The Pro115Gln and Pro12Ala PPAR gamma gene mutations in obesity and type 2 diabetes

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Abstract

OBJECTIVE: Peroxisome-proliferator-activated receptors γ (PPAR γ), is a key regulator of adipocyte differentiation and energy balance. Two naturally occurring mutations in the PPAR γ gene, Pro115Gln and Pro12Ala, have recently been shown to impair the function of the PPARγ2 isoform of the receptor and to be associated with obesity or diabetes- related phenotypes in different populations.

SUBJECTS: We studied the occurrence and possible associations of the Pro115Gln and Pro12Ala in the PPAR γ2 gene with several clinical and metabolic phenotypes in three independent large populations of non-obese non-diabetic, type 2 diabetic, and morbidly obese French Caucasians.

RESULTS: The Pro115Gln mutation was not found in any of the 1069 subjects screened including 626 obese patients. The frequency of the Pro12Ala mutation was similar in all groups (0.08, 0.11,0.09) and was not associated with BMI or any of the clinical parameters tested.

CONCLUSIONS: We conclude that the Pro115Gln mutation is not a frequent cause of morbid obesity in Caucasians and that the Pro12Ala mutation is not associated with clinically significant changes in these populations.

Main

The peroxisome-proliferator-activated receptor γ (PPAR γ), is a member of the nuclear hormone receptor superfamily.1 This regulator of transcription has been identified as a functional receptor for the Thiazolinedione class of insulin sensitizing drugs.2,3 PPAR γ2, the predominantly adipose isoform of PPAR γ, is expressed selectively and at higher level in adipose tissue,4 where it modulates the expression of target genes implicated in adipocyte differentiation and glucose homeostasis.1,5,6 The PPAR γ2 gene is therefore a major candidate gene for type 2 diabetes or obesity, both complex phenotypes determined by the combination of multiple genetic and environmental factors.7,8

Two naturally occurring mutations in this gene have recently been shown to impair the function of the PPARγ2 isoform of the receptor and to be associated with obesity and/or diabetes and insulin sensitivity related phenotypes in different populations.9,10

The Pro115Gln10 mutation affects the serine 114 phosphorylation of PPARγ2 leading to a constitutively more active protein as assessed by the rate of adipocyte differentiation and fat accumulation in tissue-culture model of adipogenesis. This mutation was found in 1% (4/358) of subjects of a German population recruited for a study on the genetic of type 2 diabetes and was only present in a subgroup of obese patients accounting for 3% (4/121) of those cases in this population. All four Pro115Gln carriers described were severely obese (body mass index, BMI, range 37.9–47.3kg/m2). This result suggests that this mutation could account for a significant number of severe obesity cases in Caucasians. To date, no study has investigated the prevalence of this mutation in independent populations.

The Pro12Ala mutant was found to lower the transactivation capacity of PPARγ2.9 This mutant occurs with extremely variable frequencies in populations of different ethnic origins.11 It has been associated with a modest reduction in BMI and slightly increased insulin sensitivity in two large Finnish populations and with a reduced prevalence of diabetes in Japanese Americans.9 In contrast one study did not find an association with markers of adiposity and insulin resistance in a smaller population of Japanese men,12 two studies did not find an association of this variant with type 2 diabetes in Caucasians13,14 and one study found a positive relationship with BMI.15

Given the important role of PPARγ as a therapeutic target in metabolic ddiseases and given the physiologically meaningful data supporting a functional role for the PPARγ2 Pro115Gln and Pro12Ala mutations, more independent replications are clearly needed to evaluate the potential impact of those mutations in clinically relevant populations.

In this study, 1069 individuals from three independent populations of obese, diabetic and control Caucasians of comparable geographical origin were screened for the Pro12Ala and Pro115Gln PPARγ2 mutations. Associations between those mutations and obesity related phenotypes were evaluated.

Non-obese, non-diabetic controls (n=295) were randomly chosen among 3000 individuals participating in a prospective study evaluating the efficacy of daily supplementation with antioxidant vitamins and minerals in reducing the major health problems and the causes of premature death in a large population of healthy volunteer (SU-VI-MAX study).16 Subject characteristics were as follows: gender (f/m) 195/100; age (y) 53.2±6.7 (range 40–66), BMI (kg/m2) 22±1.8 (range 15–25); max BMI reached (kg/m2) 23.5±2.2 (range 16.5–29.7); waist/hip ratio 0.78±0.08 (range 0.6–1.04); none of those subjects had diabetes.

