基于PPARγ/NF-κB信号通路探讨四妙勇安汤对脂肪细胞炎症的作用机制研究

目的:从PPARγ/NF-κB信号通路探讨四妙勇安汤对脂多糖(lipopolysaccharide,LPS)诱导的脂肪细胞炎症的影响及作用机制。方法:采用LPS诱导成熟脂肪细胞制备脂肪细胞炎症模型。给予四妙勇安汤冻干粉干预24 h后观察脂肪细胞炎症信号通路蛋白的表达,给予PPARγ抑制剂GW9662干预后明确四妙勇安汤是否是通过关键因子PPARγ发挥抗炎作用。CCK-8法检测四妙勇安汤对细胞活力的影响;蛋白质免疫印迹法(Western blot,WB)检测细胞内炎性因子IL-6、NLRP3及相关调控因子PPARγ、NF-κB p65、LXRα和GLUT4的蛋白表达。结果:CCK-8检测结果显示四妙勇安汤冻干粉在浓度为0~500μg/mL范围内对细胞活力无显著影响。WB结果显示:经1μg/mL LPS刺激脂肪细胞造模,与对照组相比,模型组细胞内炎症因子IL-6、NLRP3蛋白表达增加,PPARγ蛋白表达降低、NF-κB p65蛋白表达增加,LXRα、GLUT4蛋白表达均降低;与模型组相比,四妙勇安汤组细胞内炎症因子IL-6、NLRP3蛋白表达降低,且呈剂量依赖性变化趋势,并明确四妙勇安汤干预最佳药物浓度为250μg/mL;给予中剂量四妙安勇汤干预后,脂肪细胞内PPARγ、LXRα、GLUT4蛋白表达增加,NF-κB蛋白表达减少,给予PPARγ抑制剂GW9662后四妙勇安汤调控关键因子和抗炎作用相应减弱。结论:四妙勇安汤通过PPARγ/NF-κB信号通路发挥抑制脂肪细胞炎症的作用。

基于PPARγ/NF-κB信号通路探讨防己黄芪消肿方干预代谢综合征表型骨关节炎作用机制

目的基于PPARγ/NF-κB信号通路探究防己黄芪消肿方对代谢综合征表型骨关节炎(MS-OA)大鼠的治疗作用及机制。方法SD大鼠随机分为假手术组、骨关节炎(OA)组、MS-OA组、西药组和中药高、低剂量组,采用改良Hulth法建立OA模型,改良Hulth法+高碳水高脂肪饮食建立MS-OA模型。中药高、低剂量组分别予防己黄芪消肿方药液15.12、7.56 g/kg灌胃,西药组给予洛索洛芬钠16.2 mg/kg灌胃,其余组予生理盐水灌胃,每日1次,连续6周。测量大鼠体质量,生化检测血脂及血糖,ELISA检测血清肿瘤坏死因子-α(TNF-α)、白细胞介素(IL)-1β、IL-10及瘦素含量,番红O-固绿及HE染色观察软骨组织形态,免疫组化染色检测软骨组织蛋白聚糖(Acan)、Ⅹ型胶原(ColⅩ)、基质金属蛋白酶13(MMP13)、TNF-α、IL-1β、过氧化物酶体增殖物激活受体γ(PPARγ)表达,Western blot检测软骨组织PPARγ、核因子(NF)-κBp65、p-NF-κBp65蛋白表达。结果与假手术组比较,MS-OA组大鼠体质量及血清总胆固醇(TC)、三酰甘油(TG)、低密度脂蛋白胆固醇(LDL-C)、TNF-α、IL-1β、瘦素含量均升高(P<0.01),高密度脂蛋白胆固醇(HDL-C)、IL-10含量降低(P<0.01),软骨组织退变明显,Mankin评分增加(P<0.01),软骨组织ColⅩ、MMP13、TNF-α、IL-1β、p-NF-κBp65蛋白表达升高(P<0.05,P<0.01),Acan、PPARγ蛋白表达降低(P<0.01);与MS-OA组比较,中药高剂量组大鼠血清TC、TG、LDL-C、TNF-α、瘦素含量降低(P<0.05,P<0.01),IL-10含量增加(P<0.05),软骨组织病理损伤改善,Mankin评分降低(P<0.01),软骨组织ColⅩ、MMP13、TNF-α、IL-1β、p-NF-κBp65蛋白表达降低(P<0.05,P<0.01),Acan、PPARγ蛋白表达升高(P<0.01,P<0.05)。结论防己黄芪消肿方可改善MS-OA大鼠脂代谢紊乱、改善关节内炎症环境、平衡软骨代谢、延缓软骨退变,其作用机制可能与调节PPARγ/NF-κB信号通路有关。

