替米沙坦对2 型糖尿病合并高血压患者胰岛β 细胞功能影响 的临床观察

目的:观察替米沙坦对糖尿病合并高血压患者胰岛β细胞功能的影响,探索血管紧张素Ⅱ阻断剂降压以外的胰岛功能修复作用.方法:70例糖尿病合并轻、中度高血压的患者随机分为替米沙坦治疗组和氨氯地平治疗组,每组35例患者;替米沙坦治疗组在控制血糖的治疗上给予替米沙坦进行降压治疗;氨氯地平治疗组在控制血糖的治疗上给予氨氯地平进行降压治疗;两组的观察周期均12周,每2周观察1次血压、空腹血糖、并记录低血糖及其它不良反应.治疗前后测糖化血红蛋白(HbAlc)、餐后2小时血糖,并按照HOMA稳态模型公式计算胰岛β细胞功能指数(HOMA-β)和胰岛素抵抗指数(HOMA-IR).结果:两组患者在降压效果方面无显著性差异,收缩压、舒张压对比,P＞0.05;与氨氯地平组比较,替米沙坦组胰岛素峰值和HOMA-3显著升高,HOMA-IR则显著降低.结论:替米沙坦可更显著改善糖尿病合并高血压的患者的胰岛β细胞功能,具有降压以外的改善胰岛细胞功能的作用.     

Distinct Cell Clusters Touching Islet Cells Induce Islet Cell Replication in Association with Over-Expression of Regenerating Gene (REG) Protein in Fulminant Type 1 Diabetes

Background

Pancreatic islet endocrine cell-supporting architectures, including islet encapsulating basement membranes (BMs), extracellular matrix (ECM), and possible cell clusters, are unclear.

Procedures

The architectures around islet cell clusters, including BMs, ECM, and pancreatic acinar-like cell clusters, were studied in the non-diabetic state and in the inflamed milieu of fulminant type 1 diabetes in humans.

Result

Immunohistochemical and electron microscopy analyses demonstrated that human islet cell clusters and acinar-like cell clusters adhere directly to each other with desmosomal structures and coated-pit-like structures between the two cell clusters. The two cell-clusters are encapsulated by a continuous capsule composed of common BMs/ECM. The acinar-like cell clusters have vesicles containing regenerating (REG) Iα protein. The vesicles containing REG Iα protein are directly secreted to islet cells. In the inflamed milieu of fulminant type 1 diabetes, the acinar-like cell clusters over-expressed REG Iα protein. Islet endocrine cells, including beta-cells and non-beta cells, which were packed with the acinar-like cell clusters, show self-replication with a markedly increased number of Ki67-positive cells.

Conclusion

The acinar-like cell clusters touching islet endocrine cells are distinct, because the cell clusters are packed with pancreatic islet clusters and surrounded by common BMs/ECM. Furthermore, the acinar-like cell clusters express REG Iα protein and secrete directly to neighboring islet endocrine cells in the non-diabetic state, and the cell clusters over-express REG Iα in the inflamed milieu of fulminant type 1 diabetes with marked self-replication of islet cells.     

Islet Endothelial Activation and Oxidative Stress Gene Expression Is Reduced by IL-1Ra Treatment in the Type 2 Diabetic GK Rat

Background

Inflammation followed by fibrosis is a component of islet dysfunction in both rodent and human type 2 diabetes. Because islet inflammation may originate from endothelial cells, we assessed the expression of selected genes involved in endothelial cell activation in islets from a spontaneous model of type 2 diabetes, the Goto-Kakizaki (GK) rat. We also examined islet endotheliuml/oxidative stress (OS)/inflammation-related gene expression, islet vascularization and fibrosis after treatment with the interleukin-1 (IL-1) receptor antagonist (IL-1Ra).

