Sex-dependent modulation of T and NK cells and gut microbiome by low sodium diet in patients with primary aldosteronism

Abstract

Background: High dietary sodium intake is a major cardiovascular risk factor and adversely affects blood pressure control. Patients with primary aldosteronism (PA) are at increased cardiovascular risk, even after medical treatment, and high dietary sodium intake is common in these patients. Here, we analyze the impact of a moderate dietary sodium restriction on microbiome composition and immunophenotype in patients with PA.

Methods: Prospective two-stage clinical trial including two subgroups: 15 treatment-naive PA patients compared to matched normotensive controls; and 31 PA patients on mineralocorticoid receptor antagonist treatment before and three months after sodium restriction. Patients underwent blood pressure measurements, laboratory tests, analysis of peripheral blood mononuclear cells via flow cytometry and microbiome analysis.

Results: We observed a higher percentage of Tregs in treatment-naive PA patients (p = 0.0303), while the abundance of Bacteroides uniformis was higher in PA patients compared to normotensive controls (p = 0.00027) and the abundance of Lactobacillus species however was higher in the subgroup of normotensive controls (p = 0.0290). Sodium restriction was accompanied by a decrease in pro-inflammatory Tc17 cells in male patients (p = 0.0081, females p = 0.3274). Bacteroides uniformis abundance was higher in female patients (0.01230, p = 0.0016) and decreased upon sodium restriction (0.002309, p = 0.0068).

Conclusion: Dietary sodium restriction in patients with PA modulates the peripheral immune cell composition toward a less inflammatory phenotype. This suggests a potential mechanism by which sodium reduction modulates immune cell composition, leading to blood pressure reduction and positively impacting cardiovascular risk.

Keywords: Tc17; Tregs (regulatory T cells); microbiome; primary aldosteronism (PA); sodium.

Figure 1

Study protocol. The study is subdivided in two different parts with a total of three different analysis steps. Part A – 1 focuses on comparison of PA patients before MRA treatment (n = 15) to sex, age, and BMI-matched healthy controls (n = 15). PA patients were again evaluated three months after start of MRA therapy (Part A – 2, see Supplementary). In part B – 3, 13 of these 15 patients on three months of stable MRA treatment and another 18 PA patients with over three months of stable MRA treatment were analyzed before and after sodium restriction.

Figure 2

Comparison of immune cell subsets and microbiome between patients with PA and healthy controls. Immune cell subsets and abundance of microbiota of part A (n = 15 patients with PA before MRA treatment and 15 matched healthy controls). (A) Analysis of the percentage of CD25+ T cells of all CD4+ T cells. (B) Analysis of the percentage of CD25+Foxp3+ Tregs of all CD25+ Th cells. Mann-Whitney test was used for statistic analysis with p < 0.05. Presented is the median and IQR. (C) Abundance of Bacteroides uniformis ASV. The p value indicated in the boxplot is generated from MaAsLin2 results with adjusting age and gender. The adjusted p-values (q-values) is 0.058. (D) Analysis of abundance of Lactobacillus genus.

Figure 3

Changes in immune cell subsets dependent on sodium restriction. Immune cell subsets of part B (n = 31 patients with PA before (PA – SR) and after (PA + SR) sodium restriction) after 4 hours stimulation with PMA/ionomycin by multicolor flow analysis. (A) Analysis of the percentage of Tc17 cells of all Tc cells. (B) Sex-specific analysis of Tc17 cells of all Tc cells (males = 13, females = 17). (C) Analysis of Tc17/CD25+Foxp3+ Treg ratio before and after sodium restriction. (D) Sex-specific analysis of Tc17/Treg ratio (males = 13, females = 17). (E) Analysis of the percentage of CD3-CD56+ NK cells in percentage of CD3- cells. (F) Analysis of the percentage of CD107a+ NK cells of CD3-CD56+ NK cells in PA patients before and after intervention. Wilcoxon test was used for statistic analysis with p < 0.05. Presented is the median and IQR.

Figure 4

Alpha diversity and microbial composition in PA patients depending on treatment status. (A, B) Abundance of Bacteroides uniformis was analyzed in PA patients without PA-specific medical treatment (PA), after at least three months of a stable MRA dose (PA – SR, n = 31) and three months after additional sodium restriction (PA + SR) and compared to a subgroup of healthy controls (n = 15). (C) Alpha diversity (i.e. Shannon effective numbers) was analyzed in PA patients without PA-specific medical treatment (PA), after at least three months of MRA treatment on a stable dose (PA – SR) and three months after additional sodium restriction (PA + SR) and compared to a subgroup of healthy controls (n = 15). (D) Principal coordinates analysis (PCoA) analysis indicated the difference in overall composition of gut microbial community among the groups using Bray-Curtis dissimilarity. In 4A and C Kruskal-Wallis test was used for statistic analysis with p < 0.05. P-values in 4B refer to results of Mann-Whitney test.