MODY 2: Mutation identification and molecular ancestry in a Brazilian family

Maturity Onset Diabetes of the Young (MODY) is a heterogeneous group of genetic diseases characterized by a primary defect in insulin secretion and hyperglycemia, non-ketotic disease, monogenic autosomal dominant mode of inheritance, age at onset less than 25 years, and lack of auto-antibodies. It accounts for 2–5% of all cases of non-type 1 diabetes. MODY subtype 2 is caused by mutations in the glucokinase (GCK) gene. In this study, we sequenced the GCK gene of two volunteers with clinical diagnosis for MODY2 and we were able to identify four mutations including one for a premature stop codon (c.76C>T). Based on these results, we have developed a specific PCR-RFLP assay to detect this mutation and tested 122 related volunteers from the same family. This mutation in the GCK gene was detected in 21 additional subjects who also had the clinical features of this genetic disease. In conclusion, we identified new GCK gene mutations in a Brazilian family of Italian descendance, with one due to a premature stop codon located in the second exon of the gene. We also developed a specific assay that is fast, cheap and reliable to detect this mutation. Finally, we built a molecular ancestry model based on our results for the migration of individuals carrying this genetic mutation from Northern Italy to Brazil.     

Functional levels of mitochondrial anion transport proteins in non-insulin-dependent diabetes mellitus

The effect of non-insulin-dependent diabetes mellitus (i.e., NIDDM; type 2 diabetes) on the levels of functional mitochondrial anion transport proteins has been determined utilizing a chemically-induced neonatal model of NIDDM. We hypothesized that moderate insulin deficiency exacerbated by the insulin resistance, which is characteristic of NIDDM, would cause changes in mitochondrial anion transporter function that were similar to those we have previously shown to occur in insulin-dependent diabetes mellitus (i.e., IDDM; type 1 diabetes) (Arch. Biochem. Biophys. 280: 181–191, 1990). Our experimental approach consisted of the extraction of the pyruvate, dicarboxylate and citrate transport proteins from the mitochondrial inner membrane with Triton X-114 using rat liver mitoplasts (prepared from diabetic and control animals) as the starting material, followed by the functional reconstitution of each transporter in a proteoliposomal system. This strategy permitted the quantification of the functional levels of these three transporters in the absence of the complications that arise when such measurements are carried out with intact mitochondria (or mitoplasts). We found that experimental NIDDM did not cause significant changes in the extractable and reconstitutable specific (and total) transport activities of the pyruvate, dicarboxylate, and citrate transporters. These results are in marked contrast to our previous findings obtained using rats with IDDM and negated our hypothesis. The present results, in combination with our earlier findings, allow us to conclude that insulin plays an important role in the regulation of mitochondrial anion transporter function. Accordingly, in this model of NIDDM, where the level of insulin is not profoundly deficient, transporter function is unaltered, whereas in IDDM, where a profound insulinopenia exists, transporter function is altered. Furthermore, the present studies suggest that in the neonatal model of NIDDM the three mitochondrial transporters investigated are neither affected by, nor are they the sites of the well documented hepatic post-receptor insulin resistance which is characteristic of this disease.     

Effect of chromium nicotinic acid supplementation on selected cardiovascular disease risk factors

The effects of daily supplemental chromium (200 μg) complexed with 1.8 mg nicotinic acid on plasma glucose and lipids, including total cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides, were assessed in 14 healthy adults and 5 adults with noninsulin-dependent diabetes mellitus (NIDDM) using a double-blind crossover study with 8-wk experimental periods. Eight of the 14 healthy subjects and all 5 subjects with NIDDM also underwent an oral glucose tolerance test with assessment of 90 min postprandial plasma glucose and insulin concentrations. No statistically significant effects of chromium nicotinic acid supplementation were found on plasma insulin, glucose, or lipid concentrations, although chromium nicotinic acid supplementation slightly lowered fasting plasma total and LDL cholesterol, triglycerides, and glucose concentrations, and 90-min postprandial glucose concentrations in individuals with NIDDM.     

