescence and atherosclerosis in apolipoprotein E-deficient (ApoE) mice fed a high-fat (HF) diet. METHODS:ApoE mice fed the HF diet were assigned to non-stressed and immobilized-stress groups for 12 weeks. Mice fed the HF diet were divided into 2 groups and administered vehicle or exenatide for 12 weeks under stress conditions. RESULTS:Chronic stress enhanced vascular endothelial senescence and atherosclerotic plaque growth. The stress increased the levels of plasma depeptidyl peptidase-4 activity and decreased the levels of plasma GLP-1 and both plasma and adipose adiponectin (APN). As compared with the mice subjected to stress alone, the exenatide-treated mice had decreased plaque microvessel density, macrophage accumulation, broken elastin, and enhanced plaque collagen volume, and lowered levels of peroxisome proliferator-activated receptor-α, gp91phox osteopontin, C-X-C chemokine receptor-4, toll-like receptor-2 (TLR2), TLR4, and cathepsins K, L, and S mRNAs and/or proteins. Exenatide reduced aortic matrix metalloproteinase-9 (MMP-9) and MMP-2 gene expression and activities. Exenatide also stimulated APN expression of preadipocytes and inhibited ox-low density lipoprotein-induced foam cell formation of monocytes in stressed mice. CONCLUSIONS:These results indicate that the exenatide-mediated beneficial vascular actions are likely attributable, at least in part, to the enhancement of APN production and the attenuation of plaque oxidative stress, inflammation, and proteolysis in ApoE mice under chronic stress.
Anti-atherogenic and anti-inflammatory properties of glucagon-like peptide-1, glucose-dependent insulinotropic polypepide, and dipeptidyl peptidase-4 inhibitors in experimental animals.
Hirano Tsutomu,Mori Yusaku
Journal of diabetes investigation
We reported that native incretins, liraglutide and dipeptidyl peptidase-4 inhibitors (DPP-4i) all confer an anti-atherosclerotic effect in apolipoprotein E-null (Apoe (-/-)) mice. We confirmed the anti-atherogenic property of incretin-related agents in the mouse wire injury model, in which the neointimal formation in the femoral artery is remarkably suppressed. Furthermore, we showed that DPP-4i substantially suppresses plaque formation in coronary arteries with a marked reduction in the accumulation of macrophages in cholesterol-fed rabbits. DPP-4i showed an anti-atherosclerotic effect in Apoe (-/-) mice mainly through the actions of glucagon-like peptide-1 and glucose-dependent insulinotropic polypepide. However, the dual incretin receptor antagonists partially attenuated the suppressive effect of DPP-4i on atherosclerosis in diabetic Apoe (-/-) mice, suggesting an incretin-independent mechanism. Exendin-4 and glucose-dependent insulinotropic polypepide elicited cyclic adenosine monophosphate generation, and suppressed the lipopolysaccharide-induced gene expression of inflammatory molecules, such as interleukin-1β, interleukin-6 and tumor necrosis factor-α, in U937 human monocytes. This suppressive effect, however, was attenuated by an inhibitor of adenylate cyclase and mimicked by 8-bromo-cyclic adenosine monophosphate or forskolin. DPP-4i substantially suppressed the lipopolysaccharide-induced expression of inflammatory cytokines without affecting cyclic adenosine monophosphate generation or cell proliferation. DPP-4i more strongly suppressed the lipopolysaccharide-induced gene expression of inflammatory molecules than incretins, most likely through inactivation of CD26. Glucagon-like peptide-1 and glucose-dependent insulinotropic polypepide suppressed oxidized low-density lipoprotein-induced macrophage foam cell formation in a receptor-dependent manner, which was associated with the downregulation of acyl-coenzyme A cholesterol acyltransferase-1 and CD36, as well as the up-regulation of adenosine triphosphate-binding cassette transporter A1. Our studies strongly suggest that incretin-related agents have favorable effects on macrophage-driven atherosclerosis in experimental animals.
Glucagon-like peptide-1 receptor signalling reduces microvascular thrombosis, nitro-oxidative stress and platelet activation in endotoxaemic mice.
Steven Sebastian,Jurk Kerstin,Kopp Maximilian,Kröller-Schön Swenja,Mikhed Yuliya,Schwierczek Kathrin,Roohani Siyer,Kashani Fatemeh,Oelze Matthias,Klein Thomas,Tokalov Sergey,Danckwardt Sven,Strand Susanne,Wenzel Philip,Münzel Thomas,Daiber Andreas
British journal of pharmacology
BACKGROUND AND PURPOSE:Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated with organ dysfunction and high mortality. The present studies aimed to investigate whether inhibition of dipeptidyl peptidase-4 (DPP-4) and supplementation with glucagon-like peptide-1 (GLP-1) receptor agonists improved endotoxaemia-associated microvascular thrombosis via immunomodulatory effects. EXPERIMENTAL APPROACH:Endotoxaemia was induced in C57BL/6J mice by a single injection of LPS (17.5 mg kg for survival and 10 mg kg for all other studies). For survival studies, treatment was started 6 h after LPS injection. For all other studies, drugs were injected 48 h before LPS treatment. KEY RESULTS:Mice treated with LPS alone showed severe thrombocytopenia, microvascular thrombosis in the pulmonary circulation (fluorescence imaging), increased LDH activity, endothelial dysfunction and increased markers of inflammation in aorta and whole blood (leukocyte-dependent oxidative burst, nitrosyl-iron haemoglobin, a marker of nitrosative stress, and expression of inducible NOS). Treatment with the DPP-4 inhibitor linagliptin or the GLP-1 receptor agonist liraglutide, as well as genetic deletion of DPP-4 (DPP4 mice) improved all these parameters. In GLP-1 receptor-deficient mice, both linagliptin and liraglutide lost their beneficial effects and improvement of prognosis. Incubation of platelets and cultured monocytes (containing GLP-1 receptor protein) with GLP-1 receptor agonists inhibited the monocytic oxidative burst and platelet activation, with a GLP-1 receptor-dependent elevation of cAMP levels and PKA activation. CONCLUSIONS AND IMPLICATIONS:GLP-1 receptor activation in platelets by linagliptin and liraglutide strongly attenuated endotoxaemia-induced microvascular thrombosis and mortality by a cAMP/PKA-dependent mechanism, preventing systemic inflammation, vascular dysfunction and end organ damage. LINKED ARTICLES:This article is part of a themed section on Redox Biology and Oxidative Stress in Health and Disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.12/issuetoc.
Glucagon-like peptide 1-related peptides increase nitric oxide effects to reduce platelet activation.
Barale Cristina,Buracco Simona,Cavalot Franco,Frascaroli Chiara,Guerrasio Angelo,Russo Isabella
Thrombosis and haemostasis
Glucagon-like peptide 1 (GLP-1) is object of intensive investigation for not only its metabolic effects but also the protective vascular actions. Since platelets exert a primary role in the pathogenesis of atherosclerosis, inflammation and vascular complications, we investigated whether GLP-1 directly influences platelet reactivity. For this purpose, in platelets from 72 healthy volunteers we evaluated GLP-1 receptor (GLP-1R) expression and the effects of a 15-minute incubation with the native form GLP-1(7-36), the N-terminally truncated form GLP-1(9-36) and the GLP-1 analogue Liraglutide (100 nmol/l) on: i) aggregation induced by collagen or arachidonic acid (AA); ii) platelet function under shear stress; iii) cGMP and cAMP synthesis and cGMP-dependent protein kinase (PKG)-induced Vasodilator-Stimulated-Phosphoprotein (VASP) phosphorylation; iv) activation of the signalling molecules Phosphatidylinositol 3-Kinase (PI3-K)/Akt and Mitogen Activated Protein Kinase (MAPK)/ERK-1/2; and v) oxidative stress. Experiments were repeated in the presence of the nitric oxide donor Na-nitroprusside. We found that platelets constitutively express GLP-1R and that, independently of GLP-1R, GLP-1(7-36), GLP-1(9-36) and Liraglutide exert platelet inhibitory effects as shown by: a) increased NO-antiaggregating effects, b) increased the activation of the cGMP/PKG/VASP pathway, c) reduced the activation of PI3-K/Akt and MAPK/ERK-2 pathways, d) reduced the AA-induced oxidative stress. When the experiments were repeated in the presence of the antagonist of GLP-1R Exendin(9-39), the platelet inhibitory effects were maintained, thus indicating a mechanism independent of GLP-1R. In conclusion, GLP-1(7-36), its degradation product GLP-1(9-36) and Liraglutide exert similar inhibitory effects on platelet activation, suggesting a potential protective effect on the cardiovascular system.
GLP-1 secretion is increased by inflammatory stimuli in an IL-6-dependent manner, leading to hyperinsulinemia and blood glucose lowering.
Kahles Florian,Meyer Christina,Möllmann Julia,Diebold Sebastian,Findeisen Hannes M,Lebherz Corinna,Trautwein Christian,Koch Alexander,Tacke Frank,Marx Nikolaus,Lehrke Michael
Hypoglycemia and hyperglycemia are both predictors for adverse outcome in critically ill patients. Hyperinsulinemia is induced by inflammatory stimuli as a relevant mechanism for glucose lowering in the critically ill. The incretine hormone GLP-1 was currently found to be induced by endotoxin, leading to insulin secretion and glucose lowering under inflammatory conditions in mice. Here, we describe GLP-1 secretion to be increased by a variety of inflammatory stimuli, including endotoxin, interleukin-1β (IL-1β), and IL-6. Although abrogation of IL-1 signaling proved insufficient to prevent endotoxin-dependent GLP-1 induction, this was abolished in the absence of IL-6 in respective knockout animals. Hence, we found endotoxin-dependent GLP-1 secretion to be mediated by an inflammatory cascade, with IL-6 being necessary and sufficient for GLP-1 induction. Functionally, augmentation of the GLP-1 system by pharmacological inhibition of DPP-4 caused hyperinsulinemia, suppression of glucagon release, and glucose lowering under endotoxic conditions, whereas inhibition of the GLP-1 receptor led to the opposite effect. Furthermore, total GLP-1 plasma levels were profoundly increased in 155 critically ill patients presenting to the intensive care unit (ICU) in comparison with 134 healthy control subjects. In the ICU cohort, GLP-1 plasma levels correlated with markers of inflammation and disease severity. Consequently, GLP-1 provides a novel link between the immune system and the gut with strong relevance for metabolic regulation in context of inflammation.
