A missense mutation in hepatocyte nuclear factor-4 alpha, resulting in a reduced transactivation activity, in human late-onset non-insulin-dependent diabetes mellitus.

Non-insulin-dependent diabetes mellitus (NIDDM) is a heterogeneous disorder characterized by hyperglycemia resulting from defects in insulin secretion and action. Recent studies have found mutations in the hepatocyte nuclear factor-4 alpha gene (HNF-4alpha) in families with maturity-onset diabetes of the young (MODY), an autosomal dominant form of diabetes characterized by early age at onset and a defect in glucose-stimulated insulin secretion. During the course of our search for susceptibility genes contributing to the more common late-onset NIDDM forms, we observed nominal evidence for linkage between NIDDM and markers in the region of the HNF-4alpha/MODY1 locus in a subset of French families with NIDDM diagnosed before 45 yr of age. Thus, we screened these families for mutations in the HNF-4alpha gene. We found a missense mutation, resulting in a valine-to-isoleucine substitution at codon 393 in a single family. This mutation cosegregated with diabetes and impaired insulin secretion, and was not present in 119 control subjects. Expression studies showed that this conservative substitution is associated with a marked reduction of transactivation activity, a result consistent with this mutation contributing to the insulin secretory defect observed in this family.     

Clinical application of 1,5-anhydroglucitol measurements in patients with hepatocyte nuclear factor-1alpha maturity-onset diabetes of the young

OBJECTIVE—1,5-anhydroglucitol (1,5-AG) is a short-term marker of metabolic control in diabetes. Its renal loss is stimulated in hyperglycemic conditions by glycosuria, which results in a lowered plasma concentration. As a low renal threshold for glucose has been described in hepatocyte nuclear factor-1α (HNF-1α) maturity-onset diabetes of the young (MODY), the 1,5-AG level may be altered in these patients. The purpose of this study was to assess the 1,5-AG levels in patients with HNF-1α MODY and in type 2 diabetic subjects with a similar degree of metabolic control. In addition, we aimed to evaluate this particle as a biomarker for HNF-1α MODY.RESEARCH DESIGN AND METHODS—We included 33 diabetic patients from the Polish Nationwide Registry of MODY. In addition, we examined 43 type 2 diabetic patients and 47 nondiabetic control subjects. The 1,5-AG concentration was measured with an enzymatic assay (GlycoMark). Receiver operating characteristic (ROC) curve analysis was used to evaluate 1,5-AG as a screening marker for HNF-1α MODY.RESULTS—The mean 1,5-AG plasma concentration in diabetic HNF-1α mutation carriers was 5.9 μg/ml, and it was lower than that in type 2 diabetic patients (11.0 μg/ml, P = 0.003) and in nondiabetic control subjects (23.9 μg/ml, P < 0.00005). The ROC curve analysis revealed 85.7% sensitivity and 80.0% specificity of 1,5-AG in screening for HNF-1α MODY at the criterion of <6.5 μg/ml in patients with an A1C level between 6.5 and 9.0%.CONCLUSIONS—1,5-AG may be a useful biomarker for differential diagnosis of patients with HNF-1α MODY with a specific range of A1C, although this requires further investigation. However, the clinical use of this particle in diabetic HNF-1α mutation carriers for metabolic control has substantial limitations.1,5-anhydroglucitol (1,5-AG) is a monosaccharide that shows a structural similarity to glucose. Its main source in humans is dietary ingestion, particularly meats and cereals (1). In addition, ∼10% of 1,5-AG is derived from endogenous synthesis. 1,5-AG is generally not metabolized, and in healthy subjects it achieves a stable plasma concentration that reflects a steady balance between ingestion and urinary excretion (1).1,5-AG is reabsorbed in renal tubules by an AG/fructose/mannose common transport system that is distinct from the major glucose reabsorption system (2,3). In hyperglycemic conditions, the excess of glucose is reabsorbed owing to chemical similarity not only by its own specific active transporters, but also by the AG/fructose/mannose transporter; therefore, it competes with 1,5-AG. Subsequently, 1,5-AG urinary excretion is increased during hyperglycemia, and it results in a lowered plasma concentration (4). This explains its low plasma level in patients with poorly controlled diabetes. This particle was established in clinical practice as a short-term marker of metabolic control, and recently it has been investigated as a marker of postprandial hyperglycemia (5,6). In contrast to A1C, it reflects changes in glycemic control over a period of 1–2 weeks (5).The 1,5-AG excretion rate depends on the renal threshold for glucose (7). Thus, its clinical usefulness in evaluation of some groups of patients, for example, pregnant women and subjects with end-stage renal disease, is obvious (7,8). Interestingly, the decreased renal threshold for glucose was observed in patients with hepatocyte nuclear factor-1α (HNF-1α) maturity-onset diabetes of the young (MODY), formerly classified as MODY3, and in nondiabetic mutation carriers of this gene (9,10). Diabetes that results from HNF-1α mutations is usually accompanied by extrapancreatic features. One of them is a tubulopathy that results in a low renal threshold for glucose. An animal model suggests that it is caused by decreased expression of sodium-glucose cotransporter 2, a low-affinity, high-capacity transporter in proximal renal tubules (11). Thus, hypothetically, the increased glucose load in renal tubules in these patients may cause stronger competition with 1,5-AG for reabsorption and, subsequently, increased urinary loss. Moreover, one cannot exclude the impact of HNF-1α mutations on the expression of the AG/fructose/mannose transporter. Therefore, in HNF-1α MODY, lower plasma concentrations of 1,5-AG compared with those in other types of diabetes can be expected at a similar A1C. In the scenario of a very low 1,5-AG level, this particle should also be considered as a candidate biochemical marker for this monogenic type of diabetes, which may have considerable clinical implications and enable screening for patients with HNF-1α MODY in large cohorts, potentially avoiding the expensive and laborious technique of direct gene sequencing. In this study, we compared 1,5-AG plasma concentrations in diabetic HNF-1α mutation carriers with those in patients with type 2 diabetes and in nondiabetic subjects.     

