Effect of Suan Zao Ren (Semen Ziziphi Spinosae) Extract on the TXNIP/NLRP3 Pathway in Insomniac Rats

Objectives This study aimed to investigate the therapeutic effects of Suan Zao Ren(Semen Ziziphi Spinosae,SZS)extract on insomnia induced by p-chlorophenylalanine(PCPA)in rats and its influence on the thioredoxin-interacting protein(TXNIP)/nucleotide-binding domain Leucine-rich repeat and pyrin domain-containing receptor 3(NLRP3)inflammasome pathway,and to preliminarily explore themechanism by which SZS extract improves insomnia.Methods Fifty male Sprague–Dawley(SD)rats were used,with 8 rats in the blank group and 42 rats in the modeling group.The modeling group was induced by intraperitoneal injection of PCPA at a dose of 500mg·kg^(-1) for six consecutive days,with daily cage exchange.After 6 days,40 successfully modeled rats were randomly divided into five groups:themodel group(equal volume of distilled water),the positive group(0.75 mg·kg^(-1)),and low-,medium-,and high-dose SZS extract groups(1.5,3,and 6 g·kg^(-1),respectively),with 8 rats in each group.Treatments were administered for seven consecutive days.Enzyme-linked immunosorbent assay was used to measure levels of 5-hydroxytryptamine(5-HT)and gamma-aminobutyric acid(GABA)in the rat cerebral cortex.The thiobarbituric acid(TBA)method was used to determine malondialdehyde(MDA)levels,and the hydroxylamine method was used to determine superoxide dismutase(SOD)levels.The 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)method was used to measure total antioxidant capacity(TAOC)in the cerebral cortex.Pathological changes in the cerebral cortex were observed,and Western blot was used to detect the protein expressions of TXNIP,NLRP3,apoptosis-associated speck-like protein containing a Caspase activation and recruitment domain(CARD),and cysteine–aspartate-specific protease 1(Caspase-1)in the cerebral cortex.Results Compared with the blank group,the model group showed a significantly prolonged sleep latency(p<0.001)and a significantly shortened sleep duration(p<0.001).There were no changes in serum MDA and SOD levels.MDA levels in the cerebral cortex were significantly increased(p<0.001),while SOD and TAOC levels were significantly decreased(p<0.001).The 5-HT level was increased(p<0.05),and the GABA level was significantly decreased(p<0.001).SZS extract improved these conditions to varying degrees.Light microscopy showed no significant changes in cortical neurons but transmission electron microscopy revealed intact mitochondrial structures in the blank group,while the model group showed swollen and unclear mitochondria with reduced organelles.After 7 days of treatment,these conditions improved in the SZS extract groups.Compared with the blank group,the expressions of the four proteins in the model group were increased,and the expressions of these proteins were decreased in the SZS extract groups compared with the model group.Conclusion SZS extract may exert an antioxidant effect to treat insomnia by downregulating the expression of TXNIP/NLRP3 proteins and regulating oxidative stress levels in the cerebral cortex.

Amitriptyline inhibits NLRP3 inflammasome activation via the ASM/CE pathway in a cell model of NAFLD

Background:Nonalcoholic fatty liver disease(NAFLD)is a global health concern with the acid sphingomyelinase(ASM)/ceramide(CE)pathway and the NOD-like receptor family,pyrin domain-containing protein 3(NLRP3)inflammasome identified as pivotal players in lipid disorders and inflammation.This study explores the interaction mechanism between the ASM/CE pathway and NLRP3 in NAFLD cell models,aiming to understand the impact of amitriptyline(Ami),an ASM inhibitor,on lipid deposition and hepatocyte injury by regulating the ASM/CE-NLRP3 pathway.Methods:HepG2 and HL-7702 cells were exposed to free fatty acids(FFAs)to establish the NAFLD model.The cells were divided into 5 groups:control group,model group,Ami group,tumor necrosis factoralpha(TNF-α)group,and Ami+TNF-αgroup.Intracellular lipid droplets were visualized using Oil Red O staining,and Western blot analysis quantified ASM,NLRP3,and caspase 1 protein expression.Enzyme linked immunosorbent assay(ELISA)was measured CE and ASM levels,while qRT-PCR assessed mRNA expression.The apoptotic rate was evaluated by flow cytometry(FCM).Results:Following FFAs incubation,significant increases in ASM and CE levels were observed in HepG2 and HL-7702 cells,accompanied by elevated expression of NLRP3,and caspase 1,and IL-1β.TNF-αtreatment further amplified these indicators.Ami demonstrated a reduction in lipid deposition,suppressed ASM/CE pathway activation,downregulated NLRP3 and caspase 1 expression,and improved apoptosis.Additionally,MCC950,a selective inhibitor of the NLRP3,mitigated NLRP3,caspase 1,and IL-1βexpression,alleviating lipid deposition and apoptosis in the NAFLD cell model.Conclusion:The ASM/CE-NLRP3 pathway in NAFLD cells promotes hepatocyte steatosis,inflammation,and cell damage.Ami emerges as a promising therapeutic agent by inhibiting the ASM/CE-NLRP3 pathway,underscoring its potential as a key target for NAFLD treatment.

