Meimei Shi

Serum metabolic variables associated with impaired glucose tolerance induced by high-fat-high-cholesterol diet in Macaca mulatta

Dyslipidemia caused by ‘Western-diet pattern’ is a strong risk factor for the onset of diabetes. This study aimed to disclose the relationship between the serum metabolite changes induced by habitual intake of high-fat and high-cholesterol (HFHC) diet and the development of impaired glucose tolerance (IGT) and insulin resistance through animal models of Macaca mulatta. Sixteen M. mulatta (six months old) were fed a control diet or a HFHC diet for 18 months. The diet effect on serum metabolic profiles was investigated by longitudinal research. Islet function was assessed by intravenous glucose tolerance and hyperinsulinemic-euglycemic clamp test. Metabonomics were determined by 1 H proton nuclear magnetic resonance spectroscopy. Prolonged diet-dependent hyperlipidemia facilitated visceral fat accumulation in liver and skeletal muscle and disorder of glucose homeostasis in juvenile monkeys. Glucose disappearance rate (KGlu) and insulin response to the glucose challenge effects in HFHC monkeys were significantly lower than in control monkeys. Otherwise, serum trimethylamine-N-oxide (TMAO), lactate and leucine/isoleucine were significantly higher in HFHC monkeys. Sphingomyelin and choline were the most positively correlated with KGlu (R2 = 0.778), as well as negative correlation (R2 = 0.64) with total cholesterol. The HFHC diet induced visceral fat, abnormal lipid metabolism and IGT prior to weight gain and body fat content increase in juvenile monkeys. We suggest that increased serum metabolites, such as TMAO, lactate, branched-chain amino acids and decreased sphingomyelin and choline, may serve as possible predictors for the evaluation of IGT and insulin resistance risks in the prediabetic state.

Mesenchymal stem cell-conditioned media ameliorate diabetic endothelial dysfunction by improving mitochondrial bioenergetics via the Sirt1/AMPK/PGC-1α pathway

Vasculopathy is a major complication of diabetes. Impaired mitochondrial bioenergetics and biogenesis due to oxidative stress are a critical causal factor for diabetic endothelial dysfunction. Sirt1, an NAD+-dependent enzyme, is known to play an important protective role through deacetylation of many substrates involved in oxidative phosphorylation and reactive oxygen species generation. Mesenchymal stem cell-conditioned medium (MSC-CM) has emerged as a promising cell-free therapy due to the trophic actions of mesenchymal stem cell (MSC)-secreted molecules. In the present study, we investigated the therapeutic potential of MSC-CMs in diabetic endothelial dysfunction, focusing on the Sirt1 signalling pathway and the relevance to mitochondrial function. We found that high glucose-stimulated MSC-CM attenuated several glucotoxicity-induced processes, oxidative stress and apoptosis of endothelial cells of the human umbilical vein. MSC-CM perfusion in diabetic rats ameliorated compromised aortic vasodilatation and alleviated oxidative stress in aortas. We further demonstrated that these effects were dependent on improved mitochondrial function and up-regulation of Sirt1 expression. MSC-CMs activated the phosphorylation of phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt), leading to direct interaction between Akt and Sirt1, and subsequently enhanced Sirt1 expression. In addition, both MSC-CM and Sirt1 activation could increase the expression of peroxisome proliferator-activated receptor γ co-activator-1α (PGC-1α), as well as increase the mRNA expression of its downstream, mitochondrial, biogenesis-related genes. This indirect regulation was mediated by activation of AMP-activated protein kinase (AMPK). Overall our findings indicated that MSC-CM had protective effects on endothelial cells, with respect to glucotoxicity, by ameliorating mitochondrial dysfunction via the PI3K/Akt/Sirt1 pathway, and Sirt1 potentiated mitochondrial biogenesis, through the Sirt1/AMPK/PGC-1α pathway.

1H NMR-based metabonomic analysis of serum and urine in a nonhuman primate model of diabetic nephropathy

Diabetic nephropathy (DN) is a serious metabolic disease, and comprehensive understanding of its complex mechanism will help in preventing the onset and progression of DN. To reveal the systemic metabolic changes associated with renal injury, we performed 1H NMR-based metabonomic and multivariate analyses to analyze serum and urine obtained from a nonhuman primate model of DN. Our results indicated that DN monkeys exhibited a distinct metabolic profile, including higher levels of VLDL/LDL, lipids, unsaturated lipids, uric acid, allantoin, fumarate and hippurate, as well as lower levels of HDL, alanine, glutamate, pyruvate, formate, tyrosine, histidine and NAD+. The disturbed metabolic pathways were further identified, including NAD+ metabolism, purine metabolism, oxidative stress, lipid metabolism, and renal tubular reabsorption. This study highlights that NMR-based metabonomics provides insight into the underlying pathways in the pathogenesis and progression of DN at the metabolic level.