胰岛素启动子因子1基因突变与上海地区糖尿病发病的关联

目的:对上海地区2型糖尿病患者、特发性1型糖尿病患者及健康者进行胰岛素启动子因子1基因序列筛查,探讨该基因在糖尿病发病中的作用。方法:①选择2002-05/2003-12上海交通大学附属第一人民医院内分泌科住院2型糖尿病及特发性1型糖尿病患者。2型糖尿病患者201例(2型糖尿病组),男94例,女107例;特发性1型糖尿病患者4例(特发性1型糖尿病组),男3例,女1例。纳入同期本院内分泌门诊体检健康者100例为对照组,男67例,女33例。均自愿接受相关检查,并对实验目的知情同意。②测定血生化学指标及与糖尿病发病相关抗体。对胰岛素启动子因子1基因的两个外显子序列(包括编码区及侧翼区)进行聚合酶链反应扩增,聚合酶链反应产物经纯化后,经ABI377DNA全自动测序仪对所有聚合酶链反应产物直接测序。发现突变后用特异性限制性内切酶对突变位点进行酶切,验证突变。③正态分布数据以x±s表示,偏态分布数据用M表示,C肽呈偏态分布将其转换为以10为底的对数后呈正态分布。结果:2型糖尿病患者201例,特发性1型糖尿病患者4例,健康者100例均进入结果分析。2型糖尿病组及对照组中均未发现胰岛素启动子因子1基因突变。特发性1型糖尿病组中发现1例P239Q(CCG-CAG)杂合突变,该聚合酶链反应产物可被PstⅠ部分酶切,所切的2个片段大小分别为347和252bp,与预测相吻合。结论:①胰岛素启动子因子1基因不是上海地区2型糖尿病发病的主要易感基因。②4例特发性1型糖尿病中发现1例P239Q(CCG-CAG)突变,今后需扩大样本量及对先证者作家系调查,以探讨胰岛素启动子因子1基因突变是否与特发性1型糖尿病早期胰岛功能衰竭有关,为今后对特发性1型糖尿病的病因研究开拓思路。     

胰岛素启动子因子1基因突变与上海地区糖尿病发病的关联

目的：对上海地区2型糖尿病患者、特发性1型糖尿病患者及健康者进行胰岛素启动子因子1基因序列筛查,探讨该基因在糖尿病发病中的作用.方法：①选择2002-05/2003-12上海交通大学附属第一人民医院内分泌科住院2型糖尿病及特发性1型糖尿病患者.2型糖尿病患者201例（2型糖尿病组）,男94例,女107例;特发性1型糖尿病患者4例（特发性1型糖尿病组）,男3例,女1例.纳入同期本院内分泌门诊体检健康者100例为对照组,男67例,女33例.均自愿接受相关检查,并对实验目的知情同意.②测定血生化学指标及与糖尿病发病相关抗体.对胰岛素启动子因子1基因的两个外显子序列（包括编码区及侧翼区）进行聚合酶链反应扩增,聚合酶链反应产物经纯化后,经ABI 377 DNA全自动测序仪对所有聚合酶链反应产物直接测序.发现突变后用特异性限制性内切酶对突变位点进行酶切,验证突变.③正态分布数据以x&;#177;s表示,偏态分布数据用M表示,C肽呈偏态分布将其转换为以10为底的对数后呈正态分布.结果：2型糖尿病患者201例,特发性1型糖尿病患者4例,健康者100例均进入结果分析.2型糖尿病组及对照组中均未发现胰岛素启动子因子1基因突变.特发性1型糖尿病组中发现1例P239Q（CCG-CAG）杂合突变,该聚合酶链反应产物可被PstⅠ部分酶切,所切的2个片段大小分别为347和252 bp,与预测相吻合.结论：①胰岛素启动子因子1基因不是上海地区2型糖尿病发病的主要易感基因.②4例特发性1型糖尿病中发现1例P239Q（CCG-CAG）突变,今后需扩大样本量及对先证者作家系调查,以探讨胰岛素启动子因子1基因突变是否与特发性1型糖尿病早期胰岛功能衰竭有关,为今后对特发性1型糖尿病的病因研究开拓思路.     

