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.     

Genetic evidence that HNF-1alpha-dependent transcriptional control of HNF-4alpha is essential for human pancreatic beta cell function

Mutations in the genes encoding hepatocyte nuclear factor 4alpha (HNF-4alpha) and HNF-1alpha impair insulin secretion and cause maturity onset diabetes of the young (MODY). HNF-4alpha is known to be an essential positive regulator of HNF-1alpha. More recent data demonstrates that HNF-4alpha expression is dependent on HNF-1alpha in mouse pancreatic islets and exocrine cells. This effect is mediated by binding of HNF-1alpha to a tissue-specific promoter (P2) located 45.6 kb upstream from the previously characterized Hnf4alpha promoter (P1). Here we report that the expression of HNF-4alpha in human islets and exocrine cells is primarily mediated by the P2 promoter. Furthermore, we describe a G --> A mutation in a conserved nucleotide position of the HNF-1alpha binding site of the P2 promoter, which cosegregates with MODY. The mutation results in decreased affinity for HNF-1alpha, and consequently in reduced HNF-1alpha-dependent activation. These findings provide genetic evidence that HNF-1alpha serves as an upstream regulator of HNF-4alpha and interacts directly with the P2 promoter in human pancreatic cells. Furthermore, they indicate that this regulation is essential to maintain normal pancreatic function.     

'I don't feel like a diabetic any more': the impact of stopping insulin in patients with maturity onset diabetes of the young following genetic testing

Hepatocyte nuclear factor-1alpha (HNF-1alpha) maturity onset diabetes of the young (MODY) is the commonest cause of monogenic diabetes but is frequently misdiagnosed as type 1 diabetes. The availability of genetic testing in MODY has improved diagnosis. Sulphonylurea sensitivity in HNF-1alpha patients means that those on insulin from diagnosis can transfer to sulphonylureas and may improve glycaemic control. To gain insight into the implications for patients of stopping insulin, in-depth interviews were conducted with eight HNF-1alpha patients transferred to sulphonylureas after a median of 20 years on insulin. Thematic content analysis highlighted four key themes: Fear, anxiety and excitement regarding stopping insulin, particularly among those who had been on insulin for many years or had never omitted insulin in the past. Improved lifestyle and self image accompanied by feelings of relief and 'increased normality'. Reflections on their time on insulin, including feelings of annoyance, particularly when the need for insulin treatment had been questioned at diagnosis. Difficulty 'letting go' of insulin treatment--some patients found it hard to believe that they no longer required injections as this conflicted with messages previously received from healthcare professionals. Transferring from insulin to sulphonylureas had a positive impact on lifestyle but support was needed for patients to adjust, many having grown up with the belief they would be on insulin for life.     

Mutations in the genes for hepatocyte nuclear factor (HNF)-1alpha, -4alpha, -1beta, and -3beta; the dimerization cofactor of HNF-1; and insulin promoter factor 1 are not common causes of early-onset type 2 diabetes in Pima Indians

OBJECTIVE: Maturity-onset diabetes of the young (MODY) is a genetically heterogeneous subtype of type 2 diabetes characterized by an early age at onset and autosomal dominant inheritance. MODY can result from heterozygous mutations in at least five genes. The purpose of this study was to determine whether alterations in known MODY genes and two MODY candidate genes contribute to the development of early-onset type 2 diabetes in Pima Indians. RESEARCH DESIGN AND METHODS: The coding regions of the known MODY genes hepatocyte nuclear factor (HNF)-1alpha, HNF-4alpha, HNF-1beta, and insulin promoter factor 1 and the coding regions of two MODY candidate genes, HNF-3beta and the dimerization cofactor of HNF-1, were sequenced in genomic DNA from Pima Indians. The primary "affected" study population consisted of 46 Pima Indians whose age at onset of type 2 diabetes was < or =20 years. DNA sequence variants identified in the affected group were then analyzed in a group of 80 "unaffected" Pima Indians who were at least 40 years old and had normal glucose tolerance. RESULTS: A total of 11 polymorphisms were detected in these genes. However, none of the polymorphisms differed in frequency among Pima Indians with an early age at onset of diabetes compared with older Pima Indians with normal glucose tolerance. CONCLUSIONS: Mutations in these known MODY or MODY candidate genes are not a common cause of early-onset diabetes in Pima Indians.     

