Abstract Background CD4+ T cell hyperactivation is a pivotal driver of systemic autoimmunity in systemic lupus erythematosus (SLE), yet the molecular regulators that set its activation threshold remain poorly defined. Methods PPP2R3C expression was measured by qRT‐PCR and Western blotting in CD4+ T cells from 45 SLE patients and 37 healthy controls, as well as in pristane‐induced lupus (PIL) mice. Jurkat cells with PPP2R3C knockdown or overexpression were generated by lentiviral transduction. Signaling mechanisms were dissected using transcriptome sequencing and calcium flux assays. In vivo, PPP2R3C was restored in PIL mice via T cell‐targeted Ark313 vector or systemic AAV9 delivery; disease progression was assessed at 24 and 48 weeks after pristane induction. Results We identified the protein phosphatase 2A regulatory subunit PPP2R3C as a critical and selective negative regulator of T cell receptor (TCR) signaling, which was downregulated in CD4+ T cells from SLE patients and pristane‐induced lupus (PIL) mice. Reduced PPP2R3C expression was also observed in the kidneys of both PIL mice and lupus nephritis patients. In PIL mice, this reduction was evident in podocytes, endothelial cells, and mesangial cells, indicating widespread downregulation across kidney resident cell populations. Mechanistically, PPP2R3C deficiency enhanced T cell activation, cytokine production, and calcium flux by potentiating PLCγ1 phosphorylation and subsequent TCR‐driven JNK/c‐Jun signaling, whereas its overexpression produced opposing effects. To assess therapeutic potential, we restored PPP2R3C expression in PIL mice using two complementary gene delivery strategies. T cell‐targeted reconstitution via the Ark313 vector potently suppressed T cell activation and autoantibody production at an early stage (24 weeks), culminating in markedly attenuated proteinuria and glomerular immune complex deposition by late‐stage disease (48 weeks). Systemic delivery of AAV9‐PPP2R3C provided comprehensive therapeutic effects, ameliorating renal pathology while also normalizing immune dysregulation in both the thymus and spleen across both time points. Conclusions Our findings identify PPP2R3C as an important contributor to SLE pathogenesis and support its restoration as a gene therapy approach worthy of further exploration for its ability to rebalance immunity and mitigate tissue injury in SLE. Key points PPP2R3C is selectively downregulated in SLE CD4+ T cells and correlates with disease activity. It restrains the PLCγ1‐JNK axis to limit T cell hyperactivation and cytokine production. Gene therapy restoring PPP2R3C suppresses autoimmunity and lupus nephritis in mice, and its reduction in kidney resident cells indicates a direct renoprotective effect, positioning PPP2R3C as a promising therapeutic target for SLE.

This study aims to investigate the role of protein phosphatase 2A (PP2A) dysfunction in the hyperinflammatory response observed in systemic lupus erythematosus (SLE) monocytes. The focus is on elucidating the functional connection between dysregulation of PP2A subunits, loss of enzymatic activity, and aberrant innate immune activation through the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon response cGAMP interactor 1 (STING) pathway in monocytes. CD14+ monocytes were isolated from 88 SLE patients and 40 healthy controls (HCs). PP2A subunits (PPP2R1A, PPP2CA, PPP2CB) mRNA expression was analyzed and correlated with autoantibody profiles. PP2A phosphatase activity and inhibitory phosphorylation (PP2Ac-Y307) were assessed. Functional effects of PP2A modulation were evaluated by pharmacologically inhibiting PP2A (LB-100) in HCs monocytes or activating it (FTY720) in SLE monocytes, followed by cGAS-STING stimulation with cGAMP. Inflammatory parameters, including interferon-stimulated gene (ISG15, ISG20, IFIT3) expression and cytokine (IL-6, IL-1β, IFN-α) secretion, were measured. Monocytes derived from patients with SLE demonstrated a selective downregulation of the PP2A structural subunit, PPP2R1A, which led to a reduction in PP2A phosphatase activity and an increase in phosphorylation at PP2Ac-Y307. Functionally, the pharmacological inhibition of PP2A in monocytes from HCs resulted in an amplification of cGAS–STING–dependent expression of ISGs and hypersecretion of cytokines, thereby mimicking the hyperinflammatory phenotype observed in SLE. In contrast, activation of PP2A through FTY720 in SLE monocytes significantly curtailed this exaggerated immune response. These findings position PP2A hypofunction as a critical pathogenic driver in SLE and highlight the therapeutic potential of targeting this phosphatase. The restoration of PP2A activity mitigates dysregulated innate immune responses, presenting a promising strategy for immunomodulation in SLE.

