Multiple Low Doses Of Streptozotocin Research Articles

Antioxidant Potency of Okra (Abelmoschus esculentus Moench) Pods Extract on SOD Level and Tissue Glucose Tolerance in Diabetic Mice

The purpose of this study was to determine the effect of various fractions of okra pods extract (VOPE) on superoxide dismutase (SOD) level and tissue glucose tolerance in the streptozotocin (STZ) induced diabetic mice. This study was using 35 male Balb/c strain mice. The samples were divided into seven groups, namely KN (normal control), KD (diabetic control), KA (acarbose control), EK (crude extract), NP (non-polar), SP (semipolar), and EP (polar). VOPE and acarbose suspension were administered orally for 14 days in the STZ-induced diabetic mice. Intraperitoneal injection of multiple low-dose STZ (30mg/kg body weight) induced diabetes daily for five consecutive days. On the 15th day, the mice were terminated. Data of SOD level and tissue glucose tolerance were analyzed by ANOVA, at α = 0.05. This study concluded that the administration of VOPE could increase SOD level and tissue glucose tolerance in the STZ-induced diabetic mice. In conclusion, VOPE is a promising antioxidant agent due to its antihyperglycemic properties.

The Influence of Ethanolic Root Extracts of Ruellia tuberosa L. on Pancreatic Protease Activity and MDA Level of Rats (Rattus norvegicus) Induced by MLD-STZ

Diabetes mellitus (DM) is a metabolism disorder, indicated by increasing of blood glucose levels, due to lack of insulin secretion or insulin resistance. This study aimed to determine the influence of ethanolic root extracts of Ruellia tuberosa L on protease activity and MDA levels on pancreatic diabetic rats. This experiment used 20 rats in randomized design, that were divided into five groups: negative control, positive control, and three groups of treatments, with doses of 250, 375, and 500 mg/kg bw, respectively for 21 days. The positive control group and therapeutic group were induced by MLD-STZ (multiple-low dose streptozotocin) at dose of 20 mg/kg bw for 5 consecutive days, in order to induce diabetic states. Protease activity and MDA levels were determined by spectrophotometric method. The results showed that ethanolic root extracts of Ruellia tuberosa L. at dose of 250 mg/kg bw was the best dose to decrease significantly protease activity and MDA level in the pancreas of diabetic rats with the percentages of 52.11% and 50.54%, respectively.

IL-33 Prevents MLD-STZ Induction of Diabetes and Attenuate Insulitis in Prediabetic NOD Mice.

Type 1 diabetes is an autoimmune disease caused by the immune-mediated destruction of pancreatic β-cells. Prevention of type 1 diabetes requires early intervention in the autoimmune process against beta-cells of the pancreatic islets of Langerhans, which is believed to result from disordered immunoregulation. CD4+Foxp3+ regulatory T cells (Tregs) participate as one of the most important cell types in limiting the autoimmune process. The aim of this study was to investigate the effect of exogenous IL-33 in multiple low dose streptozotocin (MLD-STZ) induced diabetes and to delineate its role in the induction of protective Tregs in an autoimmune attack. C57BL/6 mice were treated i. p. with five doses of 40 mg/kg STZ and 0.4 μg rIL-33 four times, starting from day 0, 6, or 12 every second day from the day of disease induction. 16 weeks old NOD mice were treated with 6 injections of 0.4 μg/mouse IL-33 (every second day). Glycemia and glycosuria were measured and histological parameters in pancreatic islets were evaluated at the end of experiments. Cellular make up of the pancreatic lymph nodes and islets were evaluated by flow cytometry. IL-33 given simultaneously with the application of STZ completely prevented the development of hyperglycemia, glycosuria and profoundly attenuated mononuclear cell infiltration. IL-33 treatment was accompanied by higher number of IL-13 and IL-5 producing CD4+ T cells and increased presence of ST2+Foxp3+ regulatory T cells in pancreatic lymph nodes and islets. Elimination of Tregs abrogated protective effect of IL-33. We provide evidence that exogenous IL-33 completely prevents the development of T cell mediated inflammation in pancreatic islets and consecutive development of diabetes in C57BL/6 mice by facilitating the induction Treg cells. To extend this finding for possible relevance in spontaneous diabetes, we showed that IL-33 attenuate insulitis in prediabetic NOD mice.