Identifying hepatic nuclear factor 1alpha mutations in children and young adults with a clinical diagnosis of type 1 diabetes

OBJECTIVE: HNF-1alpha gene mutations (MODY3) present with marked hyperglycemia in lean young adults and may, therefore, be mistaken for type 1 diabetes, with implications for individual treatment and risk of diabetes in other family members. We examined the prevalence of HNF-1alpha mutations in families with three generations of diabetes identified in a population-based study of childhood diabetes, representing a subpopulation in which misclassification was likely. RESEARCH DESIGN AND METHODS: In a study population of 1,470 families, 36 families (2.4%) with three affected generations were identified. In the 18 families in whom DNA samples were available, islet autoantibody testing, HLA class II genotyping, and HNF-1alpha sequencing were performed. RESULTS: At least one islet autoantibody was found in 13 of 14 probands, and diabetes-associated HLA class II haplotypes were found in 17 of 18. One proband, who had no islet autoantibodies and was homozygous for the protective HLA haplotype DRB1*02-DQB1*0602, had a novel HNF-1alpha heterozygous nonsense mutation (R54X). This mutation cosegregated with diabetes in the family. The proband, his brother, mother, and maternal grandmother were diagnosed with type 1 diabetes aged 14-18 years and treated with insulin (0.39-0.74 units/kg) from diagnosis. The mother has since been successfully transferred to sulfonylurea treatment. CONCLUSIONS: Family history alone is of limited value in identification of individuals with HNF-1alpha mutations, and we propose a stepwise approach that restricts sequencing of the HNF-1alpha gene to those with a family history of diabetes who also test negative for islet autoantibodies.     

Adult-onset atypical (type 1) diabetes: additional insights and differences with type 1A diabetes in a European Mediterranean population

OBJECTIVE: In 1997, the American Diabetes Association proposed two subcategories for type 1 diabetes: type 1A or immunomediated diabetes and type 1B or idiopathic diabetes characterized by negative beta-cell autoimmunity markers, lack of association with HLA, and fluctuating insulinopenia. The aim of this study was to examine clinical characteristics, beta-cell function, HLA typing, and mutations in maturity-onset diabetes of the young (MODY) genes in patients with atypical type 1 diabetes (type 1 diabetes diagnosed at onset, without pancreatic autoantibodies and fluctuating insulinopenia). RESEARCH DESIGN AND METHODS: Eight patients with atypical type 1 diabetes (all men, 30.7 +/- 7.6 years) and 16 newly diagnosed age- and sex-matched patients with type 1A diabetes were studied retrospectively. Islet cell, GAD, tyrosine phosphatase and insulin antibodies, and basal and stimulated plasma C-peptide were measured at onset and after 1 year. HLA-DRB1-DQA1-DQB1 typing and screening for mutations in the HNF-1alpha and HNF-4alpha genes were performed from genomic DNA. RESULTS: Atypical patients displayed significantly higher BMI and better beta-cell function at onset and after 12 months. Three patients carried protective or neutral type 1 diabetes haplotypes, five patients displayed heterozygosity for susceptible and protective haplotypes, and seven patients showed Asp(beta57). We found a nondescribed variant Pro436Ser in exon 10 of the HNF-4alpha gene in one atypical patient without susceptible haplotypes. CONCLUSIONS: In our population, there are atypical forms of young adult-onset ketosis-prone diabetes initially diagnosed as type 1 diabetes, differing from type 1 diabetes in the absence of beta-cell autoimmunity, persistent beta-cell function capacity, fluctuating insulin requirements and ketosis-prone episodes, as well as clinical features of type 2 diabetes. Only one subgroup could be strictly classified as having type 1B diabetes. Additional information is still needed to improve our understanding of the mechanisms that finally lead to the disease.     