Our previous study has shown that ATP action on P2X7R could be the second signal to induce the onset of gouty arthritis. However, the functional changes of P2X7R single nucleotide polymorphisms (SNPs) on the effects of ATP-P2X7R-IL-1β signaling pathway and uric acid remained unknown. We aimed to investigate the association between the functional change of P2X7R containing the Ala348 to Thr polymorphisms (rs1718119) and the pathogenesis of gout. First, 270 gout patients and 70 hyperuricemic patients (without gout attack history in recent 5 years) were recruited for genotyping. In addition, the changes of ATP-induced pore formation were assessed in HEK-293T cells overexpressing different mutants in P2RX7, and the effects on P2X7R-NLRP3-IL-1β pathway activation were explored in P2RX7 overexpression THP-1 cells. The risk allele for gout was A at rs1718119, and the AA and AG genotypes exhibited a higher risk of gout. Furthermore, Ala348 to Thr mutants increased P2X7-dependent ethidium+ bromide uptake, upregulated IL-1β and NLRP3 levels as compared to the wild-type. We suggest that genetic polymorphisms of P2X7R containing the Ala348 to Thr are associated with the increased risk of gout, showing an enhanced gain-of-function effect on the development of this disease.

Abstract Gout is a common disease of inflammatory arthritis caused by hyperuricemia and the deposition of MSU crystals. Our previous study has shown that ATP action on P2X7R could be the second signal to induce the onset of gouty arthritis. However, the biological function changes of SNPs in the P2X7R, which affect the ATP-P2X7R-IL-1β signaling pathway with high uric acid, remained unknown. To further research P2RX7 gene variant (encoded by the SNP as rs1718119) polymorphism association between the gout susceptibility and the functional effect, 270 patients with gout and 70 hyperuricemia patients without history of gout flare (over 5-years course) were recruited in this study. The current study first time analyzed the genotyping study in genomic DNA samples from gout and hypeluricemia patients, then flow cytometry was used to test the transfected Ala348to Thr mutant in HEK-293T cells. Expression levels of IL-1β, NLRP3, ASC, caspase-1 mRNA in THP-1 cells were analyzed by quantitative real-time polymerase chain reaction. Expression levels of IL-1β were measured by enzyme-linked immunosorbent assay. The gout-sensitivity allele at rs1718119 was A. The AA and AG genotypes exhibited a higher risk of gout and the gout-sensitivity allele at rs1718119 was A. Moreover, Ala348to Thr increased P2X7-dependent ethidium+ bromide uptake. What’s more, Ala348 to Thr significantly up-regulated the serum and mRNA levels of IL-1β compared with wild type (P=0.0007;P=0.0334,respectively). Expression levels of NLRP3 mRNA in cells with Ala348 to Thr also showed a higher level than wild type(p=0.0003). However, no statistical significance were found in the mRNA expression of ASC and caspase-1 between Ala348to Thr and wild type in all three groups(P＞0.05). Our study revealed that the biological function changes of one SNP (rs1718119) mutation affected the ATP-P2X7R-IL-IL-1β signaling pathway with high uric acid. And, both rs1718119 was dominant gene. The genetic variability of the P2X7R rs1718119 gene might, in part, be associated with susceptibility for gout onset.

