T2DM Rat Research Articles

Targeting microbiota–immune–synaptic plasticity to explore the effect of tea polyphenols on improving memory in the aged type 2 diabetic rat model

ABSTRACT Objectives The study aimed to explore whether TP could improve memory in the aged type 2 diabetic rat model by regulating microbiota-immune-synaptic plasticity axis. Methods The experiment was divided into two parts. Firstly, to investigate the effects of TP on the physiopathology of the aged T2DM model rats, rats were randomly divided into the Normal control group, the aged group, the Aged T2DM model group, the TP 75, 150, 300 mg/kg groups, the 150 mg/kg Piracetam group and the 3 mg/kg Rosiglitazone group. Then, to further verify whether TP improved memory in aged T2DM rat model by regulating intestinal flora, the fecal microbiota transplantation (FMT) from the rats in the 300 mg/kg TP group into the rats in the aged T2DM model group was carried out. Effects on gut microbiota, colonic integrity (epithelial tight junction proteins), and endotoxemia (serum LPS) were examined, along with synaptic structure, synaptic plasticity-related structural proteins and inflammation signaling of the hippocampus in our study. Results Our results demonstrated that TP alleviated memory impairments in the aged T2DM rat model. The specific outcomes were as follows: TP 300 mg/kg corrected the gut dysbacteriosis, alleviated intestinal permeability reduction and peripheral/central inflammation, inhibited the TLR4/NF-κB signaling pathway. Meanwhile, TP improved the synaptic plasticity in the hippocampus of the aged T2DM model rats, whose expressions of SYN, PSD 95, NMDAR1 and GluR1 in hippocampus were significantly up-regulated. Surprisingly, rats of the FMT group displayed the same changes. Discussion TP improves the memory in aged T2DM rat model. The mechanism may be related to the alteration of gut flora, which can inhibit hippocampal TLR4/NF-κB signaling to attenuate neuroinflammation, then improve synaptic plasticity. The study proposes that TP interventions aimed at manipulating the gut microbiota may hold great potential as an effective approach for preventing and treating this disease.

Therapeutic potential of ADSCs in diabetic wounds: a proteomics-based approach.

Diabetes mellitus (DM), a chronic metabolic disease characterized by elevated blood sugar, leads to delayed or non-healing wounds, increasing amputation risks, and placing a significant burden on patients and society. While extensive research has been conducted on adipose-derived stem cells (ADSCs) for promoting wound healing, there is a scarcity of studies focusing on diabetic wounds, particularly those employing proteomics and bioinformatics approaches. This study aimed to investigate the mechanisms by which ADSCs promote diabetic wound healing using proteomics and bioinformatics techniques. Healthy rat fat tissue was used to isolate ADSCs. A T2DM rat model with back wounds was established. The experimental group received ADSC injections around the wound, while the control group received PBS injections. Wound healing rates were documented and photographed on days 0, 3, 7, 10, and 14. On day 7, wound tissues were excised for HE and Masson's staining. Additionally, on day 7, tissues were analyzed for protein quantification using 4D-DIA, with subsequent GO and KEGG analyses for differentially expressed proteins (DEPs) and protein-protein interaction (PPI) network analysis using STRING database (String v11.5). Finally, Western blot experiments were performed on day 7 wounds to verify target proteins. In all measured days postoperatively, the wound healing rate was significantly higher in the ADSC group than in the PBS group (day 7: p < 0.001, day 10: p = 0.001, day 14: p < 0.01), except on day 3 (p > 0.05). Proteomic analysis identified 474 differentially expressed proteins, with 224 key proteins after PPI analysis (78 upregulated and 146 downregulated in the ADSC group). The main cellular locations of these proteins were "cellular anatomical entity" and "protein-containing complex", while the biological processes were "cellular processes" and "biological regulation". The primary molecular functions were "binding" and "catalytic activity", with GO enrichment focused on "Wnt-protein binding", "neural development", and "collagen-containing extracellular matrix". Further analysis of PPI network nodes using LASSO regression identified Thy1 and Wls proteins, significantly upregulated in the ADSC group, as potentially crucial targets for ADSC application in diabetic wound treatment.

Untargeted metabolomics reveals dynamic changes in metabolic profiles of rat supraspinatus tendon at three different time points after diabetes induction.