通脉消斑汤上调PPARα抑制NF-кB减轻缺氧复氧受损心肌细胞炎症反应

目的:探讨通脉消斑汤上调过氧化物酶体增生因子活化受体α(PPARα)抑制NF-кB保护缺氧复氧受损心肌细胞炎症反应。方法:建立大鼠H9C2细胞缺氧复氧模型,将细胞分为3组:对照组(Ⅰ组)、缺氧复氧组(Ⅱ组)、缺氧复氧加通脉消斑汤组(Ⅲ组)。CCK8检测不同浓度通脉消斑汤对大鼠H9C2活力影响;缺氧复氧以及加用通脉消斑汤对细胞活力的影响。免疫荧光观察PPAR缺氧复氧前后及加用通脉消斑汤后变化,NF-кB P65核转移以及磷酸化NF-κB P65(p-NF-κB P65)胞核内表达变化。Western Blot检测PPAR、p-NF-κB、NF-κB蛋白表达的变化。结果:与空白对照组比较,缺氧复氧组H9C2细胞活力降低,加用通脉消斑汤后,细胞活力升高(P﹤0.05)。免疫荧光显示缺氧复氧组细胞PPAR核内表达较空白对照组降低,加用通脉消斑汤组细胞,PPAR较前升高(P﹤0.05);缺氧复氧组细胞出现NF-κB P65核转移,通脉消斑汤组核转移减少;与空白对照组比较,磷酸化NF-κB在缺氧复氧组表达升高,加用通脉消斑汤组降低。Western Blot结果显示PPAR在缺氧复氧组表达较空白对照组降低,在通脉消斑汤升高;磷酸化NF-κB、细胞间粘附分子1(ICAM1)在缺氧复氧组表达升高,同样在通脉消斑汤组降低。结论:通脉消斑汤上调PPARα,抑制NF-κB,保护缺氧复氧受损心肌细胞。

Downregulation of MUC1 Inhibits Proliferation and Promotes Apoptosis by Inactivating NF-κB Signaling Pathway in Human Nasopharyngeal Carcinoma

Objective To investigate the effect of mucin 1(MUC1)on the proliferation and apoptosis of nasopharyngeal carcinoma(NPC)and its regulatory mechanism.Methods The 60 NPC and paired para-cancer normal tissues were collected from October 2020 to July 2021 in Quanzhou First Hospital.The expression of MUC1 was measured by real-time quantitative PCR(qPCR)in the patients with PNC.The 5-8F and HNE1 cells were transfected with siRNA control(si-control)or siRNA targeting MUC1(si-MUC1).Cell proliferation was analyzed by cell counting kit-8 and colony formation assay,and apoptosis was analyzed by flow cytometry analysis in the 5-8F and HNE1 cells.The qPCR and ELISA were executed to analyze the levels of TNF-αand IL-6.Western blot was performed to measure the expression of MUC1,NFкB and apoptosis-related proteins(Bax and Bcl-2).Results The expression of MUC1 was up-regulated in the NPC tissues,and NPC patients with the high MUC1 expression were inclined to EBV infection,growth and metastasis of NPC.Loss of MUC1 restrained malignant features,including the proliferation and apoptosis,downregulated the expression of p-IкB、p-P65 and Bcl-2 and upregulated the expression of Bax in the NPC cells.Conclusion Downregulation of MUC1 restrained biological characteristics of malignancy,including cell proliferation and apoptosis,by inactivating NF-κB signaling pathway in NPC.

Apatinib reduces liver cancer cell multidrug resistance by modulating NF-κB signaling pathway

Objectives:This investigation aimed to elucidate the inhibitory impact of apatinib on the multidrug resistance of liver cancer both in vivo and in vitro.Methods:To establish a Hep3B/5-Fu resistant cell line,5-Fu concentrations were gradually increased in the culture media.Hep3B/5-Fu cells drug resistance and its alleviation by apatinib were confirmed via flow cytometry and Cell Counting Kit 8(CCK8)test.Further,Nuclear factor kappa B(NF-κB)siRNA was transfected into Hep3B/5-Fu cells to assess alterations in the expression of multidrug resistance(MDR)-related genes and proteins.Nude mice were injected with Hep3B/5-Fu cells to establish subcutaneous xenograft tumors and then categorized into 8 treatment groups.The treatments included oxaliplatin,5-Fu,and apatinib.In the tumor tissues,the expression of MDRrelated genes was elucidated via qRT-PCR,immunohistochemistry,and Western blot analyses.Results:The apatinibtreated mice indicated slower tumor growth with smaller size compared to the control group.Both the in vivo and in vitro investigations revealed that the apatinib-treated groups had reduced expression of MDR genes GST-pi,LRP,MDR1,and p-p65.Conclusions:Apatinib effectively suppresses MDR in human hepatic cancer cells by modulating the expression of genes related to MDR,potentially by suppressing the NF-κB signaling pathway.