Methodology/Principal Findings

Gene expression was analyzed by quantitative RT-PCR on islets isolated from 10-week-old diabetic GK and control Wistar rats. Furthermore, GK rats were treated s.c twice daily with IL-1Ra (Kineret, Amgen, 100 mg/kg/day) or saline, from 4 weeks of age onwards (onset of diabetes). Four weeks later, islet gene analysis and pancreas immunochemistry were performed. Thirty-two genes were selected encoding molecules involved in endothelial cell activation, particularly fibrinolysis, vascular tone, OS, angiogenesis and also inflammation. All genes except those encoding angiotensinogen and epoxide hydrolase (that were decreased), and 12-lipoxygenase and vascular endothelial growth factor (that showed no change), were significantly up-regulated in GK islets. After IL-1Ra treatment of GK rats in vivo, most selected genes implied in endothelium/OS/immune cells/fibrosis were significantly down-regulated. IL-1Ra also improved islet vascularization, reduced fibrosis and ameliorated glycemia.

Conclusions/Significance

GK rat islets have increased mRNA expression of markers of early islet endothelial cell activation, possibly triggered by several metabolic factors, and also some defense mechanisms. The beneficial effect of IL-1Ra on most islet endothelial/OS/immune cells/fibrosis parameters analyzed highlights a major endothelial-related role for IL-1 in GK islet alterations. Thus, metabolically-altered islet endothelium might affect the β-cell microenvironment and contribute to progressive type 2 diabetic β-cell dysfunction in GK rats. Counteracting islet endothelial cell inflammation might be one way to ameliorate/prevent β-cell dysfunction in type 2 diabetes.     

Association of Levels of Mannose-Binding Lectin and the MBL2 Gene with Type 2 Diabetes and Diabetic Nephropathy

Objective

To investigate the association of Mannose-binding lectin (MBL) and the MBL2 gene with type 2 diabetes and diabetic nephropathy and the influence of MBL2 polymorphisms on serum MBL levels.

Methods

The study population included 675 type 2 diabetic patients with or without nephropathy and 855 normoglycemic controls. The single nucleotide polymorphisms (SNPs) of rs1800450, rs1800451, and rs11003125 of the MBL2 gene were determined by the Multiplex Snapshot method. Serum MBL levels were measured by enzyme-linked immune sorbent assay.

Results

Rs1800450 and rs11003125 SNPs demonstrated strong linkage disequilibrium in the study population (r² = 0.97). The haplotypes constructed from the G allele of rs1800450 and the C allele of rs11003125 increased the risk for type 2 diabetes (OR = 1.2, 95% CI = 1.1–1.4, P = 0.01). For rs1800450, GG and GA genotypes were associated with type 2 diabetes (P = 0.02, 0.01, respectively). For rs11003125, the GC genotype frequency was significantly different between patients and controls (18.1% vs. 24.9%, P = 0.001). Analyses of genotypes and allele frequency distributions among patients with normal UAE, microalbuminuria, and macroalbuminuria showed that there was no obvious evidence of association between the MBL2 gene and diabetic nephropathy. Subjects with the GG genotype of rs1800450 and the CC genotype of rs11003125 had much higher serum MBL levels.

Conclusions

The rs1800450 and rs11003125 SNPs of the MBL2 gene have strong linkage disequilibrium and are associated with type 2 diabetes in the North Chinese Han population. No association was observed between the MBL2 gene and diabetic nephropathy. Subjects with the GG genotype of rs1800450 and the CC genotype of rs11003125 had much higher serum MBL levels. An association between elevated serum MBL and diabetic nephropathy was also observed.     