The association between the total antioxidant potential of plasma and the presence of coronary heart disease and renal dysfunction in patients with NIDDM

Oxidative stress may be an important pathogenetic factor in the development of diabetic vascular complications. The total antioxidative potential of plasma reflects the ability of an individual to resist oxidative stress. We measured the plasma total peroxyl radical-trapping potential (TRAP) and the concentrations of four plasma chain-breaking antioxidants in 81 patients with non-insulin-dependent diabetes mellitus (NIDDM) nine years after diagnosis and in 102 well-matched non-diabetic control subjects. The association between the total antioxidative potential and the presence of coronary heart disease (CHD) and diabetic kidney disease were also studied. There were no significant differences in plasma TRAP between NIDDM patients and control subjects (1250 ± 199 vs. 1224 ± 198 μM). Nor were there any significant differences in the concentrations of plasma uric acid, ascorbic acid, α-tocopherol, and protein thiols between NIDDM patients and control subjects. Patients with a low glomerular filtration rate and/or high urinary albumin excretion had elevated plasma uric acid. Plasma TRAP was not, however, associated with renal dysfunction. The plasma of NIDDM patients with CHD had a significantly higher value of unidentified antioxidative potential than that of patients without CHD. This relation was strongly dependent upon smoking. In conclusion, these data demonstrate that there are no major defects in the antioxidative potential of plasma caused by NIDDM per se. CHD and diabetic renal dysfunction were not associated with changes in plasma TRAP.     

长期高脂饮食加小剂量链脲佐霉素建立人类普通2型糖尿病大鼠模型的研究

目的　建立类似人类普通 2型糖尿病大鼠模型。方法　采用长期高脂高能量饮食结合小剂量链脲佐霉素 (STZ)腹腔注射的方法。结果　模型大鼠有高血糖 (18 78± 1 97)mmol L、高甘油三酯血症 (10 95 4± 5 5 4 )mmol L、高胆固醇血症 (2 4 4 2± 0 74 )mmol L、脂肪肝、高胰岛素血症 (0 937± 0 2 6 )mmol L、胰岛素敏感性下降及不低于正常对照组的体重。结论　这是一种类似人普通 2型糖尿病大鼠模型     

2型糖尿病患者外周血中T淋巴细胞亚群变化的研究

以免疫荧光双标记单克隆抗体及流式细胞仪检测外周血淋巴细胞亚群 ,在 2型糖尿病患者尤其是非肥胖型患者中 ,外周血CD3 CD8 阳性细胞数显著低于健康对照组 ,而CD4/CD8比值增高。 2型糖尿病患者特别是非肥胖型可能存在明显的细胞免疫功能异常。     

诺沙坦改善非胰岛素依赖型糖尿病大鼠胰岛素敏感性的作用机制

本文研究血管紧张素Ⅱ受体拮抗剂诺沙坦对非胰岛素依赖型糖尿病(non-insulin-dependent diabetes mellitus,NIDDM)大鼠胰岛素敏感性的改善作用,并探讨其作用机制。从饮水中给予正常或高脂喂养加小剂量链脲佐菌素(STZ)诱发的NIDDM大鼠诺沙坦(4 mg/kg),连续6周。分离骨骼肌,用免疫印迹法检测诺沙坦对胰岛素受体底物1(insulin receptor substrate 1,IRS-1)、蛋白激酶B(protein kinase B,PKB)和葡萄糖转运因子4(glucose transporter 4,GLUT4)的表达,以及IRS-1的磷酸化、IRS-1与磷脂酰肌醇3激酶(phosphatidylinositol(PI)3-kinase)的结合。口服葡萄糖耐量试验表明,口服诺沙坦可改善糖尿病大鼠胰岛素敏感性。在骨骼肌组织,NIDDM和正常大鼠的IRS-1、PKB和GLUT4蛋白表达无差异,且不受诺沙坦处理的影响。NIDDM大鼠胰岛素刺激后的骨骼肌IRS-1酪氨酸磷酸化水平、PI 3-kinase结合IRS-1的活性和PKB活性较对照组显著降低(P<0.01),且不能被诺沙坦改善。诺沙坦显著增加NIDDM大鼠肌细胞质膜(plasma membrane,PM)和T管(T-tubules,TT)胰岛素诱导的GLUT4的 含量(P<0.05)。与该结果一致的是,诺沙坦处理的NIDDM大鼠血糖水平较未处理NIDDM大鼠下降(P<0.05)。结果表明,诺沙坦可改善胰岛素抵抗状态,主要是通过非PI 3-kinase依赖的     

Effect of long-term treatment with vanadate in drinking water on KK mice with genetic non-insulin-dependent diabetes mellitus

