Gut microbiota dysbiosis contributes to the development of hypertension

Abstract

Background: Recently, the potential role of gut microbiome in metabolic diseases has been revealed, especially in cardiovascular diseases. Hypertension is one of the most prevalent cardiovascular diseases worldwide, yet whether gut microbiota dysbiosis participates in the development of hypertension remains largely unknown. To investigate this issue, we carried out comprehensive metagenomic and metabolomic analyses in a cohort of 41 healthy controls, 56 subjects with pre-hypertension, 99 individuals with primary hypertension, and performed fecal microbiota transplantation from patients to germ-free mice.

Results: Compared to the healthy controls, we found dramatically decreased microbial richness and diversity, Prevotella-dominated gut enterotype, distinct metagenomic composition with reduced bacteria associated with healthy status and overgrowth of bacteria such as Prevotella and Klebsiella, and disease-linked microbial function in both pre-hypertensive and hypertensive populations. Unexpectedly, the microbiome characteristic in pre-hypertension group was quite similar to that in hypertension. The metabolism changes of host with pre-hypertension or hypertension were identified to be closely linked to gut microbiome dysbiosis. And a disease classifier based on microbiota and metabolites was constructed to discriminate pre-hypertensive and hypertensive individuals from controls accurately. Furthermore, by fecal transplantation from hypertensive human donors to germ-free mice, elevated blood pressure was observed to be transferrable through microbiota, and the direct influence of gut microbiota on blood pressure of the host was demonstrated.

Conclusions: Overall, our results describe a novel causal role of aberrant gut microbiota in contributing to the pathogenesis of hypertension. And the significance of early intervention for pre-hypertension was emphasized.

Keywords: Fecal transplant; Gut microbiota; Hypertension; Metabolism; Pre-hypertension.

Figures

Fig. 1

Decreased diversity and shift of gut enterotypes in human adults with pHTN and HTN. a Rarefaction curves for gene number in control (n = 41), pHTN (n = 56), and HTN (n = 99) after 100 random sampling. The curve in each group is near smooth when the sequencing data are great enough with few new genes undetected. b, c Comparison of the microbial gene count and α diversity (as accessed by Shannon index) based on the genera profile in the three groups. C, control; P, pHTN; H, HTN. P = 0.024, C vs P; P = 0.04, C vs H; for gene count. P = 0.023, C vs P; P = 0.016, C vs H; for α diversity. P values are from Kruskal-Wallis test. d A total of 196 samples are clustered into enterotype 1 (blue) and enterotype 2 (red) by PCA of Jensen-Shannon divergence values at the genus level. The major contributor in the two enterotypes is Prevotella and Bacteroides, respectively. e Relative abundances of the top genera (Prevotella and Bacteroides) in each enterotype. P = 6.31e−31 and P = 2.09e−15, respectively; Wilcoxon rank sum test. f The percentage of control, pHTN and HTN samples distributed in two enterotypes. 26.83% normotensive controls, 48.21% pHTN, and 45.45% HTN are found in enterotype 1. P = 0.02, C vs P; P = 0.03, C vs H; Fisher’s exact test. Boxes represent the inter quartile ranges, the inside line or points represent the median, and circles are outliers

Fig. 2

Genera strikingly different across groups. a Relative abundance of the top 44 most different genera across groups at the criteria of P value <0.1 by Wilcoxon rank sum test. C, control; P, pHTN; H, HTN. The abundance profiles are transformed into Z scores by subtracting the average abundance and dividing the standard deviation of all samples. Z score is negative (shown in blue) when the row abundance is lower than the mean. Genera at P value <0.01 are marked with dark green star, P value <0.05 with light green star, and P value ≥0.05 with gray circle. b The box plot shows the relative abundance of four genera enriched in pHTN and HTN patients, and 11 genera abundant in control. Genera are colored according to the phylum. Boxes represent the inter quartile ranges, lines inside the boxes denote medians, and circles are outliers

Fig. 3

Comparative analysis of GM enrichment across groups based on CAGs. a CAGs are defined as a minimum of 50 linked genes, and the correlation network of CAGs differentially enriched in pHTN and the control group is performed by Spearman’s correlation based on the abundance. b The network of CAGs enriched in HTN is compared to controls. CAGs are colored according to the taxonomic assignment as labeled, and the node size is scaled with the number of genes within the CAG. Edges between nodes denote Spearman correlation >0.8 (red) or between 0.7 and 0.8 (gray)

