TRPV4 Mediates the Macrophage Phagocytic and Cytokine Response to Leptin

Abstract

Abstract Obesity is characterized by chronic activation of the innate immune system. Leptin is an adipokine upregulated in obesity that has been shown to stimulate macrophage function through an unidentified signal. Our lab recently showed that the mechanosensitive calcium channel, transient receptor potential vanilloid 4 (TRPV4) mediates macrophage phagocytosis in response to infection. This study seeks to determine if TRPV4 also mediates the macrophage response to leptin. Bone marrow derived macrophages were isolated from 7–10 week old WT and TRPV4 KO mice and incubated in the presence of pathophysiological range leptin concentrations (0.1–1μg/mL; 24 hr) ± LPS (E coli 0111:B4 as a positive control) ± TRPV4 inhibitor. Macrophage phagocytosis was measured as uptake of fluorescently-labeled E coli particles. Intracellular calcium was measured using the FLIPR Calcium 5 Assay in the presence of a selective TRPV4 agonist (GSK). MIP-2 and IL-8 secretion was measured by ELISA. Leptin increases phagocytosis of E coli particles by 17% (59% of the LPS response) and TRPV4 activity by 24% (51% of the LPS response). The phagocytic response and calcium influx are abrogated by either pharmacologic inhibition or deletion of TRPV4. Secretion of MIP-2 and IL-8 in response to leptin increases by 8-fold after pharmacologic inhibition of TRPV4. Our data show that TRPV4 mediates macrophage phagocytosis of E coli particles, and suppresses proinflammatory cytokine release, in response to leptin. In addition, we show that TRPV4 function is increased by leptin. In vitro results predict that TRPV4 enhances host defense and suppresses inflammation in the setting of obesity-induced hyperleptinemia, thus potentially modulating pneumonia severity in obese patients.