Integrating machine learning algorithms and single-cell analysis to identify gut microbiota-related macrophage biomarkers in atherosclerotic plaques.

Objective: The relationship between macrophages and the gut microbiota in patients with atherosclerosis remains poorly defined, and effective biological markers are lacking. This study aims to elucidate the interplay between gut microbial communities and macrophages, and to identify biomarkers associated with the destabilization of atherosclerotic plaques. The goal is to enhance our understanding of the underlying molecular pathways and to pave new avenues for diagnostic approaches and therapeutic strategies in the disease. Methods: This study employed Weighted Gene Co-expression Network Analysis (WGCNA) and differential expression analysis on atherosclerosis datasets to identify macrophage-associated genes and quantify the correlation between these genes and gut microbiota gene sets. The Random Forest algorithm was utilized to pinpoint PLEK, IRF8, BTK, CCR1, and CD68 as gut microbiota-related macrophage genes, and a nomogram was constructed. Based on the top five genes, a Non-negative Matrix Factorization (NMF) algorithm was applied to construct gut microbiota-related macrophage clusters and analyze their potential biological alterations. Subsequent single-cell analyses were conducted to observe the expression patterns of the top five genes and the interactions between immune cells. Finally, the expression profiles of key molecules were validated using clinical samples from atherosclerosis patients. Results: Utilizing the Random Forest algorithm, we ultimately identified PLEK, IRF8, CD68, CCR1, and BTK as gut microbiota-associated macrophage genes that are upregulated in atherosclerotic plaques. A nomogram based on the expression of these five genes was constructed for use as an auxiliary tool in clinical diagnosis. Single-cell analysis confirmed the specific expression of gut microbiota-associated macrophage genes in macrophages. Clinical samples substantiated the high expression of PLEK in unstable atherosclerotic plaques. Conclusion: Gut microbiota-associated macrophage genes (PLEK, IRF8, CD68, CCR1, and BTK) may be implicated in the pathogenesis of atherosclerotic plaques and could serve as diagnostic markers to aid patients with atherosclerosis.

Impact of Probiotic Lactiplantibacillus plantarum ATCC 14917 on atherosclerotic plaque and its mechanism.

Atherosclerosis is viewed as not just as a problem of lipid build-up in blood vessels, but also as a chronic inflammatory disease involving both innate and acquired immunity. In atherosclerosis, the inflammation of the arterial walls is the key characteristic that significantly contributes to both the instability of plaque and the occlusion of arteries by blood clots. These events ultimately lead to stroke and acute coronary syndrome. Probiotics are living microorganisms that, when consumed in the right quantities, offer advantages for one's health. The primary objective of this study was to investigate the influence of Lactiplantibacillus plantarum ATCC 14917 (ATCC 14917) on the development of atherosclerotic plaques and its underlying mechanism in Apo lipoprotein E-knockout (Apoe-/- mice). In this study, Apoe-/- mice at approximately 8 weeks of age were randomly assigned to three groups: a Normal group that received a normal chow diet, a high fat diet group that received a gavage of PBS, and a Lactiplantibacillus plantarum ATCC 14917 group that received a high fat diet and a gavage of 0.2 ml ATCC 14917 (2 × 109 CFU/mL) per day for a duration of 12 weeks. Our strain effectively reduced the size of plaques in Apoe-/- mice by regulating the expression of inflammatory markers, immune cell markers, chemokines/chemokine receptors, and tight junction proteins (TJPs). Specifically, it decreased the levels of inflammatory markers (ICAM-1, CD-60 MCP-1, F4/80, ICAM-1, and VCAM-1) in the thoracic aorta, (Ccr7, cd11c, cd4, cd80, IL-1ß, TNF-α) in the colon, and increased the activity of ROS-scavenging enzymes (SOD-1 and SOD-2). It also influenced the expression of TJPs (occludin, ZO-1, claudin-3, and MUC-3). In addition, the treatment of ATCC 14917 significantly reduced the level of lipopolysaccharide in the mesenteric adipose tissue. The findings of our study demonstrated that our strain effectively decreased the size of atherosclerotic plaques by modulating inflammation, oxidative stress, intestinal integrity, and intestinal immunity.

Role of periodontal pathogens in atherosclerotic plaque development and progression: An overview.

