Low-dose Streptozotocin-induced Diabetes In Mice Research Articles

Dose-dependent progression of multiple low dose streptozotocin-induced diabetes in mice.

This study investigated the effects of different multiple low doses of streptozotocin (STZ), namely 35 and 55 mg/kg, on the onset and progression of diabetes in mice. Both doses are commonly used in research, and while both induced a loss of beta cell mass, they had distinct effects on whole glucose tolerance, beta cell function and gene transcription. Mice treated with 55 mg/kg became rapidly glucose intolerant, whereas those treated with 35 mg/kg had a slower onset and remained glucose tolerant for up to a week before becoming equally glucose intolerant as the 55 mg/kg group. Beta cell mass loss was similar between the two groups, but the 35 mg/kg-treated mice had improved glucose-stimulated insulin secretion in gold-standard hyperglycemic clamp studies. Transcriptomic analysis revealed that the 55 mg/kg dose caused disruptions in nearly five times as many genes as the 35 mg/kg dose in isolated pancreatic islets. Pathways that were downregulated in both doses were more downregulated in the 55 mg/kg-treated mice, whereas pathways that were upregulated in both doses were more upregulated in the 35 mg/kg treated mice. Moreover, we observed a differential downregulation in the 55 mg/kg-treated islets of beta cell characteristic pathways, such as exocytosis or hormone secretion. On the other hand, apoptosis was differentially upregulated in 35 mg/kg-treated islets, suggesting different transcriptional mechanisms in the onset of STZ-induced damage in the islets. This study demonstrates that the two STZ doses induce distinctly mechanistic progressions for the loss of functional beta cell mass.

Abstract 15322: O-glcnac Modification on Runx2 Promotes Vascular Calcification

Vascular calcification is prevalent in patients with diabetes mellitus and well documented in animal models of diabetes. We have previously reported that vascular calcification in diabetes is associated with elevated vascular protein O -linked GlcNAc modification ( O -GlcNAcylation). The present studies sought to determine a causative link of O -GlcNAcylation with diabetic vascular calcification and the underlying molecular mechanisms. Using coronary arteries from diabetic subjects, we identified increased O -GlcNAcylation in the highly calcified lesions compared to those in the surrounding uncalcified areas. By generating a new mouse model with inducible SMC-specific deletion of the O -GlcNAc transferase (OGT), we demonstrated that smooth muscle cell (SMC)-specific OGT deletion significantly inhibited protein O -GlcNAcylation and calcification, and decreased aortic stiffness in the low-dose streptozotocin-induced diabetic mice. Consistent with the observations in human tissues, inhibition of vascular calcification by OGT ablation was associated with down-regulation of Runx2, the essential osteogenic regulator for vascular calcification. OGT deficiency in VSMC further attenuated Runx2-induced VSMC calcification. Immunoprecipitation analysis uncovered a direct O-GlcNAc modification on Runx2, which was abolished in the OGT-deficient VSMC. With a serial of Runx2 truncation mutants and point mutations on putative O -GlcNAcylation sites, we localized the essential Runx2 osteogenic functional domain between amino acids 391 and 432, and identified Runx2 T412 was responsible for Runx2 O -GlcNAcylation and Runx2-induced VSMC calcification. Inhibition of Runx2 O -GlcNAcylation by T412A mutation decreased Runx2 transcription activity and inhibited Runx2 binding with its key co-factors. Taken together, our studies have provided the first genetic proof demonstrating the causative effects of O -GlcNAcylation on vascular calcification in diabetes, and identified a novel mechanism underlying the essential role of O -GlcNAcylation of Runx2 in Runx2 osteogenic activity and Runx2-induced VSMC calcification. These results may shed lights on novel targets that are amenable to drug discovery for diabetic vascular calcification.

Protective effects of acarbose against insulitis in multiple low-dose streptozotocin-induced diabetic mice

AimsThe development of type 1 diabetes is associated with inflammatory lesion of the pancreatic islets, known as insulitis. In this study, we focused on the protective effects of acarbose against insulitis in streptozotocin (STZ)-induced diabetic mice and the underlying mechanisms. Main methodsThe mouse models were established via intraperitoneal injection of multiple low-dose STZ. Blood glucose level and body weight were measured. The severity of insulitis and inflammatory parameters in pancreatic tissues were evaluated. Insulin levels in pancreas and serum were also assessed. In vitro, MIN6 β cells were exposed to pro-inflammatory cytokines to assess the protective effects of acarbose. Cell function and apoptosis were evaluated. Key findingsWe found that acarbose administration by gavage reduced the severity of insulitis and improved insulin levels in the experimental diabetic mice. ELISA revealed decreased levels of the inflammatory response markers IL-1β and TNF-α in mouse pancreatic tissues following acarbose treatment. In vitro, acarbose increased cell viability, decreased cell apoptosis, and improved GSIS in MIN6 β cells exposed to pro-inflammatory cytokines. In addition, caspase-3 level and p-p53/p53 ratio in β cells were reduced by acarbose treatment. SignificanceTaken together, these results revealed a novel function of acarbose in attenuating insulitis. The protective effects of acarbose elicited in vitro and in vivo were shown to be mediated, at least in part, through its anti-inflammatory action.

Ethanolic extracts of Pluchea indica (L.) leaf pretreatment attenuates cytokine-induced β-cell apoptosis in multiple low-dose streptozotocin-induced diabetic mice.

Loss of β-cell mass and function is a fundamental feature of pathogenesis for type 1 and type 2 diabetes. Increasing evidence indicates that apoptosis is one of the main mechanisms of β-cell death in both types. Ethanolic extracts of Pluchea indica leaf (PILE) have been reported to possess blood glucose lowering actions in vivo. Nevertheless, further study is required to determine the underlying mechanisms. In this report, we have investigated the preventive effects of PILE on multiple low doses of streptozotocin (MLDS)-induced β-cell apoptosis. Mice were pre-treated with PILE at 50 mg/kg (PILE 50) or 100 mg/kg (PILE 100) for 2 weeks before streptozotocin (STZ) stimulation, and the treatment continued for 4 or 8 weeks. Results revealed that PILE 100 mice exhibited improved blood biochemistry, maintained a higher body weight, had decreased hyperglycemia, and restored islet architectures compared to non-treated STZ mice. Significantly, PILE 100 decreased levels of inflammatory response markers interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interlukin1-β (IL-1β), concomitant with the inhibition of caspase-3, caspase-8, capsepase-9, phosphorylation of signal transducer and activator of transcription 1 (pSTAT1), nuclear factor-κBp65 (NF-κBp65), and inducible nitric oxide synthase (iNOS). Additionally, survival and proliferative ability of β-cells was mediated by up-regulated Bcl-2 and Ki67, respectively. These results provide strong evidence that pretreatment with PILE 100 effectively attenuated STZ-induced diabetes-related symptoms and these effects could be associated with the inhibition of cytokine-induced β-cell apoptosis.

γ-Oryzanol protects pancreatic β-cells against endoplasmic reticulum stress in male mice.

Endoplasmic reticulum (ER) stress is profoundly involved in dysfunction of β-cells under high-fat diet and hyperglycemia. Our recent study in mice showed that γ-oryzanol, a unique component of brown rice, acts as a chemical chaperone in the hypothalamus and improves feeding behavior and diet-induced dysmetabolism. However, the entire mechanism whereby γ-oryzanol improves glucose metabolism throughout the body still remains unclear. In this context, we tested whether γ-oryzanol reduces ER stress and improves function and survival of pancreatic β-cells using murine β-cell line MIN6. In MIN6 cells with augmented ER stress by tunicamycin, γ-oryzanol decreased exaggerated expression of ER stress-related genes and phosphorylation of eukaryotic initiation factor-2α, resulting in restoration of glucose-stimulated insulin secretion and prevention of apoptosis. In islets from high-fat diet-fed diabetic mice, oral administration of γ-oryzanol improved glucose-stimulated insulin secretion on following reduction of exaggerated ER stress and apoptosis. Furthermore, we examined the impact of γ-oryzanol on low-dose streptozotocin-induced diabetic mice, where exaggerated ER stress and resultant apoptosis in β-cells were observed. Also in this model, γ-oryzanol attenuated mRNA level of genes involved in ER stress and apoptotic signaling in islets, leading to amelioration of glucose dysmetabolism. Taken together, our findings demonstrate that γ-oryzanol directly ameliorates ER stress-induced β-cell dysfunction and subsequent apoptosis, highlighting usefulness of γ-oryzanol for the treatment of diabetes mellitus.

