Single nucleotide polymorphisms of leptin and its receptor gene in type 2 diabetes patients among three major Malaysia ethnics

Abstract

Type 2 diabetes mellitus (T2DM) is a chronic metabolic syndrome that is rapidly increasing across the world, especially in Malaysia. Leptin and its receptor play vital role in the regulation of glucose metabolism and insulin sensitivity. Variations in the LEP and LEPR genes have been associated with insulin resistance, leptin level, and T2DM across different populations, but have not been extensively reported within the Malaysian population. This study aimed to investigate the genetic impacts of LEP and LEPR gene polymorphisms (A19G, G2548A, K109R, and Q223R, respectively) on serum leptin levels and insulin resistance among T2DM patients. This case-control study involved 150 T2DM patients and 150 non-diabetic volunteers from ethnic Malays, Chinese and Indians. The genotyping analysis of LEP and LEPR gene polymorphisms was carried out using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) followed by genotyping of a few samples for each SNP by Sanger sequencing method for validation. Serum leptin and insulin levels were determined via Enzyme Linked Immunosorbent Assay (ELISA). A homeostasis model assessment-insulin resistance (HOMA-IR) was calculated. Chi-square test was used to determine the distribution of genotypes and allelic frequencies and ANOVA was used to determine the association of clinical and biochemical parameters with each SNP. The study shows that the frequency of the AG genotype of the LEPR Q223R variant was significantly higher in T2DM patients compared to the control group (58.66% vs. 42%, p = 0.013). The A allele frequency was significantly higher in patients with T2DM than in non-diabetic individuals (36.66% and 29%, respectively, P = 0.046). Besides, T2DM patients with GG genotype had significantly higher serum leptin, insulin, BMI, and HOMA-IR index (P < 0.05). The frequency of the AA genotype and the A allele of the LEP G2548A variant were significantly (P < 0.05) higher in T2DM patients compared to the control groups. Furthermore, elevated serum leptin, insulin levels, and BMI in diabetic patients were found to be associated with the AA genotype of the LEP G2548A variant, compared to GG and GA genotypes (P < 0.05). However, no statistical significant differences were found in the genotype and allele frequencies of LEPR K109R and LEP A19G variants between T2DM patients and non-diabetic volunteers (P < 0.05). Furthermore, no significant differences in anthropometrical and biochemical parameters were observed between the genotypes of the LEPR K109R and LEP A19G polymorphisms. The AGAG haplotype combination of four SNPs was significantly different (OR = 0.633, 95% CI: 0.423 – 0.947, p = 0.025). Generally, fasting serum leptin levels were significantly (P < 0.001) higher in T2DM patients compared to non-diabetic subjects (166.78 pg/ml and 101.94 pg/ml, respectively). This study suggests a significant association between LEPR Q223R polymorphism and T2DM patients among Malay and Chinese ethnic groups, and it is significantly correlated with higher serum leptin, insulin, BMI, and HOMA-IR index. LEP G2548A polymorphism was significantly associated with T2DM among Malay and Indian ethnics and it is markedly associated with elevated serum leptin, insulin levels, and BMI in diabetic patients. Whereas, the other two SNPs of LEPR K109R and LEP A19G may not be useful markers for diabetes among Malaysian population but may have synergistic effects on diabetes.