NLRP3 x TRPV1

Emerging evidence underscores the pivotal role of the NLRP3 inflammasome in the pathogenesis of inflammatory pain, positioning it as a promising therapeutic target. Herein, we reported the structure-based design, synthesis, and optimization of novel transient receptor potential vanilloid 1 (TRPV1) antagonist bearing a 1-((2,2-difluorobenzo[d][1,3] dioxol-5-yl) methyl) piperazine scaffold, aiming to simultaneously address TRPV1-mediated neurogenic inflammation and NLRP3-dependent inflammatory signaling. A total of 48 compounds were synthesized and lead compound 3q demonstrated potent in vitro TRPV1 antagonism (IC₅₀ = 63.1 ± 9.6 nM) and inhibition of NLRP3 inflammasome activation in THP-1 cells, as evidenced by reduced IL-1β secretion (IC₅₀ = 348.9 ± 69.62 nM). In vivo, 3q significantly alleviated formalin-induced inflammation pain and LPS/ATP-induced acute peritonitis in mice, markedly reducing IL-1β by 51 %. Furthermore, compound 3q exhibited superior efficacy in DSS-induced colitis, resulting in a lower DAI and histological score. Preliminary pharmacokinetic profiling revealed favorable oral bioavailability (F = 34.4 %) and a half-life of 11.04 h in mice. These findings highlight the potential of 3q as an alternative anti-inflammatory analgesic, advancing the development of TRPV1/NLRP3 dual modulatory therapies for inflammatory pain management.

The pathology of early MIRI (myocardial ischemia/reperfusion injury) is characterized by sterile inflammation. TRPV1 (transient receptor potential vanilloid 1) and NLRP3 inflammasome sense harmful stimulation and modulate inflammation in cardiomyocytes. We recently demonstrated morphine downregulated p-TRPV1 and exacerbated MIRI. In this study, we investigate the potential crosstalk of TRPV1 and NLRP3 inflammasome activities and a potential modulatory effect of morphine on the interaction in MIRI. In vivo and in vitro experiments were conducted. Coding RNA and pharmacological modulations were used in this study. We found MI/R (myocardial ischemia and reperfusion) upregulated p-TRPV1 without change in expression of NLRP3 inflammasome. Giving morphine during myocardial ischemia increased ventricular arrythmia, reduced heart rate and +dp/dt Max in reperfusion, and increased serum cTnI (cardiac troponin I) and infarct size. Suppression of p-TRPV1 but enhancement of NLRP3 inflammasome activity at the end of MI/R were detected. The alterations were reversed by an opioid μ-receptor antagonist or a NLRP3 inhibitor. Giving TRPV1 antagonist or knockdown of TRPV1 in cultured primary cardiomyocytes inhibited p-TRPV1 but increased NLRP3 inflammasome and the downstream cytokines and LDH (lactate dehydrogenase) in the supernatants. Conversely, treatment with capsaicin (a TRPV1 agonist) or upregulation of TRPV1 via transfection of Ad-TRPV1 elevated p-TRPV1 and reduced NLRP3 and LDH. The results indicated morphine treatment during myocardial ischemia aggravates MIRI by increasing the activity of NLRP3 inflammasome, via suppressing the inhibitory effect of p-TRPV1 on NLRP3 inflammasome. Targeting the signal chain of opioid μ-receptor agonist/TRPV1/NLRP3 inflammasome may find a novel way to improve MIRI.

Migraine is a prevalent neurological disorder accompanied by a considerable economic burden. Xiongshao Zhitong granules (XSZT) have anti-inflammatory and analgesic functions in the clinic and are used for migraine therapy. However, the mechanisms by which XSZT treats migraine remain unclear.

To discover the underlying mechanism and active ingredients of XSZT in the treatment of migraine.

The nitroglycerin (NTG)-induced chronic migraine (CM) model was established and used to detect the therapeutic effect of XSZT on migraine. To elucidate the mechanism, we detected transient receptor potential vanilloid 1 (TRPV1) -mediated NOD-like receptor protein 3 (NLRP3) inflammasome activation in the CM rat model and the LPS-induced inflammatory BV-2 cell model using Western blotting, immunofluorescence and ELISA techniques. The potentially active ingredients of XSZT were determined by UHPLC-LTQ-Orbitrap MS, molecular docking, and surface plasmon resonance.

Our findings revealed that XSZT reduced the number of head scratching, increased the periorbital pain threshold and shortened the time spent in the dark box, decreased c-Fos expression in the CM rat model, suggesting an analgesic effect of XSZT on migraine. XSZT inhibited neurogenic inflammation, including downregulating CGRP, TNF-α, IL-1β and IL-18 levels and decreasing the degranulation rate of mast cells. Additionally, XSZT suppressed the expression and activation of TRPV1 and the NLRP3 inflammasome in the trigeminal nucleus caudalis. In vitro experiments confirmed that activated TRPV1 increased the level of the NLRP3 inflammasome by increasing intracellular calcium levels. Galloylpaeoniflorin, isogastrin, ellagic acid and salvianolic acid A interacted with TRPV1 and inhibited IL-1β secretion.

XSZT plays a therapeutic role in migraine through regulating TRPV1-mediated NLRP3 inflammatory activation and galloylpaeoniflorin, isogastrin, ellagic acid and salvianolic acid A might be the active ingredients of XSZT, which provides an experimental basis for the clinical treatment of migraine.