help button home button Endocrine Society JCEM
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Urhammer, S. A.
Right arrow Articles by Pedersen, O.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Urhammer, S. A.
Right arrow Articles by Pedersen, O.
The Journal of Clinical Endocrinology & Metabolism Vol. 83, No. 12 4506-4509
Copyright © 1998 by The Endocrine Society

Original Studies

The Ala/Val98 Polymorphism of the Hepatocyte Nuclear Factor-1{alpha} Gene Contributes to the Interindividual Variation in Serum C-Peptide Response during an Oral Glucose Tolerance Test: Evidence from Studies of 231 Glucose-Tolerant First Degree Relatives of Type 2 Diabetic Probands1

Søren A. Urhammer, Torben Hansen, Claus T. Ekstrøm, Hans Eiberg and Oluf Pedersen

Steno Diabetes Center and Hagedorn Research Institute (S.A.U., T.H., C.T.E., O.P.), and University Institute of Medical Biochemistry and Genetics, Department of Medical Genetics, University of Copenhagen (H.E.), Copenhagen, Denmark

Address all correspondence and requests for reprints to: Søren Urhammer, M.D., Steno Diabetes Center, Niels Steensens Vej 2, DK-2820 Gentofte, Copenhagen, Denmark.


   Abstract
Top
 Abstract
Introduction
 Subjects and Methods
 Results
 Discussion
 References
 
The third form of maturity-onset diabetes of the young is caused by mutations in the hepatocyte nuclear factor-1{alpha} gene. Recently, we demonstrated an association between a prevalent polymorphism at codon 98, Ala/Val98, of this gene and a 20% decreased insulin release during an oral glucose tolerance test (OGTT) in middle-aged glucose-tolerant Danish Caucasian subjects. The major objective of the present study was to replicate this finding among glucose-tolerant first degree relatives of type 2 diabetic patients of the same ethnic origin. All participants, 231 glucose-tolerant offspring of 62 type 2 diabetic probands, underwent an OGTT with measurements of plasma glucose, serum insulin, and serum C peptide during the test. Thirty-three heterozygous carriers of the Ala/Val variant were identified, whereas no subjects had the variant in its homozygous form. Ala/Val carriers had a 20% reduction in serum C peptide at 30 min during the OGTT (1225 ± 636 vs. 1507 ± 624 pmol/L; P = 0.02) compared to wild-type carriers. No significant differences in serum insulin levels during the OGTT were observed between carriers of the variant and Ala/Ala homozygotes. In conclusion, among Danish glucose-tolerant first degree relatives of type 2 diabetic patients the Ala/Val98 polymorphism of the hepatocyte nuclear factor-1{alpha} gene is associated with a decreased serum C-peptide secretion during an OGTT. This finding confirms our previously reported observation of the functional importance of the variant to insulin secretion during an OGTT among middle-aged healthy subjects.


   Introduction
Top
 Abstract
 Introduction
Subjects and Methods
 Results
 Discussion
 References
 
MATURITY-ONSET diabetes of the young (MODY) is a subset of type 2 diabetes showing an autosomal dominant mode of inheritance, an early age of disease onset, and an impairment of ß-cell function. To date, five MODY genes have been identified: MODY1 is caused by mutations in the hepatocyte nuclear factor-4{alpha} (HNF-4{alpha}) gene (1), MODY2 is caused by mutations in the glucokinase gene (2), MODY3 is caused by mutations in the HNF-1{alpha} gene (3), MODY4 is caused by mutations in the insulin promoter factor gene (4), and MODY5 is caused by mutations in the HNF-1ß gene (5). Mutations in the HNF-1{alpha} gene are a common cause of MODY among Caucasians (6, 7). The HNF-1{alpha} gene might, however, also be implicated in the more common late-onset form of type 2 diabetes. Evidence of linkage between the MODY3 region on chromosome 12 and common type 2 diabetes has been obtained in a subset of Finnish type 2 diabetic families with low insulin secretion (8) and in Caucasian type 2 diabetic sibling pairs with a history of diabetic nephropathy (9). By performing mutational analysis of the HNF-1{alpha} gene in patients with late-onset type 2 diabetes, we previously found that the gene is not a frequent cause of the common form of type 2 diabetes in Danish Caucasians (10). However, we observed that middle-aged healthy heterozygous carriers of a prevalent amino acid polymorphism, Ala/Val98, exhibit a 20% reduction in serum C peptide and insulin responses at 30 min during an oral glucose tolerance test (OGTT) compared to wild-type carriers, suggesting that this amino acid replacement might influence ß-cell function during an oral glucose challenge (11). Hence, the objective of the present study was an attempt to reproduce our recent findings in an independent sample of Danish Caucasian ancestry.