Syntaxin 4 Expression in Pancreatic β-Cells Promotes Islet Function and Protects Functional β-Cell Mass.

Syntaxin 4 (Stx4) enrichment in human and mouse islet grafts improves the success of transplants in reversing streptozotocin (STZ)-induced diabetes in mice, although the underlying molecular mechanisms remain elusive. Toward a further understanding of this, human islets and inducible transgenic mice that selectively overexpress Stx4 in islet β-cells (βTG-Stx4) were challenged with proinflammatory stressors in vitro and in vivo. Remarkably, βTG-Stx4 mice resisted the loss of β-cell mass and the glucose intolerance that multiple low doses of STZ induce. Under standard conditions, glucose tolerance was enhanced and mice maintained normal fasting glycemia and insulinemia. Conversely, Stx4 heterozygous knockout mice succumbed rapidly to STZ-induced glucose intolerance compared with their wild-type littermates. Human islet β-cells overexpressing Stx4 exhibited enhanced insulin secretory capability; resilience against proinflammatory cytokine–induced apoptosis; and reduced expression of the CXCL9, CXCL10, and CXCL11 genes coordinate with decreased activation/nuclear localization of nuclear factor-κB. Finding ways to boost Stx4 expression presents a novel potential therapeutic avenue for promoting islet function and preserving β-cell mass.

Effect of 4-Phenylbutyric Acid and Tauroursodeoxycholic Acid on Magnesium and Calcium Metabolism in Streptozocin-Induced Type 1 Diabetic Mice

Recent evidence has identified a role of micronutrients, such as magnesium (Mg2+) and calcium (Ca2+), in glycemic control. 4-Phenylbutyric acid (PBA) and tauroursodeoxycholic acid (TUDCA) are molecular chaperones that can improve protein folding and alleviate endoplasmic reticulum (ER) stress. Increasingly, research is focusing on the association between molecular chaperones and micronutrients. This study established and characterized a mouse model of type 1 diabetes (T1D) and investigated the effect of PBA and TUDCA on Mg2+ and Ca2+ metabolism in these mice. T1D was established in Friend virus B-type mice using multiple low doses of streptozotocin. Mice were administered chaperones. Mg2+and Ca2+ levels in tissues and serum were detected using acid digestion and ICP-MS. At 2 weeks and 2 months after chaperone administration was initiated, Mg2+ levels in the heart, liver, kidney, and serum and Ca2+ levels in spleen and serum of T1D mice were significantly decreased compared with controls; Ca2+ levels in the kidney and muscle of T1D mice were significantly increased; Mg2+ and Ca2+ levels in the heart, liver, kidney, muscle, spleen, and serum were positively correlated in control and T1D mice; and PBA restored renal Mg2+ levels to normal values and TUDCA restored hepatic, renal, and serum Mg2+ levels and renal and serum Ca2+ levels to normal values in T1D mice. PBA restored muscular Ca2+ levels to normal values in T1D mice at 2 months after chaperone or vehicle administration was initiated. Further research is required to investigate the underlying mechanisms by which chaperones regulate micronutrients in diabetes.

Nanoceria Suppresses Multiple Low Doses of Streptozotocin-Induced Type 1 Diabetes by Inhibition of Nrf2/NF-κB Pathway and Reduction of Apoptosis

The present study was designed to probe the antidiabetic effects of nanoceria (NC) in Type 1 diabetes (T1DM). NC was characterized using scanning electron microscopy, Fourier transform IR, powder x-ray diffraction and zeta sizer. Multiple low doses of streptozotocin (40 mg/kg, intraperitoneally, five consecutive days) induced Type 1 diabetic Swiss mice were treated with NC at two doses (0.2 and 2 mg/kg, ip.). NC treatment significantly reduced glucose levels and diabetogenesis to 50% (4/8 animals) at 0.2 mg/kg and 37.5% (3/8 animals) at 2.0 mg/kg doses. Cytokines (IL-6 and TNF-α; p < 0.048 at 2 mg/kg) and p65-NF-κB expression were diminished by NC treatment whereas the Nrf2 expression was enhanced by NC intervention indicating the role of modulation of NF-κB/Nrf2 pathway. NC exhibited promising superoxide dismutase 1 mimetic and anti-apoptotic activity. Considered together, our data establishes the antidiabetic potential of NC which may become a novel strategy to combat T1DM in the near future.