To investigate the dynamic changes of metabolite composition in rat supraspinatus tendons at different stages of diabetes by untargeted metabolomics analysis. A total of 80 Sprague-Dawley rats were randomly divided into normal (NG, n = 20) and type 2 diabetes mellitus groups (T2DM, n = 60) and subdivided into three groups according to the duration of diabetes: T2DM-4w, T2DM-12w, and T2DM-24w groups; the duration was calculated from the time point of T2DM rat model establishment. The three comparison groups were set up in this study, T2DM-4w group vs. NG, T2DM-12w group vs. T2DM-4w group, and T2DM-24w group vs. T2DM-12w group. The metabolite profiles of supraspinatus tendon were obtained using tandem mass spectrometry. Metabolomics multivariate statistics were used for metabolic data analysis and differential metabolite (DEM) determination. The intersection of the three comparison groups' DEMs was defined as key metabolites that changed consistently in the supraspinatus tendon after diabetes induction; then, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed. T2DM-4w group vs. NG, T2DM-12w group vs. T2DM-4w group, and T2DM-24w group vs. T2DM-12w group detected 94 (86 up-regulated and 8 down-regulated), 36 (13 up-regulated and 23 down-regulated) and 86 (24 up-regulated and 62 down-regulated) DEMs, respectively. Seven key metabolites of sustained changes in the supraspinatus tendon following induction of diabetes include D-Lactic acid, xanthine, O-acetyl-L-carnitine, isoleucylproline, propoxycarbazone, uric acid, and cytidine, which are the first identified biomarkers of the supraspinatus tendon as it progresses through the course of diabetes. The results of KEGG pathway enrichment analysis showed that the main pathway of supraspinatus metabolism affected by diabetes (p < 0.05) was purine metabolism. The results of the KEGG metabolic pathway vs. DEMs correlation network graph revealed that uric acid and xanthine play a role in more metabolic pathways. Untargeted metabolomics revealed the dynamic changes of metabolite composition in rat supraspinatus tendons at different stages of diabetes, and the newly discovered seven metabolites, especially uric acid and xanthine, may provide novel research to elucidate the mechanism of diabetes-induced tendinopathy.

Mulberry leaf multi-components exert hypoglycemic effects through regulation of the PI-3K/Akt insulin signaling pathway in type 2 diabetic rats

Ethnopharmacological relevancePhytochemicals have unique advantages in the treatment of diabetes due to their multi-target activity and low toxicity. Mulberry leaves, a traditional Chinese herbal medicine, have been used in the prevention and treatment of diabetes for centuries. The main active ingredients in mulberry leaves with regards to the hypoglycemic effect are 1-deoxynojirimycin, flavonoids, and polysaccharides. However, the combined hypoglycemic effects and mechanisms of mulberry leaf multi-components remain unclear. Aim of the studyThis study explored the anti-diabetic effects of mulberry leaf multi-components (MMC) and the role of the PI-3K/Akt insulin signalling pathway in improving insulin resistance. Materials and methodsThe main chemical components of MMC were analyzed using the phenol-sulfuric acid method, aluminum nitrate-sodium nitrite method, and HPLC-ultraviolet/fluorescence detection method. The T2DM rat model was created via feeding a high-fat diet and peritoneal injection of streptozotocin. T2DM rats were divided into four groups: model, model plus metformin, model plus low-dose, and model plus high-dose MMC groups (100 and 200 mg/kg body weight/day, respectively), and plus normal group for a total of five groups. MMC was administered by oral gavage for six weeks. Fasting blood glucose and serum lipid profiles were measured using a glucometer and an automatic biochemistry analyzer, respectively. Serum insulin and adipocytokine levels were analyzed by ELISA. Hepatic glucose metabolizing enzyme activity was evaluated by ELISA and the double antibody sandwich method. Expression of PI-3K/Akt signalling pathway proteins was analyzed by RT-PCR and Western blotting. ResultsExtracted 1-deoxynojirimycin, flavonoid, and polysaccharide purity was 70.40%, 52.34%, and 32.60%, respectively. These components were then mixed at a ratio of 1:6:8 to form MMC. MMC significantly reduced serum glucose, insulin, and lipid levels. In diabetic rats, MMC enhanced insulin sensitivity and alleviated inflammatory and oxidative damage by lowing adipocytokine levels and increasing anti-oxidative enzyme activity. Insulin resistance was also mitigated. MMC regulated the activity of key downstream enzymes of hepatic glucose metabolism via activating the expression of PI-3K, Akt, PDX-1, and GLUT4 at the mRNA and protein levels, thereby correcting hepatic glucolipid metabolism disorders and exerting a hypoglycemic effect. ConclusionMMC ameliorated hepatic glucolipid metabolism disorders and improved insulin resistance in T2DM rats by activating the PI-3K/Akt signaling pathway. These results highlight the multi-component, multi-target, and combined effects of MMC, and suggest it may be further developed as a hypoglycemic drug.

Pharmacokinetic interaction of saxagliptin with andrographolide and their hypoglycemic effect in type 2 diabetes rats

Controlling blood glucose is the primary therapeutic strategy for T2DM. Both saxagliptin and andrographolide possess hypoglycemic effects, which makes the combination easy. This study aimed to evaluate the co-administration of saxagliptin and andrographolide in rats and revealed their combined impact on type 2 diabetes (T2DM). T2DM rat models were established by high-fat diet and the injection of nicotinamide and streptozotocin. The blood glucose and insulin resistance index were monitored after modeling. The pharmacokinetics of saxagliptin was assessed by orally administrating 10 mg/kg saxagliptin. The co-administration was performed with the pre-treatment of 30 or 100 mg/kg andrographolide. In vitro, the metabolic stability of saxagliptin was assessed in rat liver microsomes. The co-administration of saxagliptin with andrographolide induced significant changes in the pharmacokinetics of saxagliptin, behaving as the increasing AUC(0-t), Cmax, t1/2, and the decreasing clearance rate. The effect of andrographolide was enhanced with the growing concentration. Combination with andrographolide enhanced the hypoglycemic effect and alleviated insulin resistance of saxagliptin in T2DM rats. Co-administration of saxagliptin with 100 mg/kg andrographolide induced hypoglycemia. In vitro, andrographolide significantly improved the metabolic stability of saxagliptin and showed a significant inhibitory effect on the activity of CYP3A4. Combining saxagliptin with andrographolide could increase the systemic exposure of saxagliptin via inhibiting CYP3A4 and improve the hypoglycemic effect, but the high concentration of andrographolide is the risk of inducing hypoglycemia.