Endothelial GLP-1 (Glucagon-Like Peptide-1) Receptor Mediates Cardiovascular Protection by Liraglutide In Mice With Experimental Arterial Hypertension.
Helmstädter Johanna,Frenis Katie,Filippou Konstantina,Grill Alexandra,Dib Mobin,Kalinovic Sanela,Pawelke Franziska,Kus Kamil,Kröller-Schön Swenja,Oelze Matthias,Chlopicki Stefan,Schuppan Detlef,Wenzel Philip,Ruf Wolfram,Drucker Daniel J,Münzel Thomas,Daiber Andreas,Steven Sebastian
Arteriosclerosis, thrombosis, and vascular biology
OBJECTIVE:Cardiovascular outcome trials demonstrated that GLP-1 (glucagon-like peptide-1) analogs including liraglutide reduce the risk of cardiovascular events in type 2 diabetes mellitus. Whether GLP-1 analogs reduce the risk for atherosclerosis independent of glycemic control is challenging to elucidate as the GLP-1R (GLP-1 receptor) is expressed on different cell types, including endothelial and immune cells. Approach and Results: Here, we reveal the cardio- and vasoprotective mechanism of the GLP-1 analog liraglutide at the cellular level in a murine, nondiabetic model of arterial hypertension. Wild-type (C57BL/6J), global (), as well as endothelial () and myeloid cell-specific knockout mice () of the GLP-1R were studied, and arterial hypertension was induced by angiotensin II. Liraglutide treatment normalized blood pressure, cardiac hypertrophy, vascular fibrosis, endothelial dysfunction, oxidative stress, and vascular inflammation in a GLP-1R-dependent manner. Mechanistically, liraglutide reduced leukocyte rolling on the endothelium and infiltration of myeloid Ly6GLy6C and Ly6GLy6C cells into the vascular wall. As a consequence, liraglutide prevented vascular oxidative stress, reduced S-glutathionylation as a marker of eNOS (endothelial NO synthase) uncoupling, and increased NO bioavailability. Importantly, all of these beneficial cardiovascular effects of liraglutide persisted in myeloid cell GLP-1R-deficient () mice but were abolished in global () and endothelial cell-specific () GLP-1R knockout mice. CONCLUSIONS:GLP-1R activation attenuates cardiovascular complications of arterial hypertension by reduction of vascular inflammation through selective actions requiring the endothelial but not the myeloid cell GLP-1R.
Glucagon-like peptide-1 (GLP-1) analog liraglutide inhibits endothelial cell inflammation through a calcium and AMPK dependent mechanism.
Krasner Nadia M,Ido Yasuo,Ruderman Neil B,Cacicedo Jose M
Liraglutide is a glucagon-like peptide-1 (GLP-1) mimetic used for the treatment of Type 2 diabetes. Similar to the actions of endogenous GLP-1, liraglutide potentiates the post-prandial release of insulin, inhibits glucagon release and increases satiety. Recent epidemiological studies and clinical trials have suggested that treatment with GLP-1 mimetics may also diminish the risk of cardiovascular disease in diabetic patients. The mechanism responsible for this effect has yet to be determined; however, one possibility is that they might do so by a direct effect on vascular endothelium. Since low grade inflammation of the endothelium is an early event in the pathogenesis of atherosclerotic cardiovascular disease (ASCVD), we determined the effects of liraglutide on inflammation in cultured human aortic endothelial cells (HAECs). Liraglutide reduced the inflammatory responses to TNFα and LPS stimulation, as evidenced by both reduced protein expression of the adhesion molecules VCAM-1 and E-Selectin, and THP-1 monocyte adhesion. This was found to result from increased cell Ca2+ and several molecules sensitive to Ca2+ with known anti inflammatory actions in endothelial cells, including CaMKKβ, CaMKI, AMPK, eNOS and CREB. Treatment of the cells with STO-609, a CaMKK inhibitor, diminished both the activation of AMPK, CaMKI and the inhibition of TNFα and LPS-induced monocyte adhesion by liraglutide. Likewise, expression of an shRNA against AMPK nullified the anti-inflammatory effects of liraglutide. The results indicate that liraglutide exerts a strong anti-inflammatory effect on HAECs. They also demonstrate that this is due to its ability to increase intracellular Ca2+ and activate CAMKKβ, which in turn activates AMPK.
Mechanism of Glucagon-Like Peptide 1 Improvements in Type 2 Diabetes Mellitus and Obesity.
Nolen-Doerr Eric,Stockman Mary-Catherine,Rizo Ivania
Current obesity reports
PURPOSE OF REVIEW:The purpose of this review is to emphasize the pivotal role of glucagon-like peptide 1 (GLP-1) in tackling the parallel epidemics of obesity and type 2 diabetes (T2DM). RECENT FINDINGS:GLP-1-based therapies and in particular GLP-1 receptor agonists (GLP-1 RA) have proven to be effective in lowering blood glucose and decreasing weight. GLP-1 RA not only mitigate these significant medical burdens but also result in weight loss and weight loss independent factors that decrease cardiovascular disease (CVD) and microvascular complications of T2DM, such as diabetic nephropathy. GLP-1-based therapies are critical for a patient-centered approach in choosing appropriate pharmacotherapy for T2DM and obesity while also taking into consideration comorbidities, such as cardiovascular and chronic kidney diseases.
Bone disease in diabetes.
Isidro M Luisa,Ruano Belén
Current diabetes reviews
The relationship between diabetes and bone disease is complex. While low bone mineral density (BMD) is consistently observed in type 1 diabetes (T1DM), in type 2 diabetes (T2DM) bone mineral density is similar to or higher than in non diabetic subjects. Yet, for both types of diabetes bone appears to be more fragile for a given density. Recent meta-analyses and cohort studies confirm that T1DM and T2DM are associated with higher fracture risk. Many factors influence the probability of fractures. Diabetes can affect bone through multiple pathways including obesity, changes in insulin levels, higher concentrations of advanced glycation end products in collagen, increased urinary excretion coupled with lower intestinal absorption of calcium, inappropriate homeostatic response of parathyroid hormone secretion, complex alterations of vitamin D regulation, reduced renal function, lower insulin-like growth factor-I, microangiopathy, and inflammation. Data on cellular mechanisms and experimental models are extensive, but the relevance of each one of these factors to the clinical situation is unclear. In this article we review the pathophysiological mechanisms potentially involved in the altered BMD found in diabetic patients, show data on the increased risk of fractures, and speculate on the potential causes of the increased risk of fractures in this context. Finally, we comment on the prevention and treatment of osteoporosis in diabetes, although the lack of trials testing the use of pharmacotherapy on preventing fractures in this context is emphasized.
Novel skeletal effects of glucagon-like peptide-1 (GLP-1) receptor agonists.
Mabilleau Guillaume,Pereira Marie,Chenu Chantal
The Journal of endocrinology
Type 2 diabetes mellitus (T2DM) leads to bone fragility and predisposes to increased risk of fracture, poor bone healing and other skeletal complications. In addition, some anti-diabetic therapies for T2DM can have notable detrimental skeletal effects. Thus, an appropriate therapeutic strategy for T2DM should not only be effective in re-establishing good glycaemic control but also in minimising skeletal complications. There is increasing evidence that glucagon-like peptide-1 receptor agonists (GLP-1RAs), now greatly prescribed for the treatment of T2DM, have beneficial skeletal effects although the underlying mechanisms are not completely understood. This review provides an overview of the direct and indirect effects of GLP-1RAs on bone physiology, focusing on bone quality and novel mechanisms of action on the vasculature and hormonal regulation. The overall experimental studies indicate significant positive skeletal effects of GLP-1RAs on bone quality and strength although their mechanisms of actions may differ according to various GLP-1RAs and clinical studies supporting their bone protective effects are still lacking. The possibility that GLP-1RAs could improve blood supply to bone, which is essential for skeletal health, is of major interest and suggests that GLP-1 anti-diabetic therapy could benefit the rising number of elderly T2DM patients with osteoporosis and high fracture risk.
The dataset of methylglyoxal activating p38 and p44/42 pathway in osteoclast.
Lee Kwan Ming,Lee Cheuk Yan,Zhang Ge,Lyu Aiping,Yue Kevin Kin Man
Data in brief
Diabetes mellitus (DM) is a kind of chronic metabolic disease that could be characterized by uncontrollable high blood glucose (hyperglycemia) over a prolonged period and diverse complications in various organs. These complications include activation of stress responses in bone such as oxidative stress and inflammation, which have been implicated in various bone diseases, including osteoporosis. Non-enzymatic glycation of proteins form and accumulate in patients under hyperglycemia condition. Methylglyoxal (MG) is a reactive advanced glycation end-product precursor. Abnormal high concentration of MG was in serum of diabetic patients. It was proven that MG induces various stress responses. This indicates that it might possibly the key metabolite leading to diabetes-associated bone loss. In this data report, using cell models, the underlying mechanism of methylglyoxal on osteoclast that may lead to bone loss was investigated. In cell cultures, RAW264.7, Macrophages, was treated with methylglyoxal and gene expressions of osteoclast bone biomarkers were investigated. Furthermore, the inhibitions of p38 and p44/42 activities were employed to investigate the osteoclast biomarkers CTSK, OSCAR, and TRACP5 gene expressions. These data implied that MG activated the p38 and p44/42, which was reported to regulate proliferation and differentiation of osteoclast. However, the decreasing MAPK though siRNA knockdown did not change expression of those target markers, TRACP5, OSCAR, and CTSK, in mRNA level. The effects of MG to other osteoclast markers through p38 and p44/42 would be worth to be investigated. For more insight please see Methylglyoxal Activates Osteoclasts through JNK Pathway leading to Osteoporosis.