Insulin gene in diabetes. Analysis through RFLP

Insulin deficiency is a prominent feature of non-insulin-dependent (NIDDM) and insulin-dependent (IDDM) diabetes mellitus that could result from defects in the insulin gene. Cloning of this gene has permitted molecular-genetic studies including the definition of multiple-DNA-sequence polymorphisms detected with restriction endonucleases, or restriction-fragment-length polymorphisms (RFLPs), and the mapping of the insulin gene to the short arm of chromosome 11 adjacent to the insulinlike growth factor II (IGF-II) and tyrosine hydroxylase genes. The combined RFLPs for the insulin, IGF-II, and tyrosine hydroxylase genes make this a highly informative locus for genetic studies of the insulin gene in diabetes. Early studies of an RFLP consisting of variable-number tandem repeats (VNTR) of DNA near the insulin gene suggested an association of certain alleles with approximately 170 copies of the repeat unit with NIDDM. Although subsequent studies in NIDDM did not confirm this association, an association of different alleles defined by approximately 40 copies of the repeat unit in this VNTR region with IDDM has been demonstrated in multiple studies. This VNTR region and the multiple other RFLPs for this region have been used in linkage analysis to study the segregation of insulin genes in families. These studies have failed to demonstrate a major significant role for insulin-gene defects in NIDDM, maturity-onset diabetes of the young, or IDDM in American Blacks and Whites and under various models of inheritance. Several pedigrees with diabetes and defects of the insulin gene have been described, however, and a minor role for this gene in NIDDM cannot be eliminated from available studies. Similarly, the association studies of the insulin gene and IDDM suggest a minor modifying role undetectable in pedigree studies. The role of defects in or near the insulin gene in a small subset of NIDDM or in IDDM must await direct investigation of the insulin gene in diabetic individuals with the most recent methods for gene amplification and sequence analysis.     

Etiological investigation of diabetes in young adults presenting with apparent type 2 diabetes

OBJECTIVE: Young adults with newly diagnosed apparent type 2 diabetes present the clinician with a wide differential diagnosis of possible etiology, including autoimmune and genetic causes as well as young-onset type 2 diabetes (YT2D). The characteristics of these groups have been described, but it is not known in which subjects investigation for etiology may be beneficial. RESEARCH DESIGN AND METHODS: A total of 268 unselected U.K. Caucasian subjects diagnosed at ages 18-45 years and not treated with permanent insulin for < or =6 months were studied. All subjects underwent clinical assessment and screening for GAD antibodies (GADA) and tyrosine phosphatase IA-2 antibodies (IA-2A). Screening for a common mutation in the hepatocyte nuclear factor-1 alpha (HNF-1 alpha) gene and the common mitochondrial mutation was performed in the antibody-negative subjects. Subjects without insulin resistance were selected for sequencing of the HNF-1 alpha gene. RESULTS: A specific etiology was defined in 11.6% of the 268 subjects and in 24.7% of the lean subjects. Twenty-six subjects (9.7%) were positive for a beta-cell antibody, one subject had familial partial lipodystrophy and the lamin A/C mutation R482W, and two subjects had the mitochondrial mutation A3243G. Two of 15 selected subjects had HNF-1 alpha mutations, the novel missense mutation A501T, and the previously reported R583Q. CONCLUSIONS: This unselected series shows that there is considerable heterogeneity in apparent YT2D. beta-Cell autoantibodies should be performed in all those presenting at ages 18-45 years. Genetic investigations can be targeted to phenotypically defined subjects. The finding of a specific etiology will allow individualization of management and give patients valuable information about their condition.     

Pathogenesis of impaired insulin secretion in NIDDM

The most important role of pancreatic beta-cells is the insulin secretion responding to the plasma glucose level. Molecular biological and electrophysiological approaches have been revealing the molecular mechanism of glucose-stimulated insulin secretion. A lot of key molecules of the systems, including GLUT2, glucokinase, SUR1, Kir6.2 and CD38, have been cloned and characterized whether the mutations in these genes are responsible for the pathogenesis of non-insulin-dependent diabetes mellitus (NIDDM). In this paper, we summarized the recent advances concerning pathogenesis of NIDDM in respect of impaired insulin secretion from pancreatic beta-cells.     

Beta-cell dysfunction, insulin sensitivity, and glycosuria precede diabetes in hepatocyte nuclear factor-1alpha mutation carriers

