Principal findings
In this Mendelian randomization study, we investigated the causal effects of glucokinase activator (GKA) on cardio-cerebrovascular diseases (CVDs) and the mediating role of circulating metabolites. GKA was found to be significantly associated with a reduced risk of atrial fibrillation (AF) (OR = 0.71, 95% CI: 0.54–0.95, P = 0.019) and stroke (OR = 0.97, 95% CI: 0.96–0.98, P = 4.82 × 10-5), with no evidence of heterogeneity or horizontal pleiotropy. However, no definitive causal relationship was identified between GKA and heart failure, coronary artery disease, or myocardial infarction; although the IVW P—value for CAD was < 0.05, the weighted median result weakened the statistical significance. In exploring GKA’s interactions with circulating metabolites, 36 metabolites were found to be significantly associated with GKA after Bonferroni correction (threshold P = 2.98 × 10−4). In particular, within the category of circulating particle sizes, GKA was observed to increase the average diameter of VLDL particles (β = 0.33 [95% CI 0.18, 0.48]; P = 1.85 × 10−5) and decrease the average diameter of HDL particles (β = − 0.30 [95% CI − 0.43, − 0.16]; P = 1.27 × 10−5). Further analysis of these metabolites’ effects on CVDs showed that, after Bonferroni correction, only one metabolite was associated with AF, with little evidence for a stroke association. Notably, a larger average VLDL particle diameter was inversely related to AF risk (OR 0.90 [95% CI 0.85, 0.96]; P = 8.85 × 10−4), and mediation analysis revealed a significant indirect effect of GKA on AF risk mediated by VLDL particle diameter (OR 0.88 [95% CI 0.78, 0.99]; P = 0.047), accounting for 11.64% of the total effect. Collectively, these findings highlight the potential cardiovascular benefits of GKA, particularly in relation to AF, and underscore the mediating role of specific circulating metabolites in this pathway.
The association between GKA and CVD incidence
The relationship between glucokinase activator (GKA) and atrial fibrillation (AF) has attracted increasing attention in the field of cardiovascular research. Multiple previous studies have explored this association from different perspectives and provided insights into the underlying potential mechanisms. In terms of glucose metabolism, some studies have confirmed that abnormal glucose homeostasis can trigger electrical and structural remodeling in the atria, thereby increasing susceptibility to AF [30]. Oxidative stress and inflammation induced by hyperglycemia play crucial roles in the development of AF and lead to alterations in ion channel function and atrial fibrosis [31]. Research has shown that hyperglycemia-induced changes in the atrial extracellular matrix promote the development of AF, and improving glycemic control with drugs such as GKA might mitigate these structural alterations [32]. As a regulator of glucose metabolism, GKA is highly likely to intervene in these pathophysiological processes. The increase in glucokinase activity promoted the efficient utilization of glucose in key tissues, such as the heart and pancreas, which might contribute to maintaining normal electrical activity in the atria and reducing the risk of AF.
Some studies have also explored the potential role of GKA in modulating the risk of AF. Research has indicated that GKA can regulate the activity of ion channels in cardiomyocytes, which is essential for maintaining normal cardiac electrical conduction [33]. By stabilizing the cardiac electrical conduction system, GKA might reduce the incidence of arrhythmias such as AF. In addition, Garcia et al [34] proposed that GKA might have a protective effect on the endothelial function of the heart. Endothelial dysfunction is associated with the occurrence of AF, and the improvement in endothelial function mediated by GKA might help reduce the risk of AF. Previous reports have suggested that GKA might affect the regulation of the heart by the autonomic nervous system, which plays a key role in the initiation and maintenance of AF [35]. By modulating autonomic nerve tension, GKA might contribute to maintaining a stable cardiac rhythm. Moreover, at the gene expression level, some studies have confirmed that GKA can influence the expression of certain genes involved in cardiac electrophysiology and provide a potential molecular mechanism for its anti-AF effect [36]. In our Mendelian randomization study, which used genetic variants as instrumental variables, we found that GKA was significantly associated with a reduced risk of AF. Compared with previous studies, this causal relationship provides more robust evidence for the potential protective effect of GKA against AF.
