Toshikatsu Shigihara

CXC Chemokine Ligand 10 Neutralization Suppresses the Occurrence of Diabetes in Nonobese Diabetic Mice through Enhanced β Cell Proliferation without Affecting Insulitis

We have shown that neutralization of IFN-inducible protein 10/CXCL10, a chemokine for Th1 cells, breaks Th1 retention in the draining lymph nodes, resulting in exacerbation in Th1-dominant autoimmune disease models induced by immunization with external Ags. However, there have been no studies on the role of CXCL10 neutralization in Th1-dominant disease models induced by constitutive intrinsic self Ags. So, we have examined the effect of CXCL10 neutralization using a type 1 diabetes model initiated by developmentally regulated presentation of β cell Ags. CXCL10 neutralization suppressed the occurrence of diabetes after administration with cyclophosphamide in NOD mice, although CXCL10 neutralization did not significantly inhibit insulitis and gave no influence on the trafficking of effector T cells into the islets. Because both CXCL10 and CXCR3 were, unexpectedly, coexpressed on insulin-producing cells, CXCL10 was considered to affect mature and premature β cells in an autocrine and/or paracrine fashion. In fact, CXCL10 neutralization enhanced proliferative response of β cells and resultantly increased β cell mass without inhibiting insulitis. Thus, CXCL10 neutralization can be a new therapeutic target for β cell survival, not only during the early stage of type 1 diabetes, but also after islet transplantation.

CXCL10 DNA Vaccination Prevents Spontaneous Diabetes through Enhanced β Cell Proliferation in NOD Mice

CXCL10, a chemokine for Th1 cells, is involved in the pathogenesis of various Th1-dominant autoimmune diseases. Type 1 diabetes is considered to be a Th1-dominant autoimmune disease, and a suppressive effect of CXCL10 neutralization on diabetes development has been reported in a cyclophosphamide-induced accelerated diabetes model through induction of β cell proliferation. However, intervention in a diabetes model might bring about opposite effects, depending on the timing, amount, or method of treatment. In the present study, we examined the effect of CXCL10 neutralization in a “spontaneous diabetes” model of NOD mice, using CXCL10 DNA vaccination (pCAGGS-CXCL10). pCAGGS-CXCL10 treatment in young NOD mice induced the production of anti-CXCL10 Ab in vivo and suppressed the incidence of spontaneous diabetes, although this treatment did not inhibit insulitis or alter the immunological response. pCAGGS-CXCL10 treatment enhanced the proliferation of pancreatic β cells, resulting in an increase of β cell mass in this spontaneous diabetes model as well. Therefore, CXCL10 neutralization is suggested to be useful for maintaining β cell mass at any stage of autoimmune diabetes.

Efficacy and safety of liraglutide monotherapy compared with metformin in Japanese overweight/obese patients with type 2 diabetes

There is little information on direct comparison between metformin and glucagon-like peptide-1 (GLP-1) receptor agonists in the Asian population. This study examined the efficacy and safety of liraglutide monotherapy compared with metformin monotherapy in overweight/obese Japanese patients with type 2 diabetes (T2DM). The study was a 24-week, open-labeled, randomized controlled study. Overweight or obese patients with T2DM aged 20-75 years with suboptimal glycemic control were randomized to liraglutide or metformin monotherapy. The primary endpoint was change in HbA1c at week 24. Secondary endpoints included changes in daily glycemic profile, body weight, incidence of hypoglycemia and other adverse events. The study, which was originally planned to enroll 50 subjects in each group, was ended with insufficient recruitment. A total of 46 subjects completed the study, and analysis was conducted in this cohort. Reduction in HbA1c at week 24 was comparable between the metformin (n = 24) and liraglutide (n = 22) groups (-0.95 ± 0.80% vs. -0.80 ± 0.88%, p = 0.77), while the liraglutide group reached maximal reduction more rapidly than did the metformin group. There was no significant difference in weight gain or incidence of hypoglycemia between the groups. Diarrhea was more frequent in the metformin group, while constipation was more frequent in the liraglutide group. There was no significant difference in treatment satisfaction between the groups. In conclusion, liraglutide and metformin monotherapy showed similar reduction in HbA1c during 24 weeks, with no difference in weight gain or incidence of hypoglycemia in overweight or obese Japanese patients with T2DM.

Pioglitazone may accelerate disease course of slowly progressive type 1 diabetes.

