CDKAL1 and HHEX are well-accepted pathogenesis-related key genes for T2DM, they play pivotal role in regulating the insulin secretion function of pancreatic β cells. This study evaluated 8 SNPs on CDKAL1 and HHEX genes to determine the significant association with T2DM. Of those, 4 SNPs (rs4712524, rs10946398 and rs7754840 on CDKAL1, rs5015480 in HHEX) were associated with T2DM in Chinese population.
CDKAL1 expression in human pancreatic β-cells participates in the correlation of CDKAL1- and CDK5-mediated pathways[17]. It has been shown that CDKAL1 could increase insulin secretion in pancreatic β-cells by inhibiting CDK5[5, 7, 8]. In 2007, rs7754840C was confirmed to be associated with T2DM in genome-wide association analysis for Finnish and Sweden populations[18, 19]. The following year, Unoki, H. et al. conducted a genome-wide association study and demonstrated the association of rs4712524G allele with T2DM in Japanese population, which has a similar minor allele frequency to the current study[20]. At the same year, Herder, C. et al. reported that the rs10946398C and the rs7754840C were risk alleles of T2DM in German population[21]. In 2009, Hu, Cheng et al. detected the rs10946398, rs7754840 in CDKAL1 and identified their moderate association with T2DM in Chinese population(< P0.05)[22]. In 2009, Lyssenko, V. et al. reported that CDKAL1 rs7754840C allele was the risk predictor for the incidence of T2DM in Swedish population during a median follow-up period of 23.5 years (P = 0.004, OR = 1.11; 95%CI: 1.03–1.19)[23]. In 2010, Han, X. et al. replicated the CDKAL1 rs10946398C was associated with T2DM in Chinese Han population[24]. Furthermore, the results of the meta-analysis of three Chinese Han populations also showed the association of the CDKAL1 rs10946398C with T2DM[24]. In 2016, Kommoju, U. J. et al. reported that the rs7754840C allele was associated with T2DM in Indian population[25]. In 2017, Nikitin, A. G. et al. identified the association of rs10946398C with T2DM in Russian population[26]. In the current study, the rs4712524G, rs10946398C and rs7754840C allele and genotype were also confirmed to be associated with T2DM. -Indeed, the rs7754840 has been proved to be associated with glucose metabolic traits. In 2014, Klimentidis, Y. C. et al. identified that the rs7754840C was associated with higher HbA1C in Alaska population, which has the similar minor allele frequency to Chinese Han population[27]. It was possible that these variants in CDKAL1 have effects on β-cell insulin secretion and furtherly functioned as risk variants in T2DM. For rs10946398, the C allele was conferred to be associated with decreased 30-min insulin response, an index of β-cell function, in European population[14] and lower homeostasis model assessment β-cell function in Chinese population[24]. Similarly, the rs7754840C as a risk allele was nominally associated with reduced insulin secretion in Finland and Sweden population [18]. It was very interesting to note that the different rs10946398T risk allele was proved in Chinese She population. The minor allele frequency of Chinese She population is C with 36.3–48.6%, which is opposite to Chinese Han population[28]. However, in Chinese Uyghur population, Song, Manshu et al. replicated the association of rs7754840C allele with T2DM in 2015 as the Uyghur population shared a similar allele frequency to Chinese Han population[29].
HHEX gene encode a transcription factor which is involved in a fundamental pathway called Wnt signaling pathway[11]. It is required for cell growth and development. A study in 2014 showed that transcription factor is required for δ cells differentiation [30]. If the somatostatin level increases the secretion level of insulin release from β cells will be impaired[30] In 2008, Herder, C. et al has validated the association of the rs5015480C in HHEX gene with T2DM in the German populations[21]. In the same year, Wu et al. reported that the associations of the rs5015480C allele with T2DM in Chinese Shanghai population[14]. In 2010, Zhou et al. reported that the HHEX rs5015480 C allele frequency showed a marginally significant statistics difference between T2DM and control group in Chinese population [31]༎In the same year, Han et al. also reported that the rs5015480C allele was significantly associated with T2DM in the Chinese population [24]. However, a study in 2011 reported that further analysis revealed that the sequence variant (rs5015480) on HHEX was associated with the susceptibility of T2DM in females, but not in males Korean population [32].Few years later, in 2013, this association of rs5015480T allele, with T2DM had been validated in Chinese She population [28]. In Chinese She population, the rs5015480T allele frequency with 52.7–59.7% is higher than other Chinese population [28]. In current study we found C allele of rs5015480 in HHEX (P < 0.001, OR = 1.295; 95%CI: 1.124–1.493) functioned as risk allele in incidence of T2DM and 2C/C + C/T was the risk genotype. The role of rs5015480 in T2DM might be explained by the associations of the rs5015480T allele was associated with lower homeostatic model assessment of β-cell function in Alaska population[27].
The rs1111875C allele, as a diabetes risk allele identified in previous GWAS was confirmed to be associated with decreased 30-min insulin response not only in European population[33], but also in Pima Indians[34]. In 2007, the rs1111875C in the HHEX gene was reported to be associated with T2DM in Finland and Sweden populations[18]. In 2008, Jana et al.[35] conducted an association study, their data confirmed the association of the common variant rs1111875C in the HHEX gene, identified by Sladek et al[36] in French population, with an increased risk of T2DM in a Dutch population. In 2008, Wu et al. reported that the associations of the HHEX rs1111875C allele with T2DM in Chinese Shanghai population[14]. In 2009, the rs1111875C in HHEX gene were detected to have not be associated with T2DM in German population. In 2010, Zhou et al. reported that the associations of the HHEX rs1111875C allele with T2DM in Chinese population[31]. The rs1111875C allele, as a risk allele identified in previous GWAS, showed different frequency from Caucasian population and African American population. In the current study, the C minor allele is 17.5–21.6%. While in Finland and Sweden population, the HHEX rs1111875 C allele is 53%[36]. And the C allele of the rs1111875 was 67% in African American population which did not detected the association of rs1111875 with T2DM[37]. The different genetic backgrounds might be responsible for the absent association of rs1111875 alone with T2DM in our study. The second reason might be the medium sample size which was the limitation of our study.
However, in our study we did not observed any association between the rs7923837 and T2DM(P = 0.181) which may be explained by different environmental risk profiles between Europeans and Asians, different body composition and genetic backgrounds, different linkage disequilibrium patterns. This possibility also be supported by association study done for Thai population in 2018, Plengvidhya, N. et al. found that no significant association could be concluded between rs7923837 on HHEX and T2DM[38].
In the current study, the rs4712524-rs10946398-rs7754840-rs9460546 were in LD, we observed that the distribution of the rs4712524-rs10946398-rs7754840-rs9460546GCCG (P = 0.001, OR = 1.210; 95%CI: 1.076–1.360) haplotype was associated with a higher risk of T2DM development. Conversely, the haplotype rs4712524-rs10946398-rs7754840-rs9460546 AAGT functioned as protector to against the development of T2DM (P = 0.001, OR = 0.827; 95%CI: 0.735–0.930). The HHEX rs1111875-rs5015480 haplotype analysis with D’>0.9 showed that rs1111875-rs5015480CC was associated with development of T2DM. It was noted that CDKAL1 rs4712524-rs10946398-rs7754840-rs9460546 and HHEX rs1111875-rs5015480 haplotype were associated with T2DM in our study, while the rs9460546 and rs1111875 failed to have shown the association with T2DM alone, respectively. The haplotype has more powerful than single variants in detecting the association of variants with complex diseases. Its efficiency in finding genetic factors in disorders lied in the representative for multi single nucleotide polymorphisms as the haplotype characterized in LD pattern and tend to inherited as a whole block. Otherwise, it was specific in a studied population as it was affected by population history, gene conversion and natural selection[39]. Therefore, the associations of the haplotype the rs4712524-rs10946398-rs7754840-rs9460546 and the HHEX rs1111875-rs5015480 with T2DM provided a powerful evidence in assessing the underlying genetic preposition in T2DM besides the genetic haplotype information in specific population. It was also help to study the function of CDKAL1 and HHEX gene in T2DM through influencing insulin secretion. Therefore, the effects of CDKAL1 and HHEX gene haplotype on β-cell insulin secretion should be done in the future.