Objective:To investigate the effect of P2X7R on MSU crystal-induced acute gouty arthritis in rats and its mechanism on inflammatory responses. Methods:In vivo activation or inhibition of P2X7R was examined in the ATP group or the BBG group of rats, and the control group were injected with PBS. All three groups of rats were injected with MSU in the right joint cavity. The development of acute gouty arthritis was observed and evaluated at 6h, 12h, 24h, 48h and 72h. The clinical manifestations of acute arthritis, the expression level of P2X7R in spleen macrophages, the ability of macrophages to take up YO-PRO-1, and the level of Tregs, Th17 cells and inflammatory cytokines were assessed. Besides, mRNA expression levels of P2X7R, NLRP3 and IL-1β were also detected. Results:After 12h and 24h administration, P2X7R agonist ATP significantly accelerated the development of acute gouty arthritis, while the P2X7R inhibitor BBG had the opposite effect on this process. Activation of P2X7R significantly aggravated the ankle joint arthritis of the rat and promoted the infiltration of neutrophils and macrophages in the synovial tissue. In addition, the expression of P2X7R in macrophages of ATP group, the uptake of YO-PRO-1 and the expression of NLRP3 mRNA were significantly higher than that in other two groups. At 12h or 24h, activation or inhibition P2X7R had a significant effect on the IL-1β, IL-6, IL-17, IL-10 and TGF-β1. The ratios of Treg/Th17 gradually decreased in the First three time points, it was the lowest at 24h. Conclusion:Activation of P2X7R by ATP aggravated the development of acute gouty arthritis through P2X7R/NLRP3 pathway, promoted the secretion of related inflammatory cytokines, which affected radio of Tregs/Th17 cells. The whole pathogenesis process appeared a pattern from acute attack to remission in time-dependent trend.

Objective:To determine the role of nerve injury-induced protein 1 (NINJ1) introduced plasma membrane rupture (PMR) and damage-associated molecular patterns (DAMPs) release in the pathogenesis and progression of gout and to explore the potential of NINJ1 as a therapeutic target in gout.Methods:Both peripheral blood mononuclear cells (PBMCs) and serum sample from gout patients (n = 58) and healthy controls (n = 16) were collected and processed to NINJ1 expression, lactate dehydrogenase (LDH) detection, NINJ1 inhibition, and NINJ1 expression experiments, respectively. NINJ1 knockdown was carried out by lentivirus in a monosodium urate (MSU) induced rat model, and NINJ1 neutralizing antibody was applied in a MSU induced mouse model.Results:Our results found that NINJ1 was upregulated during a gout flare, and the resulting induction of PMR correlated with gout progression. NINJ1 knockdown significantly reduced the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation and joint swelling in the rat model, and NINJ1 neutralizing antibody also significantly reduced gout flare in the mouse model and PBMCs. Moreover, NINJ1 expression is under NLRP3 inflammasome produced interleukin (IL)-1β control.Conclusion:These results support the notion of a pathogenic role of NINJ1 introduced PMR in gout and provide a detailed mechanism for gout pathogenesis involving inflammatory cell death and DAMPs release introduced by IL-1β. In addition, targeting NINJ1 might be a potential therapeutic approach for gout.

Systemic lupus erythematosus (SLE) is a multisystem autoimmune disease with complex genetic predisposing factors involved. PU.1 is an important member of the ETS transcription factors family which has diverse functions such as regulating the proliferation, differentiation of immune cells and multiple inflammatory cytokines. Previous studies preliminary explored the relation between PU.1 and SLE. To further explain the potential role of PU.1 in the pathogenesis of SLE, 40 SLE patients and 20 age-sex matched healthy controls (HC) were recruited in this study. Flow cytometry was used to test the percentages of CD4+PU.1+T cells in peripheral blood mononuclear cells (PBMCs) from patients with SLE and HC. Expression levels of PU.1 mRNA in CD4+T cells from SLE patients and HC were analyzed by real-time transcription-polymerase chain reaction. Expression levels of plasma IL-1β, IL-9, IL-18, IL-6, IFN-α, TNF-α, IL-10 and TGF-β1 were measured by enzyme-linked immunosorbent assay. The percentage of CD4+PU.1+T cells in PBMCs from patients with SLE was significantly higher than that from HC (P < 0.001). In addition, the PU.1 mRNA expression in CD4+T cells from SLE patients was increased than that from HC (P = 0.002). In SLE patients, no significant correlation was found between the percentage of CD4+PU.1+T cells and the expression of PU.1 mRNA in CD4+T cells (P > 0.05). Associations of PU.1 mRNA expression in CD4+T cells with major clinical and laboratory parameters of SLE patients were also analyzed, but no significant correlations were found. Consistent with previous studies, SLE patients had increased IL-1β, IL-18, IL-6, IFN-α, TNF-α and IL-10 plasma concentrations than HC (P < 0.01). The expression level of plasma TGF-β1 was significantly decreased in SLE patients than in HC (P < 0.001). In SLE patients, the expression level of IL-1β was positive correlated with PU.1 mRNA expression in CD4+T cells (P = 0.001). Our study first time evaluated the expression profile of PU.1 in CD4+T cells from SLE patients confirming that PU.1 may participate in the pathogenesis of SLE.

