1. Introduction
Folate is a vital water-soluble micronutrient, and includes naturally occurring folate in the body as well as synthetic folic acid in supplements and fortified foods [1]. Folate plays a crucial role in cell development and differentiation, DNA and histone methylation in epigenetics, and the regulation of immune cell functions [2]. Current epidemiologic studies have identified an association of inadequate folate concentrations with elevated all-cause and cardiovascular mortality [3,4,5,6].
Given its involvement in one-carbon metabolism, specifically concerning homocysteine (Hcy) [7], which is recognized by the American Heart Association to be associated with stroke and other vascular diseases [8], it was hypothesized that folic acid intake or plasma/serum folate concentrations may affect stroke risk by modulating Hcy concentrations. The recent systematic review of randomized controlled intervention trials has demonstrated that adequate folic acid supplementation could reduce stroke risk [9]. However, the evidence concerning the association between folate concentrations and stroke risk remains scarce and contradictory, especially across different stroke types. One nested case-referent study suggested that folate might have a protective effect on hemorrhagic rather than ischemic stroke [10]. A prospective population-based study found no significant association between plasma folate concentrations and ischemic stroke risk, whereas dietary folic acid was negatively correlated with ischemic stroke [11].
Methylenetetrahydrofolate reductase (MTHFR) is an essential enzyme in folate and Hcy metabolism. It facilitates the irreversible conversion of 5,10-methylenetetrahydrofolate into 5-methylenetetrahydrofolate, a circulating form of folate involved in the remethylation of Hcy to methionine [12]. MTHFR C677T polymorphism (rs1801133) disrupts methylation capacity, activates inflammatory responses, induces lipid metabolism disorders, and promotes endothelial dysfunction and thrombosis, serving as a representative of constant exposure to low folate and high Hcy concentrations [13]. Either a whole-genome sequencing analysis or another prospective study uncovered a link between the C677T mutation and elevated stroke risk in the low-folate Chinese population [14,15]. A recent meta-analysis of case–control studies revealed that the T allele of MTHFR C677T might be a risk factor for ischemic stroke [16].
To date, the roles of folate concentrations in stroke remain unestablished, and the impact of Hcy and potential modification of the MTHFR C677T gene polymorphism on the folate–stroke relationship in the general population have yet to be investigated. The current study aims to analyze the relationship between serum folate concentrations and a 10-year risk of stroke among a Chinese community cohort without a history of cardiovascular disease (CVD). Importantly, the mediation effect of Hcy and potential inheritance–metabolism interaction on stroke will also be investigated.
2. Materials and Methods
2.1. Study Design and Population
The participants were recruited from an atherosclerosis cohort in two communities in Shijingshan District, Beijing, China. A total of 9540 individuals aged 40 and above were initially enrolled in the baseline survey conducted between December 2011 and April 2012. A 10-year follow-up survey was then carried out from 2011 to 2021. Participants with missing follow-up information, lacking serum folate and plasma Hcy concentrations, ineligible MTHFR C677T genotype and a history of baseline CVD, defined as self-reported myocardial infarction or stroke (including transient ischemic attack), were excluded. A total of 4903 subjects were included for subsequent data analysis (Figure S1). This study adhered to the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) guidelines [17].
2.2. Demographic and Clinical Characteristics
Demographic data, such as baseline age, sex, lifestyle, and medical history, were gathered by trained healthcare practitioners. Body mass index (BMI) was computed as dividing weight in kilograms by the square of height in meters. In-depth interviews were conducted about alcohol consumption (drinking once or more weekly for over six months) and tobacco usage (smoking at least one cigarette daily for over six months). Blood pressure was measured by an electronic sphygmomanometer (Omron HEM-7117, Omron, Kyoto, Japan). Hypertension was defined as systolic blood pressure ≥ 140 mmHg, diastolic blood pressure ≥ 90 mmHg, self-reported history or usage of antihypertensive medication. Dyslipidemia was identified by low-density lipoprotein cholesterol ≥ 3.4 mmol/L, high-density lipoprotein cholesterol < 1.0 mmol/L, triglycerides ≥ 1.7 mmol/L, total cholesterol ≥ 5.2 mmol/L, self-reported history or usage of lipid-lowering medication. Diabetes was characterized by fasting blood glucose level ≥ 7 mmol/L, postprandial blood glucose level ≥ 11.1 mmol/L, self-reported history or usage of antidiabetic medication.
2.3. Determination for Biochemical Variables
Serum concentrations of glucose, lipids and creatinine were measured by the Roche C8000 Automatic Biochemical Analyzer (Roche, Basel, Switzerland). The estimated glomerular filtration rate (eGFR) was calculated based on the Chronic Kidney Disease Epidemiology Collaboration equation [18], and abnormal renal function was defined as eGFR below 90 mL/min/1.73 m2. Employing the Maglumi4000 automated chemiluminescent immunoassay analyzer (SNIBE, Shenzen, China), serum folate concentrations were measured by the electrochemiluminescence method. Utilizing the Beckman Coulter AU480 automated biochemical analyzer (Beckman Coulter, Brea, CA, USA), plasma Hcy concentrations were determined through the enzymatic cycling method. The MTHFR C677T genotype was tested using the Asian ExomeChip, a custom exome array built upon the Infinium HumanExome BeadChip (Illumina, San Diego, CA, USA), as previously documented [19].
