Dannesboe, J. | bioRxiv Science

bioRxiv2024

Low-dose colchicine treatment improves vasorelaxation, reduces arterial remodeling and attenuates blood pressure increases in spontaneously hypertensive rats

Colchicine, a microtubule depolymerizing agent, is an effective therapy for the secondary prevention of cardiovascular disease and has been approved recently as a novel treatment for atherosclerosis associated with coronary artery disease. Hypertension is a leading cause of cardiovascular disease, yet the impact of colchicine on hypertension has not been studied. We hypothesized that low-dose colchicine could be used to treat hypertension to reduce cardiovascular disease risk. The aim of this study was to administer daily, low dose (0.05 mg/kg/day) oral colchicine for 4 weeks to spontaneously hypertensive rats (SHR) and normotensive controls (WKY) and determine the effect on blood pressure, vascular reactivity, remodeling and inflammation, and left ventricular hypertrophy. Daily blood pressure measurements recorded by telemetry in conscious rats showed colchicine prevented increases in mean arterial pressure observed in the SHRs receiving vehicle over the 4-week treatment period. After the 4-weeks of treatment, 3rd order mesenteric artery vasorelaxations to isoproterenol, sodium nitroprusside and the Kv7.2-5 channel activator, ML213, were enhanced in the SHRs receiving colchicine compared to vehicle. The improved isoproterenol-mediated relaxation was also observed in WKY rats receiving colchicine, and in both the SHR and WKY, this improved effect was attenuated by the {beta}2 adrenoceptor antagonist, ICI118,551. Proteomic analysis of the mesenteric arteries by mass spectrometry revealed that colchicine treatment prevented changes observed when comparing the SHR vehicle group with the WKY vehicle group in proteins associated with extracellular matrix pathways. Immunostaining of 3rd order mesenteric arteries with Sirius red found that colchicine treatment attenuated the increased media thickness of the artery wall observed in SHRs receiving vehicle. Multiplex immunoassay and Western blots revealed colchicine reduced certain inflammatory mediators in the wall of the SHR mesenteric arteries, particularly the nucleotide-binding domain and leucine-rich repeat pyrin containing protein-3 (NLRP3), IL-18, CXCL10, and CXCL2, as well as reducing phosphorylated STAT3. Finally, in the left ventricle of the SHR, colchicine treatment attenuated a number of inflammatory mediators, including NLRP3, IL-1{beta}, and IL-18, and reduced fibrosis and cell size, which are indicative of left ventricular hypertrophy. Overall, we show colchicine has the potential to elicit cardiovascular protective effects in hypertension by targeting multiple cell types.

physiology↗

bioRxiv2023

Dynamic conformational changes of acid-sensing ion channels in different desensitizing conditions

Acid-sensing ion channels (ASICs) are proton-gated cation channels that contribute to fast synaptic transmission and have roles in fear conditioning and nociception. Apart from activation at low pH, ASIC1a also undergoes several types of desensitization, including acute desensitization that terminates activation, steady-stated desensitization that occurs at sub-activating proton concentrations and limits subsequent activation, and tachyphylaxis that results in a progressive decrease in response during a series of activations. Structural insights from a desensitized state of ASIC1 have provided great spatial detail, but dynamic insights into conformational changes in different desensitizing conditions are largely missing. Here, we use electrophysiology and voltage-clamp fluorometry to follow the functional changes of the pore along with conformational changes at several positions in the extracellular and upper transmembrane domain via cysteine-labeled fluorophores. Acute desensitization terminates activation in wild-type but introducing an N414K mutation in the {beta}11-12 linker of mouse ASIC1a interfered with this process. The mutation also affected steady-state desensitization and led to pronounced tachyphylaxis. Common to all types of desensitization was that the extracellular domain remained sensitive to pH and underwent pH-dependent conformational changes. These conformational changes did, however, not necessarily lead to desensitization. N414K-containing channels remained sensitive to known peptide modulators that increased steady-state desensitization, indicating that the mutation only reduced, but not precluded, desensitization. Together, this study contributes to understanding the fundamental properties of ASIC1a desensitization, emphasizing the complex interplay between the conformational changes of the ECD and the pore during channel activation and desensitization. Statement of significanceAcid-sensing ion channels (ASICs) are proton-gated ion channels that contribute to synaptic activity and play roles in acidosis-related diseases. Prolonged acidosis can lead to desensitization in ASIC1a, and modulators that affect this desensitization have shown beneficial effects in pain and stroke. In this study, we investigated the functional and conformational changes during acute desensitization, steady-state desensitization, and tachyphylaxis through a mutation in the {beta}11-12 linker of ASIC1a. We found that the mutation retained pH-dependent conformational changes of the extracellular domain (ECD) but largely disconnected these movements from the channel pore. Collectively, our work emphasizes the critical role of the {beta}11-12 linker for the pH-dependent conformational interplay between the ECD and the channel pore.

molecular biology↗