Intrauterine hyperglycemia is associated with an earlier diagnosis of diabetes in HNF-1alpha gene mutation carriers

OBJECTIVE: In animals, experimentally induced maternal hyperglycemia during pregnancy results in hyperglycemic offspring. Similarly, Pima Indian offspring with mothers who are diabetic at the time of pregnancy have increased risk of early-onset diabetes. We hypothesized that exposure to hyperglycemia in utero would decrease the age at diagnosis of diabetes in patients with maturity-onset diabetes of the young (MODY) due to a mutation in the hepatocyte nuclear factor 1alpha (HNF-1alpha) gene. RESEARCH DESIGN AND METHODS: We analyzed the affect of maternal diabetes on age at diagnosis of diabetes in 150 HNF-1alpha gene mutation carriers from 55 families. RESULTS: Age at diagnosis in HNF-1alpha mutation carriers was younger when the mother was diagnosed before pregnancy compared with when the mother was diagnosed after pregnancy (15.5 +/- 5.4 vs. 27.5 +/- 13.1 years, P < 0.0001). This is unlikely to represent a generalized familial decrease in age at diagnosis due to a more severe mutation, because no difference was seen in age of the offspring at diagnosis of diabetes when the father was diagnosed at a young age, and a similar trend was seen when only the single common mutation, P291fsinsC, was analyzed. CONCLUSIONS: We conclude that maternal hyperglycemia during pregnancy probably increases the penetrance of HNF-1alpha mutations. The potential role of exposure to hyperglycemia in utero in a monogenic diabetic subgroup warrants prospective study.     

Beta-cell dysfunction in classic transient neonatal diabetes is characterized by impaired insulin response to glucose but normal response to glucagon

OBJECTIVE: To investigate beta-cell function and the long-term health of four case subjects presenting with chromosome 6-associated transient neonatal diabetes (TND). RESEARCH DESIGN AND METHODS: Two unrelated case subjects presenting with paternal uniparental isodisomy of chromosome 6 (UPD6) and two siblings with a paternally inherited duplication of 6q24 were studied. Three case subjects presented with neonatal diabetes that recurred at 4-17 years, while diabetes was incidentally discovered in the other case subject at 14 years of age. beta-Cell function was investigated after diabetes relapse by means of an oral glucose tolerance test (OGTT), an intravenous glucose tolerance test (IVGTT), and glucagon tests. The quantitative insulin sensitivity check index (QUICKI) was calculated from fasting blood samples as an estimate of insulin sensitivity. RESULTS: beta-Cell function was investigated at diabetes relapse in two case subjects: the insulin response to both an OGTT and IVGTT was low, whereas the basal levels of C-peptide were normal. No evidence of insulin resistance was found. Residual beta-cell function was further explored by a glucagon test in all subjects at the age of 16-28 years and was found to be normal. Final height was within the normal percentiles, whereas one case, who had been poorly controlled since puberty, presented with diabetes-related microvascular complications. CONCLUSIONS: In patients with chromosome 6-associated TND, the beta-cell is preserved and able to secrete insulin through the stimulatory G protein pathway while exhibiting a specific defect of insulin secretion after glucose stimulation. This form of diabetes can be managed with insulin or diet, although new therapeutic agents (glucagon-like synthetic analogs) may prove useful in the future. Lack of treatment leads to long-lasting hyperglycemia without the risk of ketoacidosis but associated with microangiopathy in adult life.     

Mitochondrial regulation by c-Myc and hypoxia-inducible factor-1 alpha controls sensitivity to econazole

Econazole is an azole antifungal with anticancer activity that blocks Ca(2+) influx and stimulates endoplasmic reticulum (ER) Ca(2+) release through the generation of mitochondrial reactive oxygen species (ROS), resulting in sustained depletion of ER Ca(2+) stores, protein synthesis inhibition, and cell death. c-Myc, a commonly activated oncogene, also promotes apoptosis in response to growth factor withdrawal and a variety of chemotherapeutic agents. We have investigated the role of c-myc in regulating sensitivity to econazole. Here, we show that c-myc-negative cells are profoundly resistant to econazole. c-Myc-negative rat fibroblasts failed to generate mitochondrial ROS in response to econazole and consequently failed to deplete the ER of Ca(2+). HL60 cells knocked down for c-myc expression also displayed decreased ROS generation and decreased econazole sensitivity. Addition of H(2)O(2) restored sensitivity to econazole in both c-myc-negative rat fibroblasts and c-myc knocked-down HL60 cells, supporting a role for ROS in cell death induction. c-Myc-negative cells and HL60 cells knocked down for c-myc have reduced mitochondrial content compared with c-myc-positive cells. The hypoxia sensor, hypoxia-inducible factor-1alpha (HIF-1alpha), interacts antagonistically with c-myc and also regulates mitochondrial biogenesis. Knockdown of HIF-1alpha in c-myc-negative cells increased mitochondrial content restored ROS generation in response to econazole and increased sensitivity to the drug. Taken together, these results show that c-myc and HIF-1alpha regulate sensitivity to econazole by modulating the ability of the drug to generate mitochondrial ROS.