Background: Acute gouty arthritis is a common inflammatory arthropathy resulting from urate deposition in joints during persistent hyperuricemia. Nevertheless, effective therapeutic strategies are still unavailable. Here, we propose the crucial role of bromodomain-containing protein 4 (BRD4) in acute gouty arthritis.

Nod-like receptor protein 3 (NLRP3)-mediated pyroptosis has a causal role in the pathogenesis of gout. P2Y₁₄ receptor (P2Y₁₄R) distributed in immune cells including macrophages is a Gi-coupled receptor that inhibits the synthesis of cAMP, which has been regarded as a potential regulator of inflammatory response. Nevertheless, the role of P2Y₁₄R in MSU-induced pyroptosis of macrophages involved in acute gouty arthritis is still unclear. In our present study, P2Y₁₄R knockout (P2Y₁₄R-KO) disrupted MSU-induced histopathologic changes in rat synoviums, accompanied with a significant inhibition of pyroptotic cell death characterized by Caspase-1/PI double-positive and blockade of NLRP3 inflammasome activation in synovial tissues, which was consistent with that observed in in vitro studies. Owing to the interaction of NLRP3 inflammasome and cAMP, we then investigated the effect of adenylate cyclase activator (Forskolin) on macrophage pyroptosis and gout flare caused by MSU stimulation. The reversal effect of Forskolin verified the negative regulatory role of cAMP in MSU-induced pyroptosis. More importantly, adenylate cyclase inhibitor (SQ22536) intervention led to a reversal of protection attributed to P2Y₁₄R deficiency. Findings in air pouch animal models also verified aforementioned experimental results. Our study first identified the role of P2Y₁₄R/cAMP/NLRP3 signaling pathway in acute gouty arthritis, which provides a novel insight into the pathological mechanisms of pyroptosis-related diseases.

Macrophage-derived foam cells play a causal role during the pathogenesis of atherosclerosis. P2Y₆ receptor (P2Y₆R) highly expressed has been considered as a disease-causing factor in atherogenesis, but the detailed mechanism remains unknown. This study aims to explore P2Y₆R in regulation of macrophage foaming, atherogenesis, and its downstream pathways. Furthermore, the present study sought to find a potent P2Y₆R antagonist and investigate the feasibility of P2Y₆R-targeting therapy for atherosclerosis. The P2Y₆R expression was examined in human atherosclerotic plaques and mouse artery. Atherosclerosis animal models were established in whole-body P2Y₆R or macrophage-specific P2Y₆R knockout mice to evaluate the role of P2Y₆R. RNA sequencing, DNA pull-down experiments, and proteomic approaches were performed to investigate the downstream mechanisms. High-throughput Glide docking pipeline from repurposing drug library was performed to find potent P2Y₆R antagonists. The P2Y₆R deficiency alleviated atherogenesis characterized by decreasing plaque formation and lipid deposition of the aorta. Mechanically, deletion of macrophage P2Y₆R significantly inhibited uptake of oxidized low-density lipoprotein through decreasing scavenger receptor A expression mediated by phospholipase C_β/store-operated calcium entry pathways. More importantly, P2Y₆R deficiency reduced the binding of scavenger receptor A to CALR, accompanied by dissociation of calreticulin and STIM1. Interestingly, thiamine pyrophosphate was found as a potent P2Y₆R antagonist with excellent P2Y₆R antagonistic activity and binding affinity, of which the pharmacodynamic effect and mechanism on atherosclerosis were verified. Macrophage P2Y₆R regulates phospholipase C_β/store-operated calcium entry/calreticulin signalling pathway to increase scavenger receptor A protein level, thereby improving foam cell formation and atherosclerosis, indicating that the P2Y₆R may be a potential therapeutic target for intervention of atherosclerotic diseases using P2Y₆R antagonists including thiamine pyrophosphate. Macrophage P2Y₆ receptor deficiency inhibits foam cell and atherosclerosis via the phospholipase C_β/store-operated calcium entry/calreticulin/scavenger receptor A axes. P2Y₆ receptor regulates the formation of foam cells by coupling G protein-coupled receptors Gα_q/11 to target the activation of phospholipase C_β, and then phospholipase C_β isoforms hydrolyse phosphatidylinositol 4,5-bisphosphate to inositol-14,5-trisphosphate inositol 1,4,5-trisphosphate and diacylglycerol. STIM1 is localized in the endoplasmic reticulum membrane and forms a complex with calreticulin. Depletion of Ca²⁺ stores triggers Ca²⁺ release from the endoplasmic reticulum through inositol 1,4,5-trisphosphate receptors and consequent Ca²⁺ dissociation from STIM-calreticulin complex. Consequently, STIM oligomerizes and translocates next to the plasma membrane and STIM1 opens Orai1 channels, a process known as store-operated calcium entry, promoting extracellular Ca²⁺ entry. Moreover, calreticulin and scavenger receptor A form a complex. Thereby, increasing expression of scavenger receptor A facilitates the uptake of oxidized low-density lipoprotein and accumulation of cholesterol within macrophages and further results in the promotion of atherosclerosis. Thiamine pyrophosphate is an antagonist of P2Y₆ receptor, which inhibits P2Y₆ receptor and its downstream response, thereby improving atherosclerosis. CALR, calreticulin; IP3R, inositol 1,4,5-trisphosphate receptor; ox-LDL, oxidized low-density lipoprotein; P2Y₆R, P2Y₆ receptor; PLC_β, phospholipase C_β; SOCE, store-operated calcium entry; SR-A, scavenger receptor A; TPP, thiamine pyrophosphate.

