Amongst the array of afflictions categorized as cardiovascular diseases (CVD), atherosclerosis reigns as a prevailing force. Characterized by the accumulation of lipids and pronounced inflammation within pivotal arteries, atherosclerosis advances steadily, potentially resulting in dire clinical consequences, such as myocardial infarction (MI) and stroke. Despite its gradual nature and declining prevalence in certain nations, this insidious malady continues to claim a prominent position among the leading causes of mortality within our global community [1]. Atherosclerosis has emerged as a matter of international apprehension, and despite remarkable progress in comprehending the origin and management of ailments, there still exists an inadequacy in its prevention [2, 3]. The discovery of shielding or instigating elements in atherosclerosis carries paramount importance and necessitates further exploration into newly found treatment objectives.

The “microbiota” (referring to the myriad of bacteria, viruses, and fungi comprising the human gut) plays a vital role in maintaining a harmonious and flourishing human ecological system that abundantly thrives within the intestinal tract. Specifically, bacteria contribute to the processes of food digestion, fortify the immune system, and generate unique metabolites capable of permeating the host’s bloodstream [1]. Consequently, a more all-encompassing perception of metabolism is presently being proffered, asserting that modifications in the host’s metabolic processes, in conjunction with interactions between the gut microbiota and the host’s remote organs, collectively impact the entirety of human metabolism [4]. Mounting evidence suggests that the gut microbiota (GM) exerts regulatory control over the host’s immune responses, inflammatory processes, metabolism, and cardiovascular function [5]. For instance, it has been reported in some studies that the metabolites of gut microbiota, such as trimethylamine N-oxide (TMAO), short-chain fatty acid butyrate, and indole-3-propionic acid (IPA), have indirect influences on the occurrence of atherosclerosis [4, 6, 7]. Despite extensive epidemiological research, the underlying cause of the association between the composition of the gut microbiota and various diseases, such as CVD, remains largely elusive [8]. The intricacy arises from the presence of various supplemental factors, such as gender, age, and ethnicity, which can potentially impact both the progression of atherosclerosis and the composition of the gut microbiota. It poses a considerable challenge to adequately encompass these variables within an observational study, thereby limiting the capacity to definitively establish a causal relationship between the gut microbiome and atherosclerosis.

However, additional investigations are necessary to ascertain the precise role that various gut microbiota taxa (GM taxa) play in the development of atherosclerosis. Mendelian randomization (MR), a novel method for exploring the relationship between gut microbiota and atherosclerosis in this context, bears a resemblance to randomized controlled trials (RCT) [9]. Single nucleotide polymorphisms (SNPs) are referred to as instrumental variables (IVs) in MR investigations, serving to quantify the causal connection between exposure and the outcome of interest [10]. SNPs adhere to the concept that genetic variation is randomly assigned during meiosis, thereby circumventing confounding influences and potential reverse causation, thereby ensuring that relationships between genetic variants and outcomes remain unaffected [11]. Consequently, MR analysis can swiftly identify the causal link between a specific exposure and an outcome, more so than RCTs. Thus, to determine the potential impact of GM taxa on atherosclerosis in three distinct vascular locations: atherosclerosis (excluding cerebral, coronary, and peripheral arterial disease), coronary atherosclerosis, and cerebral atherosclerosis, we conducted MR analysis by utilizing the genome-wide association study (GWAS) summary statistics from the FinnGen and MiBioGen consortia. This approach may provide validation for existing evidence and yield novel insights pertaining to the treatment and prevention of atherosclerosis.

The MR method was utilized to investigate a potential causal relationship between various GM taxa and atherosclerosis across three distinct vascular sites. The findings revealed a total of 20 causal connections, with one demonstrating strong causality and the remaining 19 showing nominal causality.