益母草碱调节IRE1α/TXNIP/NLRP3信号通路对急性呼吸窘迫综合征大鼠炎症反应的影响

目的探究益母草碱调节IRE1α/TXNIP/NLRP3信号通路对急性呼吸窘迫综合征(ARDS)大鼠炎症反应的影响。方法将大鼠随机分为空白组、模型组、益母草碱低剂量组、益母草碱高剂量组、联用HY-N2485组(高剂量益母草碱+NLRP3激活剂)。检测大鼠肺功能;血气分析仪检测氧分压(PaO_(2))和二氧化碳分压(PaCO_(2));ELISA法检测肺泡灌洗液白细胞介素-10(IL-10)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)水平;检测肺组织干湿比;检测肺组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)活性;HE染色观察大鼠肺组织病理形态;Western blotting检测肺组织中IRE1α、TXNIP、NLRP3、Caspase-1、Cleaved Caspase-3蛋白表达。结果与空白组相比,模型组肺组织形态有明显损伤,PEF、FVC和FEV0.3/FVC水平、PaO_(2)、OI水平、肺泡灌洗液IL-10水平、肺组织CAT和SOD活性降低,PaCO_(2)水平、肺泡灌洗液IL-1β、IL-6、TNF-α水平、肺组织病理评分、肺组织MDA含量、IRE1α、TXNIP、NLRP3、Caspase-1、Cleaved Caspase-3蛋白表达水平升高(P<0.05);与模型组相比,益母草碱低、高剂量组大鼠肺组织损伤明显减轻,PEF、FVC和FEV0.3/FVC水平、PaO_(2)、OI水平、肺泡灌洗液IL-10水平、肺组织CAT和SOD活性升高,PaCO_(2)水平、肺泡灌洗液IL-1β、IL-6、TNF-α水平、肺组织病理学评分、肺组织MDA含量、IRE1α、TXNIP、NLRP3、Caspase-1、Cleaved Caspase-3蛋白表达水平降低(P<0.05);HY-N2485可减弱益母草碱对ARDS大鼠的改善作用(P<0.05)。结论益母草碱可降低ARDS大鼠炎症和氧化应激,减轻肺组织损伤,改善肺功能,可能与抑制IRE1α/TXNIP/NLRP3信号通路有关。

Porphyromonas gingivalis Induces Chronic Kidney Disease through Crosstalk between the NF-κB/NLRP3 Pathway and Ferroptosis in GMCs

Objective Porphyromonas gingivalis(P.gingivalis)is a gram-negative bacterium found in the human oral cavity and is a recognized pathogenic bacterium associated with chronic periodontitis and systemic diseases,including chronic kidney disease(CKD),but the roles and molecular mechanism of P.gingivalis in CKD pathogenesis are unclear.Methods In this study,an animal model of oral P.gingivalis administration and glomerular mesangial cells(GMCs)cocultured with M1-polarized macrophages and P.gingivalis supernatant were constructed.After seven weeks of P.gingivalis gavaged,peripheral blood was collected to detect the changes in renal function.By collecting the teeth and kidneys of mice,H&E staining and IHC were used to analyze the expression of periodontal inflammatory factors in mice,PAS staining was used to analyze glomerular lesions.The supernatant of macrophages was treated with 5%P.gingivalis supernatant.H&E staining,IHC,Western blot and RT-PCR were applied to analyze renal inflammatory factors,macrophage M1 polarization,NF-κB,NLRP3 and ferroptosis changes in vitro.Results We found that oral P.gingivalis administration induced CKD in mice.P.gingivalis supernatant induced macrophage polarization and inflammatory factor upregulation,which triggered the activation of the NF-κB/NLRP3 pathway and ferroptosis in GMCs.By inhibiting the NF-κB/NLRP3 pathway and ferroptosis in GMCs,cell viability and the inflammatory response were partially alleviated in vitro.Conclusion We demonstrated that P.gingivalis induced CKD in mice by triggering crosstalk between the NFκB/NLRP3 pathway and ferroptosis in GMCs.Overall,our study suggested that periodontitis can promote the pathogenesis of CKD in mice,which provides evidence of the importance of periodontitis therapy in the prevention and treatment of CKD.