Total flavonoids of Hippophae rhamnoides L. improves type 2 diabetes symptoms in rats through down-regulating of the DAG/PRKCA/MAPK10/p65/TNF-α signalling pathway

Ethnopharmacological relevanceDry mature fruits of Hippophae rhamnoides L. (HRL), Elaeagnaceae, have traditional functions of invigorating spleen and improving spleen insufficiency. Traditional Chinese medicine (TCM) clinics have been proved that HRL is in favor of diabetes treatment. Modern pharmacological studies demonstrated that total flavones of Hippophae rhamnoides (TFH) are the main substance for HRL to develop anti-inflammation and anti-diabetes functions. However, chemical features, active ingredients and anti-diabetes pharmacological mechanism of HRL still remain unclear. Aim of the studyKey targets and metabolites in anti-type-II diabetes mellitus (T2DM) of TFH have been explored based on AGE-RAGE signaling pathway in diabetic complications. The anti-T2DM mechanism of TFH has been elaborated from comprehensive perspectives, including target prediction, metabolites, potential metabolic pathways, and so on. Materials and methodsIn this study, a qualitative test of chemical composition of HRL was carried out based on ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The anti-T2DM targets and pathways of HRL were predicted through network pharmacological approach. The T2DM rat model was induced by high-fat and high-glucose diet combined with streptozotocin (STZ). The T2DM model was evaluated through fasting blood glucose level, body weight, serum biochemical indicators, insulin levels and homeostatic model assessment of insulin resistance. The key metabolic pathways were screened through the correlation between metabolites and key targets. Finally, the quantitative analysis of key targets and metabolites was verified through experiments. ResultsAfter TFH intervention, the fasting blood-glucose level of T2DM rats induced by high-fat and high-glucose diet combined with streptozotocin (STZ) was downregulated significantly, while body weight, serum liquid level, insulin levels and homeostatic model assessment of insulin resistance (HOMA-IR) were improved. According to ELISA, Western blotting (WB) and reverse transcriptase polymerase chain reaction (RT-PCR), TFH significantly downregulates expression levels of diglyceride (DAG)-activated protein kinase C (PRKCA), mitogen activated protein kinase 10 (MAPK10), human nuclear factor κB subunit p65 (NF-κB p65) and tumor necrosis factor-α (TNF-α) in pancreas of STZ-induced rats. ConclusionsTFH downregulates expressions of PRKCA, MAPK10 and p65 TNF-α as well as level of the key metabolite DA in the DAG/PRKCA/MAPK10/TNF-α/p65 pathways, improves lipid metabolism disorder, inhibits inflammatory response and thereby relieves symptoms of T2DM.

Assessment of Antidiabetic and Anti-Inflammatory Activities of Carissa carandas Linn Extract: In Vitro and In Vivo Study

This study investigated the effects of aqueous fruit extracts of Carissa carandas (CCA) on inflammation and insulin resistance using an in vitro cellular model, in vivo high-fat diets, and a streptozotocin-induced type 2 diabetic (T2DM) rat model. CCA significantly ameliorated inflammation by decreasing nitric oxide production in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. Interestingly, CCA showed anti-insulin resistance activities, as it significantly improved glucose uptake and decreased glycerol release in LPS-induced 3T3-L1 adipocytes. In vivo studies showed that a high dose of 12-week oral supplementation of CCA (400 mg/kg BW/day) significantly reduced visceral fat, triglycerides, and cholesterol level in the blood of diabetic rats. Importantly, the metabolic parameters in both fasting and postprandial states, including fasting plasma glucose, HOMA-IR, and glucose intolerance, significantly improved, indicating its antihyperglycemic benefit in diabetic rats. Moreover, the results of the HOMA-β and histological examination suggested that pancreatic β-cell function and pancreatic morphological changes of the CCA and metformin treatments appeared to be better than those in non-treated diabetes, indicating the protective effect of CCA against pancreatic damage caused by hyperglycemia. In conclusion, the present study first reported that the C. carandas fruit extract has anti-inflammation and anti-insulin resistance, and subsequently improved glycemic control in the T2DM rat model.