The glucose-lowering effect of vanadate, ammonium metavanadate (AMV), on diabetic KK mice was examined. Five-week-old male KK mice were administrated with a solution of AMV via drinking water at concentrations of vanadium (V) with 0.1, 1.0, 10 and 100 μg/mL for a period of 10 wk, respectively. Body weight, consumption of food and water, and blood glucose levels was measured every week for 10 wk. The results showed that food consumption and body weight in the experimental groups were similar to those in the control group. A statistically significant decrease of drinking water consumption and blood glucose levels in the group treated with 100 μg V/mL was observed. The glucose tolerance in the vanadate-treated mice with 10 and 100 μg V/mL was remarkably improved compared with the control group. Biochemical analyses at the end of experiments demonstrated that a distinct tendency for the glucose and hemoglobin A1c (HbA1c) levels to decrease with vanadate treatment in the blood was also observed. The glutamic pyruvic transaminase, glutamic oxaloacetate transaminase, blood urea nitrogen, triglyceride, high-density lipoprotein, and total cholesterol levels in plasma were lower in the higher vanadium groups than those in the control group. These results indicate that vanadium effectively produced the glucose-lowering effect at a higher dose than that at a low dose of vanadium in drinking water, without any overt signs of toxicity.     

磺酰脲类受体基因多态性与2型糖尿病的相关性研究

研 究磺酰脲类受体1（SUR1）基因外显子16-3c/t多态性在中国某南方汉族人群中是否为2型糖尿病的致病基因座。采用聚合酶链反应-限制酶酶切片段长度多态性（PCR-RFLP）方法对南方汉族46个2型糖尿病高发家系成员的SUR1基因外显子16的多态性进行分析。利用Mantel-Haenszel分层分析研究该基因座多态性与2型糖尿病的关系。在高发家系人群中,SUR1基因外显子16-3c/t多态性的基因型频率为：cc型29.3%、ct型507%、tt型20%,c等位基因频率为54.7%;患者组基因型频率为：cc型30.2% 、ct型53.8%、tt型16.0% ,c等位基因频率为57.1% ;未患病亲属组基因型频率为：cc型28.3% 、ct型47.2%、tt型24.5%,c等位基因频率为519%,两组间基因型和等位基因的差异经检验无统计学意义（分别为χ2=3.224,P=0.199;χ2=1.250,P=0264）。在性别、吸烟、饮酒、肥胖、高血压等混杂因素中的频率差异亦无显著性。c等位基因频率低于北方汉族人。在中国某南方汉族2型糖尿病高发家族人群中,未发现SUR1基因外显子16-3c/t多态性与2型糖尿病存在关联,该基因座可能不是该人群的致病基因。 Abstract:To study whether the 3c/t polymorphism of the sulfonylurea receptor 1 (SUR1) gene exon16 increased the risk of type 2 diabetes mellitus in type 2 diabetes mellitus pedigrees in Han population in south area of China.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used in 46 type 2 diabetes mellitus pedigrees.The polymorphism in SUR1 was tested and analyzed by Mantel-Haenszel χ2 test.Frequencies of SUR1-3c/t polymorphism had no significant difference between type 2 diabetes mellitus and normal relatives（genotypes χ2=3.224,P=0.199;frequency of allele χ2=1.250,P=0.264）.In all subjects,type 2 diabetes mellitus and normal relatives,SUR1-3c/t genotypes were listed (cc:29.3%,30.2%,28.3%;ct:50.7%,53.8%,47.2%;tt:20%,16.0%,24.5% respectively).The frequencies of c were 54.7%,57.1% and 51.9% respectively.The frequency of c is lower than Han population in northern China.The results show that SUR1 exon16-3c/t polymorphism is not associated with type 2 diabetes mellitus in the population.     

Coagulation inhibitors and glycaemic control in non-insulin-dependent diabetes mellitus

The relationship between long-term glycaemic control and the activity of coagulation inhibitors was investigated in 60 non-insulin-dependent diabetes mellitus (NIDDM) patients not on insulin therapy. Overall, the activities of antithrombin III (AT III) (median 96%, range 65–133%), protein C (127%, 24–190%) and protein S (130%, 54–163%) were not reduced. Patients in poor long-term glycaemic control as verified by increased glycated haemoglobin (HbA1c) demonstrated significantly decreased median AT III activity in comparison with patients in good glycaemic control (92% vs 101%,P=0.016). However, individual values for AT III activity were not below the critical limit of 60%. An inverse correlation between AT III activity and long-term glycaemic control (HbA1c) was calculated (r=–0.378,P=0.0029). As AT III concentrations were found to be normal, we propose that non-enzymatic glycation leads to reduced activity of AT III without affecting its concentration.