Advanced glycation end products and diabetic complications.
Singh Varun Parkash,Bali Anjana,Singh Nirmal,Jaggi Amteshwar Singh
The Korean journal of physiology & pharmacology : official journal of the Korean Physiological Society and the Korean Society of Pharmacology
During long standing hyperglycaemic state in diabetes mellitus, glucose forms covalent adducts with the plasma proteins through a non-enzymatic process known as glycation. Protein glycation and formation of advanced glycation end products (AGEs) play an important role in the pathogenesis of diabetic complications like retinopathy, nephropathy, neuropathy, cardiomyopathy along with some other diseases such as rheumatoid arthritis, osteoporosis and aging. Glycation of proteins interferes with their normal functions by disrupting molecular conformation, altering enzymatic activity, and interfering with receptor functioning. AGEs form intra- and extracellular cross linking not only with proteins, but with some other endogenous key molecules including lipids and nucleic acids to contribute in the development of diabetic complications. Recent studies suggest that AGEs interact with plasma membrane localized receptors for AGEs (RAGE) to alter intracellular signaling, gene expression, release of pro-inflammatory molecules and free radicals. The present review discusses the glycation of plasma proteins such as albumin, fibrinogen, globulins and collagen to form different types of AGEs. Furthermore, the role of AGEs in the pathogenesis of diabetic complications including retinopathy, cataract, neuropathy, nephropathy and cardiomyopathy is also discussed.
Bone disorders associated with diabetes mellitus and its treatments.
Cortet Bernard,Lucas Stéphanie,Legroux-Gerot Isabelle,Penel Guillaume,Chauveau Christophe,Paccou Julien
Joint bone spine
Both type 1 and type 2 diabetes mellitus are associated with bone disorders, albeit via different mechanisms. Early studies in patients with type 1 diabetes suggested a 10-fold increase in the hip fracture risk compared to non-diabetic controls. Meta-analyses published more recently indicate a somewhat smaller risk increase, with odds ratios of 6 to 7. Diminished bone mineral density is among the contributors to the increased fracture risk. Both types of diabetes are associated with decreased bone strength related to low bone turnover. The multiple and interconnected pathophysiological mechanisms underlying the bone disorders seen in type 1 diabetes include insulin deficiency, accumulation of advanced glycation end products, bone microarchitecture alterations, changes in bone marrow fat content, low-grade inflammation, and osteocyte dysfunction. The bone alterations are less severe in type 2 diabetes. Odds ratios for hip fractures have ranged across studies from 1.2 to 1.7, and bone mineral density is higher than in non-diabetic controls. The odds ratio is about 1.2 for all bone fragility fractures combined. The pathophysiological mechanisms are complex, particularly as obesity is very common in patients with type 2 diabetes and is itself associated with an increased risk of fractures at specific sites (humerus, tibia, and ankle). The main mechanisms underlying the bone fragility are an increase in the risk of falls, sarcopenia, disorders of carbohydrate metabolism, vitamin D deficiency, and alterations in cortical bone microarchitecture and bone matrix. The medications used to treat both types of diabetes do not seem to play a major role. Nevertheless, thiazolidinediones and, to a lesser extent, sodium-glucose cotransporter inhibitors may have adverse effects on bone, whereas metformin may have beneficial effects. For the most part, the standard management of bone fragility applies to patients with diabetes. However, emphasis should be placed on preventing falls, which are particularly common in this population. Finally, there is some evidence to suggest that anti-fracture treatments are similarly effective in patients with and without diabetes.
Molecular Modulation of Osteoblasts and Osteoclasts in Type 2 Diabetes.
Rathinavelu Selvalakshmi,Guidry-Elizondo Crissy,Banu Jameela
Journal of diabetes research
Diabetes is a common disease affecting majority of populations worldwide. Since 1980, there has been an increase in the number of people diagnosed as prediabetic and diabetic. Diabetes is characterized by high levels of circulating glucose and leads to most microvascular and macrovascular complications such as retinopathy, nephropathy, neuropathy, stroke, and myocardial infarction. Bone marrow vascular disruption and increased adiposity are also linked to various complications in type II diabetes mellitus. In addition to these complications, type 2 diabetic patients also have fragile bones caused by faulty mineralization mainly due to increased adiposity among diabetic patients that affects both osteoblast and osteoclast functions. Other factors that increase fracture risk in diabetic patients are increased oxidative stress, inflammation, and drugs administered to diabetic patients. This review reports the modulation of different pathways that affect bone metabolism in diabetic conditions.
MLN64 deletion suppresses RANKL-induced osteoclastic differentiation and attenuates diabetic osteoporosis in streptozotocin (STZ)-induced mice.
Lei Chen,Xueming Han,Ruihang Duan
Biochemical and biophysical research communications
Diabetes mellitus is known as a main cause to induce osteoporosis. However, the pathogenesis of osteoporosis induced by diabetes has not been fully understood. MLN64 is highly homologous to the steroidogenic acute regulatory (STAR) protein, sharing the highly conserved START domain and exhibiting various biological activities. In the study, we attempted to explore the role of MLN64 in osteoporosis progression through the in vitro and in vivo studies. At first, the in vitro study suggested that MLN64 was over-expressed during the osteoclast differentiation induced by receptor activator of NF-kB ligand (RANKL). MLN64-knockdown markedly reduced the number of TRAP-positive multinucleated cells induced by RANKL, along with the down-regulation of specific genes related to osteoclastogenesis, including tartrate-resistant acid phosphatase (TRAP), nuclear factor of activated T cells (NFATc1), cathepsin K (CTSK), tartrate resistant (ACP5) and c-FOS and up-regulation of transcriptional receptor runt related transcription factor 2 (Runx2) and osteopontin (OPN). In contrast, over-expressing MLN64 significantly promoted the production of TRAP-positive multinucleated cells triggered by RANKL. Moreover, RANKL exposure led to remarkable increase in inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and MMP-1, which was attenuated by the knockdown of MLN64. Additionally, the expression of phosphorylated (p)IKKα, p-IκBα and p-nuclear factor (NF)-κB stimulated by RANKL was clearly alleviated by MLN64 silence. However, RANKL-induced inflammation was accelerated by MLN64 over-expression. Further, the in vivo study suggested that streptozotocin (STZ)-caused reduction in body weight of mice was improved by the loss of MLN64. STZ-induced diabetic mice exhibited higher levels of blood alkaline phosphatase (ALP) and TRAP, while being down-regulated in MLN64-knockout mice. MLN64 deletion reversed STZ-induced trabecular deleterious effects and stimulated bone remodeling. What's more, MLN64 knockout inhibited the expression levels of TRAP, RANKL and RANK in femoral heads of STZ mice, accompanied with the repression of inflammatory response. In conclusion, the results in our study suggested that MLN64 played a critical role in the meditation of osteoclastic differentiation, and its suppression alleviated diabetic osteoporosis in STZ-induced mice. Thus, MLN64 could be served as an essential target for developing effective therapeutic strategy to prevent diabetic osteoporosis.
Irbesartan attenuates advanced glycation end products-mediated damage in diabetes-associated osteoporosis through the AGEs/RAGE pathway.
Cheng Yan-Zhen,Yang Shuang-Li,Wang Ji-Yu,Ye Meng,Zhuo Xiao-Yun,Wang Li-Tao,Chen Hong,Zhang Hua,Yang Li
AIMS:Diabetes-associated osteoporosis is mainly caused by the formation and accumulation of advanced glycation end products (AGEs). Angiotensin II type 1 receptor blocker (ARB) has anabolic bone effects on the physicochemical properties of the bone in diabetes. We hypothesized that ARB could inhibit AGEs-induced deleterious effects. MAIN METHODS:In this study, we chose seven-week-old Leprdb/Lepr+ (db/+) and Leprdb/Leprdb (db/db) mice. After 12 week intervention by irbesartan, the microarchitecture and mechanical strength of the bone of seven-week-old db/db mice were investigated systematically. Meanwhile, the molecular mechanisms of the osteoblasts were analyzed, after AGEs or irbesartan were added to the culture. Also, intracellular formation of reactive oxygen species (ROS) was measured with DCF fluorescence. KEY FOUNDINGS:Results showed that 12-week irbesartan treatment could dramatically improve trabecular bone microarchitecture through increasing BV/TV (p = 0.003, +46.7%), Tb.N (p = 0.020, +52.0%), and decreasing that of Tb.Sp (p = 0.005, -21.2%) and SMI (p = 0.007, -26.4%), comparing with the db/db group. Irbesartan could also substantially raise biomechanical parameters including max load (p = 0.013, +20.7%), fracture load (p = 0.014, +70.5%), energy absorption (p = 0.019, +99.4%). Besides, it could inhibit AGEs-induced damage of cell proliferation and osteogenic differentiation of osteoblasts, as well as suppressing the activation of apoptosis caused by AGEs. Moreover, co-incubation with irbesartan could prevent the AGEs-induced increase of intracellular oxidative stress and RAGE expression in osteoblasts. SIGNIFICANCE:In conclusion, this study suggested that irbesartan might play a protective role in diabetes-related bone damages by blocking the deleterious effects of AGEs/RAGE-mediated oxidative stress. This may provide a revolutionary benefits to therapy with irbesartan on diabetic osteoporosis.