Previous studies have clearly demonstrated the association between glucose metabolism and stroke. Patients with impaired glucose tolerance have a greater risk of stroke [37]. With respect to the role of glucokinase activator (GKA) in stroke, multiple studies have explored its potential benefits. Some studies have shown that GKA can enhance endothelial function [38]. Endothelial cells play crucial roles in maintaining vascular health. GKA activated glucokinase in these cells, promoting the production of nitric oxide, which could relax blood vessels. These findings are highly important for preventing ischemic stroke. In ischemic stroke models, GKA can reduce neuronal damage [39] and improve glucose metabolism in neurons, enabling them to better withstand ischemic insults. Moreover, some studies have indicated that GKA can regulate lipid metabolism, reduce the accumulation of cholesterol and triglycerides in blood vessels, and thus decrease the formation of atherosclerotic plaques in the cerebral vasculature [40]. At the gene level, studies have shown that GKA can regulate the expression of certain genes related to angiogenesis in the brain [41]. This regulatory effect might contribute to the development of collateral blood vessels and enhance the blood supply to ischemic areas. Although genetically proxied GKA exposure demonstrated a statistically significant but modest reduction in stroke risk (3% relative risk reduction), the clinical impact at an individual level is likely minimal. However, this magnitude of risk reduction may yield substantial public health benefits through decreased absolute stroke events in large populations. This dichotomy underscores the necessity of evaluating interventions through both individual effect sizes and population-attributable impact for prevalent conditions like stroke. The association provides genetic evidence implicating GKA pathways in stroke risk modulation, warranting large-scale trials to quantify absolute benefits in target populations.
The neutral effects of genetically proxied GKA on HF, CAD, and MI may arise from several factors. First, GKA’s cardiometabolic effects likely exhibit pathway specificity: while robustly influencing VLDL size and AF/stroke risk (linked to lipoprotein metabolism and atrial remodeling), its impact on atherosclerosis-related pathways (e.g., plaque inflammation, thrombosis) or myocardial contractility appears limited. Second, limited statistical power cannot be overlooked. The smaller effect sizes of GKA on these outcomes, combined with potential heterogeneity in genetic instruments across diverse cardiovascular endophenotypes, might have constrained our ability to detect true causal effects. Finally, GKA-associated metabolites may not fully capture key biological intermediates driving coronary or myocardial pathogenesis, as GKA’s primary hepatic action may influence pathways less relevant to these endpoints than to AF and stroke. Larger GWAS and expanded biomarker profiling are needed to clarify these relationships.
Associations between GKA and circulating metabolites
The relationship between glucokinase activator (GKA) and the 36 identified circulating metabolites has been a subject of great concern in the field of metabolic research. In in vitro cell models and in vivo animal models, some scholars have shown that GKA treatment significantly increases the levels of cholesteryl esters in large HDL, cholesterol in large HDL, and total lipids in large HDL. Mechanistically, they discovered that GKA upregulated the expression of key genes involved in HDL biosynthesis, such as APOA1, which encodes apolipoprotein A-I, a major component of HDL. This upregulation enhanced the assembly and secretion of HDL particles, thereby leading to an increase in HDL-related metabolite levels. These results are consistent with our findings, further validating the positive impact of GKA on HDL-related lipid metabolism [42]. The use of isotope-labeled lipid tracers revealed that GKA treatment elevated the levels of triglycerides in very large VLDLs, triglycerides in chylomicrons, extremely large VLDLs, and total lipids in chylomicrons and extremely large VLDLs. The mechanism underlying this increase was related to enhanced lipid synthesis in the liver and increased lipid uptake by the intestine. Moreover, GKA promoted the expression of lipid transporters in the intestine, which facilitated the absorption of dietary lipids into chylomicrons [43]. In a clinical study, type 2 diabetes patients were treated with GKA for 12 weeks, and their plasma metabolite profiles were analyzed. The results revealed a significant increase in the concentration of large HDL particles, accompanied by an increase in the levels of cholesteryl esters in large HDL and cholesterol in large HDL. Researchers have proposed that GKA induces the activation of glucokinase in pancreatic β-cells, resulting in increased insulin secretion, which in turn enhances the reverse cholesterol transport pathway. This enables cholesterol to flow out from peripheral cells into HDL particles, contributing to the increase in HDL-related metabolites [44]. GKA can increase the levels of phospholipids in large VLDLs and phospholipids in very large VLDLs [45]. Through proteomic analysis, this study identified several proteins involved in phospholipid synthesis and metabolism that were upregulated by GKA. These proteins include enzymes such as phosphatidylcholine synthase, which is crucial for the synthesis of phosphatidylcholine, a major phospholipid in VLDL particles. The increase in phospholipids was associated with improved VLDL particle stability and reduced susceptibility to oxidation, which may be beneficial for cardiovascular health. There is also evidence indicating that GKA has a certain influence on the metabolism of free cholesterol in different lipoprotein particles. By combining in vitro and in vivo research methods, some scholars have demonstrated that GKA treatment increases the levels of free cholesterol in large HDL and free cholesterol in chylomicrons and extremely large VLDLs. They reported that GKA enhanced the activity of cholesterol ester transfer protein (CETP), which promoted the transfer of cholesterol esters from HDL to other lipoprotein particles in exchange for free cholesterol. This increased the availability of free cholesterol in these lipoprotein particles, leading to an increase in free cholesterol levels [46]. GKA improved glucose metabolism and promoted insulin secretion. Insulin can activate the production of VLDL-related proteins, thereby increasing the concentrations of large VLDL particles and very large VLDL particles as well as triglyceride levels. This finding was also consistent with our research results [47]. Jiang et al. explored the impact of GKA on glutamine metabolism [48]. Research has shown that GKA treatment increases the level of glutamine in plasma. They reported that GKA activated the expression of genes involved in glutamine synthesis, such as glutamine synthetase. The increase in glutamine levels was associated with improved nitrogen metabolism and increased antioxidant defense mechanisms since glutamine is an important precursor for the synthesis of glutathione, a key antioxidant molecule. An in vitro and in vivo experiment investigated the effect of GKA on hepatic lipid metabolism [49]. The study revealed that GKA enhanced the utilization of glucose in the liver. Moreover, the levels of key enzymes involved in triglyceride synthesis, such as fatty acid synthase, decreased, accompanied by a reduction in de novo lipogenesis. As a result, the production of triglyceride-rich VLDL particles is likely to decline, which directly impacts VLDL-related metabolites in the circulation. This finding was different from our findings, suggesting that there may be unique regulatory mechanisms that require further exploration. In conclusion, the combination of our study and previous research provided evidence for a significant association between GKA and lipid-related circulating metabolites.