Potential synergistic effects of quercetin with other phytoconstituents of Costus pictus (insulin plant) extract in the control of hyperglycemia and prevention of NSAID-induced gastroenteropathy in diabetic rats

There is a need of multifactorial management to treat T2DM. Till date, no clinically simulated animal model and therapy for NSAID-induced gastroenteropathic damage (NSAID-iGD) in T2DM patients. T2DM was developed using high-fat diet plus multiple low doses of streptozotocin (30 mg/kg, IP). Rats treated with ethanolic extract of Insulin plant (EIP; 125, 250 and 500 mg/kg, PO; b.i.d.)/Quercetin (QCT; 50 mg/kg)/vehicle for total 10 days. Diclofenac sodium (DCF; 7.5 mg/kg, PO, b.i.d.) administered for final five days of EIP/vehicle administration. Rats fasted after last dose on the 9th day; water was provided ad libitum. 12 h after the last dose on 10th day, GI tracts assessed for haemorrhagic damage, XO activity, LPO, intestinal permeability, luminal pH alterations along with haematological, biochemical and histological parameters. The evidence suggested that DCF administration caused significant gastroenteropathic damage. In presence of T2DM, NSAID-iGD significantly exacerbated. Whereas, QCT/EIP treatment significantly attenuated T2DM dependent exacerbation of NSAID-iGD, and also efficiently managed T2DM in a dose-dependent manner. Low amount of QCT in EIP(190.96 ± 7.5 ng/mg) than its effective dose(50 mg/kg) indicates that EIP's other phytoconstituents (e.g. Kaempferol, Ascorbic acid, Lupeol, Diosgenin, β-sitosterol, Stigmasterol, β-amyrin, etc.) giving synergistic actions. Costus pictus/QCT has potential to be promising candidate to treat patient with T2DM and NSAID-gastroenteropathy in T2DM.

Withaferin-A attenuates multiple low doses of Streptozotocin (MLD-STZ) induced type 1 diabetes

Type 1 diabetes mellitus (T1DM) is one of the major metabolic disorders with life-long dependence on insulin. The present study was designed to evaluate the antioxidant and anti-diabetic potential of Withaferin A (WA), the active constituent of Withania somnifera in multiple low doses of Streptozotocin (MLD-STZ) induced T1DM. STZ (40 mg/Kg) was administered intraperitoneally (i.p.) for 5 consecutive days to male Swiss albino mice to induce T1DM. Mice were concurrently treated with WA (2 & 10 mg/Kg). Blood glucose levels, intraperitoneal glucose tolerance test, oxidative stress parameters were estimated biochemically (MDA, GSH) and immunohistochemically (Nrf2, NFκB). In addition, inflammatory cytokines, and insulin levels were quantified by ELISA method. Apoptosis was assessed by immunohistochemical staining for cleaved-caspase-3 and TUNEL assay. WA treatment significantly reduced the blood glucose levels and improved glucose clearance. Strikingly, we observed a significant reduction in the incidence of diabetes upon WA treatment and only 2 out of 8 (2/8 = 25%) animals were diabetic. WA ameliorated the MLD-STZ induced oxidative and nitrosative stress. Furthermore, WA exhibited promising anti-inflammatory effect as evident from reduction in the levels of IL-6 (p < 0.05) and TNF-α (p < 0.05) compared to diabetic mice. In addition, insulitis scoring and IHC for Nrf2 and NFκB indicated promising anti-diabetic effect. WA reduced MLD-STZ induced DNA fragmentation and apoptosis, further supporting the observed protective effect. We, to the best of our knowledge, report for the first time that WA can effectively combat MLD-STZ induced T1DM via modulation of Nrf2/NFκB signaling and holds substantial potential for therapy of T1DM.

