Interaction between<i>Akkermansia muciniphila</i>and β-estradiol, and sex-differential blood pressure regulation in spontaneously hypertensive rats

Background: Akkermansia muciniphila (Akk), a Gram-negative bacterium, is recognized as a next generation probiotic due to its efficacy to improve metabolic disorders including obesity, diabetes, and cardiovascular diseases. Akk-derived extracellular vesicles have been reported to confer protective antihypertensive effects in male spontaneously hypertensive rats (SHR). Blood pressure (BP) is a sexually dimorphic trait, and the prevalence of hypertension varies between males and females. We recently reported sex differences in salt-induced remodeling of gut microbiota and its impact on BP regulation, demonstrating sex specific contributions of gut microbiota to hypertension. Moreover, ovariectomy resulted in depletion of Akk in ‘four core genotype’ female mice (XX and XY), suggesting a strong connection between Akk and female sex hormones. Therefore, we hypothesized that Akk regulates BP differently in males and females. Methods: 10-13-week-old SHR of both sexes (n=5-6 rats/group) were gavaged with or without ~10 8 CFU/ml of Akk in PBS every other day for 4 weeks. Hungate tubes were used to minimize exposure to oxygen during gavages. BP was monitored using radiotelemetry. The proximal colon was examined for the gene expression of inflammatory markers by quantitative PCR. Akk abundance and cecal microbiota diversity was evaluated using 16S rRNA sequencing. The Basic Local Alignment Search Tool was used to query the Akk proteome for potential homology to estrogen receptors. Akk was treated in vitro with β-estradiol (10µM) and quantitative PCR was used for gene expression. Results: In male SHR rats, oral administration of Akk resulted in a significant decrease in systolic BP (176.7 vs. 170.6 mmHg, p&lt;0.0001) and mean BP (148.0 vs. 144.7 mmHg, p=0.0004). In contrast, female SHR rats showed a significant increase in Systolic BP (161.2 vs. 164.5 mmHg, p=0.01) and mean BP (137.3 vs. 139.6 mmHg, p=0.0032) following oral ingestion of Akk. At baseline, female SHR exhibited a higher abundance of Akk compared to males. Akk treatment resulted in a lower microbial α diversity in female SHR, compared to control females and Akk-treated males. This was associated with an increase in the pro-inflammatory cytokine Il1b exclusively in female SHR rats treated with Akk. A hypothetical protein RS02615 of Akk was identified with 29% identity and 50% homology (E-value 0.001) to the ligand binding domain of human estrogen receptor 1 (ESR). The expression level of gene rs02615 was significantly increased in the Akk-treated female SHR, compared to treated males. In vitro, β-estradiol increased the expression of rs02615, and two other genes (i.e., rs02610, rs02605 ) clustered with rs02615 in the same operon, suggesting that β-estradiol may bind to RS02615 protein to enhance the expression of this operon which is described as related to lipopolysaccharide (LPS) production according to the function of RS02610 and RS02605 proteins. Conclusion: We report sex specific roles of Akk in BP regulation and identified rs02615 as a novel bacterial gene encoding a putative ESR-like protein RS02615. Although the rs02615 -containing operon seems involved in LPS production, future studies are warranted to investigate the binding of RS02615 to β-estradiol and its role in LPS production, and Akk-mediated sex specific BP regulation. The University of Toledo Startup funds, the American Heart Association CDA (852969), and the National Institutes of Health (R21AG079357) to Dr. Tao Yang. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.