Angiotensin-(1-7) and Gut-Bone Marrow Axis in Aging

Abstract

<b>Abstract ID 22254</b> <b>Poster Board 363</b> <b>Background:</b> Aging compromises the colon epithelial barrier integrity resulting in chronic systemic inflammation, which increases risk for cardiovascular diseases. Aging-associated gut microbial dysbiosis is implicated in this pathology. Metabolites derived from dysbiotic bacteria in aging are known to stimulate myelopoiesis in bone marrow, which directly contribute to the systemic inflammation. We have previously showed that the protective peptide of Renin Angiotensin System (RAS), Angiotensin-(1-7) (Ang-(1-7)), restored epithelial barrier integrity in Old mice via Mas receptor (MasR) activation. This study tested the hypothesis that Ang-(1-7) modulates gut-bone marrow axis to ameliorate systemic inflammation. <b>Methods:</b> Young (3-4 months) and Old (20-24 months) male C57Bl/6 mice were treated with saline or Ang-(1-7) by using osmotic pumps (1 μg/Kg/min for 4 weeks). Colonic organoids were cultured and Wnt3a expression was determined. Gut microbiome was analyzed by 16S rRNA sequencing of bacterial DNA using MiSeq followed by analysis by Qiime 2.0 and Past 4.10 software. Myelopoiesis in the bone marrow cells was determined by using colony forming unit-granulocytes-macrophages (CFU-GM) assay followed by flow cytometric enumeration of monocytes (Ly6G^-CD11b⁺Ly6C^hi) and macrophages (Ly6G^-CD11b⁺F4/80⁺Ly6C^hi). Inflammatory cells in the colon wall were characterized by immunohistochemistry. Plasma levels of Trimethylamine Oxide (TMAO), IL-1β, IL-6, HMGB1, MCP1 and TNFα were analyzed. <b>Results:</b> Organoid cultures showed decreased size of organoids and expression of Wnt3a in the Old group compared to the Young (<i>p</i> &lt;0.05, n=6), which were restored by Ang-(1-7). Aging was associated with decreased richness shown by Shannon and Chao1 diversity indices and altered beta diversity (Bray-Curtis (<i>p</i> &lt; 0.0001) or Jaccard (<i>p</i> &lt; 0.0001)). Microbial dysbiosis is evident based on altered relative abundance of 222 ASVs in aging. Plasma TMAO levels (6.99 ± 0.44 μmol/L) were higher in aging compared to the Young (3.08 ± 0.23 μmol/L, <i>p</i> &lt; 0.001). Ang-(1-7) modified microbial dysbiosis, reversed changes in alpha and beta diversity in aging and decreased TMAO levels (3.30 ± 0.24 μmol/L, <i>p</i> &lt; 0.001 vs Old). Bone marrow cells from old mice showed increased myelopoiesis compared to young (<i>p</i> &lt; 0.01, n=6) that were associated with increased infiltration of inflammatory cells in the colon. Ang-(1-7) decreased the number of monocytes (18 ± 4/10⁶ WBC) than the old group (47 ± 4/10⁶ WBC, <i>p</i> &lt; 0.001, n=6). Pro-inflammatory macrophages were decreased by Ang-(1-7) in the bone marrow and reduced the infiltration of the colon wall. Plasma levels of IL-1β, IL-6, HMGB1, MCP1 and TNFα were higher in the old group compared to the young (<i>p</i> &lt; 0.05, n=5), which were lowered by Ang-(1-7) (<i>p</i> &lt; 0.05, n=5). <b>Conclusion:</b> Ang-(1-7)/MasR pathway modulates gut-bone marrow axis by reversing microbial dysbiosis and myelopoietic bias in aging thereby ameliorates systemic inflammation.