Glycosphingolipids promote pro-atherogenic pathways in the pathogenesis of hyperglycemia-induced accelerated atherosclerosis.

Three out of four people with diabetes will die of cardiovascular disease. However, the molecular mechanisms by which hyperglycemia promotes atherosclerosis, the major underlying cause of cardiovascular disease, are not clear. Three distinct models of hyperglycemia-associated accelerated atherosclerosis were used to identify commonly altered metabolites and pathways associated with the disease. Normoglycemic apolipoprotein-E-deficient mice served as atherosclerotic control. Hyperglycemia was induced by multiple low-dose streptozotocin injections, or by introducing a point-mutation in one copy of insulin-2 gene. Glucosamine-supplemented mice, which experience accelerated atherosclerosis to a similar extent as hyperglycemia-induced models without alterations in glucose or insulin levels, were also included in the analysis. Untargeted plasma metabolomics were used to investigate hyperglycemia-associated accelerated atherosclerosis in three disease models. The effect of specific significantly altered metabolites on pro-atherogenic processes was investigated in cultured human vascular cells. Hyperglycemic and glucosamine-supplemented mice showed distinct metabolomic profiles compared to controls. Meta-analysis of three disease models revealed 62 similarly altered metabolite features (FDR-adjusted p < 0.05). Identification of shared metabolites revealed alterations in glycerophospholipid and sphingolipid metabolism, and pro-atherogenic processes including inflammation and oxidative stress. Post-multivariate and pathway analyses indicated that the glycosphingolipid pathway is strongly associated with hyperglycemia-induced accelerated atherosclerosis in these atherogenic mouse models. Glycosphingolipids induced oxidative stress and inflammation in cultured human vascular cells. Glycosphingolipids are strongly associated with hyperglycemia-induced accelerated atherosclerosis in three distinct models. They also promote pro-atherogenic processes in cultured human cells. These results suggest glycosphingolipid pathway may be a potential therapeutic target to block or slow atherogenesis in diabetic patients.

Tofogliflozin and Pemafibrate Combination Therapy Prevents Nonalcoholic Steatohepatitis (NASH) Development of Novel Metabolic Syndrome–Related NASH Model Mice

Nonalcoholic steatohepatitis (NASH) is closely associated with metabolic syndrome and widely defined by the presence of steatosis with inflammation and progressive fibrosis that causes to the liver transplantation and hepatocellular carcinoma (HCC). Here, we developed a novel NASH/HCC model represented metabolic syndrome like phenotypes (obesity, hyperglycemia, and glucose intolerance), and evaluated the therapeutic potential of Tofogliflozin and Pemafibrate combination therapy on NASH development. Male C57BL/6J mice were injected multiple low dose STZ (40 mg/kg) for 5 days and fed high-fat diet (HFD). These mice revealed hyperglycemia and higher body weight gain, and glucose intolerance compared with wild type mice. In addition, novel NASH/HCC model shows sequential histological changes from fatty liver to NASH (20 weeks), and all mice developed hepatocellular carcinoma after 52-week HFD feeding. Using our NASH/HCC model, we conducted the prevention study of Tofogliflozin and Pemafibrate (mono-therapy or combination therapy) against NASH development. The combination therapy has significantly prevented body weight gain from 3 weeks after treatment and this effect was persisted throughout study. While Tofogliflozin or Pemafibrate treatment markedly decreased blood glucose and triglyceride levels, the combination therapy showed the synergistic effect compared to mono-therapy groups. Histological analysis revealed Tofogliflozin or combination therapy group markedly suppressed ER-TR7 (fibroblast) and ballooning degeneration. Furthermore, these treatment groups significantly improved NAS score (NAFLD activity score). These results suggested that Tofogliflozin/Pemafibrate combination therapy would be a beneficial for NASH/NAFLD treatment. Disclosure Y. Sasaki: Employee; Self; Kowa Company, Ltd. M. Asahiyama: Employee; Self; Kowa Company, Ltd.. Employee; Spouse/Partner; Kowa Company, Ltd.. W. Kamiya: None. J. Sakai: None. T. Kodama: Consultant; Self; Kowa Company, Ltd. T. Tanaka: Research Support; Self; Kowa Company, Ltd..