Kuicolong-yu enema decoction retains traditional Chinese medicine enema attenuates inflammatory response ulcerative colitis through TLR4/NF-κB signaling pathway

BACKGROUND Ulcer colitis(UC)is a chronic,nonspecific,and noninfectious inflammatory bowel disease.Recently,Toll-like receptors(TLRs)have been found to be closely associated with clinical inflammatory diseases.Achieving complete remission in patients with intermittent periods of activity followed by dormancy is challenging.Moreover,no study has explored the mechanism by which Kuicolong-yu enema decoction retains traditional Chinese medicine enemas to attenuate the inflammatory response in UC.AIM To explore the mechanism by which Kuicolong-yu enema decoction retains traditional Chinese medicine enemas to attenuate the inflammatory response in UC.METHODS This prospective clinical study included patients who met the exclusion criteria in 2020 and 2021.The patients with UC were divided into two groups(control and experimental).The peripheral blood of the experimental and control groups were collected under aseptic conditions.The expression of TLR4 protein,NF-κB,IL-6,and IL-17 was detected in the peripheral blood of patients in the experimental group and control group before and 1 month after taking the drug.Linear co rrelation analysis was used to analyze the relationship between the expression level of TLR4 protein and the expression levels of downstream signal NF-κB and inflammatory factors IL-6 and IL-17,and P<0.05 was considered statistically significant.RESULTS There were no significant differences in the patient characteristics between the control and experimental groups.The results showed that the expression levels of TLR4 and NF-κB in the experimental group were significantly lower than those in the control group(P<0.05).The levels of IL-6 and IL-17 in the experimental group were significantly lower than those in the control group(P<0.05).The TLR4 protein expression in the experimental group was positively correlated with the expression level of downstream signal NF-κB and was positively correlated with the levels of downstream inflammatory cytokines IL-6 and IL-17(r=0.823,P<0.05).CONCLUSION Kuicolong-yu enema decoction retains traditional Chinese medicine enema attenuates the inflammatory response of UC through the TLR4/NF-κB signaling pathway.

Aszonapyrone A Isolated from Neosartorya spinosa IFM 47025 Inhibits the NF-κB Signaling Pathway Activated by Expression of the Ependymoma-Causing Fusion Protein ZFTA-RELA

Ependymoma is a rare and chemotherapy-resistant brain tumor, which has resulted in a delay in the development of drugs to treat it. A subclass of supratentorial ependymomas (ST-EPN), designated ST-EPN-zinc finger-translocation-associated (ZFTA, ST-EPN-ZFTA), exhibits the expression of a fusion protein comprising ZFTA and v-rel reticuloendotheliosis viral oncogene homolog A (RELA), an effector transcription factor of the nuclear factor-kappa B (NF-κB) pathway (ZFTA-RELA). The expression of ZFTA-RELA results in the hyperactivation of the oncogenic NF-κB signaling pathway, which ultimately leads to the development of ST-EPN-ZFTA. To identify inhibitors of the NF-κB signaling pathway activated by the expression of ZFTA-RELA, we used a doxycycline-inducible ZFTA-RELA-expressing NF-κB reporter cell line and found that extracts of the fungus Neosartorya spinosa IFM 47025 exhibited NF-κB inhibitory activity. We identified eight compounds [aszonapyrone A (2), sartorypyrone A (3), epiheveadride (4), acetylaszonalenin (5), (R)-benzodiazepinedione (6), aszonalenin (7), sartorypyrone E (8) and (Z, Z)-N,N’-(1,2-bis[(4-methoxyphenyl)methylene]-1,2-ethanediyl)bis-formamide (9)] from N. spinosa IFM 47025 culture extract using a variety of chromatographic techniques. The structures of these compounds were identified through the analysis of various instrumental data (1D, 2D-NMR, MS, and optical rotation). The NF-κB responsive reporter assay indicated that compounds 2, 3, 5, 7, and 9 exhibited inhibitory activity. We further evaluated the inhibitory activity of these compounds against the expression of endogenous NF-κB responsive genes (CCND1, L1CAM, ICAM1, and TNF) and found that compound 2 showed significant inhibitory activity. Further studies are required to elucidate the mechanism of action of compound 2, which may serve as a lead compound for the development of a novel therapy for ST-EPN-ZFTA.