Morbidly obese patients (n=372) were randomly chosen from a panel of 800 French morbidly obese patients of an already described population.17 Main relevant clinical and metabolic parameters were as follows: gender (f/m) 295/77, age (y) 43.5±11.8 (range 15–74); BMI (kg/m2) 47.3±7.5 (range 40–82); maximum reached BMI (kg/m2) 49.5±8.5 (range 40–99), waist/hip ratio 0.93±0.11 (range 0.7–1.4). Their glucose tolerance status was evaluated by a 75 g oral glucose tolerance test. After the 0GTT, 165 were strictly normo-glycemic subjects (NG) and 207 had abnormalities of glucose tolerance; either an impaired fasting glucose (n=31), an impaired glucose tolerance (n=72) or type 2 diabetes (n=104).

Type 2 diabetic patients (n=402) were consecutively enrolled in an ongoing study on the genetics of diabetes complications. Those patients had all been referred to the same hospital located in the Paris area. Patient characteristics were: gender (f/m) 160/242; age (y) 57.5±10.7 (range 30–86), age of diabetes onset 44.74±11.1; HbAlc (%) 8.46±1.44 (range 4.5–14.7); BMI (kg/m2) 29.6±5.5 (range 17–58); max BMI reached (kg/m2) 32.74±6.5 (range 17–68); waist/hip ratio 0.96 ± 0.1 (range 0.67–1.5); hypertension (SBP>140 or DBP>85 or antihypertensive treatment) 72%.

DNA extracted from peripheral lymphocyte was used to genotype for the two variants by PCR-RFLP assays as described.10,15 To test for the presence of the rare Pro115Gln mutation, a plasmid containing the mutated PPAR γ2 cDNA was kindly provided by M. Ristow as a positive control.10

The Pro115Gln mutation in the PPAR γ2 gene was not found in any of the 1069 individuals tested. This population included a total of 626 subjects with a phenotype comparing well to that of the carriers of this mutation in the German population of Caucasian origin (372 morbidly obese patients, and 254 obese patients from the type 2 diabetes cohort). While our result does not argue against a causal effect of the Pro115Gln mutation in the previously described severely obese carriers, these data indicate that the Pro115Gln mutation does not account for a clinically significant number of morbidly obese cases in Caucasians with or without type 2 diabetes.

The Pro12Ala genotyping results, as well as some of the clinical and biological characteristics of the patient subgroups are indicated in Table 1. The observed genotype frequency of this mutant was in agreement with Hardy-Weinberg expectations in all three populations studied. Allelic frequencies of the Pro12Ala missense mutation in the PPAR γ2 was not different between controls and morbidly obese or type 2 diabetic patient respectively. In the analysis of phenotypical data according to PPAR γ2 genotype within the three groups, we found no significant difference in BMI or any obesity related traits in any of the three groups (Table 2). In addition, no significant difference in glucose homeostasis related phenotypes was found in obese subjects analyzed together or separated by their diabetes status or when men and women were analyzed separately.

Table 1 For obese patients, diabetes, glucose intolerance (IG), impaired fasting glucose (IFG) and normoglycemia were defined according to WHO criterias. For type 2 diabetics,ob 2,3 (severe and extreme obesity — BMI≥35), and ob 1 (obesity — 35>BMI≥30) were defined according to NHANES III criterias. In this group the maximum lifetime BMI were used to avoid biases due to acute hyperglycamia at the time of enrollement of the patients. Chi-square tests were used to compare allelic frequencies between the controls and the other two groups (results in bold characters) as well as within groups (results in plain characters)
Table 2 Results are mean±s.d. Continuous clinical and biological variables were analyzed using either one-way analysis of variance or Wilcoxon and Kruskal–Wallis test depending on the shapes of the distribution curves. All statistics were performed with the JMP software (SAS Institute Inc., Cary, NC)

In summary, our results show that the Pro12Ala and the Pro115Gln mutations in the PPAR γ2 gene are unlikely to serve as clinically useful predictor of type 2 diabetes and/or obesity in Caucasians. Lack of association of the two most frequent missense variants in the PPARγ gene with type 2 diabetes and/or obesity extends the results of previous linkage studies18 indicating that the PPARγ loci is not a major contributor to obesity or type 2 diabetes in French Caucasians.

References

  1. 1

    Spiegelman B . PPARγ: a adipogenic regulator and thiazolinedione receptor Diabetes 1998 47:: 507–513.

  2. 2

    Lehmann J, Morre L, Smith-Olivier T, Wilkison W, Wilson T, Kliewer S . An antidiabetic thiazolidinedione is a high affinity ligand for peroxisome proliferator-activated receptor γ Bio Chem 1995 270:: 12953–12956.

  3. 3

    Forman B, Tontonoz P, Chen J, Brun R, Spiegelman B, Evans R . 15-Deoxy-Δ12,14 prostaglandin J2 is a ligand for the adipocyte determination factor PPARγ Cell 1995 83:: 803–812.

  4. 4

    Vidal-Puig A, Considine R, Jimenez-Linan M, Werman A, Pories W, Caro J, Flier J . Peroxisome proliferator-activated receptort gene expression in human tissues: effects of obesity, weight loss, and regulation by insulin and glucocorticoids. J Clin Invest 1997 99:: 2416–2422.

