Sequencing analysis of ghrelin gene 5′ flanking region: relations between the sequence variants, fasting plasma total ghrelin concentrations, and body mass index

Abstract

Ghrelin is a 28-amino-acid peptide with several functions linked to energy metabolism. Low ghrelin plasma concentrations are associated with obesity, hypertension, and type 2 diabetes mellitus, whereas high concentrations reflect states of negative energy balance. Several studies addressing the hormonal and neural regulation of ghrelin gene expression have been carried out, but the role of genetic factors in the regulation of ghrelin plasma levels remains unclear. To elucidate the role of genetic factors in the regulation of ghrelin expression, we screened 1657 nucleotides of the ghrelin gene 5′ flanking region (promoter and possible regulatory sites) for new sequential variations from patient samples with low (n = 50) and high (n = 50) fasting plasma total ghrelin concentrations (low- and high-ghrelin groups). Eleven single nucleotide polymorphisms (SNPs), 3 of which were rare variants (allelic frequency less than 1%) were found in our population. The genotype distribution patterns of the SNPs did not differ between the study groups, except for SNP −501A>C ( P = .039). In addition, the SNP −501A>C was associated with body mass index (BMI) ( P = .018). This variant was studied further in our large and well-defined Oulu Project Elucidating Risk for Atherosclerosis (OPERA) cohort (n = 1045) by the restriction fragment length polymorphism (RFLP) technique. No significant association of SNP −501A>C genotypes with fasting ghrelin plasma concentrations was found in the whole OPERA population. However, the association of this SNP with BMI and with waist circumference reached statistical significance in OPERA ( P = .047 and .049, respectively), remaining of borderline significance for BMI after adjustments ( P = .055). The results indicate that factors other than the 11 SNPs found in this study in the 5′ flanking region of ghrelin gene are the main determinants of ghrelin plasma levels. However, SNP −501 A>C genotype distribution seems to be different in subjects having the highest compared with those with the lowest ghrelin levels, and the SNP may be associated with BMI and waist circumference.