Parthenolide enhances the metronomic chemotherapy effect of cyclophosphamide in lung cancer by inhibiting the NF-kB signaling pathway

BACKGROUND Parthenolide(PTL),a sesquiterpene lactone derived from the medicinal herb Chrysanthemum parthenium,exhibits various biological effects by targeting NF-kB,STAT3,and other pathways.It has emerged as a promising adjunct therapy for multiple malignancies.AIM To evaluate the in vitro and in vivo effect of PTL on cyclophosphamide(CTX)metronomic chemotherapy.METHODS The cytotoxicity of PTL and CTX on Lewis lung cancer cells(LLC cells)was assessed by measuring cell activity and apoptosis.The anti-tumor efficiency was evaluated using a tumor xenograft mice model,and the survival of mice and tumor volume were monitored.Additionally,the collected tumor tissues were analyzed for tumor microenvironment indicators and inflammatory factors.RESULTS In vitro,PTL demonstrated a synergistic effect with CTX in inhibiting the growth of LLC cells and promoting apoptosis.In vivo,metronomic chemotherapy com-bined with PTL and CTX improved the survival rate of tumor-bearing mice and reduced tumor growth rate.Furthermore,metronomic chemotherapy combined with PTL and CTX reduced NF-κB activation and improved the tumor immune microenvironment by decreasing tumor angiogenesis,reducing Transforming growth factorβ,andα-SMA positive cells.CONCLUSION PTL is an efficient compound that enhances the metronomic chemotherapy effects of CTX both in vitro and in vivo,suggesting its potential as a supplementary therapeutic strategy in metronomic chemotherapy to improve the chemotherapy effects.

大黄素对高尿酸环境下人肾小管上皮细胞PPARγ/NF-κB通路的影响

目的观察大黄素(emodin,EM)对高尿酸(uric acid,UA)环境下人肾小管上皮细胞氧化物酶体增殖物激活受体γ(PPARγ)/核因子-κB(NF-κB)通路的影响,探讨其抑制尿酸性肾病(UAN)的分子机制。方法体外培养人肾小管上皮细胞株,分为正常对照组、高UA组、EM低(EML,5μmol·L^(-1))、中(EMM,10μmol·L^(-1))、高(EMH,20μmol·L^(-1))剂量组,设立24、36、48 h 3个时间进行观察。QPCR检测PPARγ、NF-κB mRNA表达水平;Western blot检测PPARγ、IκBα蛋白表达水平;ELISA检测肿瘤坏死因子-α(TNF-α)蛋白表达水平。结果与正常对照组比较,高UA组24 h时间点PPARγmRNA及蛋白、IκBα蛋白下调,TNF-α蛋白上调;36 h时间点PPARγ及IκBα蛋白下调,NF-κB mRNA及TNF-α蛋白上调;48 h时间点IκBα蛋白下调,NF-κB mRNA及TNF-α蛋白上调。与高UA组比较,24 h时间点EML组PPARγmRNA及IκBα蛋白上调,TNF-α蛋白下调,EMM组PPARγmRNA上调,TNF-α蛋白下调,EMH组TNF-α蛋白下调;36 h时间点EML组IκBα蛋白上调,TNF-α蛋白下调,EMM组NF-κB mRNA及TNF-α蛋白下调,IκBα蛋白上调,EMH组NF-κB mRNA及TNF-α蛋白下调,PPARγ蛋白上调;48 h时间点EML组NF-κB mRNA及TNF-α蛋白下调,IκBα蛋白上调,EMM组PPARγmRNA及IκBα蛋白上调,TNF-α蛋白下调,EMH组PPARγmRNA上调,NF-κB mRNA及TNF-α蛋白下调。结论EM可通过调控PPARγ/NF-κB信号通路,抑制下游炎性因子表达,从而改善UAN肾脏炎性损伤。