2.4. Ascertainment of Outcomes
The Chinese Center for Disease Control and Prevention and the Beijing Municipal Health Commission, through their National Mortality Surveillance System and Inpatient Medical Record Home Page System, respectively, contained stroke information for all participants until the occurrence of events or completion of follow-up (31 December 2021). Events were categorized by the International Classification of Diseases, 10th Revision (ICD-10). The primary endpoint was the first stroke (fatal and non-fatal), comprising ischemic stroke (I63), hemorrhagic stroke (I60–I61) and other types of stroke (I64). The secondary endpoints were the first ischemic stroke (I63) and the first hemorrhagic stroke (I60-I61).
2.5. Statistical Analysis
We employed descriptive statistics to outline the characteristics of the entire cohort and by quartiles of serum folate concentrations. Mean (standard deviation, SD) or median (interquartile range, IQR) were computed for continuous variables, while frequencies were tabulated for categorical variables. Baseline differences across folate quartiles were assessed using a one-way analysis of variance alongside the Kruskal–Wallis test for continuous variables and chi-squared analysis for categorical variables, accordingly. The Kaplan–Meier method was employed to assess the cumulative stroke hazards stratified by folate concentrations, with group differences determined through log-rank tests. The association between folate concentrations and endpoints was explored in Cox proportional hazard regression models, adjusting for various covariates such as baseline age, sex, BMI, eGFR, drinking, smoking, hypertension, dyslipidemia, diabetes and the use of antihypertensive, lipid-lowering and hypoglycemic drugs in Model 1, with further adjustment for Hcy concentrations in Model 2.
We conducted mediation analyses utilizing Hcy to evaluate both the direct and indirect relationship between folate and stroke. The mediated proportion of Hcy was computed: mediation effect (%) = (βa × βb/βTotal) × 100% (βa = indirect effect1, βb = indirect effect2, βTotal = total effect). This methodology has been previously reported on and employed [20,21]. Based on stratified analyses and interaction tests, we examined the potential modification of MTHFR C677T gene polymorphism on the association between folate concentrations and stroke. Restricted cubic spline (RCS) analyses after full adjustment were used to assess the dose–response relationship between folate and stroke in the overall population, as well as in the subgroups stratified by MTHFR C677T genotype.
A two-sided p value < 0.05 was regarded as statistically significant. All analyses were completed via R 4.3.3 (
3. Results
3.1. Baseline Characteristics
Baseline characteristics of the final cohort and different subsets stratified by quartiles of folate concentrations are presented in Table 1. The mean (SD) age of the 4903 subjects was 56.7 (8.8) years old, and 37.0% (n = 1815) were male. The median (IQR) values of serum folate and plasma Hcy were 6.2 (5.0, 8.2) ng/mL and 11.9 (10.0, 14.8) μmol/L. The distribution of the MTHFR C677T genotype (CC, CT and TT) within the total cohort was 18.7% (n = 915), 46.8% (n = 2295) and 34.5% (n = 1693), respectively. The median (IQR) values of serum folate across various quartiles were 4.4 (4.0, 4.7) ng/mL in Q1, 5.5 (5.2, 5.8) ng/mL in Q2, 7.0 (6.6, 7.6) ng/mL in Q3 and 10.3 (9.1, 12.3) ng/mL in Q4. As serum folate concentrations reduced, it was logical to observe an increase in plasma Hcy concentrations, along with a gradient decrease in the proportion of the MTHFR 677CC genotype.
3.2. Primary and Secondary Endpoints
A total of 407 strokes, 375 ischemic strokes and 47 hemorrhagic strokes occurred during the mean (SD) follow-up of 9.3 (1.8) years. The Kaplan–Meier survival curves (Figure 1) showed a significant visual relationship of folate concentrations with the 10-year incidence of stroke and ischemic stroke (log-rank tests p < 0.05 for both) but no significant cumulative hazard for hemorrhagic stroke.
The unadjusted and adjusted Cox regression analyses of folate concentrations for stroke are shown in Table 2. In terms of the primary endpoint, participants in the highest quartile of serum folate concentrations (≥8.2 ng/mL) exhibited a significantly lower risk of stroke compared with those in the lowest quartile (an absolute risk reduction: 3.3%; hazard ratio [HR]: 0.64, 95% confidence interval [CI]: 0.47–0.87, p = 0.005; p for trend = 0.002) after adjustment for multiple covariates in Model 1. Following additional adjustments for plasma Hcy concentrations in Model 2, the association between folate and stroke risk still existed (HR: 0.68, 95% CI: 0.50–0.93, p = 0.017; p for trend = 0.009). As for secondary endpoints, participants in the highest quartile of folate had a 36% reduced risk of ischemic stroke compared to those in the lowest quartile of folate (HR: 0.64, 95% CI: 0.47–0.88, p = 0.007; p for trend = 0.005). The results also remained stable after further adjustment for Hcy. Folate concentrations, whether as a continuous or categorical variable, showed no significant association with the risk of hemorrhagic stroke. The results of RCS analyses revealed that, after adjusting for covariates, the risk of stroke and ischemic stroke decreased initially and then leveled off as serum folate concentrations increased (Figure 2).
3.3. Mediation Analyses
Based on the persistent association between folate and stroke risk after adjusting for Hcy, we further explored the mediation effect of Hcy (Figure 3). With adjustments for baseline age and sex alone, the mediation effect of Hcy accounted for 23.37%. In the fully adjusted model, the direct effect of folate and indirect effect of Hcy on stroke were −0.028 (95% CI: −0.048–−0.005, p = 0.012) and −0.005 (95% CI: −0.009–−0.001, p = 0.030), respectively. Hcy partially mediated the relationship between folate and stroke risk, with a mediated proportion of 14.51% (p for proportion = 0.036).