The New Possibilities from "Big Data" to Overlooked Associations Between Diabetes, Biochemical Parameters, Glucose Control, and Osteoporosis.
Current osteoporosis reports
PURPOSE OF REVIEW:To review current practices and technologies within the scope of "Big Data" that can further our understanding of diabetes mellitus and osteoporosis from large volumes of data. "Big Data" techniques involving supervised machine learning, unsupervised machine learning, and deep learning image analysis are presented with examples of current literature. RECENT FINDINGS:Supervised machine learning can allow us to better predict diabetes-induced osteoporosis and understand relative predictor importance of diabetes-affected bone tissue. Unsupervised machine learning can allow us to understand patterns in data between diabetic pathophysiology and altered bone metabolism. Image analysis using deep learning can allow us to be less dependent on surrogate predictors and use large volumes of images to classify diabetes-induced osteoporosis and predict future outcomes directly from images. "Big Data" techniques herald new possibilities to understand diabetes-induced osteoporosis and ascertain our current ability to classify, understand, and predict this condition.
Methylglyoxal activates osteoclasts through JNK pathway leading to osteoporosis.
Lee Kwan Ming,Lee Cheuk Yan,Zhang Ge,Lyu Aiping,Yue Kevin Kin Man
Diabetes mellitus is characterized by chronic hyperglycemia and its diverse complications. Hyperglycemia is associated with inflammatory responses in different organs, and diabetic patients have a higher risk of bone fracture due to increased bone weakness. Methylglyoxal, a reactive advanced glycation end product precursor, is known to have increased level in diabetic patients. The accumulation of methylglyoxal promotes inflammation and it may play a role in diabetes related osteoporosis. In this study, therefore, the underlying mechanism of methylglyoxal on osteoporosis was studied using both animal and cell models. In the animal model, rats were treated with either methylglyoxal or saline as control. In the cell model, the macrophage RAW264.7 was treated with methylglyoxal or vehicle control. Following the treatment, animal samples were harvested for micro-CT and real-time polymerase chain reaction analyses. Cell samples were harvested for MTT assay, RT-PCR, and Western Blotting analyses. In both animals and cell cultures, methylglyoxal was shown to induce osteoclastogenesis by increased gene expression of osteoclast bone biomarkers CTSK, OSCAR and TRACP5. Furthermore, in methylglyoxal-treated macrophages activation of the c-Jun N-terminal kinases signaling pathway was observed, and inhibition of JNK activities resulted in down-regulation of osteoclast biomarkers gene expressions. Our results therefore suggested that methylglyoxal may contribute to the progression of diabetes-related osteoporosis and imbalanced bone remodeling through JNK pathway in osteoclasts.
Therapeutic potential of Liuwei Dihuang pill against KDM7A and Wnt/β-catenin signaling pathway in diabetic nephropathy-related osteoporosis.
Liu Ming Ming,Dong Rui,Hua Zhen,Lv Nan Ning,Ma Yong,Huang Gui Cheng,Cheng Jian,Xu Hai Yan
The effects of Liuwei Dihuang pill (LWDH) on diabetic nephropathy-related osteoporosis (DNOP) are unclear. The present study aimed to evaluate the effects of LWDH on KDM7A and Wnt/β-catenin signaling pathway in DNOP rats and the high glucose-induced MC3T3-E1 cells. A DNOP model was prepared by streptozotocin in 9-week-old male Sprague-Dawley (SD) rats to evaluate the effects of LWDH. The cell viability and differentiation capacity of high glucose-induced MC3T3-E1 cells were determined by CCK-8 assay, Alizarin Red staining, and alkaline phosphatase (ALP) staining, respectively. Furthermore, the expressions of KDM7A and Wnt1/β-catenin pathway-related proteins were determined by Western blot analysis. Treatment of DNOP rats with LWDH could significantly ameliorate the general state, degradation of renal function, and renal pathological changes. LWDH decreased the levels of TNF-α, IL-6, IL-8, IL-1β, ALP, and TRAP, and increased the calcium, phosphorus in serum, as well as decreased the level of the calcium and phosphorus in the urine. Besides, LWDH significantly improved bone mineral density (BMD), bone volume (BV), and the bone microstructure of DNOP rats. Moreover, LWDH increased the levels of the elastic modulus, ultimate load, and bending strength in the femurs. In MC3T3-E1 cells, serum-containing LWDH significantly increases in cell viability and osteoblastic differentiation capability. The expression of α-SMA, vimentin, KDM7A, Wnt1 and β-catenin were significantly down-regulated, and the E-cadherin, H3K9-Me2, H3K27-Me2, BMP-4, BMP-7, Runx2, osteocalcin, and Col1a1 were significantly up-regulated with LWDH treatment. The present study shows that LWDH has a therapeutic effect on DNOP, in part, through down-regulation of KDM7A and Wnt/β-catenin pathway.
Comparison of osteoporosis in US adults with type 1 and type 2 diabetes mellitus.
DeShields S C,Cunningham T D
Journal of endocrinological investigation
PURPOSE:We examined bone mineral density (BMD) and osteoporosis prevalence in those with type 1 compared to type 2 diabetes derived from a nationally representative sample from the civilian community in the United States. METHODS:Data from the National Health and Nutrition Examination Survey (NHANES) for 2005-2006, 2007-2008, 2009-2010, and 2013-2014 were merged to obtain a large sample of diabetics at least 20 years of age with participation in the interview and medical examination. Osteoporosis status was defined by BMD at the total femur, femoral neck, or total lumbar spine. Self-reported diabetics that were prescribed insulin within the first year of diagnosis, are currently taking insulin, and reported no prescriptions for any diabetic pills were classified as type 1. Remaining self-reported diabetics were deemed as having type 2. RESULTS:A total of 2050 diabetics were included in which 87 (4%) were classified as type 1. Type 1 diabetics were found to have a significantly lower BMD at the total femur and femoral neck, but not at the lumbar spine in the adjusted models. Diabetics with type 1 were 4.7 times more likely to have osteoporosis than those with type 2. There was no significant relationship between diabetes type and BMD or osteoporosis prior to adjustment for confounders. CONCLUSIONS:Although our results show an increased likelihood of osteoporosis among those with type 1 diabetes, future studies including a larger sample from a community population are needed. It may benefit diabetics, especially those with type 1, to initiate osteoporosis screening methods including evaluation of fracture risk, bone quality, and BMD measurements at multiple sites earlier than recommended.
An update on the potential role of C-peptide in diabetes and osteoporosis.
Pujia Arturo,Gazzaruso Carmine,Montalcini Tiziana
PURPOSE:C-peptide secretion is deficient or absent in type 1 diabetes mellitus. It is well accepted that insulin replacement therapy cannot prevent the development of long-term diabetes-related complications, which can often be disabling or even life-threatening. Several cross-sectional investigations have suggested that residual C-peptide production in patients with type 1 diabetes mellitus would help prevent a number of complications. In animal models of diabetes and in patients with type 1 diabetes mellitus, C-peptide replacement improves renal function, skin and skeletal muscle blood flow, nerve conduction, glucose utilization, and other diabetes-related complications. Recent investigations suggest a new beneficial effect of C-peptide, which to date has never been studied. It is known that osteoporosis is the most prevalent short-term complication in type 1 diabetes mellitus. This review will highlight new insights into the pathophysiology and future therapeutic modalities for osteoporosis in individuals with diabetes. METHODS:This review provides a concise summary of old and new insights into the role of C-peptide in diabetes-related complications. RESULTS:The data suggest that C-peptide is a bioactive peptide, acting independently of insulin, which binds to a G-protein-coupled membrane binding site in different cell types. By triggering Ca-dependent intracellular signaling pathways, both Na, K-ATPase and endothelial nitric oxide synthase are activated. C-peptide may act on osteoblast cells by ERK 1/2 pathway activation, modulate collagen biosynthesis and RANKL expression. Furthermore, C-peptide-deficient postmenopausal women, not affected by diabetes, have a lower bone mineral density than those with normal C-peptide levels. CONCLUSION:Taken together these studies encourage further investigations to elucidate the role of C-peptide in preventing bone loss in type 1 diabetes mellitus and in those individuals with C-peptide deficiency and osteoporosis.
Type 2 Diabetes Mellitus Increases the Risk to Hip Fracture in Postmenopausal Osteoporosis by Deteriorating the Trabecular Bone Microarchitecture and Bone Mass.
Mohsin Sahar,Kaimala Suneesh,Sunny Jens Jolly,Adeghate Ernest,Brown Eric Mensah
Journal of diabetes research
T2DM is linked to an increase in the fracture rate as compared to the nondiabetic population even with normal or raised bone mineral density (BMD). Hence, bone quality plays an important role in the pathogenesis of skeletal fragility due to T2DM. This study analyzed the changes in the trabecular bone microstructure due to T2DM at various time points in ovariectomized and nonovariectomized rats. Animals were divided into four groups: (I) control (sham), (II) diabetic (sham), (III) ovariectomized, and (IV) ovariectomized with diabetes. The trabecular microarchitecture of the femoral head was characterized using a micro-CT. The differences between the groups were analyzed at 8, 10, and 14 weeks of the onset of T2DM using a two-way analysis of variance and by post hoc multiple comparisons. The diabetic group with and without ovariectomies demonstrated a significant increase in trabecular separation and a decrease in bone volume fraction, trabecular number, and thickness. BMD decreased in ovariectomized diabetic animals at 14 weeks of the onset of T2DM. No significant change was found in connectivity density and degree of anisotropy among groups. The structural model index suggested a change towards a weaker rod-like microstructure in diabetic animals. The data obtained suggested that T2DM affects the trabecular structure within a rat's femoral heads negatively and changes are most significant at a longer duration of T2DM, increasing the risk to hip fractures.
NIPA2 regulates osteoblast function via its effect on apoptosis pathways in type 2 diabetes osteoporosis.