Investigation of the material basis of Xiexin Tang to alleviate type 2 diabetes mellitus based on spectrum-effect analysis by UPLC-Q-TOF/MS

Xiexin Tang (XXT) is a classic prescription for treating diabetes in clinical practices for thousands of years in China, which has been also proved by a large number of modern pharmacological studies. However, due to its complex composition, the bioactive ingredients of XXT is still unclear. In present researches, spectrum-effect relationship analysis is widely used to explore the material basis of traditional medical herbs, so this method was adopted in this study. Firstly, the extract of XXT was separated and enriched into 5 fractions by macroporous adsorption resin. Then, UPLC-Q-TOF/MS method was used for qualitative identification of components in each eluting part, and efficacy of each fraction was assessed by the T2DM rat model. Based on grey relational analysis and pearson bivariate correlation analysis, it was found that the components such as berberine, gallic acid, catechin, epicatechin, acteoside, berberastine and 1-O-galloyl-β-D-glucose might be the main effective basis of XXT to improve T2DM.

Ranolazine Attenuates Brain Inflammation in a Rat Model of Type 2 Diabetes.

Recent studies suggest a pathogenetic association between metabolic disturbances, including type 2 diabetes (T2DM), and cognitive decline and indicate that T2DM may represent a risk factor for Alzheimer's disease (AD). There are a number of experimental studies presenting evidence that ranolazine, an antianginal drug, acts as a neuroprotective drug. The aim of the present study was to evaluate the effects of ranolazine on hippocampal neurodegeneration and astrocytes activation in a T2DM rat model. Diabetes was induced by a high fat diet (HFD) and streptozotocin (STZ) injection. Animals were divided into the following groups: HFD/STZ + Ranolazine, HFD/STZ + Metformin, HFD/STZ + Vehicle, NCD + Vehicle, NCD + Ranolazine and NCD + Metformin. The presence of neurodegeneration was evaluated in the hippocampal cornus ammonis 1 (CA1) region by cresyl violet staining histological methods, while astrocyte activation was assessed by western blot analysis. Staining with cresyl violet highlighted a decrease in neuronal density and cell volume in the hippocampal CA1 area in diabetic HFD/STZ + Vehicle rats, while ranolazine and metformin both improved T2DM-induced neuronal loss and neuronal damage. Moreover, there was an increased expression of GFAP in the HFD/STZ + Vehicle group compared to the treated diabetic groups. In conclusion, in the present study, we obtained additional evidence supporting the potential use of ranolazine to counteract T2DM-associated cognitive decline.

Shexiang Tongxin Dropping Pills improve cardiac function in type 2 diabetic rats by promoting angiogenesis

This study aims to investigate the effect of Shexiang Tongxin Dropping Pills(STDP) on angiogenesis in type 2 diabetic rats. Thirty-two healthy male SD rats were randomized into a control group(n=6) and a type 2 diabetes mellitus(T2 D) group(n=26). The T2 DM rat model was established with a high-fat diet combined with intraperitoneal injection of streptozotocin(STZ). The model rats were randomized into a model group, a metformin group(200 mg·kg~(-1)·d~(-1)), a high-dose STDP group(40 mg·kg~(-1)·d~(-1)), and a low-dose STDP group(20 mg·kg~(-1)·d~(-1)). The corresponding agents were administered continuously for 8 weeks. The cardiac function indexes were detected by ultrasound at the experiment endpoint, and the myocardial pathological changes were observed via hematoxylin-eosin(HE) staining. The myocardial collagen fiber deposition was detected by sirius red staining, the myocardial microvascular density by immunohistochemistry, and the myocardial hypertrophy by immunofluorescence assay. Western blot was employed to measure the protein levels of vascular endothelial growth factor(VEGF), phosphorylated vascular endothelial growth factor receptor 2(p-VEGFR2), angiogenin-1(Ang1), and TEK tyrosine kinase endothelial(Tie2) in the myocardial tissue. Compared with the model group, STDP improved the left ventricular ejection fraction(EF) and fractional shortening(FS)(P&lt;0.05), alleivated the arrangement disorder of myocardial cells and local myocardial fiber rupture. Sirius red staining showed that STDP, especially the high-dose group(P&lt;0.01), repaired the deposition of myocardial collagen fibers in the model group. The results of immunohistochemistry showed that the cluster of differentiation 31(CD31) expression in myocardial tissue of the STDP groups was higher than that of the model group(P&lt;0.01). The immunofluorescence results showed that STDP mitigated the hypertrophy of myocardial cells(P&lt;0.01). Furthermore, STDP up-regulated the protein levels of VEGF, p-VEGFR2, Angl, and Tie2 compared with the model group(P&lt;0.01). In conclusion, STDP can effectively promote angiogenesis and improve cardiac function in type 2 diabetic rats by up-regulating the expression of VEGF, p-VEGFR2, Ang1, and Tie2.