   Subjects and Methods
Top
 Abstract
 Introduction
 Subjects and Methods
Results
 Discussion
 References
 
Subjects

Two hundred and sixty-seven offspring of type 2 diabetic probands from 62 families were recruited from the Danish family resource bank at the Department of Human Genetics, University of Copenhagen (Copenhagen, Denmark; 18 families), and from the out-patient clinic at Steno Diabetes Center (Copenhagen, Denmark) (44 families). The families were ascertained through one type 2 diabetic proband with 4 or more nondiabetic offspring. All were Danish Caucasians by self-identification. All participants underwent a 75g glucose OGTT with measurements of plasma glucose, serum insulin, and serum C peptide. Type 2 diabetes was diagnosed in accordance with 1985 WHO criteria. As diabetes may cause secondary changes in insulin secretion, only glucose-tolerant offspring were studied (n = 231). Before participation in the study, informed consent was obtained from all subjects studied. The study was approved by the ethical committee of Copenhagen and was in accordance with the principles of the Declaration of Helsinki II.

Biochemical variables

The concentration of plasma glucose was measured employing an automated glucose oxidation method (Granutest, Merck, Darmstadt, Germany). The concentration of specific insulin in serum was measured by enzyme-linked immunosorbent assay with a narrow specificity excluding des(31,32)- and intact proinsulin (enzyme-linked immunosorbent assay) (12), and the concentration of serum C peptide was determined by RIA (13) employing the polyclonal antibody M1230 (14, 15)

Screening for the amino acid polymorphism in the HNF- 1{alpha} gene

The Ala/Val98 polymorphism was detected using PCR-restriction fragment length polymorphism as previously described (11). This assay was effective in 229 of the available 231 patient cases.

Statistical analysis

Differences between groups of subjects were tested with Student’s t test, Mann-Whitney rank sum test or the {chi}2 analysis where appropriate. The variables [except for gender distribution, age, and body mass index (BMI)] were also analyzed applying a variance component model (random effects model) (16), where an extra source of variation is allowed to account for the fact that individuals from the same family might be correlated. The variant of interest and gender were included as fixed variables, age and BMI (in the analyses of serum insulin and serum C peptide at 30 min during the OGTT, the fasting level of plasma glucose was also included) were included as covariates, and family was included as a random effect. Data are expressed as medians (interquartile ranges). P < 0.05 (two-tailed) was considered significant.


   Results
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
Discussion
 References
 
Among 229 (of 231) glucose-tolerant offspring, we identified 33 heterozygous carriers of the Ala/Val98 substitution from 12 families and 196 wild-type carriers. No homozygous carriers were found. From Table 1Go it appears that Ala/Val98 carriers did not differ from wild-type carriers with respect to age, BMI, gender, or fasting levels of plasma glucose, serum insulin, and C-peptide. Despite the fact that Ala/Val98 heterozygotes and wild-type carriers had equal plasma glucose levels during the OGTT, carriers of the polymorphism had a 20% decrease in the serum C-peptide level at 30 min (P = 0.02) during the OGTT compared to wild-type carriers (Table 1Go and Fig. 1Go). The area under the serum C-peptide curve during 120 min of the OGTT was also reduced among Ala/Val98 carriers (P = 0.02; Table 1Go and Fig. 1Go). The variance component model (16), which allows us to model the familiarity correlation among siblings from the same family and at the same time to adjust for differences in confounding variables such as gender, age, BMI, and plasma glucose, revealed that Ala/Val98 heterozygotes had a significantly (P = 0.03) decreased serum C peptide level at 30 min during the oral glucose challenge (Table 1Go). No significant differences were observed between the two genotype groups with respect to serum insulin at 30 min or the area under the insulin curve during the 120 min of the OGTT using direct comparisons (Student’s t test or Mann-Whitney test) or the variance component model.