Activation of AKT by O-Linked N-Acetylglucosamine Induces Vascular Calcification in Diabetes Mellitus

Vascular calcification is a serious cardiovascular complication that contributes to the increased morbidity and mortality of patients with diabetes mellitus. Hyperglycemia, a hallmark of diabetes mellitus, is associated with increased vascular calcification and increased modification of proteins by O-linked N-acetylglucosamine (O-GlcNAcylation). We sought to determine the role of protein O-GlcNAcylation in regulating vascular calcification and the underlying mechanisms. Low-dose streptozotocin-induced diabetic mice exhibited increased aortic O-GlcNAcylation and vascular calcification, which was also associated with impaired aortic compliance in mice. Elevation of O-GlcNAcylation by administration of Thiamet-G, a potent inhibitor for O-GlcNAcase that removes O-GlcNAcylation, further accelerated vascular calcification and worsened aortic compliance of diabetic mice in vivo. Increased O-GlcNAcylation, either by Thiamet-G or O-GlcNAcase knockdown, promoted calcification of primary mouse vascular smooth muscle cells. Increased O-GlcNAcylation in diabetic arteries or in the O-GlcNAcase knockdown vascular smooth muscle cell upregulated expression of the osteogenic transcription factor Runx2 and enhanced activation of AKT. O-GlcNAcylation of AKT at two new sites, T430 and T479, promoted AKT phosphorylation, which in turn enhanced vascular smooth muscle cell calcification. Site-directed mutation of AKT at T430 and T479 decreased O-GlcNAcylation, inhibited phosphorylation of AKT at S473 and binding of mammalian target of rapamycin complex 2 to AKT, and subsequently blocked Runx2 transactivity and vascular smooth muscle cell calcification. O-GlcNAcylation of AKT at 2 new sites enhanced AKT phosphorylation and activation, thus promoting vascular calcification. Our studies have identified a novel causative effect of O-GlcNAcylation in regulating vascular calcification in diabetes mellitus and uncovered a key molecular mechanism underlying O-GlcNAcylation-mediated activation of AKT.

Thalidomide attenuates multiple low-dose streptozotocin-induced diabetes in mice by inhibition of proinflammatory cytokines

Thalidomide is an immunomodulatory and anti-inflammatory agent and is used in autoimmune disorders. It has been shown that thalidomide inhibits proinflammatory cytokines production. The purpose of this study was to investigate the effect of thalidomide on the prevention of autoimmune diabetes in mice. Diabetes was induced by multiple low-dose of streptozotocin (MLDS) injection. Mice were treated with thalidomide (300mg/kg/day orally) for 21days. Plasma levels of glucose, insulin and nitrate/nitrite as well as pancreatic cytokine levels were measured. Pathological examinations of the pancreas revealed that thalidomide reduced the islet inflammation (insulitis) and destruction of beta cells. Thalidomide treatment prevented hyperglycemia and preserved pancreatic insulin secretion in the diabetic mice. Thalidomide treatment also significantly decreased plasma levels of nitric oxide and pancreatic proinflammatory cytokines [tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-12, IL-17 and interferon (IFN)-γ)] while increased anti-inflammatory cytokine IL-10. In conclusion, these findings indicate that thalidomide may have a protective effect against the autoimmune destruction of the pancreatic beta-cells during the development of MLDS-induced type 1 diabetes in mice.