Branched-chain fatty acids from goat milk alleviate ulcerative colitis via the TLR4/NF-κB/NLRP3 pathway

Branched-chain fatty acids(BCFAs)are new bioactive fatty acids with anti-inflammatory properties.However,the role of BCFAs in alleviating ulcerative colitis has not been clarified.Herein,we evaluated the protective effect of BCFAs from goat milk in mice with colitis induced using dextran sodium sulfate(DSS)and explored the corresponding mechanism.These results show that BCFAs extracted from goat milk can significantly alleviate weight loss in mice,and reduce the disease activity index and the activity of myeloperoxidase while increasing the content of antioxidant enzymes in colon tissue and reducing the oxidation stress response.These data also show that BCFAs can down-regulate the gene and protein expression of the toll-like receptor 4(TLR4)/nuclear factorκB p65(NF-κB p65)/NOD-like receptor thermal protein domain associated protein 3(NLRP3)signaling pathway,and at the same time significantly reduce the expression of pro-inflammatory factors tumor necrosis factorα(TNF-α),interleukin 1β(IL-1β),and IL-18 in colon tissue,and significantly increase the expression of the anti-inflammatory factor IL-10.In conclusion,these results demonstrated that BCFAs in goat milk exerted effects on colitis-related inflammatory cytokines and inhibited inflammation by inducing the TLR4/NF-κB/NLRP3 pathway to alleviate DSS-induced ulcerative colitis.This study provides evidence for the potential of BCFAs as bioactive fatty acids in food products and to ameliorate ulcerative colitis development in mice.

姜黄素通过调控Trx-1/TXNIP/NLRP3通路减轻对乙酰氨基酚诱导的大鼠急性肾损伤

目的探究姜黄素(curcumin,CUR)对对乙酰氨基酚(acetaminophen,APAP)诱导的大鼠急性肾损伤的保护作用及机制。方法48只雄性SD大鼠随机均分为正常组、模型组、阳性药NAC组、CUR低(50 mg/kg)、中(100 mg/kg)、高(200 mg/kg)剂量组。NAC及CUR灌胃给药10 d后,一次性灌胃给予2 g/kg APAP建立急性肾损伤模型,造模24 h后,取血并分离大鼠,计算肾指数;检测血清肌酐(Cr)、尿素氮(BUN)水平;HE染色评价大鼠肾病理变化;检测大鼠肾组织谷胱甘肽(GSH)、总超氧化物歧化酶(T-SOD)、过氧化氢酶(CAT)活性及丙二醛(MDA)水平;Western blot检测大鼠肾组织Trx-1、TXNIP、NLRP3炎症小体等蛋白的表达水平。结果与正常组相比,模型组大鼠肾指数显著增加(P<0.01),血清Cr、BUN水平显著升高(P<0.01),病理切片显示大鼠肾组织出现明显的损伤,大鼠肾组织中GSH、T-SOD、CAT的活性显著降低(P<0.01),MDA的含量显著升高(P<0.01),大鼠肾组织中Trx-1的表达明显下调(P<0.05),TXNIP、NLRP3、ASC、Cleaved caspase-1、mature IL-1β蛋白的表达明显上调(P<0.01)。与模型组相比,CUR中、高剂量组肾指数显著降低(P<0.01),病理损伤改善,血清Cr、BUN水平显著降低(P<0.01),肾组织中GSH、T-SOD、CAT活性显著升高,MDA含量显著降低(P<0.05或P<0.01),Trx-1的表达明显上调(P<0.05或P<0.01)、TXNIP、NLRP3、ASC、Cleaved caspase-1、mature IL-1β蛋白表达明显下调(P<0.01)。结论姜黄素预防性给药通过调控Trx-1/TXNIP/NLRP3通路减轻APAP诱导的大鼠急性肾损伤。