Antioxidant and anti-inflammatory activities of Centratherum anthelminticum (L.) Kuntze seed oil in diabetic nephropathy via modulation of Nrf-2/HO-1 and NF-κB pathway

BackgroundType 2 diabetes mellitus (T2DM) approximately constitutes 90% of the reported cases. 30-40% of diabetics eventually develop diabetic nephropathy (DN); accounting for one of the major causes of morbidity and mortality. Increased glucose autoxidation and non-enzymatic glycation of proteins in diabetic kidneys lead to the excessive generation of reactive oxygen species (ROS) that results in lipid peroxidation and activation of inflammatory mediators which overwhelms the scavenging capacity of the antioxidant defense system (Nrf2/Keap1/HO-1). Centratherum anthelminticum commonly called as kali zeeri (bitter cumin) and its seeds are well known for culinary purposes in Asia (Pakistan). It has reported anti-inflammatory, antioxidant, and anti-diabetic activities. The present study has attempted to explore the in-vivo anti-inflammatory, antioxidant and antihyperglycemic potential of the C. anthelminticum seed’s fixed oil (FO) and its fractions in high fat-high fructose-streptozotocin (HF-HFr-STZ) induced T2DM rat model.MethodsThe T2DM rat model was developed by giving a high-fat and high-fructose diet followed by a single intraperitoneal injection of streptozotocin (STZ 60 mg/kg) on 28th day of the trial. After 72 hours of this injection, rats showing fasting blood glucose (FBG) levels≥230 mg/dL were recruited into six groups. These groups were orally administered distilled water (1 mL/kg), Gliclazide (200 mg/kg), Centratherum anthelminticum seed (FO) and its hexane (HF), chloroform (CF) and ethanol (EF) soluble fractions (200 mg/kg each), respectively for 4 weeks (i.e. 28 days). Blood, serum, and kidney tissue samples of euthanized animals were used for biochemical, pro-inflammatory, and antioxidant markers (ELISA, qRT-PCR, and spectrophotometric assays) and histology, respectively.ResultsC. anthelminticum FO and its fractions reduced the lipid peroxidation, and improved the antioxidant parameters: enzymatic (SOD, CAT, and GPx), non-enzymatic (GSH), and mRNA expression of anti-inflammatory markers (Nrf-2, keap1, and HO-1). mRNA expression of inflammatory and apoptotic markers (TNF-α, IL-1β, COX-1, NF-κB, Bax, and Bcl-2) were attenuated along with improved kidney architecture.ConclusionC. anthelminticum can mitigate inflammation and oxidative stress in early DN. The anti-nephropathic effect can be attributed to its ability to down-regulate NF-κB and by bringing the Nrf-2 expression levels to near normal.

α-lipoic acid ameliorates liver injury in rats with type 2 diabetes mellitus via activating AMPK/mTOR pathway

To investigate the effect of α-lipoic acid in ameliorating liver injury in rats with type 2 diabetes mellitus via activating adenosine 5'-monophosphate-activate protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway. The T2DM rat models were established by feeding with high-fat, high-sucrose diet and intraperitoneal injection of 27.5 mg/(kg·d) streptozotocin. The 32 rats with T2DM were randomly divided into 4 groups: T2DM group, α-lipoic acid group (LA), Compound C group (Comp C, an inhibitor of AMPK) and LA+Comp C group, with 8 rats in each group. Additionally, 8 Sprague-Dawlay (SD) rats without diabetes were set as normal control. The rats received α-lipoic acid at a dosage of 100 mg/(kg·d) or Compound C at a dosage of 20 mg/(kg·d) by intraperitoneal injection for 8 weeks as needed. The levels of relevant biochemical indexes were detected. The weight of liver was recorded to calculate liver weight index (LWI), and the pathological changes of liver tissues were detected by light and electron microscopy. The levels of AMPK, p-AMPK, mTOR, p-mTOR in rat liver were detected by Western blot. Compared with control group, the levels of LWI, homeostasis model assessment of insulin resistance, fasting blood glucose, alanine transaminase, aspartate transaminase, gamma glutamyl transferase and triglyceride in T2DM group were increased significantly (all P＜0.05). The liver tissue lesions were more serious and hepatic steatosis grade was higher. The expression of p-AMPK was decreased (P＜0.05) and the expression of p-mTOR was increased significantly(P＜0.05). α-lipoic acid could reverse the above-mentioned changes, ameliorate insulin resistance (all P＜0.05), protect the structure and function of liver, and activate the AMPK/mTOR pathway (P＜0.05). The protection of α-lipoic acid was weakened by the inhibition of AMPK with Compound C (P＜0.05). α-lipoic acid could protect the liver of rats with T2DM by activating AMPK/mTOR pathway.

Combined resveratrol and vitamin D treatment ameliorate inflammation-related liver fibrosis, ER stress, and apoptosis in a high-fructose diet/streptozotocin-induced T2DM model.