Effects of Brx-019 (Acetic Acid 3,6a,9-triacetoxy-6,6a,7,11b-tetrahydro-indeno [2,1-c]chromen-10-yl Ester), a Brazilin Derivative, on T Cell-mediated Immune Responses in Multiple Low Dose Streptozo-tocin-induced Diabetic C57BL/6 Male Mice

Brx-019 (acetic acid 3,6a,9-triacetoxy-6, 6a,7,11b-tetrahydro-indeno [2,1-c] chromen-10-yl ester) was derived from brazilin (CAS 474-07-7) during a trial designed to search for immunomodulators with lower toxicity and more effective immunomodulating activities than brazilin. Brx-019 was selected as a potential immunomodulator based on its effects on Concanavalin A (Con A)-induced proliferation of splenocytes and the 3-[14,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Intraperitoneally administered Brx-019 significantly improved delayed type hypersensitivity and increased immunoglobulin M (IgM) plaque forming cells (PFCs) in multiple low dose streptozotocin-induced diabetic mice (MLDS-diabetic mice). This finding suggests that Brx-019 may increase suppressed humoral and cell-mediated immunity in type 1 diabetes. Brx-019 also significantly increased Con A- or alloantigen-induced proliferation of splenocytes, Con A-induced interleukin 2 (IL-2) production from splenocytes, and IL-2-induced proliferation of Con A-activated splenocytes in MLDS-diabetic mice. These results suggest that Brx-019 might improve immunity in diabetic mice by increasing IL-2 production in splenocytes and responsiveness of splenocytes to IL-2, which were suppressed in MLDS-diabetes.

Increased galectin-1 expression in muscle of Astragalus polysaccharide-treated Type 1 diabetic mice

In order to investigate the immunopharmacological function of Astragalus polysaccharide (APS) in Type 1 diabetes mellitus (T1DM), multiple low-dose streptozotocin-induced diabetic mice and normal mice were administered either APS or saline intraperitoneally once daily. The changes in galectin-1 expression in different organs of the mice were detected by ELISA, Real-time fluorescence quantitative RT-PCR and Western blot. The percentages of apoptotic CD8(+) T cells from spleens of APS-treated diabetic mice were measured by flow cytometry. We found that the expression of galectin-1 was increased in serum of APS-treated diabetic mice compared to the non-treated diabetic mice (*p<0.05). Increased galectin-1 was mainly expressed in the muscle of APS-treated mice. In vitro, APS up-regulated the expression of galectin-1 in muscle cells in a dose-dependent manner. The percentage of apoptotic CD8(+) T cells in spleens of APS-treated mice was positively correlated with the concentration of APS treatment, and the blocking of galectin-1 in vivo by specific antibody reduced the percentage of apoptotic CD8(+) T cells in APS-treated mice. Our findings indicated that APS could up-regulate the expression of galectin-1 in muscle of T1DM mice, resulting in the apoptosis of CD8(+) T cells. This may be an important mechanism by which APS protects β cells of the pancreatic islets from apoptosis induced by CD8(+) T cells in T1DM in vivo.

Ranolazine Increases β-Cell Survival and Improves Glucose Homeostasis in Low-Dose Streptozotocin-Induced Diabetes in Mice