View this table:
[in this window]
[in a new window]
  		Table 1. Clinical and biochemical data of 229 glucose-tolerant offspring of 62 type 2 diabetic probands classified according to the genotype of the Ala/Val98 polymorphism of the hepatocyte nuclear factor-1{alpha} gene

 


View larger version (16K):
[in this window]
[in a new window]
  		Figure 1. Plasma glucose (A), serum insulin (B), and serum C peptide (C) levels during 120 min of the OGTT in 229 glucose-tolerant offspring of type 2 diabetic probands. Filled squares depict wild-type carriers, and filled triangles depict heterozygous carriers of the Ala/Val98 polymorphism of the HNF-1{alpha} gene. Values are medians (SE).

 

   Discussion
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
References
 
As the present study revealed approximately the same decrease, i.e. about 20%, in the same estimate of insulin secretion (serum C-peptide at 30 min during the OGTT) as we recently reported in randomly recruited middle-aged healthy carriers of the Ala/Val98 polymorphism, the primary hypothesis of this study is confirmed (11). The power to detect this 20% effect of the variant in the present study is more than 90%, indicating a low risk of a statistical type 1 error. In contrast to our previous observations, the present data did not reveal any significant effect of the variant on serum insulin levels during the OGTT. C-peptide is secreted equimolar with insulin from the ß-cells. However, where circulating C-peptide has a negligible hepatic extraction and a constant renal clearance, the hepatic insulin extraction may vary under different physiological conditions (17). Thus, alterations in peripheral serum C peptide levels might reflect alterations in insulin secretion more precisely than peripheral insulin levels, making serum C peptide a more valid marker of insulin secretion than serum insulin. The HNF-1{alpha} gene has a wide tissue distribution, including pancreatic islets, liver, and small intestine (18). Defects in the HNF-1{alpha} gene influencing the function of the protein might therefore affect glucose homeostasis at several organ levels, including small intestine, liver, and pancreatic ß-cells. The Ala98 residue of HNF-1{alpha} is located within a region of the DNA-binding domain of the protein conferring sequence specificity (18), and this amino acid is conserved in human, rat, mouse, and chicken (19), indicating a functional importance of the residue, which needs to be further elucidated in transfection experiments. In the present study an 8-fold interindividual variation in the serum C peptide level at 30 min was observed comparable with the variation estimated previously (11). How much of this variation is genetically determined is at present unknown. Taken together, it appears that the codon 98 polymorphism of HNF-1{alpha} may explain some of the genetically determined interindividual variation in serum C peptide responses to an oral glucose challenge.


   Acknowledgments
 
The authors thank Sandra Urioste, Annemette Forman, Lene Aabo, Bente Mottlau, Susanne Kjellberg, Lis Ølholm, and Marja Lis Halkjær for dedicated and careful technical assistance, and Grete Lademann for secretarial support.


   Footnotes
 
1 This work was supported by grants from the University of Copenhagen, the Velux Foundation, the Danish Diabetes Association, the Danish Medical Research Council, and EEC (BMH4-CT-950662). Back

Received August 12, 1998.

Accepted September 16, 1998.