OBJECTIVE: Patients with diabetes due to hepatocyte nuclear factor (HNF)-1alpha mutations have beta-cell deficiency, insulin sensitivity, altered proinsulin levels, and a low renal threshold for glucose. It is uncertain how many of these features precede the development of diabetes. The aim of our study was to test for these characteristics in young nondiabetic HNF-1alpha mutation carriers. RESEARCH DESIGN AND METHODS: A total of 47 offspring from 19 extended families underwent genetic testing, a standard oral glucose tolerance test, and urine testing. RESULTS: HNF-1alpha mutations were found in 20 offspring, 7 with diabetes and 13 without diabetes. The 13 nondiabetic mutation carriers were compared with 27 family control subjects, who were matched for age, sex, and BMI. There was marked beta-cell deficiency with reduced insulinogenic index (53.5 [31.5-90.9] vs. 226.0 [126.0-407.1], SD [range], P < 0.001) and area under the curve for insulin (P < 0.001). Insulin sensitivity was increased in mutation carriers (homeostatic model assessment of insulin sensitivity 144.6 [82.7-252.7] vs. 100 [66.9-149.4], P = 0.025). A total of 38% of mutation carriers had glycosuria at 2 h compared with 0% of control subjects (P = 0.0034). Those with glycosuria had peak glucose values that were higher than the mutations carriers without glycosuria (range 8.1-11.8 vs. 6.2-8.4 mmol/l, P = 0.002). The seven subjects with diabetes all showed glycosuria. CONCLUSIONS: We conclude that marked beta-cell deficiency, increased insulin sensitivity, and a low renal threshold are present in young nondiabetic HNF-1alpha mutation carriers. The presence of glycosuria post-glucose load could be used to screen children of mutation carriers as it occurs in all mutation carriers with a peak glucose in the oral glucose tolerance test >8.4 mmol/l.     

The prevalence of the HNF-1alpha G319S mutation in Canadian aboriginal youth with type 2 diabetes

OBJECTIVE: To investigate the prevalence of the unique HNF-1alpha G319S mutation in a population of aboriginal youth with type 2 diabetes and to describe the relationship between clinical and historical characteristics and the presence or absence of the HNF-1alpha G319S mutation. RESEARCH DESIGN AND METHODS: Participating youth were genotyped for the G319S mutation of the HNF-1alpha gene. Clinical, laboratory, and historical data were collected via chart review (blinded to genotype results). Comparison data were derived from another study involving young nondiabetic pregnant aboriginal women. RESULTS: A total of 51 youth seen sequentially in a type 2 diabetes clinic participated in this study. Of these, 21 (41.2%) had at least one copy of the mutant allele. The allele frequency in the study population was 0.29 (95% CI 0.20-0.38), which was significantly different from the allele frequency of 0.13 in the comparison population (chi(2) = 6.78, P = 0.009). The frequency of the homozygous mutation (S319/S319) was 0.18. Mean BMI was significantly lower (P = 0.002), mean HbA(1c) was significantly higher (P = 0.02), and acanthosis nigricans was significantly less frequent (P = 0.004) in those with the mutation compared with the wild type. Mean insulin levels were lower and insulin sensitivity (assessed by homeostasis model assessment [HOMA]) was greater in the homozygote group compared with the wild-type group (P = 0.002 and P = 0.0007, respectively). A dose-dependent gradient was observed for these characteristics. CONCLUSIONS: These data support the association between the HNF-1alpha G319S mutation and early-onset type 2 diabetes in this population. Those with the mutation lacked clinical characteristics of insulin resistance (e.g., obesity and acanthosis nigricans) and had lower insulin levels, suggesting that an insulin-secretory and/or -production defect plays an important role in the development of diabetes in this group. Further investigation of the pathophysiology of the S319 homo- and heterozygote is needed because it may impact treatment and/or prevention of this disease.     

Familial early-onset type 2 diabetes in Chinese patients: obesity and genetics have more significant roles than autoimmunity

OBJECTIVE: We examined the prevalence of different forms of diabetes in Hong Kong Chinese patients with familial early-onset type 2 diabetes and compared their clinical features with patients with familial late-onset type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 145 young patients with early-onset diabetes (age and age at diagnosis < or = 40 years) and a family history of diabetes were studied. They were screened for mutations in the genes encoding glucokinase, hepatocyte nuclear factor (HNF)-4alpha, and HNF-1alpha. The mitochondrial DNA A-->G at nucleotide 3243 (mt3243) and amyLin S20G mutations were studied, and antibodies to GAD (anti-GADs) were also examined. RESULTS: The prevalence of putative diabetogenic gene mutations and autoimmune markers were 4% for glucokinase, 0% for HNF-4alpha, 5% for HNF-1alpha, 3% for mt3243, 2% for amylin 520G, and 4% for anti-GAD. Compared with late-onset patients, the patients with early-onset diabetes had a higher prevalence of a parental history of diabetes and were generally more obese. When classified by obesity indexes (BMI and waist circumference), the obese patients, especially those with early-onset diabetes, had a clustering of cardiovascular risk factors and increased rates of retinopathy and albuminuria. CONCLUSIONS; Genetic factors (up to 14%) and obesity (55%) play more significant roles than autoimmunity (4%) in familial type 2 diabetes in young Chinese patients. The significance of obesity-related genes and other gene-gene and gene-environment interactions in these young patients remains to be determined.     