In addition to its anti-ischemic and antianginal effects, ranolazine has been shown to lower hemoglobin A(1c) (HbA(1c)) in patients with coronary artery disease and diabetes. The present study was undertaken to test the hypothesis that ranolazine lowers HbA(1c) because of improved glucose homeostasis in an animal model. Diabetes in mice was induced by giving multiple low doses of streptozotocin. Ranolazine was given twice daily via an oral gavage (20 mg/kg) for 8 weeks. Fasting plasma glucose levels were significantly lower in the ranolazine-treated group (187 ± 19 mg/dl) compared with the vehicle group (273 ± 23 mg/dl) at 8 weeks. HbA(1c) was 5.8 ± 0.4% in the vehicle group and 4.5 ± 0.2% in the ranolazine-treated group (p < 0.05). Glucose disposal during the oral glucose tolerance test (OGTT) and insulin tolerance test were not different between the two groups; however, during OGTT, peak insulin levels were significantly (p < 0.05) higher in ranolazine-treated mice. Mice treated with ranolazine had healthier islet morphology and significantly (p < 0.01) higher β-cell mass (69 ± 2% per islet) than the vehicle group (50 ± 5% per islet) as determined from hematoxylin and eosin staining. The number of apoptotic cells was significantly (p < 0.05) less in the pancreas of the ranolazine-treated group (14 ± 2% per islet) compared with the vehicle group (24 ± 4% per islet). In addition, ranolazine increased glucose-stimulated insulin secretion in rat and human islets in a glucose-dependent manner. These data suggest that ranolazine may be a novel antidiabetic agent that causes β-cell preservation and enhances insulin secretion in a glucose-dependent manner in diabetic mice.

Antidiabetic effects of dietary administration of Aloe arborescens Miller components on multiple low-dose streptozotocin-induced diabetes in mice: Investigation on hypoglycemic action and systemic absorption dynamics of aloe components

We carried out three experimental trials to determine antidiabetic effects of Aloe arborescens Miller components. Firstly, ICR mice which received frequent injections of streptozotocin (Sz) in small doses (low-dose Sz-induced diabetes mice) were fed ad libitum with basal diets supplemented with components of Aloe arborescens Miller var. natalensis Berger (Kidachi aloe) and Aloe vera Linne from 31 days before to 73 days after the Sz injections. Variation in blood glucose levels, incidence rates of insulitis and blood insulin levels were examined during the trial. As a result, groups receiving diets supplemented at the rate of 2% with whole leaf of Kidachi aloe and 10 KDa fraction powder (a fraction with less than 10 KDa molecular weight derived from Kidachi aloe leaf skin juice by ultra filtration) significantly suppressed the elevation of blood sugar as compared to a control group receiving basal diet. In contrast, there was no significant effect with Aloe vera leaf pulp powder. Insulitis emerged at the rate of 87% in the basal diet group. On the contrary, the whole aloe leaf and 10 KDa fraction groups significantly decreased the incidence of insulitis and incidence rates of whole aloe leaf and 10 KDa fraction powder were 51 and 38%, respectively. While insulin levels in the basal diet group averaged at 0.05 ng, more than four times the insulin level was observed in the 10 KDa group relative to the basal diet group. Secondary, the inhibitory effects of test materials on intestinal glucose absorption were observed using the jejunum of rats. A strong inhibitory action on intestinal glucose absorption was observed in the 10 KDa fraction powder group. Thirdly, phenol compounds derived from aloe in the blood serum and organs were quantitatively measured by a HPLC following forced administration of aloe components to rats to determine absorption kinetics of aloe components inside the body. The primary component of aloe phenol compounds is the same component of the 10 KDa fraction powder and it was found in the pancreas and liver in addition to in the blood serum. The above results indicate that fore and aft when Sz injections could cause selective toxicity to B cells of islets, the dietary administration of 10 KDa fraction powder to mice would lead to the persistence of aloe phenol compound having an antioxidant activity in the pancreas and blood, which could protect islets of Langerhans from the destruction caused by methyl radical derived from Sz. The results also suggested the possibility of the 10 KDa fraction powder to alleviate the burden of insulin secretion as it has an inhibitory action on glucose absorption in the jejunum of rats.

Beneficial effects of troglitazone on neutrophil dysfunction in multiple low-dose streptozotocin-induced diabetic mice.