3.4. Stratification Analyses
We conducted stratified analyses to investigate how MTHFR C677T gene polymorphism might modify the relationship between folate, treated as a continuous variable, and stroke risk. In individuals with the CC genotype, elevated serum folate concentrations were significantly associated with a decreased risk of stroke (HR: 0.88, 95% CI: 0.79–0.97, p for interaction = 0.026) and ischemic stroke (HR: 0.88, 95% CI: 0.80–0.98, p for interaction = 0.035), but not with the risk of hemorrhagic stroke (Table 3). RCS analyses following stratification by the MTHFR C677T genotype are demonstrated in Figure S2. The smooth curve for the CC genotype indicated a more notable reduction in the risk of stroke and ischemic stroke with folate concentrations rising compared with the smooth curve for the CT/TT genotype.
4. Discussion
To our knowledge, this is the first prospective large-scale cohort study utilizing mediation analysis to delve into the role of plasma Hcy concentrations and explore the interaction effect of the MTHFR C677T genotype in the association between folate concentrations and stroke through long-term follow-up in the general population. We confirmed that insufficient folate concentrations were linked to a heightened 10-year stroke risk, which was significantly mediated by plasma Hcy concentrations. In addition, the negative correlation between folate and the risk of stroke was solely observed in individuals with the CC genotype.
Folic acid supplementation has been demonstrated to mitigate stroke risk by diminishing oxidative stress, increasing brain-derived neurotrophic factors, promoting angiogenesis and facilitating cellular adaptation to ischemia and hypoxia [22,23]. The results of the China Stroke Primary Prevention Trial (CSPPT), conducted by our team, indicated that supplementation of folic acid combined with antihypertensive medication could reduce the risk of first stroke in hypertensive individuals by up to 21% compared to antihypertensive treatment alone [24]. The protective effect of folic acid on stroke may reflect a combined impact of various protective nutrients or dietary elements in one-carbon metabolism, while serum folate accurately reflects the body’s folate status. In our study, elevated folate concentrations were negatively associated with a ~10-year stroke risk. One prospective cohort study showed that elevated serum folate concentrations were associated with a decreased risk of all stroke and ischemic stroke in Eastern Finnish middle-aged men [25]. A Mendelian randomization study also revealed that higher genetically predicted folate concentrations were linked to a reduced stroke risk [26], which verifies that the relationship between them is consistent and solid in both East Asian and European populations.
This is the first study to quantify the mediation effect of Hcy in the folate–stroke relationship. A cohort study hypothesized that the association between combined low folate/vitamin B12 concentrations and an increased cerebrovascular ischemia risk was partially mediated by Hcy metabolism [27]. Another population study indicated that Hcy mediated the relationship between vitamin B12 and various diseases with small effects [28]. Elevated Hcy concentrations may increase the stroke risk by raising blood pressure, causing endothelial dysfunction, releasing vasoactive substances and inducing vascular injury [29,30,31,32]. Folate has a direct effect on stroke, possibly due to the involvement of multiple metabolites biosynthesis, epigenetics and redox homeostasis in the one-carbon metabolism. Further research is needed to explore the pathophysiological mechanisms by which folate influences stroke regardless of Hcy.
Innovatively, this study found that MTHFR C677T gene polymorphism could significantly modify the folate–stroke relationship, with the negative association being more pronounced in individuals with the CC genotype. Similarly, the CSPPT study revealed that among the hypertensive population with the CC/CT genotype, a negative correlation was observed between baseline folate concentrations and stroke risk. In contrast, individuals with the TT genotype consistently exhibited a high stroke risk across all folate concentrations [24]. Another post hoc analysis demonstrated that compared to participants with the CT/TT genotype, elevated concentrations of folate or vitamin B12 were notably associated with a reduced risk of first ischemic stroke among hypertensive patients with the CC genotype [33]. The MTHFR 677C → T mutation reduces efficiency in folate and Hcy metabolism. Therefore, exploring the relationship between 5-methyltetrahydrofolate concentrations and stroke risk needs to be further conducted for individuals with the insensitive CT/TT genotype, as 5-methyltetrahydrofolate can bypass the interference of low MTHFR activity on folate concentrations [34].
This study also has some limitations. Firstly, serum folate concentrations were assessed at the baseline, which might not precisely represent their long-term fluctuations. Numerous studies have encountered similar limitations, and further investigation into the longitudinal impact of folate changes is warranted. Secondly, atrial fibrillation (AF) is known to be a major risk factor for ischemic stroke, especially embolic stroke. Due to the limited data on AF and type of ischemic stroke, we were unable to adjust for AF as a confounding factor in the relationship between folate and stroke, nor could we explore the association between folate and embolic stroke. Thirdly, due to the limited incidence of hemorrhagic stroke, conclusions should be interpreted with caution, necessitating further in-depth research. Lastly, since this study was conducted at a single center in northern China, our findings should be validated in other cohorts.
5. Conclusions
In summary, among the general Chinese population, we have discovered an association between elevated serum folate concentrations and a reduced ~10-year risk of stroke, particularly ischemic stroke. Hcy had a significant mediation effect on the folate–stroke relationship. Only in participants with the MTHFR 677CC genotype were serum folate concentrations adversely correlated with the risk of stroke and ischemic stroke. This study provides insights for further understanding the crucial role of Hcy and the MTHFR C677T polymorphism in the association between folate and stroke.
