Folate is an essential vitamin for mammals that lack enzyme for de novo folate synthesis. The supply of folate is completely dependent on folate contained in dietary intake. Green leafy vegetables, mushrooms, legumes, and animal liver are good sources of folate supply for humans. Folate depletion can only be improved by the consumption of these foods. Folate is responsible for the reaction that conjugates with vitamin B12 to transfer the side chain including one carbon atom from one donor molecule to another. This type of biosynthetic reaction plays an important role in deoxyribonucleic acid (DNA) synthesis, cell division, and amino acid metabolism, and is essential for biological activity and homeostasis [1]. There is an epidemiological report that C677T polymorphism of methylenetetrahydrofolate reductase (MTHFR), an enzyme involved in folate-mediated reactions, is associated with various diseases (cardiovascular disease, cancers, neurology disease, diabetes, psoriasis, etc.). It suggested that folate plays an important role in maintaining human health [2]. In the past few decades, the association between folate deficiency and disease has been examined, with neural tube defect and cancer being examples. However, the association between folate and cardiovascular and all-cause mortality has been inconclusive, and therefore, the association is often discussed in relation to homocysteine.

Homocysteine is regenerated to methionine by folate and vitamin B12-mediated reactions in the methionine cycle (Fig. 1) [3]. Five to ten percent of homocysteine produced by cells is transferred to serum and excreted by the kidneys, keeping it in the normal serum concentration of 5-15 nmol/mL [4, 5]. As expected from the relationship in the methionine cycle, the 5-methyl tetrahydrofolate (THF) produced from dietary folate conjugates with vitamin B12 to convert homocysteine to methionine, so homocysteine accumulates if the folate circuit is stagnated (Fig. 1). Actually, serum folate concentrations have been reported to be inversely related to serum homocysteine concentrations in healthy populations [1]. An epidemiological study showed that hyperhomocysteinemia (HHCy) correlates with myocardial infarction and cerebral infarction, but the mechanism is unclear, although several hypotheses have been proposed [6]. In addition, another study reported that HHCy was independently associated with atherosclerosis [7]. These results suggested that HHCy would cause cardiovascular diseases [8]. Though several studies have examined whether interventions to reduce homocysteine concentrations by administration of B vitamins supplementation including folic acid reduce cardiovascular and overall mortality, a Cochrane review showed no difference comparing it to placebo [9]. It remains unclear whether serum or RBC folate is directly related to cardiovascular disease or whether it is related through its relationship with homocysteine as described above. Folic acid metabolism is involved in numerous metabolic reactions throughout the body. Therefore, it is speculated that the impact of simple oral folic acid supplementation on cardiovascular disease would be modulated by other effects of it including homocysteine.

Fig. 1
figure 1

Metabolism of folic acid. THF tetrahydrofolate, MTHFR methylenetetrahydrofolate reductase, SAM S-adenosylmethionine, SAH S-adenosylhomocysteine

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The present study showed the associations of serum and red blood cell (RBC) folate with cardiovascular and all-cause mortality in hypertensive adults using 13,986 hypertensive adults (mean age, 58.5 ± 16.1 years; 49.3% men) data from 1999 to 2014 National Health and Nutrition Examination Survey (NHANES) with a median of 7.0 years of follow-up [10]. The results showed that the fourth quartile of serum folate was associated with cardiovascular and all-cause mortality compared to the second quartile (hazard ratio [HR], 1.20 and 1.32, respectively) after multivariable adjustment. In contrast, the first quartile of serum folate was linked with increased all-cause mortality (HR,1.29), but not linked with cardiovascular mortality. In addition, the highest quartile of RBC folate was associated with cardiovascular and all-cause mortality compared to the second quartile (HR,1.68 and 1.30, respectively), but the lowest quartile was not associated with either outcome. The authors concluded that the associations between serum and RBC folate levels and the risk of cardiovascular and all-cause mortality in hypertensive adults looked U- or J- curve and non-linear. One hypothesis to explain this result is that low as well as high serum folate concentration also increase homocysteine concentration under vitamin B12 deficiency [11], and that additional verification of the effect of conjugation with vitamin B12 is needed. The strength points of the study compared the associations of serum and RBC folate with cardiovascular and all-cause mortality using folate quartile with the reference of second quartile of folate. Previous studies mostly checked only serum folate, but the present study checked RBC folate. The results of both serum and RBC folate assessment looked similar, and the results suggested that it is useful to assess only serum folate for the association with cardiovascular and all-cause mortality. The present study showed that not only high folate but also low folate might be associated with all-cause mortality. However, this study did not assess detailed information of background characteristics, such as food, life-style, pregnancy, comorbidities including blood cancer, medication including supplement, and so on. Folate is normally taken with food, and therefore, the information of food intake is important. Moreover, the study did not assess the medication and folate/folic acid intake over the study periods because of a single assessment of serum and RBC folate at baseline. As the authors mentioned, some subjects may have received intervention for folic acid deficiency in the study periods. This observational study can show the possibility of the association between folate and cardiovascular and all-cause mortality in hypertensive adults, but it could not show whether the association is causal or not. Therefore, further intervention studies are needed to conclude the association between folate and cardiovascular and all-cause mortality in hypertensive adults.

Several recent reports have shown that the association between serum folate concentration and cardiovascular and all-cause mortality is nonlinear in the classic risk population [12,13,14]. In 2020, Nkemjika et al. similarly reported a U-curve association between serum folate concentration and cardiovascular death in hypertensive adults based on NHANES data, but the analysis was using the Cox proportional hazard regression model [15]. When nonlinear dose-response correlations are expected, the use of Cox regression models with categorization is not preferred, and restricted cubic spline (RCS) analysis is one useful alternative method [16]. The present study showed the association between the two kinds of folate concentration and cardiovascular death and all-cause mortality among individuals with hypertension using RCS analysis. The results of this study confirm that folate concentrations in humans have an optimal range in relation to cardiovascular disease and all-cause mortality. Further interventional studies, in which individuals are classified by serum or RBC folate concentration and the amount of folic acid supplementation is adjusted, are needed to determine how folic acid supplementation affect these outcomes.