基于PERK/TXNIP/NLRP3通路探讨左归降糖通脉方对糖尿病合并脑梗死大鼠神经元细胞焦亡的影响

目的基于蛋白激酶R样内质网激酶(PERK)/硫氧还蛋白互作蛋白(TXNIP)/NOD样受体蛋白3(NLRP3)通路,探讨左归降糖通脉方对糖尿病合并脑梗死(diabetes mellitus complicated with cerebral infarction,DM-CI)大鼠神经元细胞焦亡的影响。方法将95只大鼠随机分为假手术组,模型组,左归降糖通脉方低(12 g·kg^(-1))、高剂量(24 g·kg^(-1))组,阳性对照(吡格列酮二甲双胍片+阿司匹林肠溶片)组。除假手术组外,其余各组以高糖高脂饲料喂养4周后联合35 mg·kg^(-1)1%链脲佐菌素(STZ)腹腔注射合并大脑中动脉闭塞(MCAO)术建立DM-CI大鼠模型。用蒸馏水,左归降糖通脉方低、高剂量,吡格列酮二甲双胍片和阿司匹林肠溶片进行相应干预,Zea-Longa评分法对大鼠进行神经功能缺损(NIHSS)评分,TTC染色法测量大鼠脑梗死体积,检测各组大鼠随机血糖和果糖胺水平,HE染色观察大鼠脑组织病理学变化,Western Blot法检测大鼠缺血侧脑皮质、海马区PERK、p-PERK、真核起始因子2α(eIF2α)、p-eIF2α、TXNIP、NLRP3、半胱天冬酶-1(Caspase-1)、消皮素D(GSDMD)、白细胞介素(IL)-1β和IL-18蛋白的表达,免疫荧光双染法检测缺血侧脑皮质、海马区组织中神经元特异性烯醇化酶(NSE)与NLRP3蛋白的共定位表达。结果与假手术组比较,模型组大鼠随机血糖、果糖胺水平、NIHSS评分、脑梗死体积、缺血侧脑皮质和海马区p-PERK/PERK、p-eIF2α/eIF2α、TXNIP、NLRP3、Caspase-1、GSDMD、IL-1β、IL-18蛋白水平和NSE与NLRP3双阳性信号细胞数量均明显升高(P<0.05,P<0.01)。与模型组比较,左归降糖通脉方高剂量组的随机血糖、果糖胺水平、NIHSS评分、脑梗死体积、缺血侧脑皮质和海马区p-PERK/PERK、p-eIF2α/eIF2α、TXNIP、NLRP3、Caspase-1、GSDMD、IL-1β、IL-18蛋白水平和NSE与NLRP3双阳性信号细胞数量均明显降低(P<0.05,P<0.01)。结论左归降糖通脉方可改善DM-CI大鼠糖代谢紊乱,减轻神经功能损伤,其机制可能与左归降糖通脉方抑制PERK/TXNIP/NLRP3通路,减轻神经元细胞焦亡有关。

Lacticaseibacillus rhamnosus Fmb14 ameliorates hyperuricemia-induced hepatocyte pyroptosis via NLRP3 inflammasome cascade inhibition

Hyperuricemia is a high-risk factor for the development of gout and renal fibrosis,but the adverse effects of hyperuricemia on the liver have been seriously neglected.This research investigated the ameliorating effect of Lacticaseibacillus rhamnosus Fmb14 on hyperuricemia induced liver dysfunction both in vitro and in vivo.Cell free extracts of high dose L.rhamnosus Fmb14 treatment reduced the death rate of HepG2 cell lines from 24.1%to 14.9%by inhibiting NLRP3 recruitment,which was mainly activated by reactive oxygen species release and mitochondrial membrane potential disorder.In purine dietary induced hyperuricemia(PDIH)mice model,liver oedema and pyroptosis were ameliorated after L.rhamnosus Fmb14 administration through downregulating the expression levels of NLRP3,caspase-1 and gasdermin-D from 1.61 to 0.86,3.15 to 1.01 and 5.63 to 2.02,respectively.L.rhamnosus Fmb14 administration restored mitochondrial inner membrane protein(MPV17)and connexin 43 from 2.83 and 0.73 to 0.80 and 0.98 respectively in PDIH mice,indicating that dysbiosis of mitochondrial membrane potential was restored in liver.Intriguingly,PDIH pyroptosis stimulates the process of apoptosis,which leads to severe leakage of hepatocytes,and both of pyroptosis and apoptosis were decreased after L.rhamnosus Fmb14 treatment.Therefore,L.rhamnosus Fmb14 is a promising biological resource to maintain homeostasis of the liver in hyperuricemia and the prevention of subsequent complications.

桑黄素通过抑制TXNIP/NLRP3/Caspase-1信号通路对脑缺血再灌注大鼠神经元凋亡的影响

目的探讨桑黄素(Morin)通过调控硫氧还蛋白互作蛋白(TXNIP)/核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)/胱天蛋白酶1(Caspase-1)信号通路,对脑缺血再灌注大鼠神经元凋亡的影响。方法将60只大鼠随机均分为假手术组、模型组、Morin低剂量(Morin-L,10 mg/kg)组、Morin高剂量(Morin-H,40 mg/kg)组、Morin-H+pLVXNC组(40 mg/kg Morin+pLVX-NC)、Morin-H+pLVX-TXNIP组(40 mg/kg Morin+pLVX-TXNIP)。采用线栓法制作大鼠大脑中动脉栓塞模型(除外假手术组),给药1 h后再灌注。再灌注72 h后,进行神经功能缺损评分;取出全脑进行脑含水量测定;苏木精-伊红(HE)染色观察脑皮质半暗带病理损伤情况;2,3,5-氯化三苯基四氮唑(TTC)检测脑组织梗死面积;酶联免疫吸附试验检测血清中白细胞介素(IL)-1β、IL-18含量;Western blot检测脑皮质中TXNIP/NLRP3/Caspase-1信号通路蛋白表达水平。结果与假手术组比较,模型组神经功能缺损评分,IL-1β、IL-18水平,脑含水量,梗死面积,TXNIP、NLRP3、Caspase-1蛋白表达均增加(P<0.05);与模型组比较,Morin-L组、Morin-H组神经功能缺损评分,IL-1β、IL-18水平,脑含水量,梗死面积,TXNIP、NLRP3、Caspase-1蛋白表达均降低(P<0.05);与Morin-H+pLVX-NC组比较,Morin-H+pLVX-TXNIP组神经功能缺损评分,IL-1β、IL-18水平,脑含水量,梗死面积,TXNIP、NLRP3、Caspase-1蛋白表达增加(P<0.05)。结论Morin可以减轻脑缺血再灌注大鼠脑损伤,抑制神经元凋亡,作用机制可能与抑制TXNIP/NLRP3/Caspase-1信号通路活化有关。