Atherosclerosis is a progressive disease marked by the accumulation of lipids and fibrous components in the large arteries. It is one of the primary causes of heart disease and stroke. Periodontal diseases encompass conditions like gingivitis and periodontitis, which are multifactorial diseases associated with dysbiotic plaque biofilms that trigger an immune-inflammatory host response, eventually resulting in the destruction of periodontal tissues. Links between periodontal disease and atherosclerosis may be based on direct invasion of periodontal pathogens or inflammatory mechanisms triggered by bacteria related to periodontal lesions, locally or systemically, that may impact the initiation of the atherosclerotic lesion. The presence of periodontal pathogens within an atheromatous lesion implies hematogenous dissemination. The invasion of atheroma by periodontal pathogens results in changes in the proatherogenic and proinflammatory properties of endothelial cells, leading to endothelial dysfunction, which is a hallmark of atherosclerosis. Clinical and epidemiological studies have offered sufficient evidence of periodontitis having an adverse effect on systemic health, including atherosclerosis; however, a direct causal effect has not yet been proved. This review aims to analyse scientific results regarding the mechanism by which periodontal pathogens may cause atherosclerosis as well as to describe the role of Porphyromonas gingivalis in atherosclerotic plaque development and progression.

Oral Porphyromonas gingivalis infection affects intestinal microbiota and promotes atherosclerosis.

AIM: The link between periodontitis and intestinal dysbiosis, two factors that contribute to atherosclerosis, has not been clearly defined. We investigated the integrative effects of oral infection with Porphyromonas gingivalis (PG), the major pathogen for periodontitis, on intestinal microbiota and atherosclerosis. MATERIALS AND METHODS: ApoE-/- mice were fed a normal chow diet (NC), a Western diet (WD) or a WD with oral PG infection (PG). The PG infection was investigated by placing a total of 109 CFUs of live PG into the oral cavity of each mouse using a feeding needle five times a week for 3 weeks. Atherosclerotic lesions of the aortae were measured, and blood lipoproteins and the expression of molecules related to lipid metabolism in the liver were analysed. We also performed 16S RNA sequencing and a microbiome analysis using faeces. RESULTS: En face bloc preparation of the aortae showed that the PG group had a 1.7-fold increase in atherosclerotic lesions compared with the WD group (p < .01). Serum analyses showed that oral PG infection induced a significant decrease in high-density lipoprotein (HDL) and triglyceride. Western blots of hepatic tissue lysates revealed that PG infection reduced the expression of scavenger receptor class B type 1 (SR-B1) in the liver by 50%. Faecal microbiota analysis revealed that species richness estimates (Chao1, ACE) decreased immediately after PG infection. PG infection also induced a significant decrease in Shannon diversity and an increase in Simpson's indices in the WD-fed mice. PG infection significantly increased the phyla Actinobacteria and Deferribacteres, along with the species Mucispirillum schaedleri and Lactobacillus gasseri, in the mice. The functional study showed that PG infection increased the expression of proteins that function in carbohydrate and glucose metabolism, including phosphotransferase system (PTS) proteins and the GntR family transcriptional regulator. CONCLUSIONS: Oral PG infection promotes atherosclerosis and induces significant metabolic changes, including reduced serum HDL and reduced hepatic SR-B1 and ABCA1 expression, as well as changes in intestinal microbiota. Our study suggests that intestinal dysbiosis accompanies periodontitis and could play a role in atherosclerosis.

Effects of Berries, Phytochemicals, and Probiotics on Atherosclerosis through Gut Microbiota Modification: A Meta-Analysis of Animal Studies.

Atherosclerosis is a major cause of death and disability. The beneficial effects of phytochemicals and probiotics on atherosclerosis have gained significant interest since these functional foods can improve inflammation, oxidative stress, and microbiome dysbiosis. The direct effect of the microbiome in atherosclerosis, however, needs further elucidation. The objective of this work was to investigate the effects of polyphenols, alkaloids, and probiotics on atherosclerosis using a meta-analysis of studies with mouse models of atherosclerosis. Identification of eligible studies was conducted through searches on PubMed, Embase, Web of Science, and Science Direct until November 2022. The results showed that phytochemicals reduced atherosclerosis, which was significant in male mice, but not in females. Probiotics, on the other hand, showed significant reductions in plaque in both sexes. Berries and phytochemicals modulated gut microbial composition by reducing the Firmicutes/Bacteroidetes (F/B) ratio and by upregulating health-promoting bacteria, including Akkermansia muciniphila. This analysis suggests that phytochemicals and probiotics can reduce atherosclerosis in animal models, with a potentially greater effect on male animals. Thus, consumption of functional foods rich in phytochemicals as well as probiotics are viable interventions to improve gut health and reduce plaque burden in patients suffering from cardiovascular disease (CVD).

