Gut microbiota steroid sexual dimorphism and its impact on gonadal steroids: influences of obesity and menopausal status

Abstract

Background: Gonadal steroid hormones have been suggested as the underlying mechanism responsible for the sexual dimorphism observed in metabolic diseases. Animal studies have also evidenced a causal role of the gut microbiome and metabolic health. However, the role of sexual dimorphism in the gut microbiota and the potential role of the microbiome in influencing sex steroid hormones and shaping sexually dimorphic susceptibility to disease have been largely overlooked. Although there is some evidence of sex-specific differences in the gut microbiota diversity, composition, and functionality, the results are inconsistent. Importantly, most of these studies have not taken into account the gonadal steroid status. Therefore, we investigated the gut microbiome composition and functionality in relation to sex, menopausal status, and circulating sex steroids.

Results: No significant differences were found in alpha diversity indices among pre- and post-menopausal women and men, but beta diversity differed among groups. The gut microbiota from post-menopausal women was more similar to men than to pre-menopausal women. Metagenome functional analyses revealed no significant differences between post-menopausal women and men. Gonadal steroids were specifically associated with these differences. Hence, the gut microbiota of pre-menopausal women was more enriched in genes from the steroid biosynthesis and degradation pathways, with the former having the strongest fold change among all associated pathways. Microbial steroid pathways also had significant associations with the plasma levels of testosterone and progesterone. In addition, a specific microbiome signature was able to predict the circulating testosterone levels at baseline and after 1-year follow-up. In addition, this microbiome signature could be transmitted from humans to antibiotic-induced microbiome-depleted male mice, being able to predict donor's testosterone levels 4 weeks later, implying that the microbiota profile of the recipient mouse was influenced by the donor's gender. Finally, obesity eliminated most of the differences observed among non-obese pre-menopausal women, post-menopausal women, and men in the gut microbiota composition (Bray-Curtis and weighted unifrac beta diversity), functionality, and the gonadal steroid status.

Conclusions: The present findings evidence clear differences in the gut microbial composition and functionality between men and women, which is eliminated by both menopausal and obesity status. We also reveal a tight link between the gut microbiota composition and the circulating levels of gonadal steroids, particularly testosterone. Video Abstract.

Keywords: Gender; Gonadal steroids; Microbiome; Progesterone; Sex; Sexual dimorphism; Testosterone.

Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1

Associations of gut microbiota composition with gender and menopause status. (a) Alpha diversity indices (n = 131). (b) Beta diversity measured by Bray Curtis and weighted unifrac. Overall differences in the microbiome composition among groups were assessed by PERMANOVA using 1000 permutations and pairwise differences between groups were assessed using the pairwise.adonis function adjusted for Bonferroni correction. *p < 0.05. (c) Volcano plot of differential bacterial abundance analysis between pre-menopausal women and men, (d) post-menopausal women and men, and (e) pre- and post-menopausal women, as calculated by DESeq2 from shotgun metagenomic sequencing, adjusting for age and obesity status. For each taxa, the fold change and the p values corrected for multiple comparisons by the Benjamini-Hochberg procedure (pFDR) are plotted. Significantly different taxa (FC > 1.5 and pFDR < 0.05) are colored according to phylum

Fig. 2

Associations of gut microbiota functionality with gender and menopause status. (a) Fold change for the significant differential KEGG pathways between pre-menopausal women and men, and (b) pre- and post-menopausal women, identified by DESeq2 adjusting for age and obesity status. Bars are colored according to the Benjamini-Hochberg corrected p values (pFDR). (c) Spearman correlation heatmap among the abundance of identified KEGG pathways and plasma concentrations of gonadal steroids. Clustering was based on Euclidean distances and Ward linkage. Significance: +, < 0.05; ++, < 0.01; +++,< 0.001. Significant associations after adjusting for age, obesity, and sex are highlighted with a black box

Fig. 3

Gender and menopausal status differences in gonadal steroids. (a) Goodnessoffit (R2Y), goodness of prediction (Q2Y), and permutation tests for the O-PLS-DA model predicting the sex and menopause status from the circulating gonadal steroid levels. (b, c) Principal component analysis score plots for the plasma levels of gonadal steroids colored according to the gender group. (d) Boxplots for the concentrations of progestin, (e) androgens, and (f) estrogens converted to base 10 logarithmic values. Differences among groups were analyzed by a Kruskal-Wallis test, and pair-wise comparisons were assessed by Wilcoxon test. Significant differences are highlighted in bold

Fig. 4

Gut microbial associations with circulating testosterone concentrations. (a) Significant gut bacterial families predicting plasma testosterone levels in humans identified by O-PLS modeling. (b) Permutation tests for the goodness-of-fit (R2Y) and goodness of prediction (Q2Y) for the O-PLS model between bacterial families and circulating testosterone concentrations in humans. (c) Volcano plot of gut bacterial families associated with testosterone levels identified by DESeq2, adjusting for age and obesity status. For each family, the fold change and the Benjamini-Hochberg corrected p values (pFDR) are plotted. Significant families (gray dashed line: pFDR < 0.05; red dashed line: pFDR < 0.1) are colored according to phylum. (d) Experimental design for the fecal microbiota transplantation study in mice. Fecal samples from 22 human donors (11 men and 11 women) were transplanted to 22 mice after 2 weeks of antibiotic treatment. After 28 days of colonization gavage, mice fecal samples were collected and analyzed by shotgun metagenomic sequencing. (e) Principal component analysis score plot based on recipient’s mice bacterial families colored according to human donor sex and menopause status. Overall differences in the microbiome composition were assessed by PERMANOVA using 1000 permutations and Euclidean distances. Pairwise differences between groups were assessed using the pairwise.adonis function adjusted for Bonferroni correction. **p < 0.01. (f) Significant recipient’s mice bacterial families predicting human donor circulating testosterone levels identified by partial Spearman correlation adjusted by age and obesity status