Mesenchymal Stem Cells Ameliorated Glucolipotoxicity in HUVECs through TSG-6

Glucolipotoxicity is one of the critical causal factors of diabetic complications. Whether mesenchymal stem cells (MSCs) have effects on glucolipotoxicity in human umbilical vein endothelial cells (HUVECs) and mechanisms involved are unclear. Thirty mM glucose plus 100 μM palmitic acid was used to induce glucolipotoxicity in HUVECs. MSCs and HUVECs were co-cultured at the ratio of 1:5 via Transwell system. The mRNA expressions of inflammatory factors were detected by RT-qPCR. The productions of reactive oxygen species (ROS), cell cycle and apoptosis were analyzed by flow cytometry. The tumor necrosis factor-α stimulated protein 6 (TSG-6) was knockdown in MSCs by RNA interference. High glucose and palmitic acid remarkably impaired cell viability and tube formation capacity, as well as increased the mRNA expression of inflammatory factors, ROS levels, and cell apoptosis in HUVECs. MSC co-cultivation ameliorated these detrimental effects in HUVECs, but no effect on ROS production. Moreover, TSG-6 was dramatically up-regulated by high glucose and fatty acid stimulation in both MSCs and HUVECs. TSG-6 knockdown partially abolished the protection mediated by MSCs. MSCs had protective effects on high glucose and palmitic acid induced glucolipotoxicity in HUVECs, and TSG-6 secreted by MSCs was likely to play an important role in this process.

Gene expression profile of vascular ischemia-reperfusion injury in rhesus monkeys

UNLABELLED The vascular system particularly endothelium is sensitive to ischemia-reperfusion (I/R) injury, which is a big challenge in surgical practices and many vascular disorders. In the present study, we reported the global gene expression changes in a 2-h ischemia and 4-h reperfusion injury induced in the hind limb vessels of rhesus monkeys (Macaca mulatta) using microarray technique. RESULTS The histological results showed abnormal morphology of endothelial cells after 2-h ischemia and the hematological detection found slightly extension of coagulation time after I/R treatment. Furthermore, we found distinct alterations in gene expression patterns during I/R process. These identified genes are mostly involved in inflammation, immune response, apoptosis, and cell stress signaling pathways. The significantly up-regulated genes included IL-6, regulator of G-protein signaling 8, selectin E, and metallothionein 2A, et al. Whist, the robustly down-regulated genes included NECAP endocytosis associated 2, transglutaminase 2, and fibronectin 1, et al. CONCLUSION Our results indicate that inflammation, primarily characterized by gene expression changes of cytokines and chemokines is the most important event in the early stage of I/R injury in blood vessels.

Molecular Cloning and Characterization of Rhesus Monkey Platelet Glycoprotein Ibα, a major ligand-binding subunit of GPIb-IX-V complex

INTRODUCTION Through binding to von Willebrand factor (VWF), platelet glycoprotein (GP) Ibα, the major ligand-binding subunit of the GPIb-IX-V complex, initiates platelet adhesion and aggregation in response to exposed VWF or elevated fluid-shear stress. There is little data regarding non-human primate platelet GPIbα. This study cloned and characterized rhesus monkey (Macaca Mullatta) platelet GPIbα. MATERIALS AND METHODS DNAMAN software was used for sequence analysis and alignment. N/O-glycosylation sites and 3-D structure modelling were predicted by online OGPET v1.0, NetOGlyc 1.0 Server and SWISS-MODEL, respectively. Platelet function was evaluated by ADP- or ristocetin-induced platelet aggregation. RESULTS Rhesus monkey GPIbα contains 2,268 nucleotides with an open reading frame encoding 755 amino acids. Rhesus monkey GPIbα nucleotide and protein sequences share 93.27% and 89.20% homology respectively, with human. Sequences encoding the leucine-rich repeats of rhesus monkey GPIbα share strong similarity with human, whereas PEST sequences and N/O-glycosylated residues vary. The GPIbα-binding residues for thrombin, filamin A and 14-3-3ζ are highly conserved between rhesus monkey and human. Platelet function analysis revealed monkey and human platelets respond similarly to ADP, but rhesus monkey platelets failed to respond to low doses of ristocetin where human platelets achieved 76% aggregation. However, monkey platelets aggregated in response to higher ristocetin doses. CONCLUSIONS Monkey GPIbα shares strong homology with human GPIbα, however there are some differences in rhesus monkey platelet activation through GPIbα engagement, which need to be considered when using rhesus monkey platelet to investigate platelet GPIbα function.

MSCs protect endothelial cells from inflammatory injury partially by secreting STC1

&NA; Inflammatory factors play an important role in the pathogenesis of diabetic vascular complications. Considerable interest in the therapeutic potential of mesenchymal stem cells (MSCs) has recently arisen. The purposes of this study were to investigate the effects of MSCs on endothelial cells under inflammatory conditions and to determine the relevant mechanism underlying these effects. In vitro, after TNF‐&agr; stimulation, MSCs‐CM treatment significantly restored cell viability, reduced THP‐1 cell adhesion and enhanced tube formation capacity via inhibiting ROS overproduction and NF‐&kgr;B activation, subsequently down‐regulating adhesion molecules and chemokines. These effects may be partially due to the up‐regulation of uncoupling protein 2 (UCP2) in HUVECs that was induced by the secretion of stanniocalcin 1 (STC1) from MSCs. In vivo, MSCs transplantation ameliorated the progression of diabetes‐associated vascular dysfunction by reducing ROS production and down‐regulating the expression of adhesion molecules. These results suggest that MSCs protect HUVECs from inflammatory injury partially by secreting STC1. MSCs may be a potential therapeutic approach for the treatment of diabetic vascular complications. HighlightsMSCs ameliorate inflammatory factor induced endothelial damage.MSCs protect HUVECs from inflammatory injury partially by secreting STC1.STC1 up‐regulated UCP2 in HUVECs and subsequently reduced ROS accumulation and inflammation cascade.MSCs may be potential for diabetic vascular complications therapy.