Comparison of different doses streptozotocin-induced mouse models for the research of type 1 diabetic keratopathy

Objective To compare the rates and pathological features of diabetic keratopathy in mice induced by single high dose or multiple low dose streptozotocin (STZ) injections. Methods Eighty male C57BL/6 mice (6-8 weeks old) were randomly divided into 4 groups with each group contain 20 mice: normal control group, multiple low dose 1 month group and multiple low dose 3 months group (injected with 60 mg/kg STZ for 5 consecutive times), single high dose 1 month group (injected with 150 mg/kg STZ). The survival rate, model success rate, body weight, glycosylated hemoglobin (HbA1c) content were compared among different modeling group.The percentages of residual epithelial defect area were examined by fluorescein sodium staining after removal of central corneal epithelium.The expression of p-Akt, Sirt1 and Ki67 were evaluated by immunofluorescent staining.The corneal sensitivity were compared among different groups before corneal epithelial curettage, 3, 7, 10 and 14 days after corneal epithelial curettage.The corneal subbasal nerve density at 14 days after corneal epithelial curettage were compared among different groups.This study complied with the declaration of ARVO Results The success rate of diabetic modeling in multiple low dose 1 month group, multiple low dose 3 months group and single high dose 1 month group was 90%, 80% and 70%, respectively.The HbA1c levels in the diabetic modeling groups were significantly higher than that in the normal control group (all at P 0.05). However, the corneal sensitivity and corneal nerve density were dramatically decreased in the multiple low dose 3 months group and single high dose 1 month group before and 14 days after the corneal epithelial removal, and there were significant differences compared with normal control group (all at P<0.05). Conclusions The injection of 60 mg/kg STZ can not induce the features of diabetic keratopathy in mice within 1 month.However, the mice of both 1 month after 150 mg/kg STZ injection and 3 months after 60 mg/kg STZ injection appear the typical epithelial and nerve features of diabetic keratopathy, therefore can be the ideal animal models for research. Key words: Streptozotocin; C57BL/6 mice; Diabetes mellitus; Corneal keratopathy

Effects of Ruellia tuberosa L. Root Extract on the Pancreatics of Diabetic Rat

Diabetes mellitus type 1 (DMT1) is an insulin deficiency disorder caused by pancreatic damage characterized by chronic hyperglycemia. Animal modeling of DMT1 using inducting Multiple Low Dose Streptozotocin (MLD-STZ) elevat levels of free radicals and inflammation in pancreatic organs which then damage β cells. Ruellia tuberosa L. is a plant containing triterpenoids from the class of antioxidant and anti-inflammatory flavonoids. This study used three groups of rat: negative control group, positive control and 240 mg/kg bw therapy group. Study are based on changes MDA levels, blood glucose and histologic of pancreas. MDA level determining with TBA method, glucose levels using EasyTouch GCU and histopalogy of pancreas using Hematoxylin-Eosin staining. Statistical analysis of MDA and blood glucose (α = 0.05) showed significant decreases in MDA and blood glucose levels. Based on the results of histologic observations of pancreas descriptively showed a decreas in cell damage on Langerhans island. Thus, the provision of root extract of Ruellia tuberosa L. can inhibit the damage in rat DMT1 model so that it can be used as therapy of diabetes mellitus.

Abstract 215: Glycosphingolipids Contribute to Pro-atherogenic Pathways in the Pathogenesis of Hyperglycemia-induced Atherosclerosis