右美托咪定通过调控PPARγ/NF-κB通路改善糖尿病脑损伤的机制探讨

目的:研究右美托咪定(DEX)对2型糖尿病大鼠脑损伤的影响及其机制。方法:选择48只SPF级雄性SD大鼠随机分为3组:正常组(control组)、糖尿病组(DM组)、DEX组,每组各16只;除control组外,其余组28只SD大鼠以高脂饲料喂养4周后以40 mg/kg单次腹腔注射链脲佐菌素(STZ)制备2型糖尿病模型;4周后检测随机血糖≥16.7 mmol/L为造模成功;DEX组在DM组造模成功后腹腔注射40μg/kg DEX,每日1次,连续注射2周,DM组给予等量的0.9%生理盐水腹腔注射,观察并记录各组大鼠空腹血糖、体重及生存情况;造模结束后处死大鼠,收集大脑皮层和海马组织,通过苏木精—伊红(HE)法检测大鼠脑组织病理学损伤变化,采用蛋白质免疫印迹(western blotting)和免疫组化法检测p-PPARγ、NF-κB的表达。结果:与control组相比,DM组大鼠的血糖升高、体重降低,可见一定程度的脑组织结构破坏及炎性细胞浸润。除此之外,p-PPARγ蛋白的表达量下降(P<0.05),NF-κB表达升高(P<0.05)。而在给予DEX处理后逆转了以上的变化,与DM组相比,DEX组的血糖下降、脑组织损伤较前改善,p-PPARγ蛋白的表达增加,NF-κB蛋白表达减少(均P<0.05)。结论:DEX可能通过调控PPARγ/NF-κB信号通路减轻2型糖尿病大鼠脑损伤。

Quercetin regulates depression-like behavior in CUMS rat models via TLR4/NF-κB signaling

Background:Depression is becoming increasingly prevalent around the world,imposing a substantial burden on individuals,families,as well as society.Quercetin is known to be highly effective in treating depression.However,additional research is needed to dissect the mechanisms of its anti-depressive effects.Methods:For this study,Sprague-Dawley(SD)rats were randomized into the control,model,quercetin,or fluoxetine group.The latter three groups were exposed to chronic unpredictable mild stress(CUMS)for 42 d.The first two groups received saline solution daily via oral gavage.Meanwhile,the quercetin group was orally administered a quercetin suspension(52.08 mg/kg)every day,while the fluoxetine group was orally administered a fluoxetine solution(2.08 mg/kg).Here,fluoxetine served as the positive control drug to compare the therapeutic effects of quercetin.The experimental period was 6 weeks.Depressive behaviors in rats were assessed through various physiological and behavioral measures.Additionally,pathological changes in hippocampal tissues were examined using Nissl staining.Serum cytokines were detected using an enzymelinked immunosorbent assay(ELISA),and immunohistochemistry was employed to quantify the levels and integral optical density(IOD)values of ionized calcium binding adaptor molecule-1(Iba-1)expression in the brain.Real-time fluorescence quantitative PCR(RT-qPCR)was utilized to evaluate the mRNA levels of inflammatory indicators as well as toll-like receptor 4(TLR4),and nuclear factor-κappa B P65(NF-κB P65)in hippocampus.Western blot(WB)technique was employed to observe the protein levels of TLR4,NF-κB P65,and phospho-NF-κB P65(p-NF-κB P65).Results:After 42 d of exposure to CUMS,rats exhibited a slow increase in body weight,a reduction in food intake,an abnormal preference for sugar water,and aberrant open-field behaviors.Pathological analysis revealed the disintegration,rupture,interruption,and disorganization of hippocampal neuronal cells after CUMS exposure,along with a decrease in Nissl bodies in the CA1 region.This was accompanied by the elevated expression of interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),and interleukin-6(IL-6)in the serum and the upregulation of IL-1β,IL-6,and TNF-αmRNA expression in the hippocampus.Increases in Iba-1-positive cells and the IOD values of Iba-1 were detected in hippocampal microglia.Furthermore,TLR4 and NF-κB P65 mRNA and protein levels were upregulated in hippocampal tissues.Quercetin,an antidepressant,could alleviate depression-like symptoms in rats and downregulate inflammatory factors associated with the TLR4/NF-κB signaling pathway in hippocampal microglia,and its therapeutic effect was comparable to fluoxetine.Conclusion:In rat models of CUMS,quercetin may act as an antidepressant by inhibiting inflammation in hippocampal microglia via TLR4/NF-κB signaling pathway.These results offer experimental and theoretical support for applying quercetin in the clinical management of depression.

Huangqin decoction alleviates lipid metabolism disorders and insulin resistance in nonalcoholic fatty liver disease by triggering Sirt1/NF-κB pathway