目的 探讨各因素导致的AKI患者行连续性CRRT抑制AngⅡ途径对PDGF表达的影响及意义.方法 选取2016年2月～2020年5月医院肾脏内科收治的AKI患者68例作为研究组,另选择肾功能正常的肾小球微小病变患者56例作为对照组,AKI患者采用CRRT方案(CVVH模式),评估CRRT对SCr、BUN、ALT、AST、Hb、Na+、K+、Ca2+的治疗性影响,并且分别于CRRT治疗前、治疗后12 h和24h检测AngⅡ、PDGF的表达变化,随后根据PDGF水平与多因素进行相关性分析.此外,经过40 w随访,对各组患者采用Kaplan-Meier法绘制生存曲线,评估近期预后情况.结果 基线资料提示:AKI组的SCr、BUN、AST、Na+、K冰平较对照组升高;Hb、Ca2水平降低(P＜0.05).AKI组经CRRT治疗后的SCr、BUN、Na+、K+水平降低;Hb、Ca2水平升高(P＜0.05).随CRRT治疗时间延长,AKI组的AngⅡ、PDGF表达水平显著下降(P＜0.05).结合Kaplan-Meier法生存分析提示AKI组患者经CRRT治疗后与对照组相比仍存在生存差异.结论 随CRRT治疗时间延长可显著抑制AngⅡ途径,降低PDGF的表达水平,并显著改善AKI组患者预后.

目的 比较高龄(≥80岁)终末期肾衰竭病人采取不同治疗方式的临床疗效,探讨适合高龄终末期肾衰竭病人的治疗方式.方法 收集本院5年间诊治的高龄终末期肾衰竭病人59例,其中18例采取单纯药物治疗,22例行常规血液透析(HD)治疗,19例以保留残余肾功能为目的进行低剂量透析治疗,比较3组间生命体征、血液生化指标、预后等指标的差异.结果 治疗初期(6个月),常规透析剂量组和低透析剂量组在贫血、酸中毒、心功能的纠正和部分生活质量的改善方面优于单纯药物组(P<0.05),常规透析剂量组和低透析剂量组在残余尿量方面差异有统计学意义(P<0.05).在治疗后期(24个月),低透析剂量组在残余尿量、生活质量方面优于单纯药物组和常规透析剂量组(P<0.05).多脏器功能衰竭发生率及生存率3组间差异无统计学意义(P>0.05).结论 高龄终末期肾衰竭病人选择HD治疗应尽量保留残余肾功能,这有利于提高病人的生活质量.

目的 评价在真实世界中达格列净对于糖尿病肾病(DKD)治疗的有效性和安全性.方法 选取2018年1月至2020年5月于我院接受达格列净治疗且随访超过12个月的DKD患者84例,回顾性分析达格列净对DKD患者血糖、血压、体重、血脂、尿蛋白量、肾功能的影响,比较治疗前后临床治疗效果的差异,并记录治疗过程中的不良反应.结果 84例DKD患者的平均年龄为(52.47±8.87)岁,男性56例(66.7％),糖尿病病程8(3,17)年,合并肥胖及超重者35例(41.7％),合并糖尿病性视网膜病变患者47例(56.0％),高血压69例(82.1％),24 h尿蛋白定量为3100(700,9100)mg,估算的肾小球滤过率为87.1(49.7,108.2)mL/(min·1.73 m2).在随访过程中,共有12例(14.3％)患者停用达格列净(男4例、女8例).女性更容易发生泌尿系感染[17.9％(5/28)vs 1.8％(1/56);OR=11.96,95％CI 1.32～108.07,P=0.007].随访12个月后DKD患者的体重(P=0.02)、BMI(P=0.02)、收缩压(P=0.04)、每日胰岛素总量(P<0.01)、空腹血糖(P<0.01)、糖化血红蛋白(P<0.01)、24 h尿蛋白定量(P=0.05)均下降.结论 真实世界的观察性数据支持达格列净对DKD的治疗效果较好且相对安全.

