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.

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.

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.

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。

Nucleolin Mediates LPS-induced Expression of Inflammatory Mediators and Activation of Signaling Pathways

Summary:In this study,we investigated the effects of nucleolin on lipopolysaccharide(LPS)-induced activation of MAPK and NF-KappaB(NF-kB)signaling pathways and secretion of TNF-a,IL-1βand HMGB1 in THP-1 monocytes.Immunofluorescence assay and Western blotting were used to identify the nucleolin expression in cell membrane,cytoplasm and nucleus of THP-1 monocytes.Inactivation of nucleolin was induced by neutralizing antibody against nucleolin.THP-1 monocytes were pretreated with anti-nucleolin antibody for 1 h prior to LPS challenge.The irrelevant IgG group was used as control.Secretion of inflammatory mediators(TNF-a,IL-1β and HMGB1)and activation of MAPK and NF-kB/I-kB signaling pathways were examined to assess the effects of nucleolin on LPS-mediated inflammatory response.Nucleolin existed in cell membrane,cytoplasm and nucleus of THP-1 monocytes.Pretreatment of anti-nucleolin antibody significantly inhibited the LPS-induced secretion of TNF-a,IL-1β and HMGB1.P38,JNK,ERK and NF-κB subunit p65 inhibitors could significantly inhibit the secretion of IL-1β,TNF-a and HMGB1 induced by LPS.Moreover,the phosphorylation of p38,JNK,ERK and p65(or nuclear translocation of p65)was significantly increased after LPS challenge.In contrast,pretreatment of anti-nucleolin antibody could significantly inhibit the LPS-induced phosphorylation of p38,JNK,ERK and p65(or nuclear translocation of p65).However,the irrelevant IgG,as a negative control,had no effect on LPS-induced secretion of TNF-a and IL-Iβ and phosphorylation of p38,JNK,ERK and p65(or nuclear translocation of p65).We demonstrated that nucleolin mediated the LPS-induced activation of MAPK and NF-κB signaling pathways,and regulated the secretion of inflammatory mediators(TNF-a,IL-1β and HMGB1).

Novel phenanthrene/bibenzyl trimers from the tubers of Bletilla striata attenuate neuroinflammation via inhibition of NF-κB signaling pathway

Five novel(9,10-dihydro)phenanthrene and bibenzyl trimers,as well as two previously identified biphenanthrenes and bibenzyls,were isolated from the tubers of Bletilla striata.Their structures were elucidated through comprehensive analyses of NMR and HRESIMS spectroscopic data.The absolute configurations of these compounds were determined by calculating rotational energy barriers and comparison of experimental and calculated ECD curves.Compounds 5b and 6 exhibited inhibitory effects on LPS-induced NO production in BV-2 cells,with IC_(50) values of 12.59±0.40 and 15.59±0.83μmol·L^(-1),respectively.A mechanistic study suggested that these compounds may attenuate neuroinflammation by reducing the activation of the AKT/IκB/NF-κB signaling pathway.Additionally,compounds 3a,6,and 7 demonstrated significant PTP1B inhibitory activities,with IC_(50) values of 1.52±0.34,1.39±0.11,and 1.78±0.01μmol·L^(-1),respectively.Further investigation revealed that compound 3a might inhibit LPS-induced PTP1B overexpression and NF-κB activation,thereby mitigating the neuroinflammatory response in BV-2 cells.

Taurolidine improved protection against highly pathogenetic avian influenza H5N1 virus lethal-infection in mouse model by regulating the NF-κB signaling pathway

Taurolidine(TRD),a derivative of taurine,has anti-bacterial and anti-tumor effects by chemically reacting with cell-walls,endotoxins and exotoxins to inhibit the adhesion of microorganisms.However,its application in antiviral therapy is seldom reported.Here,we reported that TRD significantly inhibited the replication of influenza virus H5N1 in MDCK cells with the half-maximal inhibitory concentration(EC_(50))of 34.45μg/mL.Furthermore,the drug inhibited the amplification of the cytokine storm effect and improved the survival rate of mice lethal challenged with H5N1(protection rate was 86%).Moreover,TRD attenuated virus-induced lung damage and reduced virus titers in mice lungs.Administration of TRD reduced the number of neutrophils and increased the number of lymphocytes in the blood of H5N1 virus-infected mice.Importantly,the drug regulated the NF-κB signaling pathway by inhibiting the separation of NF-κB and IκBa,thereby reducing the expression of inflammatory factors.In conclusion,our findings suggested that TRD could act as a potential anti-influenza drug candidate in further clinical studies.