Conceptualization, J.L. and Y.Z.; Methodology, Y.H., J.L. and Y.Z.; Software, Z.L., F.F. and J.J.; Validation, Y.H., J.L. and Y.Z.; Formal Analysis, Z.L. and F.F.; Investigation, Z.L., F.F., B.L., K.L., H.C. and J.J.; Resources, Y.H., J.L. and Y.Z.; Data Curation, F.F., J.J. and Y.Z.; Writing—Original Draft Preparation, Z.L. and F.F.; Writing—Review and Editing, Y.Z.; Visualization, Z.L., F.F. and Y.Z.; Supervision, F.F., J.J. and Y.Z.; Project Administration, F.F., Y.H., J.L. and Y.Z.; Funding Acquisition, F.F. and Y.Z. All authors gave approval of the last version to be published and agreed to be accountable for all aspects of the work. All authors have read and agreed to the published version of the manuscript.
The study was conducted according to the guidelines of the Declaration of Helsinki. Approval from the ethics committee of Peking University were obtained (IRB0000105211086, 12 December 2011).
Informed consent was obtained from all subjects involved in the study.
The corresponding authors had complete access to all the study data and are accountable for its integrity and the data analysis. The datasets analyzed in this study are available from the corresponding authors upon reasonable request. The data are not publicly available due to being part of an ongoing study.
We extend our gratitude to all members of the research team and the staff of the Pingguoyuan and Gucheng communities’ Health Centers for their support in this study.
The authors declare that they have no competing interests.
Footnotes
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.
Enlarge this image.Figure 1. Kaplan–Meier survival curves of the cumulative stroke hazards. (A) Stratified by quartiles of serum folate concentrations in stroke; (B) stratified by quartiles of serum folate concentrations in ischemic stroke; (C) stratified by quartiles of serum folate concentrations in hemorrhagic stroke.
Enlarge this image.Figure 2. Restricted cubic spline of the dose–response relationship between serum folate concentrations and stroke. Restricted cubic spline was adjusted for baseline age, sex, BMI, eGFR, drinking, smoking, hypertension, dyslipidemia, diabetes, the use of antihypertensive, lipid-lowering and hypoglycemic drugs and plasma Hcy concentrations. The blue shaded region denotes the 95% CI for the restricted cubic spline. The X-axis is bounded by the 0.5th and 99.5th percentiles of serum folate concentrations. CI: confidence interval; HR: hazard ratio.
Enlarge this image.Figure 3. Mediation effect of plasma Hcy concentrations on the association between serum folate concentrations and stroke. (A) Model 1 was adjusted for baseline age and sex. (B) Model 2 was adjusted for the covariates in model 1 plus BMI, eGFR, drinking, smoking, hypertension, dyslipidemia, diabetes and the use of antihypertensive, lipid-lowering and hypoglycemic drugs. Hcy: homocysteine.
Baseline characteristics of the participants.
| Characteristics | Overall | Quartiles of Serum Folate Concentrations | p Value | |||
|---|---|---|---|---|---|---|
| Q1 | Q2 | Q3 | Q4 | |||
| N | 4903 | 1223 | 1228 | 1226 | 1226 | |
| Age (year), mean (SD) | 56.7 (8.8) | 56.3 (9.1) | 56.8 (9.2) | 56.7 (8.6) | 57.1 (8.3) | 0.127 |
| Sex, N (%) | <0.001 | |||||
| Male | 1815 (37.0) | 739 (60.4) | 497 (40.5) | 342 (27.9) | 237 (19.3) | |
| Female | 3088 (63.0) | 484 (39.6) | 731 (59.5) | 884 (72.1) | 989 (80.7) | |
| Serum folate (ng/mL), | 6.2 | 4.4 | 5.5 | 7.0 | 10.3 | <0.001 |
| Plasma Hcy (μmol/L), | 11.9 | 15.1 | 12.2 | 11.3 | 10.3 | <0.001 |
| MTHFR C677T, N (%) | <0.001 | |||||
| CC | 915 (18.7) | 141 (11.5) | 220 (17.9) | 235 (19.2) | 319 (26.0) | |
| CT | 2295 (46.8) | 469 (38.3) | 586 (47.7) | 640 (52.2) | 600 (48.9) | |
| TT | 1693 (34.5) | 613 (50.1) | 422 (34.4) | 351 (28.6) | 307 (25.0) | |
| BMI (kg/m2), mean (SD) | 26.1 (3.4) | 26.3 (3.3) | 26.2 (3.3) | 26.0 (3.4) | 25.8 (3.5) | 0.002 |
| eGFR (mL/min/1.73 m2), mean (SD) | 94.8 (12.9) | 94.1 (12.9) | 94.5 (13.2) | 94.9 (12.8) | 95.6 (12.6) | 0.023 |
| eGFR classification | 0.005 | |||||
| ≥90 | 3464 (70.7) | 826 (67.6) | 854 (69.6) | 880 (71.8) | 904 (73.8) | |
| <90 | 1436 (29.3) | 396 (32.4) | 373 (30.4) | 346 (28.2) | 321 (26.2) | |
| Current drinking, N (%) | 1161 (23.7) | 455 (37.2) | 304 (24.8) | 241 (19.7) | 161 (13.1) | <0.001 |
| Current smoking, N (%) | 965 (19.7) | 480 (39.2) | 245 (20.0) | 164 (13.4) | 76 (6.2) | <0.001 |
| Prevalence of disease, N (%) | ||||||
| Hypertension | 2421 (49.4) | 588 (48.1) | 603 (49.1) | 617 (50.3) | 613 (50.0) | 0.684 |
| Dyslipidemia | 3501 (71.4) | 825 (67.5) | 862 (70.2) | 895 (73.0) | 919 (75.0) | <0.001 |
| Diabetes | 1156 (23.6) | 223 (18.2) | 247 (20.1) | 302 (24.6) | 384 (31.3) | <0.001 |
| Medication, N (%) | ||||||
| Antihypertensive drugs | 1485 (30.5) | 339 (27.9) | 362 (29.7) | 410 (33.6) | 374 (30.7) | 0.019 |
| Lipid-lowering drugs | 454 (9.4) | 80 (6.6) | 107 (8.8) | 127 (10.5) | 140 (11.5) | <0.001 |
| Hypoglycemic drugs | 482 (9.9) | 78 (6.4) | 85 (6.9) | 133 (10.9) | 186 (15.2) | <0.001 |
| Endpoint, N (%) | ||||||
| Stroke | 407 (8.3) | 121 (9.9) | 112 (9.1) | 93 (7.6) | 81 (6.6) | 0.014 |
| Ischemic stroke | 375 (7.6) | 110 (9.0) | 102 (8.3) | 88 (7.2) | 75 (6.1) | 0.039 |
| Hemorrhagic stroke | 47 (1.0) | 13 (1.1) | 13 (1.1) | 13 (1.1) | 8 (0.7) | 0.656 |
BMI: body mass index; eGFR: estimated glomerular filtration rate; Hcy: homocysteine; IQR: interquartile range; MTHFR: methylenetetrahydrofolate reductase; SD: standard deviation.