Fig. 4

Microbial gene functions annotation in pHTN and HTN. a PCA based on the relative abundance of KEGG orthology groups in 196 samples. Significant differences across groups are established at the first principal component (PC1) values, and shown in the box plots above. **P value <0.001, Wilcoxon rank sum test. b The average abundance of KEGG modules differentially enriched in control, pHTN, and HTN gut microbiome. Twenty nine modules enriched in control, and 11 modules overrepresented in both pHTN and HTN are shown in green and pink, respectively. The functional potential of KEGG modules are demonstrated on the right. c Heat map showing the abundance of 11 most significantly altered CAZy family in pHTN or HTN as compared to control

Fig. 5

Aberrant metabolic patterns in pHTN and HTN. a PLS-DA score plots based on the metabolic profiles in serum samples from control, pHTN, and HTN group in ES+ and ES−. n = 30 for control, n = 31 for pHTN, and n = 63 for HTN. b Score scatter plots of OPLS-DA comparing the metabolic differences identify the separation between pHTN and control, HTN and control, respectively. c Metabolites significantly changed in pHTN or HTN as compared to control at VIP >1.5 and P value (t test) <0.05 are identified. Venn diagrams demonstrate the number of altered metabolites shared between pHTN (green) and HTN (red) by the overlap. d The relative amount of 26 endogenous compounds concurrently varied in both pHTN and HTN groups is transformed into Z scores in the heat map. There are six metabolites failed to be identified. e The relationship between 26 endogenous metabolites and the 44 top altered genera (Fig. 2a) in pHTN and HTN is estimated by Spearman’s correlation analysis. And those with low correlated (|r| <0.4) are not shown. Genera and metabolites are distinguished as abundant in control (green) or HTN (pink)

Fig. 6

A classification to identify pHTN and HTN patients from controls. a, b Random forest models are constructed using explanatory variables of CAGs + species (red curve), CAGs + metabolites (green curve), metabolites (yellow curve), CAGs (blue curve), and species (purple curve). The AUC shows the classification of control versus HTN, or control versus pHTN as the numbers of variables increase. The CAGs + metabolites-based classification is more efficient as indicated by a higher AUC. c ROC of the random forest classifier using CAGs + metabolites based on the 1000 most important variables by ranking the variables by importance. AUC = 0.91 for control versus pHTN (n = 12, red curve), AUC = 0.89 for control versus HTN (n = 12, green curve), and AUC = 0.57 for pHTN versus HTN (n = 13, blue curve). d The top 50 different CAGs distinguish HTN from control based on the random forest model using explanatory variables of CAGs + metabolites. e The top 50 CAGs discriminate between pHTN and control using explanatory variables of CAGs + metabolites. The lengths of bar in the histogram represent mean decrease accuracy, which indicates the importance of the CAG for classification. The color denote the enrichment of CAG in control (blue), in HTN or pHTN (red) according to OR score

Fig. 7

Post-transplanted intestinal microbial profiles and BP of recipient mice. a Schematic representation of fecal microbiota transplantation. GF mice (n = 5 for control, n = 10 for HTN) are orally inoculated with prepared fecal contents from two patients of HTN and one normotensive control, respectively. The gut microbial profiles are analyzed at 7 days, and BP is measured at 10 weeks post-transplantation. C, control; H, HTN. b Venn diagram comparing the shared genera number in gut microbiome of human donors (n = 1 for control, n = 2 for HTN) and recipient mice (n = 3 for control, n = 6 for HTN). c Shannon index of recipient mice at the genus level demonstrate significantly reduced α diversity in HTN group. P = 0.048 from t test. Boxes represent the inter quartile ranges, lines inside the boxes denote medians, and circles are outliers. d PCoA plots of human donors and recipient mice based on microbial genera separate HTN group from the controls. e Heat map comparing the abundance of altered genera between control and HTN mice. Red, more abundant; blue, less abundant. Genera present consistent trend with the metagenomic analysis are marked with green points, while inconsistent with gray points. f SBP, DBP, MBP, and HR of the recipient mice (n = 5 for control, n = 10 for HTN) are measured by tail-cuff method. Data are presented as mean ± s.e.m. P = 0.018, SBP; P = 0.019, DBP; P = 0.014, MBP; P = 0.11, HR; t test