Project description:Diabetes mellitus is a group of chronic diseases characterized by high blood glucose levels. Diabetic patients have a higher risk of sustaining osteoporotic fractures than non-diabetic people. The fracture healing is usually impaired in diabetics, and our understanding of the detrimental effects of hyperglycemia on fracture healing is still inadequate. Metformin is the first-line medicine for type 2 diabetes (T2D). However, its effects on bone in T2D patients remain to be studied. To assess the impacts of metformin on fracture healing, we compared the healing process of closed-wound fixed fracture, non-fixed radial fracture, and femoral drill-hole injury models in the T2D mice with and without metformin treatment. Our results demonstrated that metformin rescued the delayed bone healing and remolding in the T2D mice in all injury models. In vitro analysis indicated that compromised proliferation, osteogenesis, chondrogenesis of the bone marrow stromal cells (BMSCs) derived from the T2D mice were rescued by metformin treatment when compared to WT controls. Furthermore, metformin could effectively rescue the impaired detrimental lineage commitment of BMSCs isolated from the T2D mice in vivo as assessed by subcutaneous ossicle formation of the BMSC implants in recipient T2D mice. Moreover, the Safranin O staining of cartilage formation in the endochondral ossification under hyperglycemic condition significantly increased at day 14 post-fracture in the T2D mice receiving metformin treatment. The chondrocyte transcript factors SOX9 and PGC1α, important to maintain chondrocyte homeostasis, were both significantly upregulated in callus tissue isolated at the fracture site of metformin-treated MKR mice on day 12 post-fracture. Metformin also rescued the chondrocyte disc formation of BMSCs isolated from the T2D mice. Taken together, our study demonstrated that metformin facilitated bone healing, more specifically bone formation and chondrogenesis in T2D mouse models.
Project description:BackgroundEpidemiological studies suggest that metformin reduces the risk of developing several types of cancer, including gliomas, and improves the overall survival in cancer patients. Nevertheless, while the effect of metformin on cancer cells has been extensively studied, its impact on other components of the tumour microenvironment, such as macrophages, is less understood.ResultsMetformin-treated mouse bone marrow cells differentiate into spindle-shaped macrophages exhibiting increased phagocytic activity and tumour cell cytotoxicity coupled with modulated expression of co-stimulatory molecules displaying reduced sensitivity to inflammatory cues compared with untreated cells. Transcriptional analyses of metformin-treated mouse bone marrow-derived macrophages show decreased expression levels of pro-tumour genes, including Tgfbi and Il1β, related to enhanced mTOR/HIF1α signalling and metabolic rewiring towards glycolysis.SignificanceOur study provides novel insights into the immunomodulatory properties of metformin in macrophages and its potential application in preventing tumour onset and in cancer immunotherapy.
Project description:Background and purposeCell therapy with bone marrow stromal cells (BMSCs) improves functional recovery after stroke in nondiabetic rats. However, its effect on diabetics with stroke is unknown. This study investigated the effect of BMSCs on stroke outcome in Type 1 diabetic (T1DM) rats.MethodsT1DM was induced in adult male Wistar rats by injecting streptozotocin. Nondiabetic and T1DM rats were subjected to 2 hours of middle cerebral artery occlusion (MCAO), treated with or without BMSCs (3×10(6)) at 24 hours after MCAO, and monitored for 14 days.ResultsFunctional benefit was not detected in T1DM-MCAO treated with BMSC rats compared with corresponding T1DM-MCAO controls. BMSC treatment in T1DM-MCAO rats had increased mortality, blood-brain barrier leakage, brain hemorrhage, and angiogenesis. Internal carotid artery neointimal formation and cerebral arteriole narrowing/occlusion were also observed in T1DM-MCAO+BMSCs rats compared with T1DM-MCAO controls (P<0.05), but not in nondiabetic stroke rats. We further studied the underlying mechanisms responsible for BMSC-induced blood-brain barrier leakage and accelerated vascular damage in T1DM-MCAO rats. We found that the expression of angiogenin (an angiogenic factor) and ED1 (a marker for macrophages) was significantly increased in the T1DM-MCAO+BMSC rats in the ischemic brain and internal carotid artery compared with nontreated T1DM-MCAO rats, but not in nondiabetic stroke rats.ConclusionsBMSC therapy in T1DM-MCAO rats does not improve functional outcome. On the contrary, it increases blood-brain barrier leakage and cerebral artery neointimal formation, and arteriosclerosis, which possibly is due to increased expression of angiogenin. Thus, BMSC treatment starting 24 hours after MCAO may not be beneficial for diabetic subjects with stroke.
Project description:Macrophages are highly plastic immune cells that are capable of adopting a wide array of functional phenotypes in response to environmental stimuli. The changes in macrophage function are often supported and regulated by changes in cellular metabolism. Capturing a comprehensive picture of metabolism is vital for understanding the role of metabolic rewiring in the immune response. Here we present a method for systematically quantifying the abundance of metabolites and lipids in primary murine bone marrow derived macrophages (BMDMs). This method simultaneously extracts polar metabolites and lipids from BMDMs using a rapid two-phase extraction procedure. The polar metabolite fraction and lipid fraction are subsequently analyzed by separate liquid chromatography-mass spectrometry (LC-MS) methods for optimized coverage and quantification. This allows for a comprehensive characterization of cellular metabolism that can be used to understand the impact of a variety of environmental stimuli on macrophage metabolism and function.