23例1型糖尿病初发患儿免疫学改变与胰岛β细胞功能的关系

目的：观察1型糖尿病（T1DM）初发患儿细胞免疫与胰岛β细胞功能之间的关系。方法：T1DM组23例初发患儿均测定淋巴细胞亚群（T细胞、B细胞及NK细胞）、HbA1c、INS、C-P,正常对照组20例,测定淋巴细胞亚群。结果：T1DM组CD4、CD4/CD8较正常对照组升高（P〈0.05）;CD8、CD3-CD56＋较对照组降低（P〈0.05）;CD4/CD8比值与HbA1c呈正相关（r=0.9451,P〈0.01）,而与INS呈负相关（r=-0.1020,P〈0.01）,与C-P呈负相关（r=-0.6174,P〈0.01）;CD3-CD56＋与HbA1c呈正相关（r=0.1320,P〈0.01）,而与INS呈正相关（r=-0.0846,P〈0.01）,与C-P呈负相关（r=-0.3224,P〈0.01）。结论：T1DM初发患者CD4、CD4/CD8明显增高,CD8、CD3-CD56＋明显降低,提示细胞免疫功能改变与胰岛β细胞功能损伤密切相关。     

23 例1 型糖尿病初发患儿免疫学改变与胰岛β细胞功能的关系

目的:观察1型糖尿病(T1DM)初发患儿细胞免疫与胰岛β细胞功能之间的关系。方法:T1DM组23例初发患儿均测定淋巴细胞亚群(T细胞、B细胞及NK细胞)、HbA1c、INS、C-P,正常对照组20例,测定淋巴细胞亚群。结果:T1DM组CD4、CD4/CD8较正常对照组升高(P<0.05);CD8、CD3-CD56+较对照组降低(P<0.05);CD4/CD8比值与HbA1c呈正相关(r=0.9451,P<0.01),而与INS呈负相关(r=-0.1020,P<0.01),与C-P呈负相关(r=-0.6174,P<0.01);CD3-CD56+与HbA1c呈正相关(r=0.1320,P<0.01),而与INS呈正相关(r=-0.0846,P<0.01),与C-P呈负相关(r=-0.3224,P<0.01)。结论:T1DM初发患者CD4、CD4/CD8明显增高,CD8、CD3-CD56+明显降低,提示细胞免疫功能改变与胰岛β细胞功能损伤密切相关。     

Maternal Microchimerism: Increased in the Insulin Positive Compartment of Type 1 Diabetes Pancreas but Not in Infiltrating Immune Cells or Replicating Islet Cells

Background

Maternal microchimeric cells (MMc) transfer across the placenta during pregnancy. Increased levels of MMc have been observed in several autoimmune diseases including type 1 diabetes but their role is unknown. It has been suggested that MMc are 1) effector cells of the immune response, 2) targets of the autoimmune response or 3) play a role in tissue repair. The aim of this study was to define the cellular phenotype of MMc in control (n = 14) and type 1 diabetes pancreas (n = 8).

Methods

Using sex chromosome-based fluorescence in-situ hybridization, MMc were identified in male pancreas and their phenotype determined by concomitant immunofluorescence.

Results

In normal pancreas, MMc positive for endocrine, exocrine, duct and acinar markers were identified suggesting that these cells are derived from maternal progenitors. Increased frequencies of MMc were observed in type 1 diabetes pancreas (p = 0.03) with particular enrichment in the insulin positive fraction (p = 0.01). MMc did not contribute to infiltrating immune cells or Ki67+ islet cell populations in type 1 diabetes.

Conclusion

These studies provide support for the hypothesis that MMc in human pancreas are derived from pancreatic precursors. Increased frequencies of MMc beta cells may contribute to the initiation of autoimmunity or to tissue repair but do not infiltrate islets in type 1 diabetes.     