A high fructose diet is a major cause of diabetes and various metabolic disorders, including fatty liver. In this study, we investigated the effects of resveratrol and vitamin D (VitD) treatments on endoplasmic reticulum (ER) stress, oxidative stress, inflammation, apoptosis, and liver regeneration in a rat model of type 2 diabetes mellitus, namely, T2DM Sprague-Dawley rats. This T2DM rat model was created through a combination treatment of a 10% fructose diet and 40mg/kg streptozotocin (STZ). Resveratrol (1mg/kg/day) and VitD (170/IU/week) were administered alone and in combination to both the diabetic and control groups. Immunohistochemical staining was performed to evaluate PCNA, NF-κB, TNF-α, IL-6, IL-1β, GRP78, and active caspase-3 in liver tissue. The TUNEL method and Sirius red staining were used to determine apoptosis and fibrosis, respectively. G6PD, 6-PGD, GR, and GST activities were measured to determine oxidative stress status. We found that the expressions of cytokines (TNF-α, IL-6, and IL-1β) correlated with NF-κB activation and were significantly increased in the T2DM rats. Increased GRP78 expression, indicating ER stress, increased in apoptotic cells, enhanced caspase-3 activation, and collagen accumulation surrounding the central vein were observed in the T2DM group compared with the other groups. The combination VitD + resveratrol treatment improved antioxidant defense via increasing G6PD, 6-PGD, GR, and GST activities compared to the diabetic groups. We concluded that the combined administration of resveratrol with VitD ameliorates the adverse effects of T2DM by regulating blood glucose levels, increasing antioxidant defense mechanisms, controlling ER stress, enhancing tissue regeneration, improving inflammation, and reducing apoptosis in liver cells. In conclusion, this study indicates that the combination treatment of resveratrol + VitD can be a beneficial option for preventing liver damage in fructose-induced T2DM.

The Association of lncRNA and mRNA Changes in Adipose Tissue with Improved Insulin Resistance in Type 2 Obese Diabetes Mellitus Rats after Roux-en-Y Gastric Bypass.

Objective Roux-en-Y gastric bypass (RYGB) has shown good effects in improving obesity and type II diabetes mellitus (T2DM), but the underlying mechanisms remain unclear. This study explored the changes of related lncRNAs, mRNAs, and signaling pathways in white adipose tissue of T2DM rats after RYGB based on RNA-Seq sequencing, with the aim to provide a theoretical basis for RYGB treatment. Methods T2DM rat models were established by continuous feeding with a high-fat diet and injection of streptozotocin (STZ), after which they underwent RYGB or sham surgery. After the surgery, their body weight was measured weekly. Their fasting blood glucose (FBG) and fasting serum insulin (FSI) were also measured. A homeostasis model assessment of insulin resistance (HOMA-IR) was calculated at weeks 0, 8, and 12. Besides, white adipose tissue of T2DM rats was collected for RNA-Seq sequencing and validated by qRT-PCR. A series of bioinformatics analyses, such as differential expression genes (DEGs) screening, was performed. GO and KEGG functional enrichment analysis and protein-protein interaction (PPI) network construction were conducted based on the sequencing data. Results RYGB surgery could significantly inhibit the weight growth rate and decrease the FBG, FSI, and HOMA-IR of T2DM rats. Bioinformatics analysis of RNA sequencing (RNA-Seq) results revealed that 87 DE- lncRNAs (49 upregulated and 38 downregulated) and 1,824 DEGs (896 upregulated and 928 downregulated) were present in between the RYGB group and Sham group. GO and KEGG analysis showed that the target genes of DEGs and differentially expressed lncRNAs (DE-lncRNAs) were mainly associated with amino acid metabolism, fatty acid metabolism, channel activity, and other processes. In addition, the PPI network diagram also displayed that genes such as Fasn, Grin3a, and Nog could be key genes playing a role after RYGB. qRT-PCR showed that the expression level of Grin3a in the RYGB group was significantly increased compared with the Sham group, while the expression of Fasn and Nog was significantly decreased, which was consistent with the sequencing results. Conclusion Using RNA-Seq sequencing, this study revealed the changes of related lncRNAs, mRNAs, and signaling pathways in the white adipose tissue of T2DM rats after RYGB and identified Fasn, Grin3a, and Nog as potential key genes to function after RYGB.

Mechanism of Astragali Radix-Puerariae Lobatae Radix combination in regulating type 2 diabetes mellitus through AMPK signaling pathway: based on network pharmacology and experimental verification

This study aims to explore the mechanism of Astragali Radix-Puerariae Lobatae Radix(AP) combination in the treatment of type 2 diabetes mellitus(T2 DM) based on network pharmacology and experiment. The effective components and targets of the pair were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP) and targets of T2 DM from each disease database. On this basis, the common targets of the medicinals and the disease were screened out. The protein-protein interaction(PPI) network was established based on STRING. Then Cytoscape 3.7.1 was employed for visualization of the common targets and the network topology analysis of key targets, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of core targets by DAVID. Thereby, the possible molecular mechanism was unveiled. High-fat diet was combined with streptozotocin(STZ, injected into tail vein) for T2 DM rat modeling. Rats were classified into the normal group, model group, positive control group(metformin hydrochloride), AP high-dose, medium-dose, and low-dose groups. After 4 weeks of intragastric administration, serum fasting blood glucose(FBG), fasting insulin(FINS), aspartate aminotransferase(AST), alanine aminotransferase(ALT), triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), high-density lipoprotein cholesterol(HDL-C), interleukin(IL)-6, and tumor necrosis factor(TNF)-α of rats in each group were measured. The expression of insulin receptor substrate-2(IRS-2), adenosine monophosphate-activated protein kinase(AMPK), phosphorylated AMPK(p-AMPK), glucose 6 phosphatase(G6 Pase), and phosphoenolpyruvate carboxy kinase(Pepck) in rat liver was detected by Western blot. A total of 131 core targets of the combination in the treatment of T2 DM were screened out, among which protein kinase B(AKT) 1, mitogen-activated protein kinase(MAPK) 1, TNF-α, IL-6 were more critical. KEGG enrichment analysis suggested that the combination decreased blood glucose mainly through PI3 K/AKT signaling pathway, AMPK signaling pathway, TNF signaling pathway, and MAPK signaling pathway. The levels of FBG and FINS were lower and the glycogen level was higher in the AP high-dose and medium-dose groups than in the model group. The levels of AST, ALT, TG, and LDL-C in the three AP groups and the level of TC in AP high-dose and low-dose groups decreased compared with those in the model group. Levels of IL-6 and TNF-α were lower in AP high-dose and medium-dose groups than in the model group. The expression of IRS-2, AMPK, and p-AMPK was higher and that of G6 Pase and Pepck was lower in AP high-dose group than in the model group. Thus, the combination had multi-component, multi-target, and multi-pathway characteristics in the treatment of T2 DM. It may regulate AMPK signaling pathway through IL-6 and TNF-α to influence insulin resistance, glycogen synthesis, gluconeogenesis, islet β cell transport, and inflammatory response, thereby exerting therapeutic effect on T2 DM.