Zhao Wei,Zhang Wei-Lin,Yang Bo,Sun Jun,Yang Mao-Wei
Biochemical and biophysical research communications
Type 2 diabetes osteoporosis has recently become a hot topic in the study of diabetic complications, but the specific mechanism of its development remains unclear. Non-imprinted in Prader-Willi/Angelman syndrome region protein 2 (NIPA2), a highly-selective magnesium ion transporter, has been found to be associated with type 2 diabetes. In this study we aimed to investigate the specific role and mechanism of NIPA2 in the pathogenesis of type 2 diabetes osteoporosis. We first used western blotting, PCR, immunofluorescence, and magnesium ion probes to detect changes of NIPA2 and intracellular magnesium levels in osteoblasts at different concentrations of advanced glycation end products (AGEs). We then up- or down-regulated NIPA2 using a lentivirus and analyzed apoptotic biomarkers as well as the osteogenic ability of osteoblasts. We found that AGEs dose-dependently down-regulated the expression of NIPA2 in osteoblasts. NIPA2 also regulated osteoblast apoptosis by affecting the intracellular magnesium level and further affecting the osteogenic capacity of osteoblasts. Our study revealed the changes of NIPA2 in response to AGEs in the environment, as well as its function and mechanism in osteoblasts, demonstrating its important role in the pathogenesis of type 2 diabetes osteoporosis. The study suggests that NIPA2 is a potential target for the treatment of type 2 diabetes osteoporosis.
miR-124-3p promotes BMSC osteogenesis via suppressing the GSK-3β/β-catenin signaling pathway in diabetic osteoporosis rats.
Li Zengying,Zhao Hengxia,Chu Shufang,Liu Xuemei,Qu Xin,Li Jinhua,Liu Deliang,Li Huilin
In vitro cellular & developmental biology. Animal
The purpose of this study is to investigate miRNAs' effects, targeting the Wnt signaling pathway, on osteogenic differentiation to provide new targets for diabetic osteoporosis treatments. Twelve male rats were divided into a normal rat group (NOR group) and a model rat group (MOD group). Cluster analysis of differentially expressed miRNAs and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed. Primary rat bone marrow mesenchymal stem cells (BMSCs) were divided into a high-glucose group and a low-glucose group, and osteogenic differentiation was induced. Alkaline phosphatase (ALP) staining and Alizarin Red staining were used for pathological analysis of the cells. Western blot analysis was used to measure GSK-3β, β-catenin, p-β-catenin, c-Myc, and CyclinD1 expression. Immunofluorescence (IF) was used to analyze the effect of GSK-3β inhibitor (CHIR99021) on β-catenin and CyclinD1 expressions levels in BMSCs. A total of 428 differentially expressed miRNAs were found between the NOR and MOD groups. KEGG analysis showed that the target genes were mostly enriched in signaling pathways, including PI3K-Akt, focal adhesion, AGE-RAGE, HIF-1, and Wnt. qPCR verification demonstrated that miR-124-3p exhibited the greatest difference in expression level. In BMSCs, miR-124-3p overexpression could reverse the inhibited expression of BMSC osteogenic markers, including Alpl, Bglap, and Runx2, induced by high glucose. Western blot analysis revealed that the transfection of miR-124-3p mimics could further reverse the upregulated p-β-catenin and GSK-3β levels and the downregulated c-Myc and CyclinD1 levels induced by high glucose. IF results revealed that BMSCs treated CHIR99021 under high glucose showed the reduced GSK-3β and increased β-catenin and CyclinD1 expression levels. Our research highlighted miRNAs' important roles in regulating the Wnt pathway and provided new information for the diagnosis and treatment of diabetic osteoporosis.
Zinc Supplementation Increased Bone Mineral Density, Improves Bone Histomorphology, and Prevents Bone Loss in Diabetic Rat.
Qi Shanshan,He Jia,Zheng Hongxing,Chen Chen,Jiang Hai,Lan Shiqiang
Biological trace element research
Diabetic osteoporosis (DOP) is a complication of diabetes, with the characteristics of bone mineral density (BMD) reduction and bone structure destruction. Zinc was reported has a benefit effect on postmenopausal osteoporosise, it was also has hypoglycemic effect, whether zinc was beneficial on diabetes-induced osteoporosis has not been reported. So in the present study, we established a diabetic rat model by streptozotocin injection (60 mg/kg), and administered zinc sulfate by oral gavage to investigate the protective effects of zinc on DOP and the underline possible mechanism. Thirty six Sprague Dawley rats were divided into T1DM group (diabetic rats), control group (vehicle treatment), and T1DM-Zinc group (diabetic rats administered zinc sulfate 0.25 mg/kg by oral gavage). The bone histomorphological parameters, serum bone metabolism markers (including ALP, OPG, RUNX 2, and RANKL), BMD, and bone marrow adipocyte numbers were detected after eight weeks of zinc sulfate treatment. The results showed zinc sulfate administration (0.25 mg/kg/d) decreased blood glucose, increased the BMD, decreased serum ALP, and RANKL, increased serum OPG and RUNX 2 levels, as well as OPG/RANKL ratio of T1DM rats. Meanwhile, the bone histomorphological parameters, bone marrow adipocytes numbers were returned to be normal. The RUNX 2, and OPG mRNA expression levels in bone tissues of T1DM-Zinc group rats were increased after zinc sulfate treatment compared with the diabetic rats (P < 0.05). Those indicating that zinc sulfate can prevent DOP, the protective mechanism were mainly related to its hypoglycemic effect, bone marrow lipogenesis inhibition effect, OPG/RANKL ratio and RUNX 2 up-regulation effect.
Puerarin alleviates streptozotocin (STZ)-induced osteoporosis in rats through suppressing inflammation and apoptosis via HDAC1/HDAC3 signaling.
Guo Chang-Jun,Xie Jing-Jing,Hong Rong-Hua,Pan Han-Song,Zhang Fu-Guo,Liang Yi-Min
Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie
Diabetic osteoporosis is a severe public health concern in the world. Puerarin (PU) is extensively attractive due to its superior bioactivities. In the study, we found that PU protected against streptozotocin (STZ)-induced osteoporotic changes in rats. PU treatment improved STZ-induced diabetes in rats, as evidenced by the reduced serum glucose and insulin levels. PU administration markedly attenuated bone loss and tartarate-resistant acid phosphatase (TRAP) activity in STZ-induced rats. Bone mineral density (BMD) was significantly decreased in diabetic rats, while being prevented by PU. STZ-induced impairments in microarchitecture of femoral tissues were markedly alleviated by PU treatment. In addition, bone-specific alkaline phosphatase (BALP), osteoprotegerin (OPG) and Runt-related transcription factor 2 (Runx2) levels in serum or tibia were improved by PU in STZ-injected rats; however, TRACP isoform 5b (TRACP-5b), carboxy-terminal collagen cross-links (β-CTX) and receptor activator of nuclear factor-κB ligand (RANKL) levels were decreased. Further, PU treatment inhibited inflammation and apoptosis in STZ-treated rats. Additionally, STZ injection increased histone deacetylase (HDAC)-1 and -3 expressions in femoral heads of rats, which were relieved by PU treatment. Notably, both HDAC1 and HDAC3 could enhance osteoporosis in vitro, as proved by the decreased ALP and Runx2 levels and the increased TRAP expression. Inflammation and apoptosis were exacerbated by HDAC1/3 over-expression, which were markedly diminished by PU treatment. In contrast, suppressing HDAC1/3 significantly abrogated fructose (Fru)-elicited inflammation and apoptosis in cells. Collectively, our data illustrated that PU is a potential therapeutic option to prevent diabetic osteoporosis by inhibiting HDAC1/HDAC3 signaling.
An update on therapies for the treatment of diabetes-induced osteoporosis.
Mohsin Sahar,Baniyas May Myh,AlDarmaki Reem Smh,Tekes Kornélia,Kalász Huba,Adeghate Ernest A
Expert opinion on biological therapy
: Currently, 424 million people aged between 20 and 79 years worldwide are diabetic. More than 25% of adults aged over 65 years in North America have Type 2 diabetes mellitus (DM). Diabetes-induced osteoporosis (DM-OS) is caused by chronic hyperglycemia, advanced glycated end products and oxidative stress. The increase in the prevalence of DM-OS has prompted researchers to develop new biological therapies for the management of DM-OS. : This review covered the current and novel biological agents used in the management of DM-OS. Data were retrieved from PubMed, Scopus, American Diabetes Association and International Osteoporosis Foundation websites, and ClinicalTrials.gov. The keywords for the search included: DM, osteoporosis, and management. : Several biological molecules have been examined in order to find efficient drugs for the treatment of DM-OS. These biological agents include anti-osteoporosis drugs: net anabolics (parathyroid hormone/analogs, androgens, calcilytics, anti-sclerostin antibody), net anti-resorptive osteoporosis drugs (calcitonin, estrogen, selective estrogen receptor modulators, bisphosphonates, RANKL antibody) and anti-diabetic drugs (alpha glucosidase inhibitors, sulfonylureas, biguanides, meglitinides, thiazolidinediones, GLP-1 receptor agonists, dipeptidylpeptidase-4 inhibitors, sodium glucose co-transporter-2 inhibitors, insulin). Biological medications that effectively decrease hyperglycemia and, at the same time, maintain bone health would be an ideal drug/drug combination for the treatment of DM-OS.
Chronic kidney disease-related osteoporosis is associated with incident frailty among patients with diabetic kidney disease: a propensity score-matched cohort study.
