Effects Of TRPV1on M1Macrophages And The Role Of Evodiamine In Regulation Of TRPV1

Background: Researches have shown that inflammation is the main and commonpathophysiologic characteristics in occurrence and development of different stages incardiovascular diseases such as hypertension, coronary heart disease andatherosclerosis. Cardiovascular diseases are characterized by the dysfunction ofvascular endothelial cells and monocytes/macrophages infiltration associated withchronic inflammation. Therefore, an intervention for inhibiting inflammatory responsewill be a potential therapy for cardiovascular diseases. Preliminary studies suggestthat the transient receptor potential vanilloid type1(TRPV1) channel plays aprotective role in endothelial inflammation, to explore the role of TRPV1on M1macrophages and the mechanisms undelying TRPV1-mediated anti-inflammation willprovide a basis for future study on animal experiments and clinical application.Objectives: To determine the mechanisms undrelying TRPV1andEvodiamine-mediated anti-inflammation, we explored the effects of TRPV1onlipopolysaccharide (LPS)-induced production of cytokines and inducible nitric oxidesynthase (iNOS) in the culture of cultivated human acute monocytic leukemia cell line(THP-1), we also determined the role of TRPV1in Evodiamine-mediatedanti-inflammation.Methods:1.Experimental groups⑴Vehicle group:0.5%DMSO+Tween80;⑵LPS+vehicle group: LPS1μg／ml, vehicle0.5%DMSO+Tween80;⑶CAP+LPS group: Capsincin10umol／L, LPS1μg／ml;⑷CAPZ+CAP+LPS group: Capsazepine10umol／L, Capsincin10umol／L,LPS1μg／ml;⑸EVO+LPS group: Evodiamine10umol／L, LPS1μg／ml; ⑹CAPZ+EVO+LPS group: Capsazepine10umol／L, Evodiamine10um／L,LPS1μg／ml.2.Effect of TRPV1on TNF-α, IL-6and IL-1βthat was detected by cell ELISA，the supernatant of M1macrophages which differentiated from THP-1were collectedon6hours and24hours induced by LPS.3. The expression of iNOS protein was measured by western blot, the protein ofM1macrophages which differentiated from THP-1were collected on6hours and24hours induced by LPS.4. Effect of TRPV1on TNF-α and IL-6that was mediated by evodiamine wastested by cell ELISA, the supernatant of M1macrophages which differentiated fromTHP-1were collected on6hours induced by LPS.Experiments repeated three times of each group.Results:1.Effect of TRPV1on TNF-α, IL-6and IL-1β was detected by cell ELISA.After6and24hours of LPS stimulation, the protein concentration of TNF-α, Il-6and IL-1βare significantly increased （P＜0.05）, activating TRPV1can significantlyreduce these inflammatory mediators（P＜0.05）, the effect would be restrained byblocking TRPV1.2.The expression of iNOS protein were detected by western blot．After6and24hours of LPS stimulation, iNOS protein expression weresignificantly elevated（P＜0.05）, activating TRPV1can significantly reduce iNOSprotein expression level（P＜0.05）, this function is restricted after blocking TRPV1.3. Effect of TRPV1on TNF-α and IL-6that was mediated by evodiamine wasdetected by cell ELISA.After6hours of LPS stimulation, the protein concentration of TNF-α and IL-6are significantly raised （P＜0.05）, activating TRPV1can obviously reduce thesecytokines（P＜0.05）, this role would be inhibited partially by the TRPV1antagonist.Conclusions:1. The release of cytokines and iNOS induced by LPS in M1macrophages whichdifferentiated from THP-1cell culture was inhibited by the activation of TRPV1. Theresults suggest TRPV1plays a inhibitory role in inflammation.2. Evodiamine-induced inhibition of cytokine production induced by LPS in M1macrophages which differentiated from THP-1cell culture was attenuated partially bythe TRPV1antagonist. The data suggest that Evodiamine may inhibit inflammatory response partially via the activation of TRPV1.3. We hypothesized that TRPV1can inhibit the function of the M1macrophages,and EVO could inhibit M1macrophages so as to adjust the function of macrophagesby activating TRPV1.