Patients with poorly controlled diabetes are at high risk of acquiring bacterial infections. However, conflicting results have been reported on neutrophil function in diabetes. We periodically evaluated neutrophil dysfunction in multiple low-dose streptozotocin (STZ)-induced diabetic mice, and then evaluated the effects of troglitazone and other thiazolidinediones (TZDs) on the decline of neutrophil function. Zymosan was injected intraperitoneally and neutrophil infiltration and phagocytosis were evaluated. While phagocytosis of zymosan by peritoneal neutrophils was consistently reduced in diabetic mice, neutrophil infiltration was decreased on day 30, but increased on day 40 after STZ injection. The in vitro chemotactic and phagocytic activities of blood neutrophils in mice that did not receive zymosan were consistently reduced in diabetic mice. Phorbol myristate acetate (PMA)-stimulated superoxide production by zymosan-induced peritoneal neutrophils and the levels of zymosan-induced tumour necrosis factor (TNF)-alpha and interleukin (IL)-1beta in peritoneal exudate fluids were also reduced in the diabetic mice. Treatment of the diabetic mice with troglitazone beginning 2 weeks after STZ injection did not improve hyperglycaemia but did prevent the decline of zymosan-induced neutrophil infiltration on day 30, and additionally promoted the increased infiltration on day 40. Troglitazone also promoted the chemotactic activity of blood neutrophils isolated from normal mice in vitro. Rosiglitazone but not pioglitazone induced a similar effect. Neutrophil phagocytosis was not enhanced by troglitazone either in vivo or in vitro. Taken together, neutrophil function is impaired by STZ-induced diabetes, but inflammatory infiltration does not always vary with the chemotactic disability or cytokine levels. Furthermore, troglitazone and rosiglitazone were suggested to improve at least neutrophil chemotactic activity in these animals.

Expression of pancreatic islet MHC class I, insulin, and ICA 512 tyrosine phosphatase in low-dose streptozotocin-induced diabetes in mice.

Activated immune cells contribute to the development of diabetes mellitus in multiple low-dose streptozotocin-treated mice. However, a role in the process for MHC Class I restricted T-cells remains a matter of debate. In this study, we examined by confocal microscopy the pancreatic expression of MHC Class I protein, insulin, and ICA 512 protein tyrosine phosphatase in C57BL/Ks mice given 40 mg/kg bw streptozotocin IP on 5 consecutive days. All animals were hyperglycemic from Day 7 and onwards. A loss of ICA 512 from the central portions of the islets was noted on Day 3. On Day 7, an increase in MHC Class I expression, confined primarily to immune cells in the exocrine pancreas and the periinsular areas, was detected. Later, several MHC class I/glucagon and some MHC class I/insulin double-positive cells were found. The insulitis was maximal on Day 14 and declined thereafter. The induction of MHC Class I expression in endocrine cells, occuring only after the cellular infiltration and when the animals were diabetic, indicates that the immune component of the disease does not depend on MHC Class I-restricted cytotoxic T-cells but rather comprises a non-antigen-specific process. (J Histochem Cytochem 48:761-767, 2000)

BCG vaccine prevents insulitis in low dose streptozotocin-induced diabetic mice

Autoimmune type 1 diabetes mellitus is caused by the immunologic destruction of pancreatic β-cells; therefore, there have been many attempts at immunologic modulation as a block or prevention of the underlying process. The aim of this study is to investigate the effect of BCG vaccination on low dose streptozotocin-induced diabetic (LDSD) mice. The mice were pretreated with BCG 7 days before starting low dose streptozotocin (STZ), observed body weight and blood glucose for 2 months, then analyzed the severity of the STZ-induced insulitis after the animals were sacrificed. In this experiment, the mean body weights in the BCG-STZ group on days 1, 19, 33, 47, and 61 of the experiment were 37.5±3.6, 37.3±3.6, 37.5±3.5, 39.4±3.9, and 39.3±4.5 (g), respectively. Those in the STZ group were 37.7±3.5, 38.3±4.5, 38.4±3.9, 36.2±4.5, and 36.3±4.0 (g), respectively ( P<0.05). The mean blood glucose levels in the BCG-STZ group on days 1, 19, 33, 47, and 61 were 106.5±8, 150±37, 147±54, 143±60, and 142±66 (mg/dl), respectively. Those in the STZ group were 103±12, 196±90, 261±236, 236±91, and 224±89 (mg/dl), respectively ( P<0.05). The numbers developing grade 0, I, II, III, and IV insulitis in the BCG-treated group were 63, 48, 33, 4, and 2, respectively, and in the control group were 16, 23, 31, 45, and 35, respectively. This study indicates that BCG vaccination reduces the development of insulitis and overt diabetes in LDSD mice.