热淋清颗粒对慢性尿路感染模型大鼠TXNIP/NLRP3炎症小体通路及免疫状态的影响

目的:探讨热淋清颗粒对慢性尿路感染模型大鼠TXNIP/NLRP3炎症小体通路及免疫功能的影响。方法:建立大鼠慢性尿路感染模型,随机分为模型组、热淋清低、中、高剂量组、左氧氟沙星组、假手术组。热淋清低、中、高剂量组分别灌胃7、14、28 g/kg热淋清颗粒;假手术组和模型组分别灌胃等体积蒸馏水;左氧氟沙星组灌胃20 mg/kg左氧氟沙星,连续给药14 d。HE染色观察肾脏组织病理形态;检测尿液量及尿白细胞数量;ELISA检测血清TNF-α、IL-6、IL-8、IL-1β水平;Western blot检测肾脏组织TXNIP、NLRP3、Caspase-1蛋白表达。结果:与假手术组相比,模型组尿液量明显减少(P<0.05),尿白细胞数量、血清TNF-α、IL-6、IL-8、IL-1β水平及肾脏组织TXNIP、NLRP3、Caspase-1蛋白表达明显升高(P<0.05),肾脏组织出现明显病理学损伤。与模型组相比,热淋清低、中、高剂量组和左氧氟沙星组尿液量明显增加(P<0.05),尿白细胞数量、血清TNF-α、IL-6、IL-8、IL-1β水平及肾脏组织TXNIP、NLRP3、Caspase-1蛋白表达明显降低(P<0.05),且肾脏组织病理学损伤明显改善。结论:热淋清颗粒对慢性尿路感染大鼠具有较好治疗效果,可能与抑制TXNIP/NLRP3炎症小体通路活化从而发挥炎症免疫调节作用有关。

Elaidic acid-induced intestinal barrier damage led to gut-liver axis derangement and triggered NLRP3 inflammasome in the liver of SD rats

Previous studies have shown that trans fatty acids(TFA) are associated with several chronic diseases,the gut microbiota is directly influenced by dietary components and linked to chronic diseases.Our research investigated the effects of elaidic acid(EA),a typical TFA,on the gut microbiota to understand the underlying mechanisms of TFA-related chronic diseases.16S rDNA gene sequencing on faecal samples from Sprague-Dawley rats were performed to explore the composition change of the gut microbiota by EA gavage for 4 weeks.The results showed that the intake of EA increased the abundance of well-documented harmful bacteria,such as Proteobacteria,Anaerotruncus,Oscillibacter and Desulfovibrionaceae.Plus,EA induced translocation of lipopolysaccharides(LPS) and the above pathogenic bacteria,disrupted the intestinal barrier,led to gut-liver axis derangement and TLR4 pathway activation in the liver.Overall,EA induced intestinal barrier damage and regulated TLR4-MyD88-NF-κB/MAPK pathways in the liver of SD rats,leading to the activation of NLRP3 inflammasome and inflammatory liver damage.

To investigate the effect of Shenqi Tiaoshen Formula on CSE induced inflammatory response of MH-S cells based on TLR4/NF-kB/NLRP3 pathway

Objective:To study the effects of Shenqi Tiaoshen Formula(SQTS)on the inflammatory response of MH-S cells induced by cigarette smoking extract(CSE)and its mechanism based on TLR4/NF-kB/NLRP3 pathway.Methods:MH-S cells were used as subjects to evaluate cell viability by CCK-8 method.The levels of TNF-α,IL-1βand IL-6 in the supernatant were detected by ELISA.ROS were detected by DCFH-DA fluorescence probe.Western blotting was used to detect the expression of TLR4/NF-kB/NLRP3 pathway protein,and TAK-242,a TLR4 inhibitor,was used to verify the role of SQTS in the TLR4/NF-kB/NLRP3 pathway.Results:Compared with blank group,the cell survival rate of CSE group was decreased,and the contents of inflammatory cytokines TNF-α,IL-1βand IL-6 were increased(P<0.05),ROS fluorescence expression level was significantly increased(P<0.01),TLR4/NF-kB/NLRP3 pathway protein expression was significantly increased(P<0.05);Compared with CSE group,the survival rate of cells in SQTS groups was increased,and the expression levels of the above indexes were decreased(P<0.05),and TLR4/NF-kB/NLRP3 pathway protein decreased in TAK-242 groups(P<0.05).Conclusion:SQTS can reduce the inflammatory response of MH-S cells induced by CSE by inhibiting TLR4/NF-kB/NLRP3 pathway.