BACKGROUND Nonalcoholic fatty liver disease(NAFLD)is a clinicopathological entity characterized by intrahepatic ectopic steatosis.As a consequence of increased consumption of high-calorie diet and adoption of a sedentary lifestyle,the incidence of NAFLD has surpassed that of viral hepatitis,making it the most common cause of chronic liver disease globally.Huangqin decoction(HQD),a Chinese medicinal formulation that has been used clinically for thousands of years,has beneficial outcomes in patients with liver diseases,including NAFLD.However,the role and mechanism of action of HQD in lipid metabolism disorders and insulin resistance in NAFLD remain poorly understood.AIM To evaluate the ameliorative effects of HQD in NAFLD,with a focus on lipid metabolism and insulin resistance,and to elucidate the underlying mechanism of action.METHODS High-fat diet-induced NAFLD rats and palmitic acid(PA)-stimulated HepG2 cells were used to investigate the effects of HQD and identify its potential mechanism of action.Phytochemicals in HQD were analyzed by highperformance liquid chromatography(HPLC)to identify the key components.RESULTS Ten primary chemical components of HQD were identified by HPLC analysis.In vivo,HQD effectively prevented rats from gaining body and liver weight,improved the liver index,ameliorated hepatic histological aberrations,decreased transaminase and lipid profile disorders,and reduced the levels of pro-inflammatory factors and insulin resistance.In vitro studies revealed that HQD effectively alleviated PA-induced lipid accumulation,inflammation,and insulin resistance in HepG2 cells.In-depth investigation revealed that HQD triggers Sirt1/NF-κB pathwaymodulated lipogenesis and inflammation,contributing to its beneficial actions,which was further corroborated by the addition of the Sirt1 antagonist EX-527 that compromised the favorable effects of HQD.CONCLUSION In summary,our study confirmed that HQD mitigates lipid metabolism disorders and insulin resistance in NAFLD by triggering the Sirt1/NF-κB pathway.

Calcitriol attenuates liver fibrosis through hepatitis C virus nonstructural protein 3-transactivated protein 1-mediated TGF β1/Smad3 and NF-κB signaling pathways

BACKGROUND Hepatic fibrosis is a serious condition,and the development of hepatic fibrosis can lead to a series of complications.However,the pathogenesis of hepatic fibrosis remains unclear,and effective therapy options are still lacking.Our group identified hepatitis C virus nonstructural protein 3-transactivated protein 1(NS3TP1) by suppressive subtractive hybridization and bioinformatics analysis,but its role in diseases including hepatic fibrosis remains undefined.Therefore,additional studies on the function of NS3TP1 in hepatic fibrosis are urgently needed to provide new targets for treatment.AIM To elucidate the mechanism of NS3TP1 in hepatic fibrosis and the regulatory effects of calcitriol on NS3TP1.METHODS Twenty-four male C57BL/6 mice were randomized and separated into three groups,comprising the normal,fibrosis,and calcitriol treatment groups,and liver fibrosis was modeled by carbon tetrachloride(CCl4).To evaluate the level of hepatic fibrosis in every group,serological and pathological examinations of the liver were conducted.TGF-β1 was administered to boost the in vitro cultivation of LX-2 cells.NS3TP1,α-smooth muscle actin(α-SMA),collagen I,and collagen Ⅲ in every group were examined using a Western blot and real-time quantitative polymerase chain reaction.The activity of the transforming growth factor beta 1(TGFβ1)/Smad3 and NF-κB signaling pathways in each group of cells transfected with pcDNA-NS3TP1 or siRNA-NS3TP1 was detected.The statistical analysis of the data was performed using the Student’s t test.RESULTS NS3TP1 promoted the activation,proliferation,and differentiation of hepatic stellate cells(HSCs)and enhanced hepatic fibrosis via the TGFβ1/Smad3 and NF-κB signaling pathways,as evidenced by the presence of α-SMA,collagen I,collagen Ⅲ,p-smad3,and p-p65 in LX-2 cells,which were upregulated after NS3TP1 overexpression and downregulated after NS3TP1 interference.The proliferation of HSCs was lowered after NS3TP1 interference and elevated after NS3TP1 overexpression,as shown by the luciferase assay.NS3TP1 inhibited the apoptosis of HSCs.Moreover,both Smad3 and p65 could bind to NS3TP1,and p65 increased the promoter activity of NS3TP1,while NS3TP1 increased the promoter activity of TGFβ1 receptor I,as indicated by coimmunoprecipitation and luciferase assay results.Both in vivo and in vitro,treatment with calcitriol dramatically reduced the expression of NS3TP1.Calcitriol therapy-controlled HSCs activation,proliferation,and differentiation and substantially suppressed CCl4-induced hepatic fibrosis in mice.Furthermore,calcitriol modulated the activities of the above signaling pathways via downregulation of NS3TP1.CONCLUSION Our results suggest that calcitriol may be employed as an adjuvant therapy for hepatic fibrosis and that NS3TP1 is a unique,prospective therapeutic target in hepatic fibrosis.