Determinants of the development of diabetes (maturity-onset diabetes of the young-3) in carriers of HNF-1alpha mutations: evidence for parent-of-origin effect

OBJECTIVE: To determine the distribution of the age at onset of diabetes (maturity-onset diabetes of the young-3 [MODY3]) and to identify determinants of the onset of diabetes in carriers of HNF-1alpha mutations. RESEARCH DESIGN AND METHODS: Extended families (n = 104) with type 2 diabetes inherited in a dominant pattern were recruited and screened for diabetes-causing mutations in HNF-1alpha. RESULTS: HNF-1alpha mutations cosegregated with diabetes in only 13 families, all with a mean age at onset <35 years. Insulin secretion was diminished or absent in mutation carriers (n = 101), and diabetes developed in 65% by age 25 years and in 100% by age 50 years. If the mutation was inherited from the mother, diabetes onset was very young in those exposed to diabetes in utero; 57 +/- 8% were affected by age 15 years as compared with 0.0% in those not exposed (P < 7 x 10(-6)). By age 25 years, the difference was reduced (85 +/- 6 and 55 +/- 12%, respectively; P = 0.02). If the mutation was inherited from the father, diabetes developed in 52 +/- 8% by age 25 years. Age at diagnosis was shown to be highly heritable (h(2) = 0.47, P = 0.003). When parent of origin was included in the analyses, the magnitude of genetic contribution increased markedly (h(2) = 0.91). CONCLUSIONS: Mutations in HNF-1alpha accounts for diabetes in a small proportion of families with a dominant pattern of inheritance. Age at onset of diabetes in MODY3 families varied widely and was influenced by familial factors (including modifying genes) and parent of origin (whether a mutation carrier was exposed to diabetes in utero).     

Cardiovascular Risk Factors and the Prediabetic Syndrome

Cardiovascular disease is increased 2- to 4-fold in non-insulin-dependent diabetes mellitus (NIDDM); yet in most studies, there is a relatively weak relationship between the frequency of coronary heart disease (CHD) and the duration of diabetes and severity of hyperglycaemia. A number of authors have suggested that the prediabetic stage may contribute to the risk of CHD in NIDDM. Hyperinsulinaemia and insulin resistance have been strongly associated with the development of NIDDM. Data are less conclusive about the relationship of hyperinsulinaemia to the development of CHD in nondiabetic subjects. Relatively little data are available on hyperinsulinaemia and/or insulin resistance to CHD in NIDDM subjects. Tight control of glycaemia with exogenous insulin improves cardiovascular risk factors in NIDDM subjects and therefore is unlikely to increase the risk of CHD. Although the relation of insulin to CHD in the general population is somewhat controversial, insulin is clearly related to multiple cardiovascular risk factors (especially elevated triglyceride, decreased high-density lipoprotein, small dense low-density lipoprotein, impaired glucose tolerance and increased plasminogen activator inhibitor 1 (PAI-1)). However, the relation of insulin resistance to hypertension remains controversial.     

The Etiology and Pathogenesis of Non-insulin-dependent Diabetes

Although environmental factors are important triggers of non-insulin-dependent diabetes mellitus (NIDDM), heredity plays a major role in the pathogenesis of the disease. Insulin resistance manifested as impaired activation of glycogen synthase and thereby storage of glucose as glycogen in skeletal muscle is demonstrable early on in NIDDM relatives, suggesting that NIDDM could be an inherited muscle disease. On the other hand, insulin deficiency is almost unequivocally present before manifest diabetes develops. An Intensive search for candidate genes for NIDDM has been initiated; so far it has not been possible to ascribe NIDDM to any alterations in the human genome. Given the heterogenous nature of NIDDM, its age-dependent penetrance and strong influence of environmental factors, it may not be fruitful to use NIDDM as an end-point in genetic linkage or association studies. It is more likely that DNA defects result in either insulin resistance or insulin deficiency, which in turn, can both lead to NIDDM.

In accordance with the thrifty gene hypothesis, the insulin resistance gene has protected individuals during long periods of starving by storing energy as fat rather than as glycogen in muscle. The abundance of food in Western society has made this once protective gene a deleterious one, suggesting that these individuals are not equipped with the metabolic machinery to handle overeating.