Three out of every four people with diabetes will die of cardiovascular disease. However, the molecular mechanisms by which diabetes promotes atherosclerosis are not clear. In this study, comprehensive metabolomic techniques were used to investigate the molecular mechanisms by which hyperglycemia promotes accelerated atherogenesis in three distinct disease mouse models. Normoglycemic apolipoprotein E-deficient mice served as the atherosclerotic control. Hyperglycemia was induced by multiple low-dose streptozotocin injections or by introducing a point mutation in one copy of the insulin-2 gene. Glucosamine-supplemented mice, which experience accelerated atherosclerosis to a similar extent as the hyperglycemia-induced models, without alterations in the levels of glucose or insulin, were also included in the analysis. Mice with accelerated atherosclerosis showed distinct metabolomic profiles compared to the controls. We detected 6369 metabolite features in the plasma of each mouse. Second-order analysis of pair comparisons between each disease model and the control resulted in 62 commonly altered features (p&lt;0.05). Identification of shared metabolites revealed alterations in glycerophospholipid and sphingolipid metabolisms, and pro-atherogenic processes including inflammation and oxidative stress. Post-multivariate and pathway analyses indicated glycosphingolipid metabolism is the most significantly altered pathway. Glycosphingolipid metabolites induced oxidative stress and inflammation in cultured human vascular cells including macrophages, endothelial and smooth muscle cells. Treatment with a known antioxidant, α-tocopherol, reduced oxidative stress and inflammation induced by glycosphingolipids. Our findings suggest that the glycosphingolipid pathway contributes to pro-atherogenic pathways in the pathogenesis of hyperglycemia-induced atherosclerosis, making it a potential therapeutic target to block or slow atherogenesis in diabetic patients.

Human urine-derived stem cells play a novel role in the treatment of STZ-induced diabetic mice.

Human urine-derived stem cells (hUSCs) are a potential stem cell source for cell therapy. However, the effect of hUSCs on glucose metabolism regulation in type 1 diabetes was not clear. Therefore, the aim of the study was to evaluate whether hUSCs have protective effect on streptozotocin (STZ)-induced diabetes. hUSCs were extracted and cultivated with a special culture medium. Flow cytometry analysis was applied to detect cell surface markers. BALB/c male nude mice were either injected with high-dose STZ (HD-STZ) or multiple low-dose STZ (MLD-STZ). Serum and pancreatic insulin were measured, islet morphology and its vascularization were investigated. hUSCs highly expressed CD29, CD73, CD90 and CD146, and could differentiate into, at least, bone and fat in vitro. Transplantation of hUSCs into HD-STZ treated mice prolonged the median survival time and improved their blood glucose, and into those with MLD-STZ improved the glucose tolerance, islet morphology and increased the serum and pancreas insulin content. Furthermore, CD31 expression increased significantly in islets of BALB/c nude mice treated with hUSCs compared to those of un-transplanted MLD-STZ mice. hUSCs could improve the median survival time and glucose homeostasis in STZ-treated mice through promoting islet vascular regeneration and pancreatic beta-cell survival.

Doc2b Protects β-Cells Against Inflammatory Damage and Enhances Function.

Loss of functional β-cell mass is an early feature of type 1 diabetes. To release insulin, β-cells require soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes, as well as SNARE complex regulatory proteins like double C2 domain-containing protein β (Doc2b). We hypothesized that Doc2b deficiency or overabundance may confer susceptibility or protection, respectively, to the functional β-cell mass. Indeed, Doc2b+/- knockout mice show an unusually severe response to multiple-low-dose streptozotocin (MLD-STZ), resulting in more apoptotic β-cells and a smaller β-cell mass. In addition, inducible β-cell-specific Doc2b-overexpressing transgenic (βDoc2b-dTg) mice show improved glucose tolerance and resist MLD-STZ-induced disruption of glucose tolerance, fasting hyperglycemia, β-cell apoptosis, and loss of β-cell mass. Mechanistically, Doc2b enrichment enhances glucose-stimulated insulin secretion (GSIS) and SNARE activation and prevents the appearance of apoptotic markers in response to cytokine stress and thapsigargin. Furthermore, expression of a peptide containing the Doc2b tandem C2A and C2B domains is sufficient to confer the beneficial effects of Doc2b enrichment on GSIS, SNARE activation, and apoptosis. These studies demonstrate that Doc2b enrichment in the β-cell protects against diabetogenic and proapoptotic stress. Furthermore, they identify a Doc2b peptide that confers the beneficial effects of Doc2b and may be a therapeutic candidate for protecting functional β-cell mass.