Mechanism of Sanshi decoction inhibits macrophage pyroptosis by inhibiting BRD4/NF-κB/NLRP3 pathway in the treatment of gouty arthritis

Objective:To observe the effect of Sanshi decoction on BRD4/NF-κB/NLRP3 pathwaymediated macrophage pyroptosis,so as to elucidate the molecular mechanism of Sanshi decoction in the treatment of gouty arthritis.Methods:THP-1 was induced into macrophages with foboside and the divided into the control group,model group,low-dose,medium-dose,high-dose group of Sanshi decoction,and BRD4 inhibitor group.Except for the control group,the remaining groups were induced with monosodium urate crystals to construct a gouty arthritis cell model.The activity of macrophages was detected by CCK8,the level of macrophage pyroptosis was detected by flow cytometry,the activity of LDH,the content of IL-1β and IL-18 were detected by enzyme-linked immunosorbent assay,and the expression of related proteins in the BRD4/NF-κB/NLRP3 pathway was detected by Western blot.Results:Compared with the control group,macrophage activity was decreased in the model group,and the level of pyroptosis,LDH activity,contents of IL-1β and IL-18,expression levels of BRD4,p-NF-kB p65,NLRP3,Caspase-1 p20,and IL-1β protein were significantly up-regulated,the differences were statistically significant(P<0.05 and P<0.01).Compared with the model group,macrophage activity was up-regulated in the Sanshi Decoction,and the level of pyroptosis,LDH activity,IL-1β and IL-18 contents,expression levels of BRD4,p-NF-kB p65,NLRP3,Caspase-1 p20,and IL-1β protein were significantly decreased with statistically significant differences(P<0.05 and P<0.01).Conclusion:Sanshi decoction inhibits macrophage pyroptosis by inhibiting BRD4/NF-κB/NLRP3 pathway activation,thus improving the inflammation level of gouty arthritis.

黄芪多糖调控TXNIP/NLRP3炎症小体对高糖诱导的人视网膜内皮细胞凋亡的作用机制

目的探究黄芪多糖调控TXNIP/NLRP3炎症小体对高糖诱导的人视网膜内皮细胞凋亡的作用机制。方法将hRECs细胞随机分为Control组、HG组、APS组和APS+pcDNA3.1-TXNIP组。采用CCK-8和流式细胞仪检测细胞存活率和凋亡率;DCFH-DA荧光探针检测活性氧(ROS)水平;检测细胞中氧化应激和炎症因子水平;蛋白质印迹法检测细胞中TXNIP、NLRP3、IL-1β和caspase-1蛋白表达。结果和Control组相比,HG组细胞存活率、SOD活性和GSH-Px含量明显降低,细胞凋亡率、ROS相对荧光强度、MDA含量、IL-1β和IL-18含量及细胞中TXNIP、NLRP3、IL-1β、caspase-1蛋白表达明显增加(P<0.05);和HG组相比,APS组细胞存活率、SOD活性和GSH-Px含量明显增加,细胞凋亡率、ROS相对荧光强度、MDA含量、IL-1β和IL-18含量及细胞中TXNIP、NLRP3、IL-1β、caspase-1蛋白表达明显降低(P<0.05);和APS组相比,APS+pcDNA3.1-TXNIP组细胞存活率、SOD活性和GSH-Px含量明显降低,细胞凋亡率、ROS相对荧光强度、MDA含量、IL-1β和IL-18含量及细胞中TXNIP、NLRP3、IL-1β、caspase-1蛋白表达明显增加(P<0.05)。结论黄芪多糖可抑制高糖诱导的视网膜内皮细胞凋亡、炎症反应和氧化应激损伤,其作用机制可能和抑制TXNIP/NLRP3通路激活有关。