Project description:Aims/hypothesisHypothalamic inflammation and sympathetic nervous system hyperactivity are hallmark features of the metabolic syndrome and type 2 diabetes. Hypothalamic inflammation may aggravate metabolic and immunological pathologies due to extensive sympathetic activation of peripheral tissues. Loss of somatostatinergic (SST) neurons may contribute to enhanced hypothalamic inflammation.MethodsThe present data show that leptin receptor-deficient (db/db) mice exhibit reduced hypothalamic SST neurons, particularly in the periventricular nucleus. We model this finding, using adeno-associated virus delivery of diphtheria toxin subunit A (DTA) driven by an SST-cre system to deplete these neurons in Sstcre/gfp mice (SST-DTA).ResultsSST-DTA mice exhibit enhanced hypothalamic c-Fos expression and brain inflammation as demonstrated by microglial and astrocytic activation. Bone marrow from SST-DTA mice undergoes skewed haematopoiesis, generating excess granulocyte-monocyte progenitors and increased proinflammatory (C-C chemokine receptor type 2; CCR2hi) monocytes. SST-DTA mice exhibited a 'diabetic retinopathy-like' phenotype: reduced visual function by optokinetic response (0.4 vs 0.25 cycles/degree; SST-DTA vs control mice); delayed electroretinogram oscillatory potentials; and increased percentages of retinal monocytes. Finally, mesenteric visceral adipose tissue from SST-DTA mice was resistant to catecholamine-induced lipolysis, displaying 50% reduction in isoprenaline (isoproterenol)-induced lipolysis compared with control littermates. Importantly, hyperglycaemia was not observed in SST-DTA mice.Conclusions/interpretationThe isolated reduction in hypothalamic SST neurons was able to recapitulate several hallmark features of type 2 diabetes in disease-relevant tissues.
Project description:Hyperglycemia has various adverse health effects, some of which are due to chronic oxidative and inflammatory impairment of bone marrow (BM), hematopoietic stem cells (HSCs), and mesenchymal stem cells (MSCs). Astaxanthin (ASTX) has been shown to ameliorate hyperglycemia-associated systemic complications and acute mortality, and this effect is partially associated with restoration of normal hematopoiesis. Here, the effects of ASTX on diabetes-induced complications in BM and BM stem cells were investigated, and the underlying molecular mechanisms were elucidated. Ten-week-old C57BL/6 mice received a single intraperitoneal injection of streptozotocin (STZ; 150 mg/kg) in combination with oral gavage of ASTX (12.5 mg/kg) for 30 or 60 consecutive days. Supplemental ASTX ameliorated acute mortality and restored the STZ-impaired bone mass accrual and BM microenvironment in STZ-injected mice. Oral gavage of ASTX suppressed osteoclast formation in the BM of STZ-injected mice. Specifically, supplementation with ASTX inhibited oxidative stress and senescence induction of BM HSCs and MSCs and ameliorated hematopoietic disorders in STZ-injected mice. These effects of ASTX were associated with BM restoration of angiopoietin 1, stromal cell-derived factor 1, β-catenin, and Nrf2. Long-term ASTX gavage also recovered the STZ-induced dysfunction in migration, colony formation, and mineralization of BM-derived stromal cells. Further, a direct addition of ASTX exhibited direct and dose-dependent inhibition of osteoclastic activation without cytotoxic effects. Collectively, these results indicate that ASTX protects against diabetes-induced damage in the BM microenvironment in BM, HSCs, and MSCs and restores normal hematopoiesis and bone accrual in STZ-injected mice.
Project description:IntroductionPeople with hazardous alcohol use are more susceptible to viral, bacterial, and fungal infections due to the effect of alcohol on immune system cell function. Metabolized ethanol reduces NAD+ to NADH, affecting critical metabolic pathways. Here, our aim was to investigate whether alcohol is metabolized by bone marrow cells and if it impacts the metabolic pathways of leukocyte progenitor cells. This is said to lead to a qualitative and quantitative alteration of key metabolites which may be related to the immune response.MethodsWe addressed this aim by using C57BL/6 mice under chronic ethanol administration and evaluating the metabolomic profile of bone marrow total cells by gas chromatography-coupled mass spectrometry (GC-MS).ResultsWe identified 19 metabolites. Our data demonstrated that chronic ethanol administration alters the metabolomic profile in the bone marrow, resulting in a statistically diminished abundance of five metabolites in ethanol-treated animals: uracil, succinate, proline, nicotinamide, and tyrosine.DiscussionOur results demonstrate for the first time in the literature the effects of alcohol consumption on the metabolome content of hematopoietic tissue and open a wide range of further studies to investigate mechanisms by which alcohol compromises the cellular function of the immune system.
Project description:Diabetic peripheral neuropathy (DPN) is a long-term complication associated with nerve dysfunction and uncontrolled hyperglycemia. In spite of new drug discoveries, development of effective therapy is much needed to cure DPN. Here, we have developed a combinatorial approach to provide biochemical and electrical cues, considered to be important for nerve regeneration. Exosomes derived from bone marrow mesenchymal stromal cells (BMSCs) were fused with polypyrrole nanoparticles (PpyNps) containing liposomes to deliver both the cues in a single delivery vehicle. We developed DPN rat model and injected intramuscularly the fused exosomal system to understand its long-term therapeutic effect. We found that the fused system along with electrical stimulation normalized the nerve conduction velocity (57.60 ± 0.45 m/s) and compound muscle action potential (16.96 ± 0.73 mV) similar to healthy control (58.53 ± 1.10 m/s; 18.19 ± 1.45 mV). Gastrocnemius muscle morphology, muscle mass, and integrity were recovered after treatment. Interestingly, we also observed paracrine effect of delivered exosomes in controlling hyperglycemia and loss in body weight and also showed attenuation of damage to the tissues such as the pancreas, kidney, and liver. This work provides a promising effective treatment and also contribute cutting edge therapeutic approach for the treatment of DPN.