Testicular histopathology and phosphorylated protein changes in mice with diabetes induced by multiple-low doses of streptozotocin: An experimental study

The streptozotocin (STZ)-induced diabetic model is widely used to evaluate the adverse effects of diabetes mellitus (DM) on spermatogenesis and testicular steroidogenesis. However, the actual mechanism of sub/infertility in DM males needs to be elucidated. To conduct a detailed examination of the testicular histopathology, sperm acrosome reaction (AR) status, and tyrosine-phosphorylated protein expression in the testis of male mice induced with STZ. Ten ICR mice were divided into two groups (n=5/each): control and diabetes induced by multiple low doses of streptozotocin (MLD-STZ). The control mice were intraperitoneally injected with citrate buffer, whereas MLD-STZ mice were injected with STZ at 40 mg/kg body weight for five consecutive days. At the end of the experiment (day 40), reproductive parameters, AR status, and the histopathology of the testis and epididymis were evaluated. The expression of testicular tyrosine phosphorylated proteins was examined. Blood glucose levels, AR percentages, and sperm abnormality of STZ group were significantly higher (p=0.003, 0.001, 0.000), while sperm concentration was significantly lower (p=0.001) compared to control. Histopathology of the seminiferous tubule was classified into 7 types. Additionally, abundant round cells were found in the epididymal lumen of the MLD-STZ mice. Moreover, the intensities of testicular phosphorylated proteins (170, 70, 36, 30, and 25 kDas) were markedly higher and a 120 kDa protein band was noticeably lower in the MLD-STZ mice. MLD-STZ-induced DM causes many testicular histopathologies, precocious sperm AR, and increased expression of testicular phosphorylated proteins. These findings may clarify some mechanisms of sub/infertility in DM males.

Investigation of insulin resistance in the popularly used four rat models of type-2 diabetes

Animal models are widely used to develop drugs for treating diabetes mellitus (DM). Insulin resistance (IR) is one of the main problems in type-2 DM (T2DM). Streptozotocin (STZ) is used to damage pancreatic cells for induction of DM. Many rat models were applied in research as T2DM. However, the degree of IR in each model is unknown. In the present study, IR and insulin signaling were compared in four models of type 2 diabetes: rats fed a fructose-rich chow for 8 weeks, rats feed high-fat chow for 4 weeks followed by injection with streptozotocin (35 mg/kg, i.p.), rats injected with a single low dose streptozotocin (45 mg/kg, i.p.), and rats injected with a single dose of nicotinamide followed by a single high dose of streptozotocin (60 mg/kg, i.p.). Values from these determinations in diabetic rats showing the order that insulin resistance is most marked in rats received fructose-rich chow followed by high-fat diet before STZ injection induced model (HFD/STZ rats), and rats injected with low dose of STZ but it is less marked in rats induced by nicotinamide and STZ. Additionally, insulin secretion was reduced in three rat models except the rats receiving fructose-rich chow. Western blots also showed the same changes in phosphorylation of IRS-1 or Akt using soleus muscle from each model. The obtained data suggest a lack of pronounced IR in the rats with acute diabetes induced by nicotinamide and STZ while IR is markedly identified in rats fed fructose-rich chow. However, the increase of plasma glucose levels in fructose-rich chow-fed rats was not so significant as other groups. Therefore, HFD/STZ rats is an appropriate and stable animal model which is analogous to the human T2DM through a combination of high-fat diet with multiple low-dose STZ injections.

Protection by dimethyl fumarate against diabetic cardiomyopathy in type 1 diabetic mice likely via activation of nuclear factor erythroid-2 related factor 2

