Journal Article

Cultured rat aortic vascular smooth muscle cells do not express a functional TRPV1

Ciotu, Cosmin I., Gold-Binder, Markus, Heiss, Elke H., Fischer, Michael J. M., Blažević, Tina, Dirsch, Verena M.

2023

Overview

We showed previously that capsaicin, an active compound of chili peppers, can inhibit platelet-derived growth factor-induced proliferation in primary rat vascular smooth muscle cells (VSMCs). The inhibition of BrdU incorporation by capsaicin in these cells was revoked by BCTC, which might be explained by a role of TRPV1 in VSMCs proliferation. To further pursue the hypothesis of a TRPV1-dependent effect of capsaicin, we investigated TRPV1 expression and function. Commercially available antibodies against two different TRPV1 epitopes (N-terminus and C-terminus) were rendered invalid in detecting TRPV1, as shown: i) in western blot experiments using control lysates of TRPV1-expressing (PC-12 and hTRPV1 transfected HEK293T) and TRPV1-downregulated (CRISPR/Cas gene edited A10) cells, and ii) by substantial differences in staining patterns between the applied antibodies using fluorescence confocal microscopy. The TRPV1 agonists capsaicin, resiniferatoxin, piperine and evodiamine did not increase intracellular calcium levels in primary VSMCs and in A10 cells. Using RT qPCR, we could detect a rather low TRPV1 expression in VSMCs at the mRNA level (Cp value around 30), after validating the primer pair in NGF-stimulated PC-12 cells. We conclude that rat vascular smooth muscle cells do not possess canonical TRPV1 channel activity, which could explain the observed antiproliferative effect of capsaicin.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Analysis / Animals / Antibodies / Aorta / Aorta - metabolism / Biology and Life Sciences / C-Terminus / Calcium (intracellular) / Calcium - metabolism / Capsaicin / Capsaicin - metabolism / Capsaicin - pharmacology / Capsaicin receptors / Cell culture / Cell cycle / Cell proliferation / Cells, Cultured / Cellular apoptosis susceptibility protein / Channel gating / Confocal microscopy / Coronary vessels / CRISPR / Epitopes / Fluorescence / Fluorescence microscopy / Gene expression / Growth factors / Health aspects / HEK293 Cells / Humans / Lysates / Medical examination / Medicine and Health Sciences / Methods / mRNA / Muscle, Smooth, Vascular - metabolism / Muscles / Musculoskeletal system / N-Terminus / Nerve growth factor / Peppers / Piperine / Platelet-derived growth factor / Proteins / Rats / Reagents / Research and Analysis Methods / Resiniferatoxin / Smooth muscle / TRPV Cation Channels - metabolism / Vascular smooth muscle / Viral antibodies