苏萸痛风方通过ROS/TXNIP/NLRP3信号通路抗痛风性关节炎的作用机制

目的 研究苏萸痛风方抗痛风性关节炎的作用机制。方法 将雄性SD大鼠按体质量随机分为正常对照组、模型组、秋水仙碱片组(阳性对照药,0.3 mg/kg)和苏萸痛风方高、中、低剂量组(5、2.5、1.25 g/kg),每组10只。给药组大鼠每日灌胃相应药物1次(10 mL/kg),连续给药7 d。正常对照组和模型组大鼠同法灌胃等体积水。第6天给药后1 h,除正常对照组外,其余各组大鼠于关节腔注入尿酸钠复制痛风性关节炎模型。检测大鼠造模后2、6、24 h的关节肿胀度和炎症指数评分。末次给药后1 h,检测大鼠血清中氧化应激相关指标[超氧化物歧化酶(SOD)、黄嘌呤氧化酶(XOD)、丙二醛(MDA)]活性及炎症因子[白细胞介素1β(IL-1β)、IL-18、肿瘤坏死因子α(TNF-α)]水平,观察各组大鼠踝关节组织病理学变化,检测大鼠踝关节组织中硫氧还蛋白互作蛋白(TXNIP)、NOD样受体热蛋白结构域相关蛋白3(NLRP3)、凋亡相关斑点样蛋白(ASC)的表达水平。结果 经苏萸痛风方干预后,大鼠关节肿胀度减轻、炎症指数评分降低、滑膜组织水肿改善、炎症细胞数量减少。苏萸痛风方高剂量组大鼠血清中SOD活性显著升高(P<0.01),XOD、MDA活性以及IL-1β、IL-18、TNF-α水平均显著降低(P<0.01),踝关节组织中ROS水平和TXNIP、NLRP3、ASC蛋白表达水平均显著降低(P<0.05或P<0.01)。苏萸痛风方中、低剂量组上述部分指标活性/水平也发生显著逆转(P<0.05或P<0.01)。结论 苏萸痛风方可通过ROS/TXNIP/NLRP3信号通路,抑制NLRP3炎性小体激活,进而发挥抗痛风性关节炎的作用。

绿原酸通过ROS/TXNIP/NLRP3信号通路介导的细胞焦亡途径减轻脓毒症小鼠急性肺损伤

目的:探究绿原酸对脓毒症小鼠急性肺损伤的影响及活性氧/硫氧还蛋白互作蛋白/NOD样受体蛋白3(ROS/TXNIP/NLRP3)信号通路在其中的作用。方法:建立脓毒症小鼠模型,随机分为模型组、阳性对照(地塞米松)组及低剂量(5 mg/kg)、中剂量(10 mg/kg)、高剂量(20 mg/kg)绿原酸组,每组各12只,另取12只正常小鼠作为对照组。计数肺泡灌洗液中炎症细胞数量;测定肺组织湿重/干重(W/D)比值;HE染色观察肺组织病理改变;TUNEL法检测肺组织细胞凋亡;ELISA法测定肺泡灌洗液中白细胞介素(IL)-1β、IL-6和肿瘤坏死因子α(TNF-α)含量,相应试剂盒检测肺组织中丙二醛(MDA)、超氧化物歧化酶(SOD)和活性氧(ROS)水平;Westernblot法检测TXNIP、NLRP3蛋白、caspase-1及细胞焦亡关键蛋白GasderminD的N末端片段(GSDMD-N)的表达水平。结果:相较于对照组,模型组肺泡间隔增厚,肺泡内及肺泡间质水肿,炎症细胞数量显著增加,IL-1β、IL-6和TNF-α含量、W/D比值、细胞凋亡率、MDA和ROS水平以及TXNIP、NLRP3、caspase-1和GSDMD-N蛋白水平均显著升高(P<0.05);而模型组SOD水平较对照组显著降低(P<0.05)。相较于模型组,绿原酸低、中、高剂量组肺泡间隔增厚及水肿情况有所减轻,炎症细胞数量显著减少,IL-1β、IL-6和TNF-α含量、W/D比值、细胞凋亡率、MDA和ROS水平以及TXNIP、NLRP3、caspase-1和GSDMD-N蛋白水平显著降低(P<0.05);而模型组SOD水平较对照组显著升高(P<0.05);绿原酸高剂量组与阳性对照组上述指标差异无统计学意义(P>0.05)。结论:绿原酸可能通过下调ROS/TXNIP/NLRP3信号通路表达水平,抑制炎症分子产生及细胞焦亡发生,有效减轻脓毒症小鼠肺损伤。

白藜芦醇抑制TXNIP/NLRP3信号通路改善脂多糖诱导的肾小管上皮细胞炎性损伤

[目的]观察白藜芦醇对脂多糖诱导的肾小管上皮细胞(HK-2)炎性损伤的保护作用,及其对TXNIP/NLRP3炎性通路的影响。[方法]采用脂多糖(LPS,1 mg/L)体外诱导HK-2细胞炎性损伤模型,将细胞分为正常对照组、LPS诱导组、LPS+低剂量(50μmol/L)白藜芦醇组、LPS+中剂量(100μmol/L)白藜芦醇组、LPS+高剂量(200μmol/L)白藜芦醇组;CCK8法检测HK-2细胞相对存活率,酶联免疫吸附测定(ELISA)法检测炎性因子白介素-6(IL-6)、白介素-1β(IL-1β)及肿瘤坏死因子-α(TNF-α)水平,Western Blot方法分析诱导型一氧化氮合酶(iNOS)、环氧酶-2(COX-2)、人硫氧还蛋白互作蛋白(TXNIP)及人隐热蛋白(NLRP3)表达。[结果]与LPS诱导组比较,不同浓度白藜芦醇干预的HK-2细胞相对存活率明显升高(P<0.05),细胞上清炎性因子IL-6、IL-1β及TNF-α水平明显降低(P<0.05),细胞中炎性蛋白iNOS及COX-2表达水平显著降低(P<0.05);细胞中TXNIP及NLRP3蛋白表达也显著降低(P<0.05),并表现出剂量依赖性。[结论]白藜芦醇可以明显抑制LPS诱导的HK-2细胞炎症损伤,抑制炎症相关蛋白的表达,其作用机制可能与抑制TXNIP/NLRP3炎性通路活化相关。