This study also unveiled a strong causal relationship following Bonferroni correction, which indicated that the Firmicutes phylum (ID: 1672) (OR = 0.852 (0.763, 0.950), p = 0.004) markedly mitigates the risk of coronary atherosclerosis. Therefore, this MR analysis reveals the protective function of the Firmicutes phylum in relation to coronary atherosclerosis. Among the assemblages prevalent in the human gut microbiota, the Firmicutes phylum stands out as a dominant faction [27]. The abundance of Eubacterium and Roseburia, which belong to the Firmicutes phylum, in the gut microbiota of patients with atherosclerosis was found to be reduced compared to the healthy control group [8, 28, 29]. Our findings align with those from other early studies and corroborate the assertion that patients with coronary atherosclerosis have a low abundance of the phylum Firmicutes in their intestinal microbiota. Growing evidence has demonstrated the impact that gut microbiome can have on cardiovascular health [30]. The gut microbiota may exert its influence on atherosclerosis through various mechanisms, with gut metabolites being regarded as one of the primary mechanisms impacting atherosclerosis [31]. Although research on the phylum Firmicutes is relatively limited, some observational studies and animal models have reported on the impact that metabolites produced by gut microbiota belonging to the phylum Firmicutes have on the occurrence and progression of atherosclerosis. For example, bacteria in the phylum Firmicutes, Clostridium, and Peptostreptococcaceae, are capable of producing IPA, a microbial metabolite derived from tryptophan. IPA has been found to downregulate the expression of miR-142-5p in macrophages, which promotes the transportation of extracellular cholesterol. Low levels of IPA have been associated with risk factors of coronary atherosclerosis, and this study also uncovered the potential of using IPA supplementation to suppress the progression of arterial atherosclerosis [32]. Meanwhile, Karlsson et al. [28] discovered that bacteria belonging to the phylum Firmicutes, specifically Ruminococcaceae spp, Eubacterium, and Roseburia, are capable of producing short-chain fatty acids (SCFAs). It has been observed that the abundance of these bacterial populations is reduced in patients with atherosclerosis [8, 28, 29], and SCFAs have been found to possess anti-inflammatory properties toward certain epithelial cells. This may potentially slow down the progression of atherosclerosis [33]. Furthermore, Rath et al. [34] found that Clostridia bacteria degrade nutrients such as phosphatidylcholine to produce TMAO, while the levels of TMAO are associated with the occurrence of atherosclerosis. However, current research has yet to confirm a causal relationship between them [31].

It is noteworthy that the criteria for Bonferroni correction are excessively stringent, sacrificing some statistical efficiency and potentially resulting in false negative outcomes. Initial associations between 19 gut microbiota taxa were observed in our study; however, these associations disappeared after applying the Bonferroni correction. This may be attributed to the possibility that a single microbiota genus might not play as substantial a role in disease as previously postulated, in relation to the gut microbiome and cardiovascular disease, which are influenced by multiple factors [35]. Furthermore, our results indicated several gut microbiota taxa with preliminary causal links, thereby corroborating prior research discoveries. For example, Karlsson et al. [28] observed decreased levels of Eubacterium in the gut microbiota of atherosclerosis patients compared to healthy controls. Likewise, Liu et al. [29] found that Ruminococcaceae were reduced in atherosclerosis patients compared to healthy controls. Although these gut microbiota taxa exhibit only preliminary causal associations at the genus level, further investigations into the collaboration and interaction among diverse gut microbiota taxa remain worthwhile.

Moreover, it is imperative to acknowledge the inherent constraints in our investigation. Primarily, we employed a rather lenient criterion (p 1 $\times$ 10${}^{-5}$) to filter the IVs owing to the minuscule count of IVs that satisfied the stringent threshold (p 5 $\times$ 10${}^{-8}$). Furthermore, to augment the study’s validity, future analyses necessitate the utilization of a larger sample size of GWAS data due to the relatively sparse frequency of cerebral atherosclerosis cases. Lastly, bearing in mind the predominantly European heritage of our participants, it is essential to exercise caution when generalizing these findings to other populations.

Overall, these MR research findings align with the majority of the previously published studies. These studies collectively suggest that the impact on atherosclerosis by the phylum Firmicutes is primarily mediated through its metabolites. Thus, increasing the abundance of Firmicutes in the gut microbiota may serve as a protective measure against atherosclerosis. However, further experimental validation is required to confirm this hypothesis. The results of our MR study can provide a confident direction for future research endeavors.

Conclusively, a total of 20 nominal connections and a strong causal relationship were successfully identified by conducting a meticulous MR analysis on the risk of distinct GM taxa in relation to atherosclerosis across three diverse vascular sites. Notably, the phylum Firmicutes has been scientifically linked to a reduced occurrence of coronary atherosclerosis, as the gathered data reveals. To shed light on the significant beneficial impact of Firmicutes on coronary atherosclerosis, along with its distinct protective mechanisms, conducting future RCT studies is imperative. However, these findings present a fresh perspective on potential strategies for the prevention and treatment of atherosclerosis by targeting gut microbiota and exploring prospective therapeutic targets for this condition.

Publicly available datasets were analyzed in this study. This data can be found at: https://mibiogen.gcc.rug.nl/, https://www.finngen.fi/fi.

YL and YXC designed the research study. YL performed the research. YL and YXC provided help and advice on data curation. ZL analyzed the data. YL wrote the manuscript. LQT was also involved in supervision the manuscript and contributed to the preliminary design of the study. YRL and YCC were both involved in data collection. YRL also contributed to the writing of the manuscript, while YCC participated in a portion of the data analysis and figure creation. All authors contributed to editorial changes in the manuscript. All authors read and approved the final manuscript. All authors have participated sufficiently in the work and agreed to be accountable for all aspects of the work.

This study utilized publicly available deidentified data from participant studies that had already been approved by an ethical standards committee. As a result, no additional separate ethical approval was necessary for this study and informed consent is waived.

We thank the MiBioGen consortium for the GWAS summary data and the participants and investigators of the FinnGen study.

This research received no external funding.

The authors declare no conflict of interest.