Gut microbiota, circulating inflammatory markers and metabolites, and carotid artery atherosclerosis in HIV infection

Abstract

Background: Alterations in gut microbiota have been implicated in HIV infection and cardiovascular disease. However, how gut microbial alterations relate to host inflammation and metabolite profiles, and their relationships with atherosclerosis, have not been well-studied, especially in the context of HIV infection. Here, we examined associations of gut microbial species and functional components measured by shotgun metagenomics with carotid artery plaque assessed by B-mode carotid artery ultrasound in 320 women with or at high risk of HIV (65% HIV +) from the Women's Interagency HIV Study. We further integrated plaque-associated microbial features with serum proteomics (74 inflammatory markers measured by the proximity extension assay) and plasma metabolomics (378 metabolites measured by liquid chromatography tandem mass spectrometry) in relation to carotid artery plaque in up to 433 women.

Results: Fusobacterium nucleatum, a potentially pathogenic bacteria, was positively associated with carotid artery plaque, while five microbial species (Roseburia hominis, Roseburia inulinivorans, Johnsonella ignava, Odoribacter splanchnicus, Clostridium saccharolyticum) were inversely associated with plaque. Results were consistent between women with and without HIV. Fusobacterium nucleatum was positively associated with several serum proteomic inflammatory markers (e.g., CXCL9), and the other plaque-related species were inversely associated with proteomic inflammatory markers (e.g., CX3CL1). These microbial-associated proteomic inflammatory markers were also positively associated with plaque. Associations between bacterial species (especially Fusobacterium nucleatum) and plaque were attenuated after further adjustment for proteomic inflammatory markers. Plaque-associated species were correlated with several plasma metabolites, including the microbial metabolite imidazole-propionate (ImP), which was positively associated with plaque and several pro-inflammatory markers. Further analysis identified additional bacterial species and bacterial hutH gene (encoding enzyme histidine ammonia-lyase in ImP production) associated with plasma ImP levels. A gut microbiota score based on these ImP-associated species was positively associated with plaque and several pro-inflammatory markers.

Conclusion: Among women living with or at risk of HIV, we identified several gut bacterial species and a microbial metabolite ImP associated with carotid artery atherosclerosis, which might be related to host immune activation and inflammation. Video Abstract.

Keywords: Atherosclerosis; Gut microbiota; HIV infection; Inflammatory markers; Metabolomics.

Fig. 1

Differentially abundant species according to carotid artery plaque status. A Taxonomic linear discriminative analysis (LDA) effect size (LefSe) analysis by carotid artery plaque status. B Integrated phylogenetic tree. Taxa from inner to outer circle represent bacteria kingdom to species level. Blue and red font represent positive and negative associations between species and plaque, respectively. C Associations between species and plaque status. Data are odds ratios (ORs) and 95% confidence intervals (CIs) for carotid artery plaque per standard deviation increment of CLR transformed abundance of gut bacterial species, adjusted for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use (model 1) and further adjusted for blood pressure, total cholesterol, high-density lipoprotein cholesterol, antihypertensive medication use, and lipid-lowering medication use (model 2). In addition, we also show ANCOM2 detection level of each species, adjusting for aforementioned covariates in model 1. D Associations between gut microbiota functional enzymes, plaque-associated bacteria species, and carotid artery plaque status. The Spearman correlation heatmap include 11 microbial functional enzymes under carbon–nitrogen ligases category and the 6 plaque-associated bacterial species. Associations between the 11 microbial functional enzymes and carotid artery plaque status were estimated by linear regression models after adjustment for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use

Fig. 2

Serum proteomic inflammatory markers and carotid artery plaque. A Serum proteomic inflammatory markers and carotid artery plaque: Partial Least Square-Discriminant Analysis (PLSDA) plot by plaque status. B The major contributors of each PLSDA PCs and their associations with carotid artery plaque. Data are odds ratios (ORs) and 95% confidence intervals (CIs), adjusted for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use

Fig. 3

Associations of plaque-associated bacterial species with host proteomic inflammatory markers. A Microbial species, proteomics PLSDA PCs, and their major contributors. B Associations between microbial species and plaque status, adjusted for proteomics profiles. Data are odds ratios (ORs) and 95% confidence intervals (CIs) for carotid artery plaque per standard deviation increment of CLR transformed abundance of gut bacterial species, adjusted for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use (model 1) and further adjusted for top 5 proteomics PLSDA PCs (model 2). Abbreviations: PLSDA, partial least square-discriminant analysis

Fig. 4

Plasma metabolomic profiles, gut microbiome, and serum proteomic inflammatory markers, in relation to carotid artery plaque. A Plaque-associated microbial species and polar metabolites. B Associations between microbial-associated metabolites and carotid artery plaque, adjusted for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use. Levels: low, quartile 1 of the metabolite; medium, quartile 2 and 3; high, quartile 4. C Microbial-associated metabolites, proteomics PLSDA PCs, and their major contributors. D Associations between microbial species and plaque status, adjusted for metabolomic profiles. Data are odds ratios (ORs) and 95% confidence intervals (CIs) for carotid artery plaque per standard deviation increment of CLR transformed abundance of gut bacterial species, adjusted for age, race, study site, antibiotics use, income, education, BMI, alcohol, smoking status, HIV status, and ART use (model 1); further adjusted for Imidazolepropionic acid (model 2); and further adjusted for top 5 proteomics PLSDA PCs (model 3). Abbreviations: Ref, reference group; PLSDA, partial least square-discriminant analysis

Fig. 5

ImP, ImP-related microbial species, functional components, and serum inflammatory markers, in relation to carotid artery plaque. A Associations between ImP, gut microbiota (GMB) score, and individual microbial species, after adjustment for demographic, behavioral, clinical, and HIV-specific variables. The beta coefficients for individual microbial species were estimated in conditional analysis regression models, with mutual adjustment for other ImP-related microbial species. B Associations between the identified ImP-associated microbial species, GMB score, and carotid artery plaque. Data are odds ratios (ORs) and 95% confidence intervals (CIs) for carotid artery plaque per standard deviation increment of CLR transformed abundance of gut bacterial species, adjusted for demographic, behavioral, clinical, and HIV-specific variables. C Plasma ImP and D gut microbial enzyme hutH, according to the levels of ImP associated GMB score (low, quartile 1 of GMB score; medium, quartiles 2 and 3; high, quartile 4). Data are adjusted means and 95% CIs estimated from regression after adjustment for demographic, behavioral, clinical, and HIV-specific variables. E Plasma ImP according to the levels of gut microbial enzyme hutH (low, quartile 1 of hutH; medium, quartiles 2 and 3; high, quartile 4). Data are adjusted means and 95% CIs estimated from regression after adjustment for demographic, behavioral, clinical, and HIV-specific variables. F ImP-associated GMB score, proteomics PLSDA PCs, and their major contributors. Abbreviations: ImP, imidazole propionate; GMB, gut microbiota; PLSDA, partial least square-discriminant analysis