Glycemic Control and Diabetic Dyslipidemia in Adolescents with Type 2 Diabetes

ObjectiveThe incidence of type 2 diabetes mellitus (T2DM) is increasing at an alarming rate, especially in ethnic minorities, and T2DM is associated with significant comorbidities. The primary objective of this study was to assess glycemic control and cardiovascular risk outcomes in children with T2DM at 1 year after diagnosis. We also assessed whether insulin treatment at onset of diabetes is beneficial for overall outcome in those with elevated glycated hemoglobin (HbA1C).MethodsA retrospective electronic chart review of non-Hispanic white (NHW) and African American (AA) children with T2DM.ResultsA total of 86 patients (66.3% females, 79.1% AA, mean age, 13.8 ± 2.4 years) with T2DM were included. Analyses of therapeutic outcome measures at the 1-year follow-up showed HbA1C <8% in 27.7% of patients, low-density-lipoprotein cholesterol (LDL-C) >130 mg/dL in 12.5%, non-high-density-lipoprotein cholesterol (non-HDL-C) >160 mg/dL in 15.6%, HDL-C <35 mg/dL in 25%, systolic hypertension (HTN) in 35.6%, and diastolic HTN in 6.8% of subjects. Among those started on insulin at initial diagnosis, there was significant improvement in glycemic outcomes (P<.0001 on insulin vs. P = .02 not on insulin) and dyslipidemia (total cholesterol [TC] [P = .001], LDL-C [P = .02], HDL-C [P = .01], non-HDL-C [P = .0002], and TC/HDL-C [P = .005]) compared with no significant change among those who did not receive insulin at diagnosis.ConclusionSubstantial numbers of children with T2DM do not achieve glycemic and cardiovascular therapeutic goals 1 year after diagnosis. Insulin therapy at diagnosis has significant beneficial effects on diabetic dyslipidemia in those with higher HbA1C. (Endocr Pract. 2013; 19:972-979)     

C型产气荚膜梭菌β2毒素基因的克隆与表达

利用PCR技术,从C型产气荚膜梭菌染色体基因组中扩增了0．72kb的β2毒素基因,将纯化的PER产物与载体pGEM—T连接,转化至受体菌JM109中,经NcoI／Bam HI和Bam HI／Eco RI酶切鉴定及核苷酸序列测定证实,重组质粒pXCPB2中含有陡毒素基因。随后用Nco I／Bam HI酶切质粒pXCPB2,回收β2毒素基因片段,插入到事先经同样酶切处理的载体pET-28c中相应酶切位点,构建了表达质粒pETXB2,经Nco I／Bam HI和Nco I／Hind Ⅲ／Bam HI酶切鉴定及核苷酸序列测定证实,表达质粒含有陡毒素基因且基因序列和阅读框架正确。重组菌株BL21（DE3）（pETXB2）表达产物经ELISA检测和SDS—PAGE分析,重组菌株表达的β2毒素蛋白能够被如毒素抗体识别,其表达量占菌体总蛋白相对含量的13．26％。     

Impact of Hypertension on the Association of BMI with Risk and Age at Onset of Type 2 Diabetes Mellitus: Age- and Gender-Mediated Modifications

Aims

Given that BMI correlates with risk of Type 2 diabetes mellitus (T2DM), and that hypertension is a common comorbid condition, we hypothesize that hypertension augments significantly the impact of obesity on T2DM onset.

Methods

We obtained data on T2DM in Kuwaiti natives from Kuwait Health Network Registry. We considered 1339 comorbid individuals with onset of hypertension preceding that of T2DM, and 3496 non-hypertensive individuals but with T2DM. Multiple linear regressions, ANOVA tests, and Cox proportional hazards models were used to quantify the impact of hypertension on correlation of BMI with age at onset and risk of T2DM.

Results

Impact of increasing levels of BMI on age at onset ot T2DM is seen augmented in patients diagnosed with hypertension. We find that the slope of the inverse linear relationship between BMI and onset age of T2DM is much steep in hypertensive patients (−0.69, males and −0.39, females) than in non-hypertensive patients (−0.36, males and −0.17, females). The decline in onset age for an unit increase of BMI is two-fold in males than in females. Upon considering BMI as a categorical variable, we find that while the mean onset age of T2DM in hypertensive patients decreases by as much as 5–12 years in every higher BMI categories, significant decrease in non-hypertensive patients exists only when severely obese. Hazard due to hypertension (against the baseline of non-hypertension and normal weight) increases at least two-fold in every obese category. While males have higher hazard due to hypertension in early adulthood, females have higher hazard in late adulthood.

Conclusion

Pre-existing condition of hypertension augments the association of BMI with Type 2 diabetes onset in both males and females. The presented results provide health professionals directives on the extent of weight-loss required to delay onset of Type 2 diabetes in hypertensive versus non-hypertensive patients.     