Chao C-T,Wang J,Huang J-W,Chan D-C,Hung K-Y,Chien K-L,
Osteoporosis international : a journal established as result of cooperation between the European Foundation for Osteoporosis and the National Osteoporosis Foundation of the USA
Chronic kidney disease (CKD)-related osteoporosis is a major complication in patients with CKD, conferring a higher risk of adverse outcomes. We found that among those with diabetic kidney disease, this complication increased the risk of incident frailty, an important mediator of adverse outcomes. INTRODUCTION:Renal osteodystrophy and chronic kidney disease (CKD)-related osteoporosis increases complications for patients with diabetic kidney disease (DKD). Since musculoskeletal degeneration is central to frailty development, we investigated the relationship between baseline osteoporosis and the subsequent frailty risk in patients with DKD. METHODS:From the Longitudinal Cohort of Diabetes Patients in Taiwan (n = 840,000), we identified 12,027 patients having DKD with osteoporosis and 24,054 propensity score-matched controls having DKD but without osteoporosis. The primary endpoint was incident frailty on the basis of a modified FRAIL scale. Patients were prospectively followed-up until the development of endpoints or the end of this study. The Kaplan-Meier technique and Cox proportional hazard regression were used to analyze the association between osteoporosis at baseline and incident frailty in these patients. RESULTS:The mean age of the DKD patients was 67.2 years, with 55.4% female and a 12.6% prevalence of osteoporosis at baseline. After 3.5 ± 2.2 years of follow up, the incidence rate of frailty in patients having DKD with osteoporosis was higher than that in DKD patients without (6.6 vs. 5.7 per 1000 patient-year, p = 0.04). A Cox proportional hazard regression showed that after accounting for age, gender, obesity, comorbidities, and medications, patients having DKD with osteoporosis had a significantly higher risk of developing frailty (hazard ratio, 1.19; 95% confidence interval, 1.02-1.38) than those without osteoporosis. CONCLUSIONS:CKD-related osteoporosis is associated with a higher risk of incident frailty in patients with DKD.
Chondroitin Sulfate Prevents STZ Induced Diabetic Osteoporosis through Decreasing Blood Glucose, AntiOxidative Stress, Anti-Inflammation and OPG/RANKL Expression Regulation.
Zheng Hong Xing,Chen De Jing,Zu Yue Xin,Wang En Zhu,Qi Shan Shan
International journal of molecular sciences
Chondroitin sulfate (CS) has antioxidative, anti-inflammatory, anti-osteoarthritic and hypoglycemic effects. However, whether it has antidiabetic osteoporosis effects has not been reported. Therefore, in this study, we established a STZ-induced diabetic rat model; CS (500 mg kg d) was orally administrated for eight weeks to study its preventive effects on diabetic osteoporosis. The results showed that eight weeks of CS treatment improved the symptoms of diabetes; the CS-treated group has increased body weight, decreased water or food intake, decreased blood glucose, increased bone-mineral density, repaired bone morphology and decreased femoral osteoclasts and tibia adipocytes numbers. After CS treatment, bone histomorphometric parameters returned to normal, the levels of serum inflammatory cytokines (IL-1β, IL-6 and TNF-α) decreased significantly, serum SOD, GPX and CAT activities increased and MDA level increased. In the CS-treated group, the levels of serum ALP, CTX-1, TRACP 5b, osteocalcin and RANKL decreased and the serum RUNX 2 and OPG levels increased. Bone immunohistochemistry results showed that CS can effectively increase the expression of OPG and RUNX2 and reduce the expression of RANKL in diabetic rats. All of these indicate that CS could prevent STZ induced diabetic osteoporosis-mainly through decreasing blood glucose, antioxidative stress, anti-inflammation and regulation of OPG/RANKL expression. CS can therefore effectively prevent bone loss caused by diabetes.
The beneficial roles of insulin and parathyroid hormones in the treatment of experimentally induced diabetic osteoporosis in female rats: bone mineral density, morphometric and histological studies.
Abd El Aziz G S,Ramadan W S,El-Fark M O,Saleh H A M
BACKGROUND:Diabetes mellitus (DM) and osteoporosis are two frequent medical conditions with an increasing prevalence in elderly people and are responsible for large number of incurable fractures. This study is designed experimentally in female rats in order to determine whether combined treatment of insulin and parathyroid hormone (PTH) enhances the reversibility of the osteoporotic changes that occurred in streptozotocin (STZ)-induced DM. MATERIALS AND METHODS:In this study, 30 adult female rats aged 3 months were used, they were randomly divided into: control group (6 rats) and diabetes group (24 rats), in which experimental DM was induced by i.p. injection of a single dose of STZ (60 mg/kg/body weight). Diabetic group was further divided into four subgroups (6 rats each): non-treated diabetic, insulin-treated (8-12 units s.c./day of Humalin U-40), PTH-treated (6.0 μg s.c./kg/day) and combined insulin and PTH-treated subgroups. All tested groups were assessed for body weight, food and water consumptions. RESULTS:At the end of the experimental period, the bone mineral density (BMD) was measured for all rats of different groups; then the rats were sacrificed and blood samples were collected for measuring glucose, alkaline phosphatase and osteocalcin levels. Right femora were dissected out and subjected to measurement of diameter of neck and shaft, length of shaft, and weight. Then the femora specimens were processed and stained with haematoxylin and eosin for histological study. The results showed that there was a statistically significant, decrease in BMD, increase in the level of alkaline phosphate, and decrease in the level of osteocalcin in rats in diabetic group compared with other groups; these parameters improved in other groups, especially in diabetes/insulin/PTH group. The rats in diabetic group showed statistically significant decrease in neck and shaft diameters and weight of femur bone compared with other groups, while rats in diabetes/insulin/PTH group showed a significant improvement of these parameters. In diabetic group, there were different histopathological changes in cortical bone and Haversian canals, which improved in other groups, especially in rats in diabetes/insulin/PTH group. CONCLUSIONS:The untreated DM resulted in dramatic reduction in BMD and morphometric parameters. Treatment with insulin ameliorated these effects to some extent, while PTH co--treatment had a more positive effect. The combination of PTH and insulin resulted in stronger improvement of all parameters to approximately like those of control rats.
IGF-1R/β-catenin signaling axis is involved in type 2 diabetic osteoporosis.
Zhang Zhi-Da,Ren Hui,Wang Wei-Xi,Shen Geng-Yang,Huang Jin-Jing,Zhan Mei-Qi,Tang Jing-Jing,Yu Xiang,Zhang Yu-Zhuo,Liang De,Yang Zhi-Dong,Jiang Xiao-Bing
Journal of Zhejiang University. Science. B
Insulin-like growth factor-1 receptor (IGF-1R) is involved in both glucose and bone metabolism. IGF-1R signaling regulates the canonical Wnt/β-catenin signaling pathway. In this study, we investigated whether the IGF-1R/ β-catenin signaling axis plays a role in the pathogenesis of diabetic osteoporosis (DOP). Serum from patients with or without DOP was collected to measure the IGF-1R level using enzyme-linked immunosorbent assay (ELISA). Rats were given streptozotocin following a four-week high-fat diet induction (DOP group), or received vehicle after the same period of a normal diet (control group). Dual energy X-ray absorption, a biomechanics test, and hematoxylin-eosin (HE) staining were performed to evaluate bone mass, bone strength, and histomorphology, respectively, in vertebrae. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were performed to measure the total and phosphorylation levels of IGF-1R, glycogen synthase kinase-3β (GSK-3β), and β-catenin. The serum IGF-1R level was much higher in patients with DOP than in controls. DOP rats exhibited strikingly reduced bone mass and attenuated compression strength of the vertebrae compared with the control group. HE staining showed that the histomorphology of DOP vertebrae was seriously impaired, which manifested as decreased and thinned trabeculae and increased lipid droplets within trabeculae. PCR analysis demonstrated that IGF-1R mRNA expression was significantly up-regulated, and western blotting detection showed that phosphorylation levels of IGF-1R, GSK-3β, and β-catenin were enhanced in DOP rat vertebrae. Our results suggest that the IGF-1R/β-catenin signaling axis plays a role in the pathogenesis of DOP. This may contribute to development of the underlying therapeutic target for DOP.
Anhydroicaritin, a SREBPs inhibitor, inhibits RANKL-induced osteoclastic differentiation and improves diabetic osteoporosis in STZ-induced mice.
Zheng Zu-Guo,Zhang Xin,Zhou Ya-Ping,Lu Chong,Thu Pyone Myat,Qian Cheng,Zhang Mu,Li Ping,Li Hui-Jun,Xu Xiaojun
European journal of pharmacology
Nowadays, more and more attention has been paid to osteoporosis caused by diabetes mellitus. Elevated levels of pro-inflammatory cytokines in diabetic patients activate the activity of osteoclasts through the RANKL/OPG pathway. The nuclear transcription factor SREBP2, a master regulator of cholesterol metabolism, has been found involved in osteoclastogenesis. In our previous study, we have identified anhydroicaritin as a potent inhibitor of transcription factor SREBPs, which improves dyslipidemia and insulin resistance. In this study, we demonstrated that anhydroicaritin could also decrease the level of SREBP2 and its target genes in osteoclasts induced by RANKL without significant cytotoxicity. Moreover, anhydroicaritin suppressed RANKL-induced osteoclasts differentiation. In STZ-induced diabetic mice model, we found that the osteoclasts were largely increased accompanied with deterioration of bone structure. Anhydroicaritin decreased the level of blood glucose and alleviated insulin resistance. More importantly, anhydroicaritin inhibited osteoclast differentiation and rescued diabetes-induced bone loss in vivo. In conclusion, anhydroicaritin, a potent SREBP2 inhibitor, inhibits the osteoclasts formation and improves diabetes-induced bone loss.
Traditional Chinese medicine Bushen-Jianpi-Huoxue decoction prevents diabetic osteoporosis in rats via Wnt and nuclear factor-kappa B signaling pathways.