  5. 5

    Spiegelman B, Flier J . Adipogenesis: rounding out the big picture Cell 1996 87:: 377–389.

  6. 6

    Latruffe N, Vamecq J . Peroxisome proliferator activated receptor (PPARs) as regulators of lipid metabolism Biochimie 1997 79:: 81–94.

  7. 7

    Bloomgarden Z . Insulin resistance: current concepts Clin Ther 1998 20 (2): 216–231.

  8. 8

    Neel J, Weder A, Julius S . Type II diabetes, essential hypertension, and obesity as “syndromes of impaired genetic homeostasis”: the “thrifty genotype” hypothesis enters the 21st century Perspect Biol Med 1998 42 (1): 44–74.

  9. 9

    Deeb S, Fajas L, Nemoto M, Pihlajamäki J, Mykkänen L, Kuusisto J, Laakso M, Fujimoto W, Auwerx J . A Pro12Ala substitution in PPARγ2 associated with decreased receptor activity, lower body mass index and improved insulin sensitivity Nature Genet 1998 20:: 284–287.

  10. 10

    Ristow M, Muller-Wieland D, Pfeiffer A, Kahn C . Obesity associated with a mutation in a genetic regulator of adipocyte differentiation N Engl J Med. 1998 339 (14): 953–959.

  11. 11

    Yen C, Beamer B, Negri C, Silver K, Brown K, Yarnall D, Burns D, Roth J, Shuldiner A . Molecular scanning of the human peroxisome proliferator activated receptor γ (hPPAR γ) gene in diabetic Caucasians: identification of a Pro12Ala PPARγ2 missense mutation Biochem Biophys Res Commun 1997 24 (2):: 270–274.

  12. 12

    Mori Y, Kim-Motoyama H, Katakura T, Yasuda K, Kadowaki H, Beamer B, Shuldiner A, Akanuma Y, Yasaki Y, Kadowaki T . Effect of the Pro12Ala variant of the human peroxisome proliferator activated receptor γ gene on adiposity, fat distribution and insulin sensitivity in Japanese men Biochem Biophys Res Commun 1998 251:: 195–198.

  13. 13

    Ringel J, Engeli S, Distler A, Sharma A . Pro12Ala PPARγ2 missense mutation of the peroxisome proliferator activated receptor γ and diabetes mellitus Biochem Biophys Res Communun 1999 254:: 450–453.

  14. 14

    Mancini F, Vaccaro O, Sabatino L, Tufano A, Rivellese A, Riccardi G, Colantuoni V . Pro12Ala substitution in the peroxisome proliferator-activated receptor-γ2 is not associated with type 2 diabetes Diabetes 1999 48:: 1466–1468.

  15. 15

    Beamer B, Yen C, Andersen R, Muller D, Elahi D, Cheskin L, Andres R, Roth J, Shuldiner A . Association of the Pro12Ala variant in the peroxisome proliferator activated receptor γ gene with obesity in two Caucasian populations Diabetes 1998 47:: 1806–1808.

  16. 16

    Hercberg S, Preziosi P, Briançon S, Galan P, Triol I, Malvy D, Roussel A-M, Favier A . A primary prevention trial using nutritional doses of antioxidant vitamins and minerals in cardiovascular diseases and cancers in a general population: the SU-VI-MAX study-design, methods, and participants characteristics Controlled Clin Trials 1998 19:: 336–351.

  17. 17

    Clément K, Lahlou N, Ruiz J, Hager J, Bougères P, Basdevant A, Guy-Grand B, Froguel P . Association of poorly controlled diabetes with low serum leptin in morbid obesity Int J Obes 1997 21:: 556–561.

  18. 18

    Clément K, Dina C, Basdevant A, Chastang N, Pelloux V, Lahlou N, Berlan M, Langin D, Guy-Grand B, Froguel P . A sib-pair analysis study of 15 candidate genes in French families with morbid obesity. Indication for linkage with islet 1 locus on chromosome 5q Diabetes 1999 48:: 398–402.

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Acknowledgements

Informed consents were obtained for all subjects and the protocols were approved by the Ethics Committee of Paris (CCPPRB Hôtel-Dieu and Cochin). This work was supported by the Direction de la Recherche Clinique/Assistance Publique-Hopitaux de Paris, Programme Hospitalier de Recherche Clinique (PHRC no. AOM 96088) and the Danone Institute (Danone/Suvimax DNA Bank). We thank Genevieve Bonhomme and Annie Legal for their excellent technical help. We also thank M. Ristow who kindly provided a mutated PPARγ2 cDNA plasmid

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Correspondence to K Clement.

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Keywords

  • obesity
  • PPARγ
  • diabetes
  • association study

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