Abstract 13364: Transplantation of Mutant Clostridium_ASF356 Rescues Aging Gut-Derived Phenylacetate-Mediated Perivascular Adipose Tissue and Endothelial Senescence

Introduction: Perivascular adipose tissue (PVAT) may contribute to atherosclerosis by promoting endothelial dysfunction in aging. Gut microbiota can induce PVAT dysfunction; yet, whether it leads to endothelial cell (EC) senescence requires further investigation. Our studies revealed that gut-derived phenylacetate (PAA) and its derivative PAGln age-dependently increase ( TwinsUK Aging Cohort ). Yet, it remains unclear whether and how it contributes to PVAT-EC senescence. Methods: To circumvent our limited knowledge of PAA-(peri)vasculature pathway in aging, we performed multi-omics (fecal shotgun metagenomics and targeted metabolomics), and cell-cell (co-culture) interaction and senescence analyses. Results: Our human ( TwinsUK , n=7,303) and mouse (24 vs. 3 months, n=6) studies identified a positive association of plasma PAA and PAGln with higher abundance of VOR - and PPFOR -harboring bacteria (metabolizing dietary phenylalanine to PAA), particularly Clostridium_ASF356, in gut of old humans and mice that exhibited PVAT and vascular dysfunction. PAA aggravated PVAT dysfunction (reduced UCP1 and Prdm16) by generating excess mitochondrial H 2 O 2 and upregulating Notch1 , leading to reduced energy supply and increased senescence-messaging secretome from PVAT. It induced senescence (increased SA-β-galactosidase, telomere attrition, cell-cycle arrest ( p16 INK4a and p21 WAF1/Cip1 ), and SASP (IL6 and VCAM1)) in co-cultured aortic ECs and reduced angiogenesis (tube formation). We also showed that transplantation of genetically-engineered Δvor, Δppfor C. ASF356 double mutants to old mice (reduces plasma PAA) decreases VCAM1 secretion from PVAT and thereby inhibits senescence in aortic ECs that exhibited eNOS uncoupling. Conclusions: We conclude that age-related increase in microbiota-driven PAA regulates PVAT-mediated EC senescence via Notch1 signaling. Our bacterial engineering approach may represent a novel (peri)vascular senotherapy.