Long non-coding RNA Tug 1 regulates inflammation in microglia and in status epilepticus rats through the NF-κB signaling pathway

Background:Inflammation plays an important role in the pathogenesis of status epilepticus(SE).The long non-cod-ing RNA(lncRNA)taurine up-regulated gene1(Tug1)plays a well-defned role in inflammatory diseases.However,the molecular mechanism of Tug1 in SE progression remains unknown.In present study,we investigated whether Tug1 is involved in microglial inflammation in SE rats.Methods:The SE rat model was established via intraperitoneal injection of lithium chloride pilocarpine.RNA-binding protein immunoprecipitation(RIP)and RIP sequencing were carried out in rat microglia(RM).Tug1 cloned into the adenovirus was overexpressed in the microglia.Knockdown of Tug1 was performed via siRNA transfection.The level of Tug1 and inflammatory factors IL-1βand TNF-αwas examined by real-time polymerase chain reaction(RT-PCR)and western blotting.Protein levels of p65,P-p65,p-IKBa and IkBa were assessed by western blotting.Results:The RIP-seq result showed 14 lncRNAs that bound to the NF-κB p65 protein in RM.The lncRNA Tug1 directly interacted with p65.The level of declined Tug1 was decreased in the hippocampus of SE rats.Overexpression ofTug1 reduced the LPS-induced inflammation and M1/M2 polarization of microglia,while knockdown of Tug1 aggravated the inflammatory response in microglia.Accordingly,the protein levels of p-p65/p65 and p-IkBa/IkBa were reduced in the Tug1-overexpression microglia and elevated in the Tug1-knockdown microglia.Conclusions:These findings indicate that Tug1 modulates the inflammation in microglia through the NF-κB signal pathway,and the Tug1/P65 axis are like to play a signifcant role in the inflammatory processes,providing a valid target for the therapy of SE.

Sphingosine kinase 1 promotes growth of glioblastoma by increasing inflammation mediated by the NF-κB/IL-6/STAT3 and JNK/PTX3 pathways

Glioblastoma(GBM)is the most challenging malignant tumor of the central nervous system because of its high morbidity,mortality,and recurrence rate.Currently,mechanisms of GBM are still unclear and there is no effective drug for GBM in the clinic.Therefore,it is urgent to identify new drug targets and corresponding drugs for GBM.In this study,in silico analyses and experimental data show that sphingosine kinase 1(SPHK1)is up-regulated in GBM patients,and is strongly correlated with poor prognosis and reduced overall survival.Overexpression of SPHK1 promoted the proliferation,invasion,metastasis,and clonogenicity of GBM cells,while silencing SPHK1 had the opposite effect.SPHK1 promoted inflammation through the NF-κB/IL-6/STAT3 signaling pathway and led to the phosphorylation of JNK,activating the JNK-JUN and JNK-ATF3 pathways and promoting inflammation and proliferation of GBM cells by transcriptional activation of PTX3.SPHK1 interacted with PTX3 and formed a positive feedback loop to reciprocally increase expression,promote inflammation and GBM growth.Inhibition of SPHK1 by the inhibitor,PF543,also decreased tumorigenesis in the U87-MG and U251-MG SPHK1 orthotopic mouse models.In summary,we have characterized the role and molecular mechanisms by which SPHK1 promotes GBM,which may provide opportunities for SPHK1-targeted therapy.