Citronellal alleviate macro- and micro-vascular damage in high fat diet / streptozotocin - Induced diabetic rats via a S1P/S1P1 dependent signaling pathway

Citronellal (CT) is an acyclic monoterpene aldehyde isolated from lemon citronella, which could ameliorate vascular endothelial dysfunction in atherosclerosis in our previous study, however, whether CT can alleviate vascular endothelial dysfunction related with type 2 diabetes (T2DM) is still unknown. So, we investigated the role of CT in vascular dysfunction related to T2DM and the mechanism involved. T2DM rat model was induced by a single intraperitoneal injection of low-dose streptozotocin (STZ) (60 mg/kg) to rats fed with high-fat diet (HFD) (4 weeks). After treated with CT (150 mg/kg/d), both the thoracic aorta injury and micro-vascular pathological injury in T2DM rats ex vivo were alleviated, and the oxidative stress in T2DM rats treated with CT were attenuated, manifested as increased content of endothelial nitric oxide synthase (eNOS) and superoxide dismutase (SOD), and decreased content of malondialdehyde (MDA). Furthermore, CT (15 μg/L) increased the migration capacity of human umbilical vein endothelial cells (HUVECs) under high glucose circumstance (30 mM), and increased the endothelial-dependent relaxation in thoracic aorta isolated from T2DM rats in vitro. Finally, all of these effects of CT were blocked by fingolimod (FTY720), a sphingosine-1-phosphate receptor agonist, and the expression of sphingosine-1-phosphate receptor 1 (S1P1) was increased by CT. In conclusion, CT improved vascular function through S1P/S1P1 signaling pathway.

Exercise Moderate Interval as an Alternative Exercise Model to Improve Insulin Sensitivity in Type 2 Diabetes Mellitus

Background: Previous study had found that adherence of patient Diabetes type-2 in moderate to vigorous continuous exercise was poor. Meanwhile regularly exercise needs to control fasting plasma glucose in T2DM. Exercise moderate interval (EMI) shows a significant effect to reduce insulin resistance in metabolic syndrome patients. But, studies to find the effect of EMI or EMC to muscle insulin receptor distribution are still limited. So, the purpose of this study is to compare the effects of EMI vs EMC in increasing insulin receptors and reducing insulin resistance in T2DM rat models. Methods: An experimental study was conducted in T2DM rat models. According to the treatments, rats were randomly assigned into the following 3 groups: sedentary group (SG), exercise moderate continuous (EMC) and exercise moderate interval (EMI). Each training session was performed on treadmill for eight weeks, three times weekly. Insulin receptor was assessed by immunohistochemistry. Meanwhile, insulin resistance was determined by Homeostatic Model Assessment for Insulin (HOMA-IR) score measured before and after completing the training session. Results: This study found that the insulin receptors in EMI = 184% and EMC= 158%. Both insulin receptors in the treatment groups had a significant difference to the sedentary group (p=0.002), but there was no significant difference in insulin receptors between EMI and EMC group (p=0.222). HOMA-IR in EMI group was 16.6±4.0 significantly different with HOMA-IR in EMC group such as 43.3±8.6 (p=0.000). There was a negative correlation between insulin receptor and insulin resistance (p=0.047, r =-0.520). Conclusion: Both exercise moderate interval and exercise moderate continuous could increase insulin receptors on skeletal muscle, but exercise moderate interval showed better improve insulin sensitivity after eight weeks' exercise. We suggested exercise moderate interval as an alternative exercise for Type 2 Diabetes Mellitus.