Capsaicin Ameliorates High-Fat Diet-Induced Atherosclerosis in ApoE-/- Mice via Remodeling Gut Microbiota.

Capsaicin is a pungent alkaloid abundantly present in peppers with outstanding biological activities, including the anti-atherosclerosis effect. Previous studies revealed that gut microbiota played an important role in the beneficial effects of capsaicin, but whether it is essential for the anti-atherosclerosis effect of capsaicin is unclear. This study evaluated the anti-atherosclerosis effect of capsaicin in ApoE-/- mice and further explored the role of depleting gut microbiota in the improvement of atherosclerosis. The results showed that capsaicin administration could prevent the development of atherosclerosis and improve serum lipids and inflammation, while antibiotic intervention abolished the alleviation of atherosclerosis by capsaicin. In addition, capsaicin administration could significantly increase the abundance of Turicibacter, Odoribacter, and Ileibacterium in feces, and decrease the abundance of deoxycholic acid, cholic acid, hypoxanthine, and stercobilin in cecal content. Our study provides evidence that gut microbiota plays a critical role in the anti-atherosclerosis effect of capsaicin.

Periodontal status and the incidence of selected bacterial pathogens in periodontal pockets and vascular walls in patients with atherosclerosis and abdominal aortic aneurysms.

The aim of the study was to examine the periodontal status of patients with atherosclerosis and abdominal aortic aneurysms. The occurrence of 5 periodontopathogens was evaluated in periodontal pockets and atheromatous plaques together with specimens from pathologically changed vascular walls of aortic aneurysms. The study comprised 39 patients who qualified for vascular surgeries. Patients with periodontitis and concomitant atherosclerosis or aneurysms were enrolled in the study. Periodontal indices were evaluated, and subgingival plaque samples were examined together with atheromatous plaques or specimens from vascular walls to identify, by polymerase chain reaction (PCR), the following periodontopathogens: Porphyromonas gingivalis, Tanarella forsythia, Aggregatibacter actinomycetemcomitans, Prevotella intermedia and Treponema denticola. The majority of patients had chronic severe generalized periodontitis in stages III and IV. Laboratory investigations showed the occurrence of one or more of the five targeted periodontopathogens in 94.6% of the periodontal pockets examined. Of the examined periodontopathogens, only Porphyromonas gingivalis was confirmed in 1 atheromatous plaque sample collected from the wall of an aortic aneurysm. Therefore, the occurrence of this bacterium in these vessels was considered to be occasional in patients with chronic periodontitis.

Guanxin Xiaoban capsules could treat atherosclerosis by affecting the gut microbiome and inhibiting the AGE-RAGE signalling pathway.

Introduction. Atherosclerosis is a chronic disorder in which plaque builds up in the arteries and is associated with several cardiovascular and cerebrovascular diseases such as coronary artery disease, cerebral infarction and cerebral haemorrhage. Therefore, there is an urgent need to discover new medications to treat or prevent atherosclerosis.Hypothesis/Gap Statement. The active components of Guanxin Xiaoban capsules may have an effect on the gut microbiome of patients with atherosclerosis and have a role in their therapeutic targets.Aim. The aim of this study was to identify genes and pathways targeted by active ingredients in Guanxin Xiaoban capsules for the treatment of atherosclerosis based on network pharmacology and analysis of changes to the gut microbiome.Methods. Mice were treated with Guanxin Xiaoban capsules. The 16S rDNA genome sequence of all microorganisms from each group of faecal samples was used to evaluate potential structural changes in the gut microbiota after treatment with Guanxin Xiaoban capsules. Western blotting and real-time quantitative PCR were used to detect gene targets in aortic and liver tissues. Haematoxylin and eosin staining was used to observe improvements in mouse arterial plaques.Results. The gut microbiota of atherosclerotic mice is disturbed. After Guanxin Xiaoban treatment, the abundance of bacteria in the mice improved, with an increase in the proportion of Akkermansia and a significant decrease in the proportion of Faecalibaculum. The main ingredients of Guanxin Xiaoban capsules are calycosin, liquiritin, ferulic acid, ammonium glycyrrhizate, aloe emodin, rhein and emodin. The core genes of this network were determined to be glutathione S-transferase mu 1 (GSTM1), vascular endothelial growth factor A (VEGFA) and cyclin-dependent kinase inhibitor 1A (CDKN1A). The compound-target gene network revealed an interaction between multiple components and targets and contributed to a better understanding of the potential therapeutic effects of the capsules on atherosclerosis. In addition, expression of the AGE-receptor for the AGE (RAGE) pathway was significantly inhibited and the mice showed signs of arterial plaque reduction. Guanxin Xiaoban capsules may improve atherosclerosis and reduce the plaque area by inhibiting the AGE-RAGE signalling pathway to delay the development of atherosclerosis. This mechanism appears to involve changes in the gut microbiota. Therefore, Guanxin Xiaoban capsules have potential value as a treatment for atherosclerosis.