Objective: Iguratimod, an anti-rheumatic drug, has been widely used in the treatment of rheumatoid arthritis, but is still at an investigative stage for treatment of systemic lupus erythematosus (SLE). We examined the therapeutic effects of iguratimod and the mechanism underlying the efficacy in murine lupus model. Methods: Pristane-induced lupus model of BALB/c mice (PI mice) were treated with iguratimod and mycophenolate mofetil. Proteinuria, anti-dsDNA antibodies and immunoglobulins production were measured. Renal pathology was evaluated. The percentage of Th17 and Treg cells in spleen and the expression of cytokines and mRNAs related to Th17 and Treg cells was analyzed. Results: Iguratimod attenuated the severity of nephritis in PI mice in a dose-dependent manner. Proteinuria was continuously decreased and pathology of glomerulonephritis and tubulonephritis was significantly reduced along with reduction of glomerular immune complex deposition. Also, serum anti-dsDNA and total IgG and IgM levels were reduced by iguratimod in mice. It is worth mentioning that the efficacy of the 30 mg/kg/d iguratimod dose is comparable to, or even better than, 100 mg kg/d of mycophenolate mofetil. Furthermore, the percentage of Th17 cells was found decreased and the percentage of Treg cells increased. ROR-gamma t mRNA and serum cytokines (IL-17A and IL-22) of Th17 cells decreased accordingly. By contrast, Foxp3 mRNA and cytokines (TGF-beta and IL10) of Treg cells increased. Conclusion: Iguratimod ameliorates nephritis of SLE and modulates the Th17/Treg ratio in murine nephritis of SLE, suggesting that Iguratimod could be an effective drug in treatment of SLE.

目的:探讨不同炎性细胞因子在急性痛风炎症启动中的作用.方法:利用沉淀法分离痛风和高尿酸血症患者外周血白细胞各40例,分别用尿酸钠(MSU)和MSU+ATP(5'-三磷酸腺苷二钠盐水合物)刺激细胞并培养,采用多重液相蛋白定量技术(CBA)检测细胞培养液中IL-1β、IL-6、IL-8和TNF-α的浓度,分析不同细胞因子在痛风炎症启动中作用.结果:单独MSU刺激白细胞时,痛风和高尿酸血症患者组细胞培养液中IL-1β、IL-6、IL-8和TNF-α的浓度差别均无统计学意义(P>0.05);用MSU+ATP共同刺激白细胞时,痛风组细胞培养液中IL-1β浓度较高尿酸血症组显著升高(Z=-2.081,P=0.037),但两组细胞培养液中IL-6、IL-8和TNF-α浓度差别均无统计学意义(P>0.05).结论:IL-1β是启动痛风急性发作的主要细胞因子.

目的 研究维持性血液透析(maintenance hemodialysis,MHD)相关肌肉减少症(sarcope-nia,简称肌少症)的患病率及危险因素.方法 入选359例MHD患者,收集患者相关资料测量肌肉质量、力量及肌肉功能,将研究对象分成肌少症组和非肌少症组,分析肌少症的患病率及危险因素.结果 本研究纳入359名MHD患者,肌少症83例,非肌少症276例,肌少症患病率为23.1％.多因素Logistic回归分析显示年龄(OR=1.962,95％CI:1.264～3.044,P=0.003)、营养不良(OR=-2.697,95％CI:1.283～5.669,P=0.009)及抑郁(OR=1.707,95％CI:1.116～2.611,P=0.014)是肌少症的独立危险因素,体质量指数(BMI)是肌少症的保护因素(OR=0.040,95％CI:0.015～0.108,P＜0.001).结论 MHD肌少症患病率高达23.1％,年龄、营养不良、抑郁是其独立危险因素,BMI是其保护性因素.