Purinergic signaling plays a causal role in the pathogenesis of inflammatory bowel disease. Among purinoceptors, only P2Y₁₄R is positively correlated with inflammatory score in mucosal biopsies of ulcerative colitis patients, nevertheless, the role of P2Y₁₄R in ulcerative colitis remains unclear. Here, based on the over-expressions of P2Y₁₄R in the intestinal epithelium of mice with experimental colitis, we find that male mice lacking P2Y₁₄R in intestinal epithelial cells exhibit less intestinal injury induced by dextran sulfate sodium. Mechanistically, P2Y₁₄R deletion limits the transcriptional activity of cAMP-response element binding protein through cAMP/PKA axis, which binds to the promoter of Ripk1, inhibiting necroptosis of intestinal epithelial cells. Furthermore, we design a hierarchical strategy combining virtual screening and chemical optimization to develop a P2Y₁₄R antagonist HDL-16, which exhibits remarkable anti-colitis effects. Summarily, our study elucidates a previously unknown mechanism whereby P2Y₁₄R participates in ulcerative colitis, providing a promising therapeutic target for inflammatory bowel disease.

Purpose: To explore the clinical characteristics of Micrococcus luteus bloodstream infection in an infant and characterize the phenotype and genotype of the isolated strains, as well as seek suitable infection models for assessing virulence.Methods: Clinical data was collected from an infant patient diagnosed with M. luteus bloodstream infection. Metagenomic sequencing was performed on the isolated blood sample. The strain was isolated and underwent mass spectrometry, biochemical tests, antibiotic susceptibility assays, and whole-genome sequencing. The Galleria mellonella infection model was used to assess M. luteus virulence.Results: Patient responded poorly to cephalosporins, but recovered after Linezolid treatment. Metagenomic sequencing identified M. luteus as the predominant species in the sample, confirming infection. They were identified as M. luteus with a high confidence level of 98.99% using mass spectrometry. The strain showed positive results for Catalase, Oxidase, and Urea tests, and negative results for Mannose, Xylose, Lactose, Mannitol, Arginine, and Galactose tests, consistent with the biochemical profile of M. luteus reference standards. M. luteus susceptibility to 15 antibiotics was demonstrated and no resistance genes were detected. Predicted virulence genes, including clpB, were associated with metabolic pathways and the type VI secretion system. The infection model demonstrated dose-dependent survival rates.Conclusion: The infant likely developed a bloodstream infection with M. luteus due to compromised immunity. Although the isolated strain is sensitive to cephalosporin antibiotics and has low pathogenicity in infection models, clinical treatment with cephalosporins was ineffective. Linezolid proved to be effective, providing valuable guidance for future clinical management of such rare infections.