Association between serum folate concentrations and the risk of stroke.
| Endpoints | N | No. of Events, | Crude | Adjusted 1 * | Adjusted 2 † | |||
|---|---|---|---|---|---|---|---|---|
| HR (95% CI) | p Value | HR (95% CI) | p Value | HR (95% CI) | p Value | |||
| Stroke | ||||||||
| Linear trend | ||||||||
| Folate | 4903 | 407 (8.3) | 1.00 (0.98, 1.02) | 0.628 | 0.99 (0.97, 1.01) | 0.376 | 0.99 (0.97, 1.01) | 0.434 |
| Folate quartiles | ||||||||
| Q1 (<5.0 ng/mL) | 1223 | 121 (9.9) | ref | ref | ref | |||
| Q2 (5.0–6.2 ng/mL) | 1228 | 112 (9.1) | 0.91 (0.71, 1.18) | 0.496 | 0.92 (0.70, 1.19) | 0.523 | 0.98 (0.75, 1.29) | 0.892 |
| Q3 (6.2–8.2 ng/mL) | 1226 | 93 (7.6) | 0.75 (0.57, 0.99) | 0.039 | 0.76 (0.57, 1.01) | 0.061 | 0.82 (0.61, 1.09) | 0.174 |
| Q4 (≥8.2 ng/mL) | 1226 | 81 (6.6) | 0.65 (0.49, 0.86) | 0.003 | 0.64 (0.47, 0.87) | 0.005 | 0.68 (0.50, 0.93) | 0.017 |
| p for trend | 4903 | 407 (8.3) | <0.001 | 0.002 | 0.009 | |||
| Ischemic stroke | ||||||||
| Linear trend | ||||||||
| Folate | 4903 | 375 (7.6) | 1.00 (0.98, 1.01) | 0.701 | 0.99 (0.97, 1.01) | 0.415 | 0.99 (0.97, 1.01) | 0.461 |
| Folate quartiles | ||||||||
| Q1 (<5.0 ng/mL) | 1223 | 110 (9.0) | ref | ref | ref | |||
| Q2 (5.0–6.2 ng/mL) | 1228 | 102 (8.3) | 0.92 (0.70, 1.20) | 0.520 | 0.91 (0.69, 1.21) | 0.523 | 0.96 (0.72, 1.28) | 0.791 |
| Q3 (6.2–8.2 ng/mL) | 1226 | 88 (7.2) | 0.78 (0.59, 1.04) | 0.089 | 0.79 (0.59, 1.06) | 0.115 | 0.83 (0.62, 1.13) | 0.236 |
| Q4 (≥8.2 ng/mL) | 1226 | 75 (6.1) | 0.66 (0.49, 0.89) | 0.006 | 0.64 (0.47, 0.88) | 0.007 | 0.68 (0.49, 0.93) | 0.018 |
| p for trend | 4903 | 375 (7.6) | 0.003 | 0.005 | 0.012 | |||
| Hemorrhagic stroke | ||||||||
| Linear trend | ||||||||
| Folate | 4903 | 47 (1.0) | 0.95 (0.86, 1.06) | 0.393 | 0.97 (0.87, 1.07) | 0.540 | 0.98 (0.89, 1.08) | 0.627 |
| Folate quartiles | ||||||||
| Q1 (<5.0 ng/mL) | 1223 | 13 (1.1) | ref | ref | ref | |||
| Q2 (5.0–6.2 ng/mL) | 1228 | 13 (1.1) | 0.99 (0.46, 2.13) | 0.972 | 1.03 (0.47, 2.26) | 0.932 | 1.20 (0.53, 2.71) | 0.665 |
| Q3 (6.2–8.2 ng/mL) | 1226 | 13 (1.1) | 0.99 (0.46, 2.13) | 0.972 | 1.05 (0.47, 2.34) | 0.914 | 1.21 (0.52, 2.80) | 0.655 |
| Q4 (≥8.2 ng/mL) | 1226 | 8 (0.7) | 0.60 (0.25, 1.46) | 0.262 | 0.69 (0.27, 1.77) | 0.443 | 0.79 (0.30, 2.07) | 0.632 |
| p for trend | 4903 | 47 (1.0) | 0.308 | 0.510 | 0.693 | |||
* Model 1: Adjusted for baseline age, sex, BMI, eGFR, drinking, smoking, hypertension, dyslipidemia, diabetes and the use of antihypertensive, lipid-lowering and hypoglycemic drugs. † Model 2: Adjusted for the covariates in model 1 plus plasma Hcy concentrations. CI: confidence interval; HR: hazard ratio.
Modifying effect of MTHFR C677T genotype on the association between serum folate concentrations and stroke.
| Subgroup | N | No. of Events, N (%) | HR (95% CI) * | p Value | p for Interaction |
|---|---|---|---|---|---|
| Stroke | |||||
| MTHFR C677T | 0.026 | ||||
| CC | 915 | 68 (7.4) | 0.88 (0.79, 0.97) | 0.012 | |
| CT | 2295 | 190 (8.3) | 1.00 (0.99, 1.01) | 0.640 | |
| TT | 1693 | 149 (8.8) | 1.00 (0.96, 1.03) | 0.792 | |
| Ischemic stroke | |||||
| MTHFR C677T | 0.035 | ||||
| CC | 915 | 65 (7.1) | 0.88 (0.80, 0.98) | 0.016 | |
| CT | 2295 | 176 (7.7) | 1.00 (0.99, 1.01) | 0.666 | |
| TT | 1693 | 134 (7.9) | 0.99 (0.95, 1.04) | 0.727 | |
| Hemorrhagic stroke | |||||
| MTHFR C677T | 0.246 | ||||
| CC | 915 | 5 (0.5) | 0.72 (0.43, 1.20) | 0.203 | |
| CT | 2295 | 19 (0.8) | 0.94 (0.79, 1.13) | 0.507 | |
| TT | 1693 | 23 (1.4) | 1.01 (0.96, 1.06) | 0.720 |
* Adjusted for baseline age, sex, BMI, eGFR, drinking, smoking, hypertension, dyslipidemia, diabetes, the use of antihypertensive, lipid-lowering and hypoglycemic drugs and plasma Hcy concentrations. CI: confidence interval; HR: hazard ratio; MTHFR: methylenetetrahydrofolate reductase.
Supplementary Materials
The following supporting information can be downloaded at:
References
1. Bjorke-Monsen, A.L.; Ueland, P.M. Folate—A scoping review for Nordic Nutrition Recommendations 2023. Food Nutr. Res.; 2023; 67, 10258. [DOI: https://dx.doi.org/10.29219/fnr.v67.10258] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/38187793]
2. Dang, S.; Jain, A.; Dhanda, G.; Bhattacharya, N.; Bhattacharya, A.; Senapati, S. One carbon metabolism and its implication in health and immune functions. Cell Biochem. Funct.; 2024; 42, e3926. [DOI: https://dx.doi.org/10.1002/cbf.3926] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/38269500]
3. Xu, J.; Zhu, X.; Guan, G.; Zhang, Y.; Hui, R.; Xing, Y.; Wang, J.; Zhu, L. Non-linear associations of serum and red blood cell folate with risk of cardiovascular and all-cause mortality in hypertensive adults. Hypertens. Res.; 2023; 46, pp. 1504-1515. [DOI: https://dx.doi.org/10.1038/s41440-023-01249-3]
4. Song, S.; Song, B.M.; Park, H.Y. Associations of Serum Folate and Homocysteine Concentrations with All-Cause, Cardiovascular Disease, and Cancer Mortality in Men and Women in Korea: The Cardiovascular Disease Association Study. J. Nutr.; 2023; 153, pp. 760-770. [DOI: https://dx.doi.org/10.1016/j.tjnut.2023.01.023]
5. Liu, Y.; Geng, T.; Wan, Z.; Lu, Q.; Zhang, X.; Qiu, Z.; Li, L.; Zhu, K.; Liu, L.; Pan, A. et al. Associations of Serum Folate and Vitamin B12 Levels with Cardiovascular Disease Mortality Among Patients with Type 2 Diabetes. JAMA Netw. Open; 2022; 5, e2146124. [DOI: https://dx.doi.org/10.1001/jamanetworkopen.2021.46124]
6. Sun, Y.; Zhang, H.; Qi, G.; Tian, W. Nutrient deficiency patterns and all-cause and cardiovascular mortality in older adults with hypertension: A latent class analysis. BMC Public Health; 2024; 24, 1551. [DOI: https://dx.doi.org/10.1186/s12889-024-19003-w]
7. Wu, D.; Yin, R.; Deng, J. Homocysteine, hyperhomocysteinemia, and H-type hypertension. Eur. J. Prev. Cardiol.; 2024; 31, pp. 1092-1103. [DOI: https://dx.doi.org/10.1093/eurjpc/zwae022]
8. Kleindorfer, D.O.; Towfighi, A.; Chaturvedi, S.; Cockroft, K.M.; Gutierrez, J.; Lombardi-Hill, D.; Kamel, H.; Kernan, W.N.; Kittner, S.J.; Leira, E.C. et al. 2021 Guideline for the Prevention of Stroke in Patients with Stroke and Transient Ischemic Attack: A Guideline from the American Heart Association/American Stroke Association. Stroke; 2021; 52, pp. e364-e467. [DOI: https://dx.doi.org/10.1161/STR.0000000000000375]
9. An, P.; Wan, S.; Luo, Y.; Luo, J.; Zhang, X.; Zhou, S.; Xu, T.; He, J.; Mechanick, J.I.; Wu, W.C. et al. Micronutrient Supplementation to Reduce Cardiovascular Risk. J. Am. Coll. Cardiol.; 2022; 80, pp. 2269-2285. [DOI: https://dx.doi.org/10.1016/j.jacc.2022.09.048]
10. Van Guelpen, B.; Hultdin, J.; Johansson, I.; Stegmayr, B.; Hallmans, G.; Nilsson, T.K.; Weinehall, L.; Witthoft, C.; Palmqvist, R.; Winkvist, A. Folate, vitamin B12, and risk of ischemic and hemorrhagic stroke: A prospective, nested case-referent study of plasma concentrations and dietary intake. Stroke; 2005; 36, pp. 1426-1431. [DOI: https://dx.doi.org/10.1161/01.STR.0000169934.96354.3a]
11. Weng, L.C.; Yeh, W.T.; Bai, C.H.; Chen, H.J.; Chuang, S.Y.; Chang, H.Y.; Lin, B.F.; Chen, K.J.; Pan, W.H. Is ischemic stroke risk related to folate status or other nutrients correlated with folate intake?. Stroke; 2008; 39, pp. 3152-3158. [DOI: https://dx.doi.org/10.1161/STROKEAHA.108.524934] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/18988909]
12. Bjorklund, G.; Peana, M.; Dadar, M.; Lozynska, I.; Chirumbolo, S.; Lysiuk, R.; Lenchyk, L.; Upyr, T.; Severin, B. The role of B vitamins in stroke prevention. Crit. Rev. Food Sci. Nutr.; 2022; 62, pp. 5462-5475. [DOI: https://dx.doi.org/10.1080/10408398.2021.1885341] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/33724098]
13. Raghubeer, S.; Matsha, T.E. Methylenetetrahydrofolate (MTHFR), the One-Carbon Cycle, and Cardiovascular Risks. Nutrients; 2021; 13, 4562. [DOI: https://dx.doi.org/10.3390/nu13124562] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/34960114]
14. Cheng, S.; Xu, Z.; Bian, S.; Chen, X.; Shi, Y.; Li, Y.; Duan, Y.; Liu, Y.; Lin, J.; Jiang, Y. et al. The STROMICS genome study: Deep whole-genome sequencing and analysis of 10K Chinese patients with ischemic stroke reveal complex genetic and phenotypic interplay. Cell Discov.; 2023; 9, 75. [DOI: https://dx.doi.org/10.1038/s41421-023-00582-8]
15. Bennett, D.A.; Parish, S.; Millwood, I.Y.; Guo, Y.; Chen, Y.; Turnbull, I.; Yang, L.; Lv, J.; Yu, C.; Davey Smith, G. et al. MTHFR and risk of stroke and heart disease in a low-folate population: A prospective study of 156,000 Chinese adults. Int. J. Epidemiol.; 2023; 52, pp. 1862-1869. [DOI: https://dx.doi.org/10.1093/ije/dyad147]
16. Zhao, L.; Li, T.; Dang, M.; Li, Y.; Fan, H.; Hao, Q.; Song, D.; Lu, J.; Lu, Z.; Jian, Y. et al. Association of methylenetetrahydrofolate reductase (MTHFR) rs1801133 (677C>T) gene polymorphism with ischemic stroke risk in different populations: An updated meta-analysis. Front. Genet.; 2022; 13, 1021423. [DOI: https://dx.doi.org/10.3389/fgene.2022.1021423]
17. von Elm, E.; Altman, D.G.; Egger, M.; Pocock, S.J.; Gotzsche, P.C.; Vandenbroucke, J.P. STROBE Initiative. The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement: Guidelines for reporting observational studies. Lancet; 2007; 370, pp. 1453-1457. [DOI: https://dx.doi.org/10.1016/S0140-6736(07)61602-X]
18. Inker, L.A.; Schmid, C.H.; Tighiouart, H.; Eckfeldt, J.H.; Feldman, H.I.; Greene, T.; Kusek, J.W.; Manzi, J.; Van Lente, F.; Zhang, Y.L. et al. Estimating glomerular filtration rate from serum creatinine and cystatin C. N. Engl. J. Med.; 2012; 367, pp. 20-29. [DOI: https://dx.doi.org/10.1056/NEJMoa1114248]
19. Liang, Z.; Li, K.; Chen, H.; Jia, J.; Li, J.; Huo, Y.; Fan, F.; Zhang, Y. The Association of Plasma Homocysteine Concentrations with a 10-Year Risk of All-Cause and Cardiovascular Mortality in a Community-Based Chinese Population. Nutrients; 2024; 16, 1945. [DOI: https://dx.doi.org/10.3390/nu16121945]
20. Lee, H.; Herbert, R.D.; McAuley, J.H. Mediation Analysis. JAMA; 2019; 321, pp. 697-698. [DOI: https://dx.doi.org/10.1001/jama.2018.21973]
21. Wu, Z.; Jiang, Y.; Li, P.; Wang, Y.; Zhang, H.; Li, Z.; Li, X.; Tao, L.; Gao, B.; Guo, X. Association of impaired sensitivity to thyroid hormones with hyperuricemia through obesity in the euthyroid population. J. Transl. Med.; 2023; 21, 436. [DOI: https://dx.doi.org/10.1186/s12967-023-04276-3] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/37403157]
22. Burgess, K.; Bennett, C.; Mosnier, H.; Kwatra, N.; Bethel, F.; Jadavji, N.M. The Antioxidant Role of One-Carbon Metabolism on Stroke. Antioxidants; 2020; 9, 1141. [DOI: https://dx.doi.org/10.3390/antiox9111141] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/33212887]
23. Yahn, G.B.; Leoncio, J.; Jadavji, N.M. The role of dietary supplements that modulate one-carbon metabolism on stroke outcome. Curr. Opin. Clin. Nutr. Metab. Care; 2021; 24, pp. 303-307. [DOI: https://dx.doi.org/10.1097/MCO.0000000000000743] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/33631772]