目的 收集高尿酸血症及痛风患者的临床资料及生化结果,分析高尿酸血症及痛风患者并发肾结石的危险因素.方法 收集安徽省立医院门诊和住院及体检中心的痛风患者82例及高尿酸血症患者178例进行对照研究,所有患者统一问卷调查,收集一般资料及实验室检查数据,均进行双肾、输尿管、膀胱B超;采用组间对照及二元Logistic回归方法进行统计学分析.结果 痛风组与高尿酸血症组肾结石发生率分别为36.6％和27.5％,两组比较差异无统计学差异(P>0.05).痛风并发肾结石组相比痛风无肾结石组身高更高、体质量更重、病程更长、低密度脂蛋白(LDL)较高(P<0.05).Logistic回归分析显示:病程>9年[OR=23.493,95％CI(2.824～195.421),P=0.003]、LDL≥4.1 mmol/L[OR=10.160,95％CI(1.218～84.747),P=0.032]是痛风患者发生肾结石的危险因素;痛风并发肾结石组相比高尿酸血症并发肾结石组有吸烟史、饮酒史的比例更高(P<0.05);所有高尿酸血症患者并发结石组相比高尿酸血症无肾结石组的体质量更重,BMI更高,舒张压更高(P<0.05).结论 高尿酸血症与痛风患者均有较高的肾结石发生率.病程>9年、LDL≥4. 1 mmol/L是痛风患者肾结石发病的独立危险因素.高体质量及高体质量指数的痛风/高尿酸血症患者更应注意进行泌尿系结石的筛查以利于早期发现肾结石并及时治疗.

目的 探讨转录因子叉头蛋白p3(Foxp3)基因表达在急、慢性痛风患者外周血调节性T细胞(Treg)不同变化中的作用.方法 选取25例痛风患者(急性痛风11例,慢性痛风14例)和11例健康人为研究对象.用流式细胞术检测外周血Treg细胞,用qPCR技术检测外周血CD4+T细胞中Foxp3 mRNA表达水平.比较三组研究对象的CD4+T细胞Foxp3 mRNA表达水平,分析Foxp3 mRNA表达与急、慢性痛风患者Treg细胞不同变化的相关性.结果 急性痛风组和慢性痛风组的外周血CD4+T细胞中Foxp3 mRNA表达水平均高于健康对照组(P<0.05);急性痛风组和慢性痛风组的Foxp3 mRNA表达水平比较,差异无统计学意义(P>0.05).急性痛风患者中,外周血CD4+T细胞中Foxp3 mRNA的表达与Treg细胞百分比呈显著正相关(r=0.640,P=0.034).慢性痛风患者中,外周血CD4+T细胞中Foxp3 mRNA的表达与Treg细胞百分比呈显著负相关(r=-0.569,P=0.032).结论 急性痛风患者外周血Treg细胞增多与转录因子Foxp3表达增加有关;慢性痛风患者外周血Treg细胞减少,但Foxp3表达仍然增加,提示慢性痛风患者中Treg细胞减少可能与Foxp3蛋白降解有关.

目的 通过建立稳定表达嘌呤受体P2 X配体门控离子通道7(P2X7R)的野生型或突变体型人急性白血病单核细胞株(THP-1)源性单核细胞系,分析高尿酸背景下P2X7R的Arg 307>Gln SNP基因型的功能改变.方法 慢病毒转染THP-1细胞,建立稳定表达野生型、突变体型Arg 307>Gln和空病毒型的THP-1细胞系,设置野生型、突变体型Arg 307>Gln和空病毒型3型,每型分3组,分别加入尿酸单钠盐(MSU)(M组)、MSU+三磷酸腺苷(ATP)(MA组)、空白对照组(C组).丙二醇甲醚醋酸酯(PMA)分别刺激3组细胞,离心撤去PMA;M组、MA组中加入MSU孵育,之后MA组中加入ATP孵育;分别收集3组细胞上清液和细胞.ELISA法检测3组上清液中白细胞介素-1β(IL-1β)蛋白表达水平,qRT-PCR检测3组细胞IL-1β、核苷酸结合寡聚化结构域样受体3(NLRP3)、凋亡相关斑点样蛋白含半胱氨酸天冬氨酸-1蛋白酶结构域(ASC)和Caspase-1 mRNA表达水平.结果 单纯加入MSU晶体后,野生型、突变体型Arg 307>Gln和空病毒型3者之间均无差异(P>0.05).加入ATP孵育后,野生型中IL-1β和NLRP3水平均高于突变体型Arg 307>Gln,差异有统计学意义(P<0.05).而ATP-P2X7R-IL-1β信号通路上ASC和Caspase-1 mRNA水平在野生型和突变体型2者之间无差异(P>0.05).结论 在高尿酸背景下,突变体型Arg 307>Gln使P2X7R功能明显下调,IL-1β和NLRP3表达水平均降低.Caspase-1、ASC可能分别在激活通路过程中被降解.