MiR-27a靶向调节PPARγ/SIRT1/NF-κB信号通路对溃疡性结肠炎幼年小鼠肠道损伤的影响

目的探究miR-27a靶向调节PPARγ/SIRT1/NF-κB信号通路对溃疡性结肠炎(UC)幼年小鼠肠道损伤的影响。方法2,4,6-三硝基苯磺酸(TNBS)诱导构建UC模型,miR-27a antagomir用于在小鼠中抑制miR-27a表达。组织学和髓过氧化物酶(MPO)活性评估结肠组织损伤情况;TUNEL检测结肠组织中细胞凋亡情况;ELISA检测血清中肿瘤坏死因子α(TNF-α)、白介素1β(IL-1β)、细胞间黏附分子1(ICAM-1)和单核细胞趋化蛋白1(MCP-1)含量;qRT-PCR和Western blot检测结肠组织中miR-27a和PPARγ、SIRT1、乙酰化NF-κB p65表达水平;双荧光素酶检测miR-27a和PPARγ靶向关系。结果在TNBS诱导的UC小鼠模型中,结肠炎评分及MPO活性增高(P<0.05);细胞凋亡率增高;TNF-α、IL-1β、ICAM-1和MCP-1含量增多(P<0.05);miR-27a表达水平和乙酰化NF-κB p65蛋白水平增高(P<0.05);PPARγ和SIRT1蛋白水平降低(P<0.05)。抑制miR-27a可以减轻TNBS诱导的结肠损伤(P<0.05);降低细胞凋亡率(P<0.05);抑制炎性因子的产生(P<0.05);促进PPARγ和SIRT1表达(P<0.05);抑制乙酰化NF-κB p65蛋白表达(P<0.05)。PPARγ是miR-27a的直接靶标(P<0.05)。结论抑制miR-27a可通过靶向调节PPARγ/SIRT1/NF-κB信号通路对UC幼年小鼠肠道损伤发挥保护作用。

Acupuncture at Back-Shu point improves insomnia by reducing inflammation and inhibiting the ERK/NF-κB signaling pathway

BACKGROUND Insomnia is a disease where individuals cannot maintain a steady and stable sleep state or fail to fall asleep.Western medicine mainly uses sedatives and hypnotic drugs to treat insomnia,and long-term use is prone to drug resistance and other adverse reactions.Acupuncture has a good curative effect and unique advantages in the treatment of insomnia.AIM To explore the molecular mechanism of acupuncture at Back-Shu point for the treatment of insomnia.METHODS We first prepared a rat model of insomnia,and then carried out acupuncture for 7 consecutive days.After treatment,the sleep time and general behavior of the rats were determined.The Morris water maze test was used to assess the learning ability and spatial memory ability of the rats.The expression levels of inflammatory cytokines in serum and the hippocampus were detected by ELISA.qRTPCR was used to detect the mRNA expression changes in the ERK/NF-κB signaling pathway.Western blot and immunohistochemistry were carried out to evaluate the protein expression levels of RAF-1,MEK-2,ERK1/2 and NF-κB.RESULTS Acupuncture can prolong sleep duration,and improve mental state,activity,diet volume,learning ability and spatial memory.In addition,acupuncture increased the release of 1L-1β,1L-6 and TNF-αin serum and the hippocampus and inhibited the mRNA and protein expression of the ERK/NF-κB signaling pathway.CONCLUSION These findings suggest that acupuncture at Back-Shu point can inhibit the ERK/NF-κB signaling pathway and treat insomnia by increasing the release of inflammatory cytokines in the hippocampus.

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.

姜黄素通过调控PPARγ/NF-κB信号通路减轻CSE诱导的人气道上皮细胞氧化应激反应

目的探讨姜黄素对香烟烟雾提取物(CSE)诱导的人气道上皮16HBE细胞氧化应激和炎症反应的抑制作用及相关的分子机制。方法使用shRNA-PPARγ(shPPARγ)转染人气道上皮16HBE细胞下调PPARγ表达。将16HBE细胞分为5组即对照组、姜黄素组、CSE组、CSE+姜黄素组和CSE+姜黄素+shRNA-PPARγ组。在0 h和24 h时使用MTT法对细胞活性进行检测;干预24 h后使用q PCR对细胞TNF-α、iNOS和PPARγm RNA表达进行检测,采用western blot检测16HBE细胞中iNOS、PPARγ蛋白表达以及NF-κB p65磷酸化水平。结果 16HBE细胞在干预24 h后各组之间细胞活性未有明显统计学差异(P>0.05)。与对照组相比较,CSE干预24 h后PPARγ表达水平明显降低,TNF-α、iNOS表达及NF-κB p65磷酸化水平明显升高,差异均有统计学意义(P<0.05)。但CSE组较CSE+姜黄素组和CSE+姜黄素+shPPARγ组PPARγ表达水平下降以及TNF-α、iNOS表达和NF-κB p65磷酸化水平升高更显著,差异均有统计学意义(P<0.05);且CSE+姜黄素+shPPARγ组较CSE+姜黄素组PPARγ表达水平下降以及TNF-α、iNOS表达和NF-κB p65磷酸化水平升高更明显,差异均有统计学意义(P<0.05)。结论姜黄素可以通过抑制PPARγ/NF-κB信号通路减轻CSE诱导的人气道上皮16HBE细胞的炎症反应及氧化应激。为姜黄素应用于慢性阻塞性肺疾病等疾病的临床治疗奠定了理论基础。