TXNIP/NLRP3炎性小体通路在COPD患者机体炎症反应中的作用研究

目的:探讨TXNIP/NLRP3炎性小体通路在COPD患者发病机制中的可能作用。方法:选取2017年11月至2018年5月住院的40例COPD患者为急性加重期组,其经过治疗进入稳定期后纳入稳定期组,40例健康体检者为对照组。荧光定量PCR法测定外周血单个核细胞中TXNIP mRNA、NLRP3 mRNA水平,Western blot法检测外周血单个核细胞中TXNIP及NLRP3蛋白表达水平。结果:(1)COPD患者急性加重期组NLRP3、TXNIP的mRNA和蛋白表达水平均显著高于对照组(P<0. 05)和稳定期组(P<0. 05); COPD患者稳定期组的NLRP3 mRNA和蛋白表达水平显著高于对照组(P<0. 05)。(2)相关性分析:急性加重期组NLRP3 mRNA表达量与FEV1%pred、FEV1/FVC负相关;急性加重期组和稳定期组的NLRP3 mRNA、TXNIP mRNA表达量均与CAT评分正相关。结论:TXNIP/NLRP3炎性小体通路参与COPD患者的机体炎症反应。

TXNIP/NLRP3通路在妊娠期糖尿病胎盘滋养层细胞迁移侵袭中的作用研究体会

研究TXNIP/NLRP3通路在妊娠期糖尿病胎盘滋养层细胞迁移侵袭中的作用。方法 择选2020年7月~2021年7月收治的60例孕妇,将其中30例合并妊娠糖尿病(GDM)的产妇作为观察组,将其中30例正常孕妇作为对照组,对两组孕妇进行孕期产检,记录孕期产检数据,包括血脂、血糖、IL-β水平、IL-6水平、 Homaβ、HomaIR水平等。结果 FBG、2hPG、HbA1c水平高于对照组,P<0.05;观察组TC、TG、LDL-C水平高于对照组,HDL-C水平低于对照组,P<0.05;观察组的Homaβ低于对照组,HomaIR、IL-1β水平高于对照组,P<0.05;观察组TXNIP、NLRP3水平高于对照组,P<0.05。结论 GDM患者不但存在糖代谢,还会对血脂水平造成影响,机体炎症小体水平高于正常孕妇,TXNIP/NLRP3炎症小体/IL-1β途径与GDM胎盘组织胰岛素抵抗有一定的相关性,可通过检测孕妇体内IL-1β水平,来预测GDM的发生。

TXNIP介导的NLRP3炎症小体激活在心肌微血管内皮细胞缺氧/复氧损伤中的作用

目的研究硫氧还蛋白结合蛋白(TXNIP)介导的NLRP3炎症小体激活在心肌微血管内皮细胞(CMECs)缺氧/复氧(H/R)损伤中的作用。方法分离培养C57BL/6J小鼠CMECs,随机分为对照组、H/R损伤组、H/R+Scrambled siRNA组和H/R+TXNIP siRNA组。各组处理后使用ELISA试剂盒检测细胞IL-1β水平,采用免疫共沉淀的方法检测TXNIP和NLRP3的相互作用,Western blot检测TXNIP和NLRP3的表达水平,乳酸脱氢酶试剂盒检测细胞培养上清LDH释放情况和细胞Caspase-3活性。结果与对照组相比,H/R后CMECs的NLRP3表达水平明显升高(P<0.05),NLRP3炎症小体激活(IL-1β水平升高,P<0.01),且TXNIP和NLRP3的结合效果明显增强(P<0.01)。运用TXNIP siRNA抑制TXNIP表达后行H/R处理,与H/R+Scrambled siRNA组相比,NLRP3炎症小体激活水平明显下降(IL-1β水平降低),细胞Caspase-3活力和LDH释放水平均显著降低(均P<0.05)。结论在CMECs发生H/R损伤时,NLRP3炎症小体激活,TXNIP和NLRP3的相互作用增强;抑制TXNIP表达后,NLRP3炎症小体激活水平降低,内皮损伤减轻。说明TXNIP介导的NLRP3炎症小体激活参与CMECs的H/R损伤,可能成为CMECs的H/R损伤的新机制。