The Effect and Mechanism of Duodenal-Jejunal Bypass to Treat Type 2 Diabetes Mellitus in a Rat Model

Objectives: Bariatric surgery can treat obesity and T2DM, but the specific mechanism is unknown. This study investigated the effect and possible mechanism of duodenal-jejunal bypass (DJB) to treat T2DM. Methods: A T2DM rat model was established using a high-fat, high-sugar diet and a low dose of streptozotocin. DJB surgery and a sham operation (SO) were performed to analyze the effects on glucose homeostasis, lipid metabolism, and inflammation changes. Furthermore, the glucagon-like peptide-1 (GLP-1) in the ileum and the markers of endoplasmic reticulum stress (ERS) in the pancreas were examined after the surgery. The insulinoma cells (INS-1) were divided into three groups; group A was cultured with a normal sugar content (11.1 mmol/L), group B was cultured with fluctuating high glucose (11.1 mmol/L alternating with 33.3 mmol/L), and group C was cultured with fluctuating high glucose and exendin-4 (100 nmol/L). The cells were continuously cultured for 7 days in complete culture medium. The viability of the INS-1 cells was then investigated using the MTT method, apoptosis was detected by flow cytometry, and the ERS markers were detected by Western blot. Results: The blood glucose, lipids, insulin, and TNF-α were significantly elevated in the T2DM model. A gradual recovery was observed in the DJB group. GLP-1 expression in the distal ileum of the DJB group was significantly higher than that in the T2DM control group (DM) and the SO group (p < 0.05), and the markers of ERS expression in the pancreases of the DJB group decreased significantly more than those of groups DM and SO (p < 0.05). Compared with group A, the cell viability in group B was decreased, and the ERS and apoptosis were increased (p < 0.05). However, compared with group B, the cell viability in group C was improved, and the ERS and apoptosis declined (p < 0.05). Conclusions: DJB can be used to treat T2DM in T2DM rats. The mechanism may be that the DJB stimulates the increased expression of GLP-1 on the far side of the ileum, and then, GLP-1 inhibits ERS in the pancreas, reducing the apoptosis of β cells to create a treatment effect in the T2DM rats.

Streptozotocin-induced type 1 and 2 diabetes in rodents: a model for studying diabetic cardiac autonomic neuropathy.

Several animal models are continually being developed to study diabetic complication. Several conflicting regimen for diabetes induction exist in the literature with varying dose strength and regimen for different study interest in diabetes. This study aims to show the effect of high dose streptozotocin (STZ) on the one hand compared with multiple low doses after high fat diet induction on diabetic cardiac autonomic neuropathy (DCAN). Eighty-four Wistar rats were used to demonstrate DCAN induction using 2 approaches one for T1DM (STZ 50mg/kg) and the other for T2DM (HFD for 8 weeks with STZ 25mg/Kg daily for five days). DCAN features were assessed using invasive biomarkers, histology patterns and cardiac nerve densities. Diabetes induction rate was 76% and 89% in T1DM and T2DM model respectively. T1DM group had significant weight loss, reduced c-peptide, and insulin level post induction. The T2DM additionally showed significantly higher total cholesterol and Homeostatic model assessment (HOMA) compared with control. Serum levels of catecholamine, choactase, nerve growth factor and cardiac nerve density confirms development of DCAN. High single dose of STZ and HFD with multiple low doses of STZ may be recommended for DCAN study in T1DM and T2DM rat model respectively.

The amelioration of T2DM rat femoral bone achieved by anti-osteoporosis of caprine CSN1S2 protein through bone morphogenetic protein signaling pathway.

We investigated the potential anti-glycation and anti-osteoporosis properties of Caprine milk CSN1S2 protein on the serum AGEs and sRAGE level, osteogenic factors expressions, femoral bone mesostructure, histomorphometry, and hydroxyapatite crystals changes in T2DM rats. Varying doses of Caprine milk CSN1S2 protein (0, 375, 750, and 1500 mg/kg BW) were used to treat the control and T2DM rats. We measured AGEs and sRAGE level; RUNX2, OSX, BMP2, and Caspase-3 expressions in rats using ELISA and immunohistochemistry, respectively. The mesostructure and histomorphometry of femoral bone was analyzed using SEM Microscope and BoneJ software, then hydroxyapatite crystal size was determined using SEM-XRD. T2DM rats showed a high level of AGEs and a low level of sRAGE, the RUNX2, OSX, and BMP2 expression was down regulated, BV, BV.TV, Tb.Th, Tb.Sp, increased and SMI levels declined, respectively. Vice versa, after administration of the CSN1S2 protein to T2DM rats, improvement in all levels of molecular and cellular markers was achieved. In the CSN1S2 highest dose, AGEs level declined and sRAGE level elevated in T2DM rats. The 375 and 750 mg/kgBW of CSN1S2 protein was able to upregulate the RUNX2, OSX, and BMP2 expression in T2DM rats, thus improving the normalization of osteoclasts and osteoblasts number. The whole dose of CSN1S2 triggered the thickening of trabecular bone wall, granule formation, and normalized the trabecular thickness (Tb.Th) parameter of T2DM rats. The hydroxyapatite crystal size was increased in the highest dose of CSN1S2-treated T2DM rats. This study indicated that CSN1S2 protein had a protective effect against osteoporosis in the T2DM rat bones by means of glycation pathway inhibition, bone histomorphometry and mesostructure improvement via bone morphometric protein signaling.