24. Huo, Y.; Li, J.; Qin, X.; Huang, Y.; Wang, X.; Gottesman, R.F.; Tang, G.; Wang, B.; Chen, D.; He, M. et al. Efficacy of folic acid therapy in primary prevention of stroke among adults with hypertension in China: The CSPPT randomized clinical trial. JAMA; 2015; 313, pp. 1325-1335. [DOI: https://dx.doi.org/10.1001/jama.2015.2274]
25. Virtanen, J.K.; Voutilainen, S.; Happonen, P.; Alfthan, G.; Kaikkonen, J.; Mursu, J.; Rissanen, T.H.; Kaplan, G.A.; Korhonen, M.J.; Sivenius, J. et al. Serum homocysteine, folate and risk of stroke: Kuopio Ischaemic Heart Disease Risk Factor (KIHD) Study. Eur. J. Cardiovasc. Prev. Rehabil.; 2005; 12, pp. 369-375. [DOI: https://dx.doi.org/10.1097/01.hjr.0000160834.75466.b0]
26. Yuan, S.; Mason, A.M.; Carter, P.; Burgess, S.; Larsson, S.C. Homocysteine, B vitamins, and cardiovascular disease: A Mendelian randomization study. BMC Med.; 2021; 19, 97. [DOI: https://dx.doi.org/10.1186/s12916-021-01977-8]
27. Weikert, C.; Dierkes, J.; Hoffmann, K.; Berger, K.; Drogan, D.; Klipstein-Grobusch, K.; Spranger, J.; Mohlig, M.; Luley, C.; Boeing, H. B vitamin plasma levels and the risk of ischemic stroke and transient ischemic attack in a German cohort. Stroke; 2007; 38, pp. 2912-2918. [DOI: https://dx.doi.org/10.1161/STROKEAHA.107.486068]
28. Wang, L.; Li, X.; Montazeri, A.; MacFarlane, A.J.; Momoli, F.; Duthie, S.; Senekal, M.; Eguiagaray, I.M.; Munger, R.; Bennett, D. et al. Phenome-wide association study of genetically predicted B vitamins and homocysteine biomarkers with multiple health and disease outcomes: Analysis of the UK Biobank. Am. J. Clin. Nutr.; 2023; 117, pp. 564-575. [DOI: https://dx.doi.org/10.1016/j.ajcnut.2023.01.005]
29. Al Hageh, C.; Alefishat, E.; Ghassibe-Sabbagh, M.; Platt, D.E.; Hamdan, H.; Tcheroyan, R.; Chammas, E.; O’Sullivan, S.; Abchee, A.; Wang, B. et al. Homocysteine levels, H-Hypertension, and the MTHFR C677T genotypes: A complex interaction. Heliyon; 2023; 9, e16444. [DOI: https://dx.doi.org/10.1016/j.heliyon.2023.e16444]
30. Du, X.; Ma, X.; Tan, Y.; Shao, F.; Li, C.; Zhao, Y.; Miao, Y.; Han, L.; Dang, G.; Song, Y. et al. B cell-derived anti-beta 2 glycoprotein I antibody mediates hyperhomocysteinemia-aggravated hypertensive glomerular lesions by triggering ferroptosis. Signal Transduct. Target. Ther.; 2023; 8, 103. [DOI: https://dx.doi.org/10.1038/s41392-023-01313-x]
31. Han, L.; Miao, Y.; Zhao, Y.; Zhang, X.; Ma, X.; Du, X.; Kong, W.; Xu, Q.; Liu, J.; Dai, K. et al. The binding of autotaxin to integrins mediates hyperhomocysteinemia-potentiated platelet activation and thrombosis in mice and humans. Blood Adv.; 2022; 6, pp. 46-61. [DOI: https://dx.doi.org/10.1182/bloodadvances.2021004572] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/34559203]
32. Shi, W.; Zhang, J.; Zhao, W.; Yue, M.; Ma, J.; Zeng, S.; Tang, J.; Wang, Y.; Zhou, Z. Intracellular Iron Deficiency and Abnormal Metabolism, Not Ferroptosis, Contributes to Homocysteine-Induced Vascular Endothelial Cell Death. Biomedicines; 2024; 12, 2301. [DOI: https://dx.doi.org/10.3390/biomedicines12102301] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/39457614]
33. Qin, X.; Spence, J.D.; Li, J.; Zhang, Y.; Li, Y.; Sun, N.; Liang, M.; Song, Y.; Zhang, Y.; Wang, B. et al. Interaction of serum vitamin B(12) and folate with MTHFR genotypes on risk of ischemic stroke. Neurology; 2020; 94, pp. e1126-e1136. [DOI: https://dx.doi.org/10.1212/WNL.0000000000008932] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/31932513]
34. He, Q.; Li, J. The evolution of folate supplementation—From one size for all to personalized, precision, poly-paths. J. Transl. Int. Med.; 2023; 11, pp. 128-137. [DOI: https://dx.doi.org/10.2478/jtim-2023-0087]
© 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.
Details
; Fan, Fangfang 1
; Liu, Bo 1
; Li, Kaiyin 1 ; Chen, Hongyu 1 ; Jia, Jia 1 ; Huo, Yong 1 ; Li, Jianping 1
; Zhang, Yan 1
1 Department of Cardiology, Peking University First Hospital, No. 8 Xishiku Street, Xicheng District, Beijing 100034, China;