Zhang Yongqing,Liu Mingming,Li Haisheng,Chen Zetao,Liang Na,Xu Jianguo,Zhang Xiaoli,Zhang Yihang
International journal of rheumatic diseases
AIM:The aim of this study was to evaluate the therapeutic effect of a traditional Chinese medicine (TCM), Bushen-Jianpi-Huoxue decoction (BJHD), on diabetic osteoporosis (DOP) and the action mechanisms likely mediated by nuclear factor-kappa B (NF-κB) and Wnt signaling pathways. METHODS:Fifty-five male Wistar rats were used in this study; they were divided into normal control (n = 10) and established DOP model (n = 45) groups. The DOP model was induced using a combination high carbohydrate - high fat diet and intraperitoneal injections of streptozotocin (STZ). The successfully induced animals were randomized to the model, Western medicine, TCM and control groups. Levels of fasting blood glucose; insulin; serum Ca, P and alkaline phosphatase, and the femoral bone mineral density (BMD) were measured. Furthermore, messenger RNA (mRNA) levels of cytokines in the Wnt and NF-κB signaling pathways were measured using reverse transcription-polymerase chain reaction (RT-PCR). RESULTS:Thirty rats were successfully established as the DOP model (10/group). After treatment, the levels of fasting blood glucose, insulin resistance and alkaline phosphatase in the TCM group rats were lower, while P and BMD were higher than those in the model groups. The mRNA levels of cytokines in the Wnt signaling pathway were higher in the TCM group than those in the model group. Moreover, the expressions of factors in the NF-κB pathway were markedly lower in the TCM group than they were in the model group. CONCLUSION:Bushen-Jianpi-Huoxue decoction relieved DOP by activating the Wnt signaling pathway while inhibiting NF-κB signaling.
Investigation for GSK3β expression in diabetic osteoporosis and negative osteogenic effects of GSK3β on bone marrow mesenchymal stem cells under a high glucose microenvironment.
Chen Yu,Chen Long,Huang Runyu,Yang Wenyue,Chen Siyue,Lin Kaili,Liu Jiaqiang
Biochemical and biophysical research communications
Osteoporosis is a common skeletal complication of diabetes mellitus (DM). The mechanisms underlying the pathophysiology of diabetic osteoporosis are complex. Glycogen synthase kinase-3β (GSK-3β) is a widely expressed serine/threonine kinase and associated with both DM and bone metabolism, which arouse our concern. In this study, we established the diabetic mouse model by high-fat diet combined with streptozotocin injection. Decreased bone mass and reduced osteogenesis were observed in femurs of the mice. Besides, we identified that there is an activated expression of GSK3β in the bone marrow mesenchymal stem cells (BMSCs) of diabetic mice. To explore the link between GSK3β and diabetic osteoporosis, we exposed BMSCs to a high glucose microenvironment in vitro and discovered that the glucose-induced GSK3β activation has negative osteogenic effects on BMSCs by suppressing β-catenin/Tcf7/Ccn4 signaling axis. Inhibition of GSK3β by specific concentrations of LiCl could reverse the impaired osteogenesis of BMSCs and increase expression of β-catenin, Tcf7 and Ccn4. Our research indicated that abnormal activation of GSK3β plays a role in diabetic osteoporosis and might be a potential target to treat diabetic osteoporosis.
Association between diabetic neuropathy and osteoporosis in patients: a systematic review and meta-analysis.
Liu Chunhua,Lv Haihong,Niu Peng,Tan Jiaojiao,Ma Yuping
Archives of osteoporosis
OBJECTIVES:Many studies have explored the association between neuropathy and osteoporosis in patients with diabetes mellitus. However, the results still remain inconsistent and controversial. We aimed to estimate the association between diabetic neuropathy and osteoporosis. METHODS:Databases, including PubMed, Embase, Web of Science, the Cochrane library, Chinese Biomedical Literature Database (CBM), and Wanfang, were screened from inception to 30 March 2020. Studies were selected and data were extracted by two independent reviewers. Study characteristics and quality sections were reviewed independently. Pooled ORs and 95% CIs were calculated using random effects model when evidence of heterogeneity was present; otherwise, fixed effects model was used. Meta-regression and subgroup analyses were performed to explore the source of heterogeneity. Sensitivity analysis and publication bias were also tested. RESULTS:A total of 11 studies with 27,585 participants were included in this analysis which indicated that there was an increased odd between diabetic neuropathy and osteoporosis (overall OR 2.20, 95% CI 1.71-2.83). In the subgroup analyses and meta-regression, diabetic neuropathy has no significant difference in osteoporosis or fracture (p = 0.532). And osteoporosis also has no significant difference in type 1 or type 2 diabetic neuropathy (p = 0.668). CONCLUSIONS:This meta-analysis suggests that patients with diabetic neuropathy have a significantly increased chance of developing osteoporosis, even fragility fracture. The clinicians should pay more attention to the patients with diabetic neuropathy. Further studies were still needed to explore the confounding factors among studies and to elucidate the underlying biological mechanisms.
Microangiopathy is associated with bone loss in female type 2 diabetes mellitus patients.
Zhong Ni,Zhang Youyang,Pu Xiangling,Xu Bei,Xu Mingxin,Cai Haidong,Zhang Ge,Cui Ran,Sheng Hui,Qu Shen
Diabetes & vascular disease research
OBJECTIVE:Type 2 diabetes mellitus complicated with microvascular diseases can be used as a model to study the relationship between bone health and the microvascular situation. METHODS:A total of 2,170 patients with type 2 diabetes mellitus (1,188 postmenopausal females and 982 males aged ⩾50 years) were included in our cross-sectional study. These patients were grouped according to 24-hour urine protein level: Group I (<30 mg), Group II (30-299 mg) and Group III (≥300 mg). Bone mineral density of the lumbar spine, hip and femoral neck was evaluated by dual-energy X-ray absorptiometry. Fundus oculi photography for diabetic retinopathy and 24-h urine protein for diabetic nephropathy were used as markers of microangiopathy in type 2 diabetes mellitus. Characteristics of the patients and bone mineral density were compared. Multivariate analysis was used to study the association between bone mineral density and microangiopathy. Statistical analysis was performed using SPSS 20.0. p < 0.05 was considered statistically significant. RESULTS:Group III had the lowest bone mineral density level in both genders. Multivariate analysis revealed that microangiopathy was negatively correlated with bone mineral density in females (lumbar: r = -0.522, p < 0.001; hip: r = -0.301, p = 0.010; femoral neck: r = -0.314, p = 0.009), but not in males, after adjustment for age, body mass index, hypertension, hyperlipidemia, diabetic status, hepatic function, kidney function, sex hormones and 25(OH) vitamin D. CONCLUSION:These results demonstrate an independent negative correlation between microangiopathy and bone mineral density in postmenopausal female type 2 diabetes mellitus patients.
Anthocyanin-rich extract from black rice (Oryza sativa L. Japonica) ameliorates diabetic osteoporosis in rats.
Qi Shanshan,He Jia,Han Hao,Zheng Hongxing,Jiang Hai,Hu Ching Yuan,Zhang Zhijian,Li Xinsheng
Food & function
Diabetic osteoporosis (DOP) is a systemic endocrine-metabolic osteopathy which has the characteristics of bone mineral density (BMD) reduction and bone microstructural destruction. Although anthocyanin-rich extract from black rice (AEBR) was reported to have a beneficial effect on diabetic rats, no studies have been performed on whether black rice anthocyanins are beneficial for diabetic osteoporosis. Therefore, in this study, a streptozotocin-induced diabetic rat model was established to investigate the protective effect of AEBR on diabetes-induced osteoporosis and its possible mechanism. AEBR at three doses (0.5, 1.0, and 2.0 g kg d) were administered by oral gavage to diabetic rats for 8 weeks. The blood glucose, BMD, bone histomorphometry parameters, serum bone turnover biomarkers, bone marrow adipocyte numbers, as well as osteoprotegerin (OPG), runt-related transcription factor 2 (RUNX 2), and receptor activator of nuclear factor-κ B ligand (RANKL) protein expression in bone and serum were detected. The results indicated that AEBR dose-dependently decreased the blood glucose, increased the BMD, and decreased the serum bone turnover markers. The bone microstructure and osteoclast numbers in bone tissues returned to normal in the high AEBR dosage group; at the same time, the AEBR dose-dependently suppressed bone marrow adipogenesis. The RUNX 2 as well as the OPG/RANKL ratio in diabetic rats' bone tissues increased significantly in the AEBR treatment group. Our results indicate that AEBR administration can ameliorate bone loss caused by diabetes; this is mainly attributed to its inhibition of bone turnover, suppression of bone marrow adipogenesis, and up-regulation of RUNX 2 and the OPG/RANKL expression ratio.
Possible therapeutic potential of berberine in the treatment of STZ plus HFD-induced diabetic osteoporosis.
Xie Huanguang,Wang Qingqing,Zhang Xinyue,Wang Te,Hu Wei,Manicum Theasha,Chen Hua,Sun Liaojun
Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie
Diabetic osteoporosis is a complication of diabetes mellitus, and can result in an increased incidence of bone fractures and a delay in fracture healing. Berberine is one of the most widely distributed isoquinoline alkaloid in plants and possesses antioxidant properties. These properties can reduce the high glucose mediated in the dysfunction of human bone marrow stem cells. Therefore, the present study was designed to investigate the apparent beneficial effect of berberine on bone characteristics in streptozotocin plus HFD-induced diabetic rats. Rats were selected at random and divided into four groups: (A) control group (CG) (n = 10); (B) diabetic group (DG) (n = 10); (C) diabetic group with 50 mg kgday of berberine (Brb-50) (n = 10); and (D) diabetic group with 100 mg kgday of berberine (Brb-100) (n = 10). After 12 weeks of being treated with berberine, the femora from all rats were assessed and other blood biochemistries evaluated. Berberine at 50 mg/kg showed little effect and significance on diabetic osteopenia, while berberine at 100 mg/kg was significantly increased in diabetic rats. The same group also displayed a significantly decreased serum osteocalcin and serum alkaline phosphatase activity in diabetic rats. The impaired micro-architecture of the femurs in diabetic rats could partially be prevented by berberine with 100 mg/kg. In addition, berberine could to an extent restore the decreased bone formation and reabsorption of the femurs in diabetic rats through the histomorphometric analysis. Berberine could not only significantly lower the oxidative level of DNA damage, but also up-regulate the activity of serum antioxidants. According to our investigations and discoveries, we have found, that berberine may be a potential drug for controlling bone loss in diabetic osteoporosis.