   References
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
 References
 

  1. Yamagata K, Furuta H, Oda N, et al. 1996 Mutations in the hepatocyte nuclear factor-4{alpha} gene in maturity-onset diabetes of the young (MODY1). Nature. 384:458–460.[CrossRef][Medline]
  2. Vionnet N, Stoffel M, Takeda J, et al. 1992 Nonsense mutation in the glucokinase gene causes early-onset non-insulin-dependent diabetes mellitus. Nature. 356:721–722.
  3. Yamagata K, Oda N, Kaisaki PJ, et al. 1996 Mutations in the hepatocyte nuclear factor-1alpha gene in maturity-onset diabetes of the young (MODY3). Nature. 384:455–458.[CrossRef][Medline]
  4. Stoffers DA, Ferrer J, Clarke WL, Habener JF. 1997 Early-onset type-2 diabetes-mellitus (MODY4) linked to ipf1. Nat Genet. 17:138–139.[CrossRef][Medline]
  5. Horikawa Y, Iwasaki N, Hara M, et al. 1997 Mutation in hepatocyte nuclear factor-1-beta gene (TCF2) associated with MODY. Nat Genet. 17:384–385.[CrossRef][Medline]
  6. Hattersley AT. 1998 Maturity-onset diabetes of the young: clinical heterogeneity explained by genetic heterogeneity. Diabetic Med. 15:15–24.[CrossRef][Medline]
  7. Hansen T, Eiberg H, Rouard M, et al. 1997 Novel MODY3 mutations in the hepatocyte nuclear factor-1alpha gene: evidence for a hyperexcitability of pancreatic beta-cells to intravenous secretagogues in a glucose-tolerant carrier of a P447L mutation. Diabetes. 46:726–730.[Abstract]
  8. Mahtani MM, Widen E, Lehto M, et al. 1996 Mapping of a gene for type 2 diabetes associated with an insulin secretion defect by a genome scan in Finnish families. Nat Genet. 14:90–94.[CrossRef][Medline]
  9. Bowden DW, Sale M, Howard TD, et al. 1997 Linkage of genetic-markers on human-chromosome-20 and human-chromosome-12 to NIDDM in Caucasian sib pairs with a history of diabetic nephropathy. Diabetes. 46:882–886.[Abstract]
  10. Urhammer SA, Rasmussen SK, Kaisaki PJ, et al. 1997 Genetic variation in the hepatocyte nuclear factor-1{alpha} gene in Danish Caucasians with late-onset NIDDM. Diabetologia.40:473–475.
  11. Urhammer SA, Fridberg M, Hansen T, et al. 1997 A prevalent amino acid polymorphism at codon 98 in the hepatocyte nuclear factor-1{alpha} gene is associated with reduced serum C-peptide and insulin responses to an oral glucose challenge. Diabetes. 46:912–916.[Abstract]
  12. Andersen L, Dinesen B, Jorgensen PN, Poulsen F, Roder ME. 1993 Enzyme immunoassay for intact human insulin in serum or plasma. Clin Chem. 39:578–582.[Abstract/Free Full Text]
  13. Heding LG. 1977 Specific and direct radioimmunoassay for human proinsulin in serum. Diabetologia. 13:467–474.[CrossRef][Medline]
  14. Faber OK, Binder C, Markussen J, et al. 1978 Characterization of seven C-peptide antisera. Diabetes. 27(Suppl 1):170–177.
  15. Faber OK, Markussen J, Naithani VK, Binder C. 1976 Production of antisera to synthetic benzyloxycarbonyl-C-peptide of human proinsulin. Hoppe- Seylers Z Physiol Chem. 357:751–757.[Medline]
  16. Hopper JL, Mathews JD. 1982 Extensions to multivariate normal models for pedigree analysis. Ann Hum Genet. 46:373–383.
  17. Polonsky KS. 1995 Lilly lecture 1994. The ß-cell in diabetes: from molecular genetics to clinical research. Diabetes. 44:705–717.[Abstract]
  18. Cereghini S. 1996 Liver-enriched transcription factors and hepatocyte differentiation. FASEB J. 10:267–282.[Abstract]
  19. Horlein A, Grajer KH, Igokemenes T. 1993 Genomic structure of the pou-related hepatic transcription factor HNF-1alpha. Biol Chem Hoppe-Seyler. 374:419–425.[Medline]



This article has been cited by other articles:


Home page
 Diabetes CareHome page
S. Anuradha, V. Radha, R. Deepa, T. Hansen, B. Carstensen, O. Pedersen, and V. Mohan
A Prevalent Amino Acid Polymorphism at Codon 98 (Ala98Val) of the Hepatocyte Nuclear Factor-1{alpha} Is Associated With Maturity-Onset Diabetes of the Young and Younger Age at Onset of Type 2 Diabetes in Asian Indians
Diabetes Care, October 1, 2005; 28(10): 2430 - 2435.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
M. N. Weedon, K. R. Owen, B. Shields, G. Hitman, M. Walker, M. I. McCarthy, A. T. Hattersley, and T. M. Frayling
A Large-Scale Association Analysis of Common Variation of the HNF1{alpha} Gene With Type 2 Diabetes in the U.K. Caucasian Population
Diabetes, August 1, 2005; 54(8): 2487 - 2491.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
G. Andersen, L. Wegner, D. P. Jensen, C. Glumer, L. Tarnow, T. Drivsholm, P. Poulsen, S. K. Hansen, E.-M. D. Nielsen, J. Ek, et al.
PGC-1{alpha} Gly482Ser Polymorphism Associates With Hypertension Among Danish Whites
Hypertension, April 1, 2005; 45(4): 565 - 570.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
G. Andersen, L. Wegner, C. S. Rose, J. Xie, H. Zhu, K. Larade, A. Johansen, J. Ek, J. Lauenborg, T. Drivsholm, et al.
Variation in NCB5OR: Studies of Relationships to Type 2 Diabetes, Maturity-Onset Diabetes of the Young, and Gestational Diabetes Mellitus
Diabetes, November 1, 2004; 53(11): 2992 - 2997.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
A. E. Jackson, P. G. Cassell, B. V. North, S. Vijayaraghavan, S. V. Gelding, A. Ramachandran, C. Snehalatha, and G. A. Hitman
Polymorphic Variations in the Neurogenic Differentiation-1, Neurogenin-3, and Hepatocyte Nuclear Factor-1{alpha} Genes Contribute to Glucose Intolerance in a South Indian Population
Diabetes, August 1, 2004; 53(8): 2122 - 2125.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
S. K. Rasmussen, S. A. Urhammer, L. Berglund, J. N. Jensen, L. Hansen, S. M. Echwald, K. Borch-Johnsen, Y. Horikawa, H. Mashima, H. Lithell, et al.
Variants Within the Calpain-10 Gene on Chromosome 2q37 (NIDDM1) and Relationships to Type 2 Diabetes, Insulin Resistance, and Impaired Acute Insulin Secretion Among Scandinavian Caucasians
Diabetes, December 1, 2002; 51(12): 3561 - 3567.
[Abstract] [Full Text] [PDF]

Home page
 Diabetes CareHome page
S. C. Elbein, J. Sun, E. Scroggin, K. Teng, and S. J. Hasstedt
Role of Common Sequence Variants in Insulin Secretion in Familial Type 2 Diabetic Kindreds: The sulfonylurea receptor, glucokinase, and hepatocyte nuclear factor 1{alpha} genes
Diabetes Care, March 1, 2001; 24(3): 472 - 478.
[Abstract] [Full Text]

Home page
 J. Clin. Endocrinol. Metab.Home page
S. K. Rasmussen, S. A. Urhammer, T. Hansen, K. Almind, A. M. Møller, K. Borch-Johnsen, and O. Pedersen
Variability of the Insulin Receptor Substrate-1, Hepatocyte Nuclear Factor-1{alpha} (HNF-1{alpha}), HNF-4{alpha}, and HNF-6 Genes and Size at Birth in a Population-Based Sample of Young Danish Subjects
J. Clin. Endocrinol. Metab., August 1, 2000; 85(8): 2951 - 2953.
[Abstract] [Full Text]

Home page
 J. Clin. Endocrinol. Metab.Home page
K. C. Chiu, L.-M. Chuang, J. M. Ryu, G. P. Tsai, and M. F. Saad
The I27L Amino Acid Polymorphism of Hepatic Nuclear Factor-1{alpha} Is Associated with Insulin Resistance
J. Clin. Endocrinol. Metab., June 1, 2000; 85(6): 2178 - 2183.
[Abstract] [Full Text]

Home page
 J. Clin. Endocrinol. Metab.Home page
S. K. Rasmussen, S. A. Urhammer, J. N. Jensen, T. Hansen, K. Borch-Johnsen, and O. Pedersen
The -238 and -308 G->A Polymorphisms of the Tumor Necrosis Factor {alpha} Gene Promotor Are Not Associated with Features of the Insulin Resistance Syndrome or Altered Birth Weight in Danish Caucasians
J. Clin. Endocrinol. Metab., April 1, 2000; 85(4): 1731 - 1734.
[Abstract] [Full Text]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Urhammer, S. A.
Right arrow Articles by Pedersen, O.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Urhammer, S. A.
Right arrow Articles by Pedersen, O.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals