Molecular Epidemiology Study Of Hypertension Combined With Hyperhomocysteinemia And Efficacy Of Enalapril Maleate-Folate Tablets On Reduction Of Fasting Plasma Glucose: A Multi-center, Randomized, Double Blind, Parallel Controlled Clinical Trial

Background: Many studies suggested that elevated plasma homocysteine (Hcy) and low folate status (LFS, folate <11nmol/L) were both the independent risk factors of cardiovascular disease (CVD). Hypertension and hyper-homocysteinemia (HHcy) had a significant synergistic effect on CVD development. It will be beneficial on CVD prevention to have a significant reduction of the major determinants such as blood pressure and plasma tHcy concentration and elevation of serum folate level.Objective: To investigate the prevalences of HHcy and LFS in Chinese hypertensive patients. To explore the associations of polymorphism of MTHFR C677T, gender, age, or plasma creatinine level with HHcy and their interactions. Evaluate the efficacy of Enalapril Maleate-Folate Tablets (EMFT) on reduction of fasting plasma glucose (FPG) and the interaction between short term folic acid supplementation and hyperglycemial on promoting FPG regression to normal.Methods: This is a multi-center, randomized, double blind, parallel control clinical trial, conducted to invesgate the efficacy and safety of EMFT on reduction of blood pressure and plasma homocysteine in Chinese mild-to-moderate primary hypertensive patients. A total of 480 participants, aged 18 to 75 years, from Beijing, Harbin, Shenyang, Xi'an, Nanjing and Shanghai were enrolled and randomly assigned to receive enalapril 10.0mg daily (the control group), or EMFT 10:0.4 (Low-folate group) or EMFT 10:0.8 (High-folate group) per day for eight weeks, respectively. Participants were interviewed face-to-face with a standardized questionnaire designed specifically for the study to collect their birth date, medical history, vitamin use, family history of hypertension, DM, myocardial infarction, premature coronary heart disease and so on. Using a mercury column sphygmomanometer and a standardized protocol, blood pressure was measured at baseline, 2nd week, 4th week, 6th week and 8th week according to the unique SOP, respectively. Fasting venous blood samples were obtained from the study participants with tubes containing EDTA (for plasma) or not (for serum) at baseline, the 28th day and the end of study. FPG, HDL, TG, TC et al. were assessed at baseline and 8th week. Serum Folate and plasma homocysteine concentration were measured at baseline, 4th week, and 8th week. Serum folate level was conducted with electrochemiluminescence in the first hospital of Beijing University. plasma total homocysteine concentration and MTHFR C677T genotypes were detected with high performance liquid chromatography or PCR-RFLP in the central lab, respectively.Results: A total of 480 participants were enrolled at the first stage. 24 were removed, in which 18 were finally excluded according to enrollment and exluding criteria and 6 because of incomplete Hcy data. 456 with complete data were used in our final analysis.⑴. Plasma total homocysteine (tHcy) in our study population was 15.1±11.2μmol/L, 18.8±14.5μmol/L for male and 12.4±6.6μmol/L for the female, respectively. The tHcy level of male participants was significantly higher than that of female.⑵. It is common for Chinese hypertensive patients complicated with HHcy.(tHcy>10μmol/L). Prevalance of HHcy in this 456 participants were 73.5% in the pooled participants, 91.3% for male and 60.0% for female, respectively. It is significantly higher than that of Chinese general population, We also did not find the phenomenon that tHcy level of participants in Northern China was significantly higher than that of those in Southern China in the general population.⑶. We first report, as we know, that the frequency of T allele of MTHFR C677T in Chinese mild-to-moderate hypertensive patients was 50%. Age and plasma creatinine concentration is positively correlated with tHcy level, r=0.26 (P<0.001) for age and r=0.41 (P<0.001) for creatinine, repectively. Plasma tHcy level and the prevalence of HHcy in participants with MTHFR 677TT genotype, elder age or higher creatinine level were all significantly higher than those of their control, respectively. Prevalance of HHcy of male participants with TT genotype, elder subjects with TT genotype or those TT genotype carriers with high creatinine were also significantly higher than those of their control, respectively.⑷. The serum folate in this population was 13.7±6.0nmol/L, 12.4±5.5nmol/L for male and 14.6±6.2nmol/L for female, respective ly. Prevalence of low folate status (folate<11 nmol/L) was 36.8%, 47.4% for the male and 28.8% for the female, which is significantly higher in the male than that in the female..⑸. Prevalence of low folate status (folate<11nmol/L) of subjects with MTHFR 677TT genotype, male, high creatinine level (creatinine≥68μmol/L), and low HDL level (HDL<1.26mmol/L) were 50.9%, 47.4%, 42.9% and 43.9%, respectively. Which is significantly higher than those with MTHFR 677CC (29.8%)or CT (33.6%) genotypes, female(28.8%), low creatinine level(30.4%) and high HDL level (29.8%).⑹. Prevalence of low folate status in Male subjects with MTHFR 677TT genotype (56.9%), subjects with high creatinine and TT genotype (52.2%), participants with low HDL and TT genotype (55.6%) were significantly higher than those female with CC/CT genotype(23.4%), low creatinine and CC/CT genotype (26.4%), or high HDL and CC/CT genotype (24.0%), respectively. The OR (95% CI) were 3.9(1.9～8.1), 2.5(1.2～5.2) and 3.5(1.5～8.2), respectively.⑺. The reduction of FPG in the control, low- and high-folate groups are, -0.03±0.87 mmol/L,0.03±0.97 mmol/L and -0.02±0.95mmol/L for the pooled population, 0.02±0.71 mmol/L,0.15±0.77 mmol/L and 0.16±0.79 mmol/L for those with normoglycemia (FPG<6.1mmol/L) and -0.23±1.30 mmol/L, -0.39±1.44 mmol/L and -0.80±1.20 mmol/L for those with hyperglycemia (FPG≥6.1 mmol/L). We can not find any significant differences of FPG reduction among three intervention groups in both the pooled population and those with normoglycemia. Comparing with the baseline FPG level, We also can't find any significant decrease in FPG level in the three treatment groups after 8 weeks intervention. However, in participants with hyperglycemia, both low-folate and high-folate groups all can significantly decrease the FPG concentration after 8 weeks treatment, while the control group can't. We also can find a significant difference in FPG reduction among three intervention groups. The FPG reduction of high-folate group is significantly higher than that of the control group, while the difference between the low-folate and control groups does not achieved statistical significant level.⑻. For participants with hyperglycemia, supplementation of 0.4 mg folic acid daily has a significant synergistic effect with the hyperglycemia on FPG reduction. The partial regression coefficient (standard error) is -0.6 (0.2) (P=0.002), after adjusting age, gender, BMI, and study center. Supplementation of 0.8 mg folic acid daily also has this significant synergistic effect. After adjusting the same potencial confounding factors, the partial regression coefficient (standard error) is -0.8 (0.2) (P<0.001). When we combind the low- and the high-folate groups as a whole folate group and repeat the same explore, we can also find the same synergistic effect. The partial regression coefficient (standard error) if -0.7 (0.2) (P<0.001).Conclusion Hyperhomocysteinemia and low folate status are common in Chinese hypertensive patients and the prevalence of HHcy is significantly higher than that of Chinese general population. Plasma tHcy concentration of male subjects is significantly higher than that of female. MTHFR 677TT genotype, male, elder age, high creatinine or low HDL level are risk factors of HHcy and low folate status. We can find a significant synergistic effect on HHcy or low folate status between MTHFR 677TT genotype and male, elder age, high plasma creatinine or low HDL level.Supplementation of EMFT can significantly promote the FPG level of those with hyperglycemia regression to normoglycemia. High-folate group may have a more significant efficacy of FPG reduction.