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bioRxiv · 10.1101/2025.02.20.639393

The Pro- vs. Anti-Angiogenic Capacity of Short Chain Fatty Acids is Dependent on the Bioavailability of FFAR3 in Peripheral Artery Disease.

Kuppuswamy, S.·Patel, K.·Zhi, W.·Ganta, V. C.
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Abstract

BackgroundOur current study builds on our recent report that showed Palmitate aggravates ischemic endothelial (EC) dysfunction in vitro and aims to determine whether Palmitate is a critical determinant of PAD severity. In contrast to Palmitate, we studied the role of short-chain fatty acids (SCFAs) in regulating ischemic revascularization in PAD. MethodsFemoral artery ligation and resection was used as preclinical-PAD model. Hypoxia serum starvation was used as in vitro PAD model. ResultsPalmitate dramatically decreased ischemic-EC survival and angiogenic capacity in vitro, whereas SCFAs significantly induced their angiogenic capacity. LC-MRM MS analysis showed decreased SCFA content in ischemic-muscle. Laser Speckle perfusion imaging showed that intramuscular SCFA treatment significantly induced perfusion recovery, whereas Palmitate showed a modest but significant impairment in perfusion recovery. Immunoblot analysis showed that SCFAs preferentially induce Free fatty acid receptor (FFAR)-3, but not FFAR2. Accordingly, silencing FFAR3 decreased ischemic-EC angiogenic capacity, whereas inhibiting FFAR2 induced ischemic angiogenesis. Pharmacological inhibition of FFAR3 by beta-hydroxybutyrate (BHB) significantly decreased perfusion recovery. SCFA treatment further decreased perfusion recovery in BHB-treated ischemic-muscle. Mechanistically, inhibiting FFAR3-induced FFAR2-levels that inhibited ischemic-EC angiogenic capacity by decreasing NO and inducing ROS levels in SCFA-treated ischemic-ECs. ConclusionsOur data shows that Palmitate alone is not sufficient to drive the PAD severity. Increased FFAR3 levels in ischemic-muscle allow SCFAs to activate the AKT-NO axis to induce ischemic angiogenesis and perfusion recovery. However, loss of FFAR3 in ischemic-muscle promotes FFAR2 activation that blocks SCFA-induced NO-production and redox balance thereby inhibiting perfusion recovery in preclinical-PAD.

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Kuppuswamy, S., Patel, K., Zhi, W., Ganta, V. C.. 2025-02-27. The Pro- vs. Anti-Angiogenic Capacity of Short Chain Fatty Acids is Dependent on the Bioavailability of FFAR3 in Peripheral Artery Disease.. https://doi.org/10.1101/2025.02.20.639393

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