Deficiency of PSRC1 accelerates atherosclerosis by increasing TMAO production via manipulating gut microbiota and flavin monooxygenase 3.

Maladaptive inflammatory and immune responses are responsible for intestinal barrier integrity and function dysregulation. Proline/serine-rich coiled-coil protein 1 (PSRC1) critically contributes to the immune system, but direct data on the gut microbiota and the microbial metabolite trimethylamine N-oxide (TMAO) are lacking. Here, we investigated the impact of PSRC1 deletion on TMAO generation and atherosclerosis. We first found that PSRC1 deletion in apoE-/- mice accelerated atherosclerotic plaque formation, and then the gut microbiota and metabolites were detected using metagenomics and untargeted metabolomics. Our results showed that PSRC1 deficiency enriched trimethylamine (TMA)-producing bacteria and functional potential for TMA synthesis and accordingly enhanced plasma betaine and TMAO production. Furthermore, PSRC1 deficiency resulted in a proinflammatory colonic phenotype that was significantly associated with the dysregulated bacteria. Unexpectedly, hepatic RNA-seq indicated upregulated flavin monooxygenase 3 (FMO3) expression following PSRC1 knockout. Mechanistically, PSRC1 overexpression inhibited FMO3 expression in vitro, while an ERα inhibitor rescued the downregulation. Consistently, PSRC1-knockout mice exhibited higher plasma TMAO levels with a choline-supplemented diet, which was gut microbiota dependent, as evidenced by antibiotic treatment. To investigate the role of dysbiosis induced by PSRC1 deletion in atherogenesis, apoE-/- mice were transplanted with the fecal microbiota from either apoE-/- or PSRC1-/-apoE-/- donor mice. Mice that received PSRC1-knockout mouse feces showed an elevation in TMAO levels, as well as plaque lipid deposition and macrophage accumulation, which were accompanied by increased plasma lipid levels and impaired hepatic cholesterol transport. Overall, we identified PSRC1 as an atherosclerosis-protective factor, at least in part, attributable to its regulation of TMAO generation via a multistep pathway. Thus, PSRC1 holds great potential for manipulating the gut microbiome and alleviating atherosclerosis.

Effects of Whole Brown Bean and Its Isolated Fiber Fraction on Plasma Lipid Profile, Atherosclerosis, Gut Microbiota, and Microbiota-Dependent Metabolites in Apoe-/- Mice.

The health benefits of bean consumption are widely recognized and are largely attributed to the dietary fiber content. This study investigated and compared the effects of whole brown beans and an isolated bean dietary fiber fraction on the plasma lipid profile, atherosclerotic plaque amount, gut microbiota, and microbiota-dependent metabolites (cecal short-chain fatty acids (SCFAs) and plasma methylamines) in Apoe-/- mice fed high fat diets for 10.5 weeks. The results showed that both whole bean and the isolated fiber fraction had a tendency to lower atherosclerotic plaque amount, but not plasma lipid concentration. The whole bean diet led to a significantly higher diversity of gut microbiota compared with the high fat diet. Both bean diets resulted in a lower Firmicutes/Bacteroidetes ratio, higher relative abundance of unclassified S24-7, Prevotella, Bifidobacterium, and unclassified Clostridiales, and lower abundance of Lactobacillus. Both bean diets resulted in higher formation of all cecal SCFAs (higher proportion of propionic acid and lower proportion of acetic acid) and higher plasma trimethylamine N-oxide concentrations compared with the high fat diet. Whole beans and the isolated fiber fraction exerted similar positive effects on atherosclerotic plaque amount, gut microbiota, and cecal SCFAs in Apoe-/- mice compared with the control diets.

Differences in local immune cell landscape between Q fever and atherosclerotic abdominal aortic aneurysms identified by multiplex immunohistochemistry.

Background: Chronic Q fever is a zoonosis caused by the bacterium Coxiella burnetii which can manifest as infection of an abdominal aortic aneurysm (AAA). Antibiotic therapy often fails, resulting in severe morbidity and high mortality. Whereas previous studies have focused on inflammatory processes in blood, the aim of this study was to investigate local inflammation in aortic tissue. Methods: Multiplex immunohistochemistry was used to investigate local inflammation in Q fever AAAs compared to atherosclerotic AAAs in aorta tissue specimen. Two six-plex panels were used to study both the innate and adaptive immune systems. Results: Q fever AAAs and atherosclerotic AAAs contained similar numbers of CD68+ macrophages and CD3+ T cells. However, in Q fever AAAs, the number of CD68+CD206+ M2 macrophages was increased, while expression of GM-CSF was decreased compared to atherosclerotic AAAs. Furthermore, Q fever AAAs showed an increase in both the number of CD8+ cytotoxic T cells and CD3+CD8-FoxP3+ regulatory T cells. Finally, Q fever AAAs did not contain any well-defined granulomas. Conclusions: These findings demonstrate that despite the presence of pro-inflammatory effector cells, persistent local infection with C. burnetii is associated with an immune-suppressed microenvironment. Funding: This work was supported by SCAN consortium: European Research Area - CardioVascualar Diseases (ERA-CVD) grant [JTC2017-044] and TTW-NWO open technology grant [STW-14716].

Interaction between bacteria and cholesterol crystals: Implications for endocarditis and atherosclerosis.

BACKGROUND: The interaction between pathogenic bacteria and cholesterol crystals (CCs) has not been investigated. However, CCs are found extensively in atherosclerotic plaques and sclerotic cardiac valves. Interactions between pathogenic bacteria and CCs could provide insights into destabilization of atherosclerotic plaques and bacterial adhesion to cardiac valves. METHODS: Staphylococcus aureus and Pseudomonas aeruginosa were used to assess in vitro bacterial adhesion to CCs and proliferation in the presence of CCs compared to plastic microspheres and glass shards as controls. Ex vivo studies evaluated bacterial adhesion to atherosclerotic rabbit arteries compared to normal arteries and human atherosclerotic carotid plaques compared to normal carotid arteries. Scanning electron microscopy (SEM) was used to visualize bacterial adhesion to CCs and confocal microscopy was used to detect cholesterol binding to bacteria grown in the presence or absence of CCs. RESULTS: In vitro, S. aureus and P. aeruginosa displayed significantly greater adhesion, 36% (p<0.0001) and 89% (p<0.0001), respectively, and growth upon exposure to CCs compared to microspheres or glass shards. Rabbit and human atherosclerotic arteries contained significantly greater bacterial burdens compared to controls (4× (p<0.0004); 3× (p<0.019), respectively. SEM demonstrated that bacteria adhered and appeared to degrade CCs. Consistent with this, confocal microscopy indicated increased cholesterol bound to the bacterial cells. CONCLUSIONS: This study is the first to demonstrate an interaction between bacteria and CCs showing that bacteria dissolve and bind to CCs. This interaction helps to elucidate adhesion of bacteria to sclerotic valves and atherosclerotic plaques that may contribute to endocarditis and plaque destabilization.

Extracellular vesicles from helicobacter pylori-infected cells and helicobacter pylori outer membrane vesicles in atherosclerosis.

BACKGROUND: The role of H. pylori infection has been reported in various extragastric diseases, particularly, the correlation between H. pylori and atherosclerosis (AS) have received lots of attention. Some scholars demonstrated that the presence of H. pylori-specific DNA in the sclerotic plaques of atheromatous patients provides biological evidences, with indicating that H. pylori infection is a potential factor of AS. However, the underlying mechanism of H. pylori or their products cross the epithelial barriers to enter the blood circulation remains unclear. Recent studies have shown that the extracellular vesicles (EVs) derived from H. pylori-infected gastric epithelial cells encapsulated H. pylori virulence factor cytotoxin-associated gene A (CagA) and existed in the blood samples of patients or mice, which indicating that they can carry CagA into the blood circulation. Based on these findings, some researchers proposed a hypothesis that H. pylori is involved in the pathogenesis of AS via EVs-based mechanisms. In addition, outer membrane vesicles (OMVs) serve as transport vehicles to deliver H. pylori virulence factors to epithelial cells. It is necessary to discuss the role of H. pylori OMVs in the development of AS. OBJECTIVES: This review will focus on the correlation between H. pylori infection and AS and tried to unveil the possible role of EVs from H. pylori-infected cells and H. pylori OMVs in the pathogenesis of AS, with a view to providing help in refining our knowledge in this aspect. METHODS: All of information included in this review was retrieved from published studies on H. pylori infection in AS. RESULTS: H. pylori infection may be an atherosclerotic risk factor and drives researchers to reevaluate the role of H. pylori in the pathogenesis of AS. Some findings proposed a new hypothesis that H. pylori may be involved in the pathogenesis of AS through EVs-based mechanisms. Besides EVs from H. pylori-infected cells, whether H. pylori OMVs may play some role in the pathogenesis of AS is still remain unclear. CONCLUSION: Existing epidemiological and clinical evidence had shown that there is a possible association between H. pylori and AS. However, except for the larger randomized controlled trials, more basic research about EVs from H. pylori-infected cells and H. pylori OMVs is the need of the hour to unveil the possible role of H. pylori infection in the pathogenesis of AS.

Characterization of Salivary Microbiota in Patients with Atherosclerotic Cardiovascular Disease: A Case-Control Study.

AIMS: Oral bacteria have been reported to be associated with the pathogenesis of atherosclerosis; however, the relationship between the oral microbiota and atherosclerosis remains unclear. The present study aimed to investigate whether or not salivary microbiota of patients with atherosclerotic cardiovascular disease (ACVD) differs from that of subjects without ACVD, and to characterize the salivary microbiota of patients with ACVD. METHODS: This study included 43 patients with ACVD and 86 age- and sex-matched non-ACVD individuals. 16S rRNA metagenomic analysis were performed using DNA isolated from the saliva samples of the participants. To select unique operational taxonomic unit (OTU) sets of ACVD, we conducted the random forest algorithm in machine learning, followed by confirmation via 10-fold cross-validation Results: There was no difference in richness or evenness between the ACVD and non-ACVD groups (alpha diversity; observed OTU index, p=0.503; Shannon's index, p=0.478). However, significant differences were found in the overall salivary microbiota structure (beta diversity; unweighted UniFrac distances, p=0.001; weighted UniFrac distances, p=0.001). The Actinobacteria phylum was highly abundant in patients with ACVD, while the Bacteroidetes phylum was less abundant. The random forest classifier identified 43 OTUs as an optimal marker set of ACVD. In a 10-fold cross validation using the validation data, an area under the curve (AUC) of 0.933 (95% CI, 0.855-1.000) was obtained. CONCLUSIONS: The salivary microbiota in patients with ACVD was distinct from that of non-ACVD individuals, indicating that the salivary microbiota may be related to ACVD.

Association between helicobacter pylori infection and subclinical atherosclerosis: A systematic review and meta-analysis.

BACKGROUND: The relationship between Helicobacter pylori (H. pylori) infection and subclinical atherosclerosis has been confirmed, but these conclusions are still controversial. Therefore, we have performed a systematic review and meta-analysis to assess the association between H. pylori infection and subclinical atherosclerosis. METHODS: Databases including PubMed, Embase, Web of Science were searched for the articles on the association of carotid intima-media thickness or pulse wave velocity with H. pylori infection published up to January 1, 2020. Stata 12.0 was used to calculate standardized mean difference (SMD) and 95% confidence interval (95% CI); the I2 test was used to evaluate heterogeneity between studies and sensitivity analysis and subgroup analysis were used to explore the source of heterogeneity. Funnel plot, Begg test, and Egger test were used to estimate publication bias. RESULTS: Data were extracted from 18 studies involving 6776 subjects with H. pylori positive and 7794 with H. pylori negative. H. pylori positive subjects is significantly associated with increased subclinical atherosclerosis as determined by carotid intima-media thickness (SMD: 0.376 mm; 95% CI: 0.178, 0.574; P < .001, I2 = 90.6%), pulse wave velocity (SMD: 0.320 m/s; 95% CI: 0.242, 0.398; P < .001, I2 = 52.6%), compared with H. pylori negative. Similar results were observed when subgroups analysis were stratified according to age, male ratio, geographical location, H. pylori diagnosis, and study design. Sensitivity analyses showed that our results were robust. The Begg test or Egger test showed no significant publication bias (all P > .05). CONCLUSIONS: This meta-analysis confirmed a significant association between H. pylori and subclinical atherosclerosis, which will help H. pylori patients to establish effective strategies for the prevention and control of cardiovascular events.

Cytotoxin-associated gene A (CagA) promotes aortic endothelial inflammation and accelerates atherosclerosis through the NLRP3/caspase-1/IL-1ß axis.

Atherosclerosis is a chronic inflammatory disease. Pathophysiological similarities between chronic infections and atherosclerosis triggered interests between these conditions. The seroepidemiological study showed that Helicobacter pylori strains that express cytotoxin-associated gene A (CagA), an oncoprotein and a major virulence factor, was positively correlated with atherosclerosis and related clinical events. Nevertheless, the underlying mechanism is poorly understood. In this study, the seroprevalence of infection by H. pylori and by strains express CagA assessed by enzyme-linked immunosorbent assay (ELISA) showed that the prevalence of CagA strains rather than H. pylori in patients was positively correlated with atherogenesis. Correspondingly, we found that CagA augmented the growth of plaque of ApoE-/- mice in the early stage of atherosclerosis and promoted the expression of adhesion molecules and inflammatory cytokines in mouse aortic endothelial cells (MAECs). Mechanistically, both si-NLRP3 and si-IL-1ß mitigated the promoting effect of CagA on the inflammatory activation of HAECs. In vivo, the inhibition of NLRP3 by MCC950 significantly attenuated the promoting effect of CagA on plaque growth of ApoE-/- mice. We also propose NLRP3 as a potential therapeutic target for CagA-positive H. pylori infection-related atherosclerosis and emphasize the importance of inflammation in atherosclerosis pathology.

The role of Akkermansia muciniphila in obesity, diabetes and atherosclerosis.

Alteration in the composition of the gut microbiota can lead to a number of chronic clinical diseases. Akkermansia muciniphila is an anaerobic bacteria constituting 3-5% of the gut microbial community in healthy adults. This bacterium is responsible for degenerating mucin in the gut; its scarcity leads to diverse clinical disorders. In this review, we focus on the role of A. muciniphila in diabetes, obesity and atherosclerosis, as well as the use of this bacterium as a next-generation probiotic. In regard to obesity and diabetes, human and animal trials have shown that A. muciniphila controls the essential regulatory system of glucose and energy metabolism. However, the underlying mechanisms by which A. muciniphila alleviates the complications of obesity, diabetes and atherosclerosis are unclear. At the same time, its abundance suggests improved metabolic disorders, such as metabolic endotoxemia, adiposity insulin resistance and glucose tolerance. The role of A. muciniphila is implicated in declining aortic lesions and atherosclerosis. Well-characterized virulence factors, antigens and cell wall extracts of A. muciniphila may act as effector molecules in these diseases. These molecules may provide novel mechanisms and strategies by which this bacterium could be used as a probiotic for the treatment of obesity, diabetes and atherosclerosis.

An Explorative Study on Monocyte Reprogramming in the Context of Periodontitis In Vitro and In Vivo.

Aims: Periodontitis is an independent risk factor for cardiovascular disease, but the mechanistic link is not fully understood. In atherosclerotic cardiovascular disease, monocytes can adopt a persistent hyperresponsive phenotype, termed trained immunity. We hypothesized that periodontitis-associated bacteria can induce trained immunity in monocytes, which subsequently accelerate atherosclerosis development. Materials and Methods: We combined in vitro experiments on human primary monocytes and in vivo techniques in patients with periodontitis to test this hypothesis. Adherent peripheral blood mononuclear cells (PBMCs) were transiently exposed in vitro to Porphyromonas gingivalis for 24 hours, and restimulated with lipopolysaccharide (LPS) or Pam3CysK4 (P3C) six days later, to measure interleukin-6 (IL-6) and tumor necrosis factor α (TNFα) production. In an exploratory observational study, patients with severe periodontitis (63 ± 6 years, n=14) and control subjects with no-to-mild periodontitis (54 ± 10 years, n=14) underwent venipuncture and 2'-deoxy-2'-[18F]fluoro-D-glucose positron-emission-tomography ([18F]FDG PET/CT) scanning. Results: When adherent peripheral blood mononuclear cells (PBMCs) were transiently exposed in vitro to Porphyromonas gingivalis for 24 hours, and restimulated with LPS or P3C six days later, IL-6 and TNFα production was significantly increased (TNFα/P3C, p<0.01). Circulating leukocytes, IL-6 and interleukin-1 receptor antagonist (IL-1Ra) concentrations were generally higher in patients compared to controls (leukocytes: p<0.01; IL-6: p=0.08; IL-1Ra: p=0.10). Cytokine production capacity in PBMCs after 24h stimulation revealed no differences between groups. [18F]FDG PET/CT imaging showed a trend for increased [18F]FDG-uptake in the periodontium [mean standard uptake value (SUVmean), p=0.11] and in femur bone marrow (SUVmean, p=0.06), but no differences were observed for vascular inflammation. Positive correlations between severity of periodontitis, measured by The Dutch Periodontal Screening Index and pocket depth, with circulating inflammatory markers and tissue inflammation were found. Conclusions: P. gingivalis induces long-term activation of human monocytes in vitro (trained immunity). Patients with severe periodontitis did have signs of increased systemic inflammation and hematopoietic tissue activation. However, their circulating monocytes did not show a hyperresponsive phenotype. Together we suggest that trained immunity might contribute to local periodontal inflammation which warrants further investigation.

Enterobacter aerogenes ZDY01 inhibits choline-induced atherosclerosis through CDCA-FXR-FGF15 axis.

Atherosclerosis is the leading cause of cardiovascular diseases worldwide. Trimethylamine N-oxide (TMAO), a metabolite of intestinal flora from dietary quaternary amines, has been shown to be closely related to the development of atherosclerosis. Previous studies have shown that Enterobacter aerogenes ZDY01 significantly reduces the serum levels of TMAO and cecal trimethylamine (TMA) in Balb/c mice; however, its role in the inhibition of choline-induced atherosclerosis in ApoE-/- mice remains unclear. Here, we demonstrated that E. aerogenes ZDY01 inhibited choline-induced atherosclerosis in ApoE-/- mice fed with 1.3% choline by reducing cecal TMA and modulating CDCA-FXR/FGF15 axis. We observed that E. aerogenes ZDY01 decreased the cecal TMA and serum TMAO levels by utilizing cecal TMA as a nutrient, not by changing the expression of hepatic FMO3 and the composition of gut microbiota. Furthermore, E. aerogenes ZDY01 enhanced the expression of bile acid transporters and reduced the cecal CDCA levels, thereby attenuating the FXR/FGF15 pathway, upregulating the expression of Cyp7a1, promoting reverse cholesterol transport. Taken together, E. aerogenes ZDY01 attenuated choline-induced atherosclerosis in ApoE-/- mice by decreasing cecal TMA and promoting reverse cholesterol transport, implying that E. aerogenes ZDY01 treatment might have therapeutic potential in atherosclerosis.

The Role of Gut Microbiota on Cholesterol Metabolism in Atherosclerosis.

Hypercholesterolemia plays a causal role in the development of atherosclerosis and is one of the main risk factors for cardiovascular disease (CVD), the leading cause of death worldwide especially in developed countries. Current data show that the role of microbiota extends beyond digestion by being implicated in several metabolic and inflammatory processes linked to several diseases including CVD. Studies have reported associations between bacterial metabolites and hypercholesterolemia. However, such associations remain poorly investigated and characterized. In this review, the mechanisms of microbial derived metabolites such as primary and secondary bile acids (BAs), trimethylamine N-oxide (TMAO), and short-chain fatty acids (SCFAs) will be explored in the context of cholesterol metabolism. These metabolites play critical roles in maintaining cardiovascular health and if dysregulated can potentially contribute to CVD. They can be modulated via nutritional and pharmacological interventions such as statins, prebiotics, and probiotics. However, the mechanisms behind these interactions also remain unclear, and mechanistic insights into their impact will be provided. Therefore, the objectives of this paper are to present current knowledge on potential mechanisms whereby microbial metabolites regulate cholesterol homeostasis and to discuss the feasibility of modulating intestinal microbes and metabolites as a novel therapeutic for hypercholesterolemia.