It has been reported that intervention with insulin in slowly progressive type 1 diabetic (SPIDDM) patients delays the progression to an insulin‐dependent state compared to that with sulfonylureas. However, the rate of progression to SPIDDM with the use of insulin‐sensitizing agents is unknown. The aim of this study was to determine the effect of insulin‐sensitizing agents on SPIDDM patients.

GAD‐Reactive T Cells Were Mainly Detected in Autoimmune‐Related Type 1 Diabetic Patients with HLA DR9

Abstract: Type 1 diabetes mellitus (T1DM) is considered to be a T cell‐mediated disease, and many reports suggest that some HLA types, especially HLA DR4 and DR9, convey susceptibility to T1DM in Japanese. We investigated the association between T cell reactivity against GAD and HLA types in “islet‐associated autoantibody‐positive” T1DM in Japanese. Blood samples were obtained from 36 “autoantibody‐positive” type 1 diabetic patients with HLA DR4 or DR9 and 23 type 2 diabetic patients with HLA DR4 or DR9 as controls. They were divided into three groups, DR4/9, DR4/X, and DR9/X groups. In each HLA type group, GAD‐reactive IFN‐γ‐producing CD4+ cells were assessed by means of intracellular cytokine staining for flow cytometry. Type 1 diabetic patients with HLA DR9/X had significantly higher numbers of GAD‐reactive IFN‐γ‐producing CD4+ cells as compared to type 1 diabetic patients with DR4/X or DR4/9 (P < 0.05) and all type 2 diabetic patients. There was no significant difference in the number of GAD‐reactive IFN‐γ‐producing CD4+ cells between type 1 diabetic and type 2 diabetic patients belonging to the DR4/X and DR4/9 groups. There was an association between T cell reactivity against GAD and HLA DR9 in Japanese type 1 diabetes.

Thiazolidinediones May Not Reduce Diabetes Incidence in Type 1 Diabetes

Abstract:  Thiazolidinediones improve glycemic control by reducing insulin resistance. Some studies have demonstrated that troglitazone had a preventative effect on diabetes in NOD (non‐obese diabetic) mice. One of the mechanisms proposed for the prevention of diabetes by thiazolidinediones is an effect on T‐helper 1/T‐helper 2 (Th1/Th2) balance. In this article, we attempted to clarify whether pioglitazone is also effective in preventing diabetes as compared to metformin, which has no immunological effect. Female NOD mice were administered pioglitazone or metformin orally, and the insulitis score, cytokines secreted from splenocytes, cytokine expression levels in the pancreas, and the incidence of diabetes after acceleration by cyclophosphamide were analyzed. We could not find any advantage of pioglitazone in preventing Th1 skewing and the development of diabetes over metformin. Therefore, further research should take place before the application of thiazolidinediones to human slowly progressive insulin‐dependent diabetes mellitus (SPIDDM) patients.

Frequency of CTLA-4 Gene CT60 Polymorphism May Not Be Affected by Vitamin D Receptor Gene Bsm I Polymorphism or HLA DR9 in Autoimmune-Related Type 1 Diabetes in the Japanese

Abstract:  One of the CTLA‐4 SNPs, +6230G>A (CT60), has recently been reported to be related to susceptibility to type 1 diabetes and autoimmune thyroid disease. We have previously reported an association between acute‐onset type 1 diabetes in Japanese and the Vitamin D receptor (VDR) gene Bsm I large B polymorphism, which is related to the Th1‐type response. Moreover, we found a significant correlation between autoimmune‐related type 1 diabetes with HLA DR9 and detection of GAD‐reactive Th1 (T helper 1)‐type cells. In the present article, we tried to clarify whether the frequency of one of the CTLA‐4 SNPs, +6230G>A (CT60), is affected by the VDR gene Bsm I polymorphism or by HLA DR9 in Japanese type 1 diabetics. The frequency of the CT60 GG genotype did not appear to be affected by either the VDR gene Bsm I large B polymorphism or HLA DR9.

N-acetyl-cysteine accelerates transfer of diabetes into non-obese diabetic scid mice

Aims/hypothesisType 1 diabetes mellitus is caused by autoimmune pancreatic beta cell destruction, and the destructive process involves several molecular mechanisms including oxygen-reactive species. A cysteine derivative, N-acetyl-cysteine, is widely used as an antioxidant, but the role of N-acetyl-cysteine in the protection of pancreatic beta cells in Type 1 diabetes remains unclear. The aim of this study was to clarify the effect of N-acetyl-cysteine on beta cells using an adoptive transfer system in a murine model of Type 1 diabetes.MethodsSplenocytes from diabetic female non-obese diabetic mice were transferred into female non-obese diabetic scid/scid recipients to induce diabetes. Just after transfer, N-acetyl-cysteine was administered to non-obese diabetic scid recipients. Two weeks after transfer, the pancreas of the recipients was examined histologically, and cytokine mRNA expression in the pancreas was analysed. In vitro, CD4-positive splenocytes from diabetic donor mice were stimulated with anti-CD3 and anti-CD28 antibodies with or without N-acetyl-cysteine.ResultsTreatment with N-acetyl-cysteine significantly accelerated the transfer of diabetes into non-obese diabetic scid recipients. Treatment with N-acetyl-cysteine accelerated the infiltration of mononuclear cells accompanied by CD8-positive cells into the intra-islet region of the recipient’s pancreas, and enhanced interferon-gamma mRNA expression in the pancreas. In vitro, treatment with N-acetyl-cysteine enhanced interferon-gamma and interleukin-2 production by CD4-positive splenocytes of the diabetic donor mice.Conclusions/interpretationN-acetyl-cysteine accelerates the transfer of diabetes into non-obese diabetic scid mice and this effect is accompanied by the promotion of local infiltration and T-helper cell type 1 responses.

Insulin-like Growth Factor-1 (IGF-1)-derived Peptide Protects against Diabetes in NOD Mice

Spontaneous diabetes in non-obese diabetic (NOD) mice results from β-cell destruction by autoreactive T lymphocytes. Here, we report the significance of insulin-like growth factor-1 (IGF-1) peptide as a tool for the prevention of type 1 diabetes. Female NOD mice were immunized with a subcutaneous injection of IGF-1, glutamic acid decarboxylase (GAD), insulin or IGF-1-derived peptides (residues 8-23, 24-41 or 50-70) in incomplete Freunds adjuvant (IFA) or with IFA only as the control group at 4 weeks of age, and observed up to 36–37 weeks of age. Diabetes onset was significantly suppressed and delayed in the IGF-1 group as compared to the GAD, insulin and control groups (p<0.05), and it was significantly suppressed and delayed in the (50-70)IGF-1 group as compared to the (8-23)IGF-1 and control groups (p<0.02). Although the degree of insulitis in all treated mice was not significantly different, a significant number of IL-4-producing cells in response to IGF-1 peptides were detected in (50-70)IGF-1-treated mice in intracellular cytokine assay. In conclusion, IGF-1 peptide 50-70 immunizations of NOD mice suppressed and delayed diabetes onset, probably through amplification of the Th2-type response. It was suggested that IGF-1 peptide 50-70 immunization can be used as a tool for prevention of type 1 diabetes.

Stromal cell-derived factor-1 chemokine gene polymorphism is not associated with onset age of Japanese type 1 diabetes.

Abstract: Type 1 diabetes is characterized by cell‐mediated autoimmune destruction of pancreatic beta cells. Although the disease shows a strong association with HLA class II alleles, other genes may influence the initiation or the rate of progression of the autoimmune process. Recently, it was reported that a polymorphism of the stromal cell‐derived factor‐1 (SDF‐1) (a kind of chemokine) gene was associated with early onset of type 1 diabetes in Caucasians. Therefore, we examined SDF‐1 gene polymorphism in Japanese type 1 diabetes in this study. We examined the SDF‐1 gene polymorphism (801G→A) in 298 unrelated Japanese type 1 diabetic patients and 270 healthy subjects by the TaqMan PCR method. Allelic and genotypic frequencies of the SDF‐1 A variants were similar in overall type 1 diabetic patients and healthy subjects. We then stratified the patients by their onset pattern (acute vs. slow onset) and islet‐associated autoantibody positivity. However, no significant difference was found among each group of type 1 diabetes. Furthermore, unlike the previous report in “Caucasian” type 1 diabetics, the SDF‐1 A variant was not associated with early onset of the disease in Japanese type 1 diabetics. The SDF‐1 gene polymorphism was not associated with onset age (or onset pattern) of type 1 diabetes in Japanese. Further study is necessary to conclude whether SDF‐1 gene polymorphism affects the onset age in type 1 diabetes in general.