Chronic Kidney Disease and Diabetic Retinopathy in Patients with Type 2 Diabetes

Purpose

To explore the relationship between chronic kidney disease (CKD) and diabetic retinopathy (DR) in a representative population of type 2 diabetes mellitus (DM2) patients in Catalonia (Spain).

Methods

This was a population-based, cross-sectional study. A total of 28,344 patients diagnosed with DM2 who had recorded ophthalmologic and renal functional examinations were evaluated. Data were obtained from a primary healthcare electronic database of medical records. CKD was defined as an estimated glomerular filtration ratio (eGFR) of <60 ml/min/1.73m² and/or urine albumin to creatinine ratio (UACR) ≥30 mg/g. DR was categorized as non-vision threatening diabetic retinopathy and vision threatening diabetic retinopathy.

Results

CKD was associated with a higher rate of DR [OR], 95% confidence interval [CI], 1.5 (1.4–1.7). When we analyzed the association between different levels of UACR and DR prevalence observed that DR prevalence rose with the increase of UACR levels, and this association was significant from UACR values ≥10 mg/g, and increased considerably with UACR values ≥300mg/g (Odds ratio [OR], 95% confidence interval [CI], 2.0 (1.6–2.5). This association was lower in patients with eGFR levels 44 to 30 mL/min/1.73m² [OR], 95% confidence interval [CI], 1.3 (1.1–1.6).

Conclusions

These results show that CKD, high UACR and/or low eGFR, appear to be associated with DR in this DM2 population.     

Association of Hepatocyte Nuclear Factor 4 Alpha Polymorphisms with Type 2 Diabetes With or Without Metabolic Syndrome in Malaysia

This study investigated the association of hepatocyte nuclear factor 4 (HNF4) alpha single nucleotide polymorphisms (SNPs) with type 2 diabetes with or without metabolic syndrome in Malaysia. Nine HNF4 alpha SNPs were genotyped in 390 type 2 diabetic subjects with metabolic syndrome, 135 type 2 diabetic subjects without metabolic syndrome, and 160 control subjects. The SNPs rs4810424, rs1884613, and rs2144908 were associated with protection against type 2 diabetes without metabolic syndrome (recessive P = 0.018, OR 0.32; P = 0.004, OR 0.25; P = 0.005, OR 0.24, respectively). The 6-SNP haplotype2 CCCGTC containing the risk genotype of these SNPs was associated with higher risk for type 2 diabetes with or without metabolic syndrome (P = 0.002, OR 2.2; P = 0.004, OR 3.1). These data suggest that HNF4 alpha SNPs and haplotypes contributed to increased type 2 diabetes risk in the Malaysian population.     

Identification of KCNJ15 as a Susceptibility Gene in Asian Patients with Type 2 Diabetes Mellitus

Recent advances in genome research have enabled the identification of new genomic variations that are associated with type 2 diabetes mellitus (T2DM). Via fine mapping of SNPs in a candidate region of chromosome 21q, the current study identifies potassium inwardly-rectifying channel, subfamily J, member 15 (KCNJ15) as a new T2DM susceptibility gene. KCNJ15 is expressed in the β cell of the pancreas, and a synonymous SNP, rs3746876, in exon 4 (C566T) of this gene, with T allele frequency among control subjects of 3.1%, showed a significant association with T2DM affecting lean individuals in three independent Japanese sample sets (p = 2.5 × 10⁻⁷, odds ratio [OR] = 2.54, 95% confidence interval [CI] = 1.76–3.67) and with unstratified T2DM (p = 6.7 × 10⁻⁶, OR = 1.76, 95% CI = 1.37–2.25). The diabetes risk allele frequency was, however, very low among Europeans in whom no association between this variant and T2DM could be shown. Functional analysis in human embryonic kidney 293 cells demonstrated that the risk allele of the synonymous SNP in exon 4 increased KCNJ15 expression via increased mRNA stability, which resulted in the higher expression of protein as compared to that of the nonrisk allele. We also showed that KCNJ15 is expressed in human pancreatic β cells. In conclusion, we demonstrated a significant association between a synonymous variant in KCNJ15 and T2DM in lean Japanese patients with T2DM, suggesting that KCNJ15 is a previously unreported susceptibility gene for T2DM among Asians.     