Oxidative stress and inflammation play key roles in the development of diabetic cardiomyopathy (DCM). Dimethyl fumarate (DMF), an FDA approved medicine for relapsing multiple sclerosis, has manifested its antioxidant and anti-inflammatory function mostly in the central nervous system. In this study, we investigated whether DMF could attenuate the development of DCM. Type 1 diabetes mouse model was established using multiple low-dose streptozotocin, and the diabetic mice were treated with DMF (10 mg/kg body weight) for 3 months. Cardiac functions were determined using echocardiography. Oxidative stress, pro-inflammatory cytokines and pro-fibrotic markers were determined with commercially available kits, real-time quantitative PCR or western blot techniques. DCM was characterized by diminished cardiac function, accompanied by oxidative stress and enhanced expression of pro-inflammatory cytokines. Diabetic cardiac tissue exhibited marked fibrosis, revealed by extracellular matrix deposition as determined by Sirius red staining of the myocardial tissues. Furthermore, Nrf2 and its downstream effectors were repressed in diabetic myocardium. On the contrary, diabetic animals treated with DMF exhibited blunted oxidative stress, inflammation, fibrosis and this correlated with Nrf2 activation. Our findings suggest that DMF could potentially thwart diabetes-induced myocardial tissue injury, likely via activation of Nrf2 function, providing firm impetus for future repurposing of DMF in the management of DCM.

Inhibition of p53 prevents diabetic cardiomyopathy by preventing early-stage apoptosis and cell senescence, reduced glycolysis, and impaired angiogenesis

Elevated tumor suppressor p53 expression has been associated with heart diseases, including the diabetic heart. However, its precise role in the pathogenesis of diabetic cardiomyopathy (DCM) remains unclear. We hypothesized that the development of DCM is attributed to up-regulated p53-mediated both early cardiac cell death and persistent cell senescence, glycolytic and angiogenetic dysfunctions. The present study investigated the effect of p53 inhibition with its specific inhibitor pifithrin-α (PFT-α) on the pathogenesis of DCM and its associated mechanisms. Type 1 diabetes was induced with multiple low doses of streptozotocin. Both hyperglycemic and age-matched control mice were treated with and without PFT-α five times a week for 2 months and then sacrificed at 3 and 6 months post-diabetes. Treatment with PFT-α significantly prevented the progression of diabetes-induced cardiac remodeling and dysfunction (i.e., DCM). Mechanistically, the inhibition of p53 prevented the cardiac apoptosis during early-stage diabetes (0.5 month), attenuated diabetes-induced cell senescence (3 and 6 months), and improved both glycolytic and angiogenic defects by increasing hypoxia-induced factor (HIF)-1α protein stability and upregulating HIF-1α transcription of specific target genes at 3 and 6 months after diabetes. Therefore, the targeted inhibition of p53 in diabetic individuals may provide a novel approach for the prevention of DCM.

Protective effects of carbonyl iron against multiple low-dose streptozotocin-induced diabetes in rodents.

Particulate adjuvants have shown increasing promise as effective, safe, and durable agents for the stimulation of immunity, or alternatively, the suppression of autoimmunity. Here we examined the potential of the adjuvant carbonyl iron (CI) for the modulation of organ-specific autoimmune disease-type 1 diabetes (T1D). T1D was induced by multiple low doses of streptozotocin (MLDS) that initiates beta cell death and triggers immune cell infiltration into the pancreatic islets. The results of this study indicate that the single in vivo application of CI to MLDS-treated DA rats, CBA/H mice, or C57BL/6 mice successfully counteracted the development of insulitis and hyperglycemia. The protective action was obtained either when CI was applied 7 days before, simultaneously with the first dose of streptozotocin, or 1 day after MLDS treatment. Ex vivo cell analysis of C57BL/6 mice showed that CI treatment reduced the proportion of proinflammatory F4/80+ CD40+ M1 macrophages and activated T lymphocytes in the spleen. Moreover, the treatment down-regulated the number of inflammatory CD4+ IFN-γ+ cells in pancreatic lymph nodes, Peyer's patches, and pancreas-infiltrating mononuclear cells, while simultaneously potentiating proportion of CD4+ IL17+ cells. The regulatory arm of the immune system represented by CD3+ NK1.1+ (NKT) and CD4+ CD25+ FoxP3+ regulatory T cells was potentiated after CI treatment. In vitro analysis showed that CI down-regulated CD40 and CD80 expression on dendritic cells thus probably interfering with their antigen-presenting ability. In conclusion, particulate adjuvant CI seems to suppress the activation of the innate immune response, which further affects the adaptive immune response directed toward pancreatic beta cells.