Protective Effect of Gamma-Hydroxybutyrate and Nicotinamide on Low-Dose Streptozotocin-Induced Diabetes in Mice

The purpose of this study was to investigate the effect of a treatment by gamma-hydroxybutyrate (GHB) and nicotinamide (NA) on low-dose streptozotocin (STZ)-induced diabetes in mice. Mean plasma glucose level was significantly elevated in mice given STZ by day 12 after the first STZ injection compared to controls (15.0 +/- 4.7 vs 8.0 +/- 1.6 mmol/l, p < 0.001) and 100% of the animals were severely diabetic by day 18. Plasma glucose levels remained in the normal range and no diabetic values were found in mice treated with combined treatment by GHB and NA for 25 days. However, hyperglycemia and glycosuria appeared within one week after discontinuation of the treatment. Treatment by either GHB or NA alone had only a slight and transient effect in preventing hyperglycemia. In vitro experiment on isolated pancreatic islets demonstrated that STZ-induced loss of insulin response to glucose was also counteracted by incubation with GHB and NA (Peak insulin response to 16.4 mM glucose: 0.69 +/- 0.31 vs 3.03 +/- 0.67 microU/islet/min), but not by GHB or NA alone. These results indicate that GHB and NA have complementary effects in preventing STZ-induced beta cell damage both in vivo and in vitro. This should be taken into account for future preventive strategies in human insulin-dependent diabetes mellitus.

Administration of a nitric oxide synthase inhibitor does not suppress low-dose streptozotocin-induced diabetes in mice

Nitric oxide (NO) has been reported as being a key mediator of the autoimmune destruction of B-cells in type I diabetes, and studies have described a suppression of low-dose streptozotocin-induced (LDS) diabetes in mice after the use of NO synthase inhibitors. However, these studies disagree with regard to the outcome of hyperglycemia and insulitis after treatment with these L-arginine analogs. The present study tries to clarify this topic by administering N-nitro-L-arginine-methylester (NAME) (15 mg/d/mouse/15 d) after an LDS treatment in 108 male C57BL6/J mice. Glycemia measured at the end of the NAME treatment did show a slight, but significant, reduction when compared to LDS control animals (p < 0.001), but values returned to diabetic levels 2 wk after withdrawal of NAME. Morphological observations demonstrated that the degree of infiltration and islet B-cell damage was absolutely not inhibited by NAME. In conclusion, treatment with L-arginine analogs is not capable of protecting mice from LDS-induced diabetes.

Spontaneous glucose intolerance in the progeny of low dose streptozotocin-induced diabetic mice

Multiple low doses of streptozotocin (LDS) induce low-incidence diabetes mellitus in Balb/cHan and high-incidence diabetes in CD-1 mice. We studied offspring of diabetic parents in both strains. Group 1 consisted of litters from control mice with no streptozotocin treatment. Group 2 litters had an LDS diabetic mother and a control father, group 3 litters had control mother with LDS diabetic father, and group 4 litters had both, LDS diabetic mother and father. Diabetes was induced by 5 x 40 mg streptozotocin per kg on five consecutive days. Progeny of diabetic mothers showed a state of reduced glucose tolerance associated with reduced glucose disappearance during intravenous glucose tolerance test and increased insulin secretion of isolated islets of Langerhans. These metabolic abnormalities predominated in the male litters of both strains of mice. Amniotic insulin was increased in diabetic mothers during pregnancy. No histologic abnormalities were observed in group 2 progeny. Pancreases in male offspring of LDS diabetic CD-1 fathers (group 3) were studied for insulitis. Insulitis was found in 40% of mice with normal glucose tolerance. A single subdiabetogenic dose of streptozotocin (40 mg/kg) induced insulitis in 90% of pancreases accompanied by reduced insulin release of isolated islets. By contrast, male Balb/cHan progeny of diabetic fathers failed to develop insulitis. In conclusion, we found (1) parental LDS diabetes was transmitted more often to male offspring, (2) maternal LDS diabetes was associated with hyperinsulin secretion and glucose intolerance in the offspring and (3) paternal LDS diabetes was accompanied by insulitis and insulin secretion deficiency in CD-1 progeny.