FTO Gene Polymorphism Is Associated with Type 2 Diabetes through Its Effect on Increasing the Maximum BMI in Japanese Men

AimSeveral studies have demonstrated that polymorphisms within the fat-mass and obesity-associated gene (FTO) are associated with type 2 diabetes (T2D). However, whether the effects of the FTO locus on T2D susceptibility are independent of fat-mass increases remains controversial. To investigate this issue, we examined the association of FTO variants with T2D and various aspects of BMI history during adult life in a Japanese population.MethodsWe genotyped SNPs within FTO (rs1121980 and rs1558902) in 760 Japanese patients with T2D who had reached a lifetime maximum BMI (BMI_max) before or at the time of diagnosis and 693 control individuals with information regarding their BMI_max.ResultsThe BMI_max showed the strongest association with T2D risk among the BMIs evaluated in this study. In the sex-combined analysis, FTO SNPs were not associated with any of the BMI variables or with T2D, but in sex-stratified analyses, both SNPs were significantly associated with the BMI_max and rs1558902 was associated with T2D in men. The association of the SNPs with T2D remained significant after adjustments for the current BMI and age, whereas the T2D association of the SNP was no longer significant after adjustments for BMI_max and age.ConclusionsThese results suggest that the effects of FTO polymorphisms on T2D susceptibility in Japanese men are mediated through their effect on increasing the BMI_max before or at the time of diagnosis.     

Contrasting Roles of Islet Resident Immunoregulatory Macrophages and Dendritic Cells in Experimental Autoimmune Type 1 Diabetes

The innate immune system critically shapes diabetogenic adaptive immunity during type 1 diabetes (T1D) pathogenesis. While the role of tissue-infiltrating monocyte-derived macrophages in T1D is well established, the role of their tissue-resident counterparts remains undefined. We now demonstrate that islet resident macrophages (IRMs) from non-autoimmune mice have an immunoregulatory phenotype and powerfully induce FoxP3+ Tregs in vitro. The immunoregulatory phenotype and function of IRMs is compromised by TLR4 activation in vitro. Moreover, as T1D approaches in NOD mice, the immunoregulatory phenotype of IRMs is diminished as is their relative abundance compared to immunostimulatory DCs. Our findings suggest that maintenance of IRM abundance and their immunoregulatory phenotype may constitute a novel therapeutic strategy to prevent and/or cure T1D.     

突变型人白细胞介素-2基因在巴斯德毕赤酵母中的表达

为了提高人重组白细胞介素 2的稳定性和活性以及减少毒副作用 ,有必要定向改造rhIL 2的分子结构 .用PCR法从白细胞介素 2 (IL 2 )cDNA全序列中扩增成熟的肽基因片段 ,并利用定点突变技术将人重组白细胞介素 2第 12 5位游离的半胱氨酸编码序列突变为丙氨酸序列 .编码 18位亮氨酸的序列突变为蛋氨酸序列 ;编码 19位亮氨酸的序列突变为丝氨酸序列 .突变型人白细胞介素 2 (MvIL 2 )基因与表达载体pPIC9K重组 ,酶切线性化后用Invitrogen转化毕赤酵母试剂盒导入酵母细胞进行整合 ,经筛选得到一高表达白介素 2的克隆 .SDS PAGE显示 ,表达量约占总量的4 5 7% .经Western印迹验证 ,重组人白介素 2有免疫活性 ;与野生型IL 2相比 ,所获得的突变型IL 2纯品的比活性为 4 0× 10 7IU mg蛋白 ,比天然型IL 2高 4～ 5倍