The CaSR/TRPV4 coupling mediates pro-inflammatory macrophage function

Aim Although calcium-sensing receptor (CaSR) and transient receptor potential vanilloid 4 (TRPV4) channels are functionally expressed on macrophages, it is unclear if they work coordinately to mediate macrophage function. The present study investigates whether CaSR couples to TRPV4 channels and mediates macrophage polarization via Ca2+ signaling.Methods The role of CaSR/TRPV4/Ca2+ signaling was assessed in lipopolysaccharide (LPS)-treated peritoneal macrophages (PMs) from wild-type (WT) and TRPV4 knockout (TRPV4 KO) mice. The expression and function of CaSR and TRPV4 in PMs were analyzed by immunofluorescence and digital Ca2+ imaging. The correlation factors of M1 polarization, CCR7, IL-1 beta, and TNF alpha were detected using q-PCR, western blot, and ELISA.Results We found that PMs expressed CaSR and TRPV4, and CaSR activation-induced marked Ca2+ signaling predominately through extracellular Ca2+ entry, which was inhibited by selective pharmacological blockers of CaSR and TRPV4 channels. The CaSR activation-induced Ca2+ signaling was significantly attenuated in PMs from TRPV4 KO mice compared to those from WT mice. Moreover, the CaSR activation-induced Ca2+ entry via TRPV4 channels was inhibited by blocking phospholipases A2 (PLA2)/cytochromeP450 (CYP450) and phospholipase C (PLC)/Protein kinase C (PKC) pathways. Finally, CaSR activation promoted the expression and release of M1-associated cytokines IL-1 beta and TNFalpha, which were attenuated in PMs from TRPV4 KO mice.Conclusion We reveal a novel coupling of the CaSR and TRPV4 channels via PLA2/CYP450 and PLC/PKC pathways, promoting a Ca2+-dependent M1 macrophage polarization. Modulation of this coupling and downstream pathways may become a potential strategy for the prevention/treatment of immune-related disease.