Low-Dose Streptozotocin-Induced Diabetes in Mice

We have previously found that the development of fatal disseminated candidiasis correlates with the detection of a strong Th2 response, while protective antifungal immunity is associated with a predominant Th1 response. In the present study we verified the hypothesis that an altered antifungal Th response could be responsible for the high susceptibility of diabetic mice to systemic Candida albicans infection. Outbred CD1 mice rendered diabetic with multiple low doses of the pancreatic islet β-cell toxic, streptozotocin, develop a fatal systemic infection when injected with low-virulence C. albicans cells. Progressive disease was found to be associated with the presence in the serum of IgA, IgE, and IgG1 Candida -reactive specific antibodies, absent footpad reactions, and elevated production in vitro of the Th2 cytokines IL-4, IL-6, and IL-10 but not the Th1 cytokine IFN-γ. Both the Th2 and Th1 (IL-2 and IFN-γ cytokines were produced in vitro by CD4 1 lymphocytes from noninfected diabetic mice that, in addition, showed a noticeable footpad reaction to Candida antigens. Thus, it appears that a perturbation in the anticandidal T helper responses resulting in the induction of a biased Th2-like antifungal response renders diabetic mice highly susceptible to systemic C. albicans infection.

Low-Dose Streptozotocin-Induced Diabetes in Mice

Diabetic patients are highly susceptible to microbial and fungal infections. To better understand the immune mechanisms underlying the diabetic host-parasite relationship, we studied the course of systemic infection with Candida albicans in mice with low-dose streptozotocin-induced diabetes. For this purpose, we used a low-pathogenic strain of C. albicans, PCA-2, which causes a chronic infection in the intact host. Injection of PCA-2 cells into diabetic mice caused a lethal acute infection. The levels of interferon-γ (IFN-γ,) determined in sera and splenocyte culture supernatants from diabetic mice were significantly higher than those in control mice. Moreover, splenic macrophages from diabetic mice were functionally activated relative to normal macrophages, as evaluated by significantly augmented C. albicans killing in vitro. However, when diabetic mice were infected with PCA-2, IFN-γ levels dropped dramatically to undetectable levels during the first week of infection and there was a marked decrease in macrophage activation. These data suggest that the levels of IFN-γ production early in infection might have a crucial role in generating the susceptibility of diabetic mice to infection.

Probucol attenuated hyperglycemia in multiple low-dose streptozotocin-induced diabetic mice

The present studies were undertaken to examine the effects of probucol on the protection against pancreatic beta-cell damage by multiple low-dose streptozotocin (STZ: 40 mg/kg, ip). The degree of hyperglycemia at 7, 14 and 17 days after STZ injection was attenuated by probucol. Serum immunoreactive insulin (IRI) levels were increased in the rats fed probucol containing diet at Day 14 and 17. Serum IRI levels after intraperitoneal injection of 2.0 g/kg glucose was reduced in STZ mice and the reduction of serum IRI levels was attenuated in the rats fed probucol, accompanied with a significant reduction of the degree of hyperglycemia after bolus of glucose. Probucol attenuated the reduction of pancreatic IRI content by STZ. The percentage of Thy 1.2-positive splenocytes was increased by STZ and probucol reduced the percentage of Thy 1.2-positive splenocytes, although there were no differences in the populations of splenocytes, positive with Lyt 2 or L3/T4. These data suggest that probucol has a protective action against pancreatic insulitis by multiple low-dose STZ administration.