阿托伐他汀钙调节PPARγ和NF-κB活性参与巨噬细胞泡沫化的形成

目的探讨阿托伐他汀钙是否通过调节PPARγ和NF-κB活性来抑制Ox-LDL诱导的THP-1巨噬细胞泡沫化形成。方法THP-1巨噬细胞与10、20、40μmol/L阿托伐他汀钙分别预孵育2 h,PBS洗2次,然后与60μg/ml ox-LDL共孵育48 h后,通过酶荧光法检测细胞胆固醇酯与总胆固醇比值,油红O染色观察细胞胞浆内脂质蓄积情况,Western blot分析清道夫受体CD36、SRA的表达,p-iκB和PPARγ的表达,以及ATP结合盒转运蛋白A1(ABCA1)的表达变化。用荧光探针DIL标记ox-LDL的方法,观察阿托伐他汀钙对THP-1巨噬细胞结合ox-LDL能力的影响。结果阿托伐他汀钙浓度依赖性降低THP-1巨噬细胞脂质蓄积和抑制清道夫受体CD36和SRA的表达,同时上调ABCA1表达,并减弱PPARγ和p-iκB活性(P<0.05)。荧光探针DIL标记ox-LDL的方法显示阿托伐他汀钙浓度依赖性降低THP-1巨噬细胞结合ox-LDL能力(P<0.05)。结论阿托伐他汀钙抗动脉硬化作用可能部分通过降低PPARγ和p-iκB活性来抑制脂质蓄积。

PPARγ在脂多糖刺激牙周膜细胞中调控NF-κB信号通路的作用研究

目的:探讨PPARγ在牙周炎中调控作用机制。方法:组织块法培养健康人牙周膜细胞(human periodontal ligament cells,hPDLCs)并鉴定。细胞处理分为以下4组,A组:对照组;B组:脂多糖(lipopolysaccharide,LPS)刺激组;C组:罗格列酮对照组,即二甲基亚砜(dimethylsulfoxide,DMSO)处理组;D组:罗格列酮处理组。免疫印迹法检测过氧化物酶体增殖物激活受体γ(peroxisome proliferator activated receptorγ,PPARγ)和核因子κB(nuclear factorκB,NF-κB)p65胞核、胞浆及总蛋白含量,细胞免疫荧光检测NF-κB p65表达部位。实时定量PCR和酶联免疫法检测白细胞介素1β(interleukin-1β,IL-1β)和肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)RNA和蛋白表达。结果:LPS刺激下,胞核PPARγ表达降低NF-κB p65表达升高,相应的IL-1β和TNF-αRNA及蛋白表达量均升高。同时加入罗格列酮后,胞核PPARγ表达升高NF-κB p65表达降低,且IL-1β和TNF-αRNA及蛋白表达量均降低。差异均有统计学意义(P<0.01)。结论:PPARγ通过下调NF-κB信号通路抑制脂多糖刺激下牙周膜细胞炎症因子RNA表达和蛋白分泌,进而调节牙周炎症反应。

PPARγ和NF-κB在肺纤维化中的表达与意义

目的:观察过氧化物酶体增殖物激活受体γ(PPARγ)和核因子-κB(NF-κB)在肺纤维化患者肺组织中的表达情况,探讨其在肺纤维化发病中的作用。方法:采用免疫组织化学方法检测16例肺纤维化患者与10例正常对照肺组织标本中PPARγ、NF-κB的表达情况。结果:肺纤维化组PPARγ的阳性表达记数为0.35±0.08,低于正常对照组的0.42±0.04(P<0.05)。肺纤维化组NF-κB的阳性表达记数为0.51±0.11,高于正常对照组的0.38±0.04(P<0.05)。肺纤维化组的PPARγ和NF-κB的表达呈负相关(P<0.05)。结论:肺组织中PPARγ的表达减弱、NF-κB的表达增强在肺纤维化的发病过程中起作用,为临床治疗肺纤维化提供了新思路。