Possible osteoprotective effects of myricetin in STZ induced diabetic osteoporosis in rats.
Ying Xiaozhou,Chen Xiaowei,Wang Te,Zheng Wenhao,Chen Long,Xu Youjia
European journal of pharmacology
Myricetin is a flavonoid which has many pharmacological effects. However, to date there is no evidence study on the effect of myricetin in diabetic condition. This study was aimed to investigate whether myricetin could protect against diabetic osteoporosis in streptozotocin induced rats. Female Wistar rats were randomly allocated to four equal groups: diabetic group (DG), diabetic group with myricetin (50 mg per kilogram per day), (D) diabetic group with myricetin (100 mg/kg/day) and normal control group (CG). Body weight was recorded once a week. After treatment with myricetin for 12 weeks, serum biochemical analyses, the microarchitecture of femora, and histological changes were evaluated. We found that the bone mineral density (BMD) of myricetin (100 mg per kilogram per day)treatment group significantly increased than in the diabetic group (P < 0.05). The alkaline phosphatase and osteocalcin were markedly blocked in diabetic rats relative to normal control group (P < 0.05); however, the inhibition was prevented by the myricetin treatment group. Results also showed that myricetin treatment could dramatically improve trabecular bone microarchitecture through increasing bone mass such as trabecular number (Tb.N), bone volume per tissue volume (BV/TV), and decreasing that of structure model index (SMI) and trabecular separation (Tb.Sp), comparing with the control group. We also found that myricetin could significantly lower the oxidative damage and up-regulate the activity of superoxide dismutase (SOD) and catalase activity. In summary, we showed that myricetin can effectively improve abnormal bone metabolism in streptozotocin induced rats, which may provide a beneficial medicine on diabetic bone disease.
Inhibition of tanshinone IIA on renin activity protected against osteoporosis in diabetic mice.
Zhang Jingjing,Cai Zixuan,Yang Min,Tong Lijuan,Zhang Yan
Bge. (Labiatae) (SMB) is applied clinically for management of diabetic osteoporosis in China, and research results has suggested its potential action on renin-angiotensin system (RAS). This study screens and explores naturally occurring bioactive constituents from the root of SMB acting on renin activity and evaluates its osteoprotective efficacy in diabetic mice. Human embryonic kidney (HEK) 293 cells, engineered to express human renin, were used as an model to identify bioactive compound, tanshinone IIA, inhibiting renin activity. The C57BL/6 mice ( = 10 in each group) with diabetes induced by streptozotocin (STZ) were intraperitoneally injected with tanshinone IIA (10 and 30 mg/kg). The mice without STZ treatment and the diabetic mice treated with aliskiren were used as non-diabetic control and positive control, respectively. Tanshinone IIA was found to display inhibitory effects on renin activity of HEK-293 cells; moreover, it down-regulated protein expression of ANG II in human renin-expressed HEK-293 cells. Treatment of diabetic mice with tanshinone IIA with both doses could significantly decrease ANG II level in serum (from 16.56 ± 1.70 to 10.86 ± 0.68 and 9.14 ± 1.31 pg/mL) and reduce ANG II expression in bone, consequently improving trabecular bone mineral density and micro-structure of proximal tibial end and increasing trabecular bone area of distal femoral end in diabetic mice. This study revealed beneficial effects of tanshinone IIA on bone of diabetic mice, and potentially suggested the application of in the treatment of osteoporosis and drug development of tanshinone IIA as a renin inhibitor.
Salidroside Improves Bone Histomorphology and Prevents Bone Loss in Ovariectomized Diabetic Rats by Upregulating the OPG/RANKL Ratio.
Zheng Hongxing,Qi Shanshan,Chen Chen
Molecules (Basel, Switzerland)
Postmenopausal diabetic women have a high risk of fractures. Salidroside has preventive effects on estrogen deficiency-induced osteoporosis and has hypoglycemic effects on diabetes in rats. However, whether salidroside inhibits bone loss in postmenopausal diabetic patients is still unknown. Here, we established a rat model of osteoporosis to investigate the protective effects of salidroside on bone loss induced by ovariectomy combined with diabetes, also investigating the underlying mechanisms. Two-month-old female Sprague-Dawley rats were divided into three equal groups (10 rats in each group): control group (with sham operation, treated with drug vehicle); OVX/T1DM group (ovariectomized diabetic rats); OVX/T1DM-SAL group, comprising ovariectomized diabetic rats treated with salidroside (20 mg/kg body weight) by gavage. The results showed that after 60 consecutive days of treatment, the bone mineral density (BMD) of OVX/T1DM-SAL increased significantly compared with the OVX/T1DM group ( < 0.01). The level of serum bone turnover markers, including alkaline phosphatase (ALP), cross linked c-telopeptide of type I collagen (CTX-1), osteocalcin, N-terminal propeptide of type I procollagen (PINP), and tartrate-resistant acid phosphatase 5b (TRACP 5b) were all increased in the OVX/T1DM group compared with the control ( < 0.01), and those were decreased by salidroside treatment. Meanwhile, the bone histopathological changes were also attenuated, and the bone marrow adipogenesis was inhibited in salidroside treated rats. Moreover, protein and mRNA ratio of bone osteoprotegerin (OPG)/receptor activator of nuclear factor-κB ligand (RANKL) was upregulated in ovariectomized diabetic rats by salidroside treatment. The results above indicated that the protective effect of salidroside on bone loss induced by ovariectomy and diabetes was mainly due to its ability to suppress bone turnover, inhibit bone marrow adipogenesis, and up-regulate the OPG/RANKL ratio.
Altered gene expression involved in insulin signaling pathway in type II diabetic osteoporosis rats model.
Li Baoxin,Wang Yan,Liu Yan,Ma Jianxia,Li Yukun
It is well established that both estrogen loss and type II diabetes mellitus (DMII) can impair bone metabolism, but whether estrogen loss exacerbates the effects of DMII is unclear. Therefore, we determined if ovariectomy (OVX) of rats on a long-term high-fat/sugar diet and injection of a low dose of streptozotocin (DMII) decreased bone mineral density (BMD) more than OVX or DMII alone. Bone insulin signaling is known to support bone metabolism; therefore, we also tested the hypothesis that OVX DMII rats (DOVX) would exhibit greater reductions in the expression of proteins important in insulin signaling, including IRS-1, IRS-2, and IGF-1. As hypothesized, BMD and plasma estrogen levels were decreased more in DOVX rats than in rats following OVX (NOVX) or DMII (DS) alone. IGF-1 expression was decreased in the liver, kidney, skeletal muscle, and bone of DOVX, DS, and NOVX rats; however, the decrease was larger and occurred sooner in DOVX rats. While IRS-1 and IRS-2 decreased in most groups in all tissues examined, the expression patterns differed in both a group- and tissue-dependent fashion. In conclusion, these data demonstrate that estrogen loss and DMII induced by a high-fat/sugar diet interact to produce osteoporosis and support the hypothesis that the bone loss may be mediated at least in part by concurrent decreases in the insulin signaling proteins in bone.
Osteoporosis in the Cohen diabetic rat: correlation between histomorphometric changes in bone and microangiopathy.
Amir Gail,Rosenmann Eliezer,Sherman Yoav,Greenfeld Ziv,Ne'eman Zvi,Cohen Aharon Mordechai
Laboratory investigation; a journal of technical methods and pathology
Osteoporosis is well documented in type I diabetes, but its occurrence is controversial in type II diabetes. Microangiopathy is a major complication of type I and type II diabetes. We studied bone and microvascular changes in the Cohen diabetic rat, a unique nonobese model of noninsulin-dependent diabetes mellitus. The aim of this study was to find whether there is a temporal correlation between the onset of these two complications. The diabetic rats were divided into three groups (A, B, and C) according to duration of diabetes (2 months, 3 months, and 7 to 8 months, respectively). Trabecular bone area was assessed by computerized image analysis and microangiopathy by means of renal function tests, histologic examination of the kidneys, and ultrastructural measurement of the width of capillary basement membranes. Bone density of the distal femur and vertebra was significantly reduced in the diabetic rats relative to the control rats in all three groups (Group A femur: 11.5 +/- 1.6% versus 21.8 +/- 3.0%, p < 0.02; Group A vertebra: 15.9 +/- 1.6% versus 28.5 +/- 2.0%, p < 0.02; Group C femur: 7.9 +/- 1.1% versus 29.6 +/- 3.5%, p < 0.001; Group C vertebra: 11.4 +/- 0.7% versus 37.1 +/- 1.9%, p < 0.002). Renal function tests were normal in the Group A diabetic rats and there was marked albuminuria in the Group C diabetic rats. Histologic changes in the kidneys were seen only in the Group C diabetic rats. Five of 15 Group C diabetic rats showed no albuminuria or histologic evidence of kidney damage. The bone density in this subgroup was reduced relative to controls to the same degree as that of the rats with renal damage. There was no evidence of capillary basement membrane thickening in the Group A diabetic rats. Our findings indicate that in the Cohen diabetic rat, osteoporosis precedes the onset of microangiopathy. Microangiopathy probably does not play an important role in the pathogenesis of osteoporosis in this animal model.