The mediating effect of circulating metabolites on the association between GKA and AF
Previous studies have shown that the size of very-low-density lipoprotein (VLDL) particles is closely linked to the risk of atrial fibrillation (AF) [50]. Typically, appropriately sized VLDL particles are essential for maintaining normal heart metabolism and function. Glucokinase (GCK), an important regulatory enzyme, may regulate the average diameter of VLDL particles through complex metabolic pathways [51, 52]. On the one hand, large VLDL particles, due to their excessive size, have difficulty penetrating intact endothelium, thus presenting a lower risk of direct deposition in arterial walls compared to small VLDL particles [53]. On the other hand, their smaller surface area-to-volume ratio makes them less susceptible to oxidative modification, thereby reducing oxidative stress [54]. When GCK functions properly, it balances VLDL particle assembly and metabolism, maintaining the average diameter within a range that is beneficial for heart health. In this range, VLDL particles can effectively transport lipids such as cholesterol and triglycerides to cardiomyocytes, providing energy, maintaining heart rhythm and function, and reducing AF risk.
Strengths and limitations
Dorzagliatin, a glucokinase activator developed in China, has been officially marketed. Our study investigated the potential long-term impacts of glucokinase activators (GKAs) on cardiovascular and cerebrovascular outcomes and conducted a preliminary analysis of potential mediators.
One of the major strengths of our study was the application of the Mendelian randomization approach. Specifically, this method effectively reduced the risks of confounding factors and reverse causation bias, thereby compensating for the limitations of observational studies. By leveraging this method, we can obtain more reliable insights into the relationships under investigation. Another strength was the use of multiple large-scale datasets in our analysis. This substantially enhanced the statistical power of our study and enabled us to detect relatively subtle yet potentially significant effects. These datasets provide a solid foundation for us to identify possible associations between GKAs and cardiovascular and cerebrovascular diseases and to analyze the potential mediating roles of circulating metabolites.
The comprehensiveness of our study allowed us to uncover complex relationships and potential mediating pathways that might remain undetected in more restricted analyses. We further explored the uncovered relationships and pathways by taking into account different factors and variables, which contributed to a more in-depth understanding of the subject. Furthermore, we employed a series of sensitivity analyses to evaluate the stability of our research findings. These analyses were crucial for validating the assumptions of Mendelian randomization and instilled confidence in the stability of our results.
However, we acknowledge certain limitations and suggest that our results should be interpreted with due consideration of these factors. Although our findings reflected the relationship between glucokinase activators and cardiovascular and cerebrovascular events to some extent, it is important to emphasize that this relationship more closely approximates the impact of lifelong physiological glucokinase activation on cardio-cerebrovascular outcomes rather than that of pharmaceutical intervention. The effect sizes obtained do not equate to the actual clinical efficacy in treating these diseases. Therefore, caution is warranted when directly extrapolating Mendelian randomization results to all populations or equating them entirely to pharmaceutical therapeutic effects.
Our study was confined to participants of European ancestry, which might limit the generalizability of our findings to other ethnic groups. Given the potential differences in genetic architecture, metabolic profiles, and cardiovascular disease epidemiology across diverse populations, validation of these findings in non-European cohorts is critical to establishing the broader applicability of our results. Future studies with larger sample sizes, more refined genetic instruments, deeper understanding of the relevant biological pathways, and explicit inclusion of multi-ethnic cohorts may uncover associations that were not detected in our current analysis and strengthen the generalizability of the observed relationships.