Background: Previous studies[1] have shown that ATP acts on the sputum receptor P2X ligand-gated ion channel 7 (P2X7R) as a second signal to induce gouty arthritis. Objectives: In this study, the functional changes of three SNP genotypes, Ala348 to Thr, Glu 496 to Ala and Arg307 to Gln, in P2X7R were analyzed with high uric acid. Methods: After transfection of HEK-293T cells by lentivirus, observing the uptake ability of HEK-293T cells to ethidium bromide. The effect of three different mutants on the P2X7 receptor was thus observed on the P2X7 channel. In addition, THP-1 cells were also transfected, stable expression of a THP-1 cell line that has been transfected with a wild-type or different mutants and thus established. Then three types were set up separately, and each type was randomized into three groups: MSU(labeled M), MSU+ATP (labeled MA), and unstimulated control group (labeled C).Detection of IL-1β protein expression level in serum by ELISA and NLRP3, ASC and Caspase-1 mRNA levels in transfected THP-1 cells by qRT-PCR. Results: 1. These three variants have different effects on the uptake function of ATP-induced ethidium + bromide in transfection of HEK-293T cells by lentivirus. Ala 348 to Thr increased P2X 7 -dependent ethidium + bromide uptake (145% of wild-type P2X 7 response, P<0.001). In contrast, Absent or very reduced P2X 7 function was found in Glu 496 to Ala and Arg 307 to Gln subjects, appeared to abolish P2X 7 -dependent dye uptake (38% and 32% of wild-type P2X 7 responses, P<0.001,), who were compared with wild-type. 2. Compared the IL-1β levels of the three variants with the wide-type and empty virus in THP-1 cells, the Ala 348 to Thr mutation significantly up-regulated the serum levels of IL-1β compared with the wide-type and empty virus in group MA with high uric acid (P=0.0007;P=0.013, respectively). Moreover, similar results have also been shown in IL-1β mRNA expressions (P=0.0334; P=0.0307, respectively). The Glu 496 to Ala and Arg 307 to Gln mutations down-regulated the serum levels of IL-1β compared with the wide-type in group MA (P=0.0189;P=0.0164, respectively). 3. NLRP3 mRNA was significantly increased in the Ala 348 to Thr mutation compared with the wide-type and empty virus in group MA (p =0.0003;P=0.0001, respectively). However, NLRP3 mRNA was significantly reduced in the Glu 496 to Ala and Arg 307 to Gln mutations compared with the wide-type in group MA (p =0.0294;P=0.0279, respectively). 4. Wild-type was signifcantly higher than empty virus in the ASC gene expression in group MA(P=0.0022). Morever, the Ala 348 to Thr mutation was higher than empty virus while Arg 307 to Gln mutation was lower than that in group MA (P=0.0138;P=0.0283, respectively). 5. Unlike NLRP3 gene expression, the data showed that the expression of caspase-1 mRNA in group C, M and MA all with no statistical significance, respectively (P>0.05). Conclusion: Our data revealed that Ala 348 to Thr up-regulate the functional status of P2X7R and Glu 496 to Ala and Gln 460 to Arg down-regulate the functional status of P2X7R, which resulted in a significant increase or decrease in IL-1β and NLRP3 expression levels with high uric acid. References: [1]Tao JH, Zhang Y, Li XP, et al. P2X7R: a potential key regulator of acute gouty arthritis[J].Semin Arthritis Rheum, 2013,43(3):376–380. Disclosure of Interests: None declared