MicroRNA-23a reduces lipopolysaccharide-induced cellular apoptosis and inflammatory cytokine production through Rho-associated kinase 1/sirtuin-1/nuclear factor-kappa B crosstalk

Background:MicroRNAs are closely associated with the progression and outcomes of multiple human diseases,including sepsis.In this study,we examined the role of miR-23a in septic injury.Methods Lipopolysaccharide(LPS)was used to induce sepsis in a rat model and H9C2 and HK-2 cells.miR-23a expression was evaluated in rat myocardial and kidney tissues,as well as H9C2 and HK-2 cells.A miR-23a mimic was introduced into cells to identify the role of miR-23a in cell viability,apoptosis,and the secretion of inflammatory cytokines.Furthermore,the effect of Rho-associated kinase 1(ROCK1),a miR-23a target,on cell damage was evaluated,and molecules involved in the underlying mechanism were identified.Results:In the rat model,miR-23a was poorly expressed in myocardial(sham vs.sepsis 1.00±0.06 vs.0.27±0.03,P<0.01)and kidney tissues(sham vs.sepsis 0.27±0.03 vs.1.00±0.06,P<0.01).Artificial overexpression of miR-23a resulted in increased proliferative activity(DNA replication rate:Control vs.LPS vs.LPS+Mock vs.LPS+miR-23a:H9C2 cells:34.13±3.12 vs.12.94±1.21 vs.13.31±1.43 vs.22.94±2.26,P<0.05;HK-2 cells:15.17±1.43 vs.34.52±3.46 vs.35.19±3.12 vs.19.87±1.52,P<0.05),decreased cell apoptosis(Control vs.LPS vs.LPS+Mock vs.LPS+miR-23a:H9C2 cells:11.39±1.04 vs.32.57±2.29 vs.33.08±3.12 vs.21.63±2.35,P<0.05;HK-2 cells:15.17±1.43 vs.34.52±3.46 vs.35.19±3.12 vs.19.87±1.52,P<0.05),and decreased production of inflammatory cytokines,including interleukin-6(Control vs.LPS vs.LPS+Mock vs.LPS+miR-23a:H9C2 cells:59.61±5.14 vs.113.54±12.30 vs.116.51±10.69 vs.87.69±2.97 ng/mL;P<0.05,F=12.67,HK-2 cells:68.12±6.44 vs.139.65±16.62 vs.143.51±13.64 vs.100.82±9.74 ng/mL,P<0.05,F=9.83)and tumor necrosis factor-α(Control vs.LPS vs.LPS+Mock vs.LPS+miR-23a:H9C2 cells:103.20±10.31 vs.169.67±18.84 vs.173.61±15.91 vs.133.36±12.32 ng/mL,P<0.05,F=12.67,HK-2 cells:132.51±13.37 vs.187.47±16.74 vs.143.51±13.64 vs.155.79±15.31 ng/mL,P<0.05,F=9.83)in cells.However,ROCK1 was identified as a miR-23a target,and further up-regulation of ROCK1 mitigated the protective function of miR-23a in LPS-treated H9C2 and HK-2 cells.Moreover,ROCK1 suppressed sirtuin-1(SIRT1)expression to promote the phosphorylation of nuclear factor-kappa B(NF-κB)p65,indicating the possible involvement of this signaling pathway in miR-23a-mediated events.Conclusion:Our results indicate that miR-23a could suppress LPS-induced cell damage and inflammatory cytokine secretion by binding to ROCK1,mediated through the potential participation of the SIRT1/NF-κB signaling pathway.

Ellagic acid supplementation ameliorates cisplatin-induced liver injury in mice by inhibiting the NF-κB pathway and activating the Nrf2/HO-1 pathway

Cisplatin(cis-diaminodichloroplatinum II,CDDP),an essential chemotherapeutic agent,can cause potential hepa-totoxicity,but the underlying molecular mechanisms remain unclear.In this study,the protective effects of ellagic acid(EA)on CDDP exposure-induced hepatotoxicity and the underlying molecular mechanisms were investigated in a mouse model.Mice were randomly divided into control,CDDP model,EA100(i.e.,100 mg/kg/day),and CDDP plus 25,50,or 100 mg/kg/day EA groups.Mice in all the CDDP-treated groups were intraperitoneally injected with 20 mg/kg/day CDDP for two days.For all EA cotreatments,the mice were orally administered EA for seven days.Our results revealed that CDDP treatment resulted in liver dysfunction,oxidative stress,and caspase activation,which were effectively attenuated by EA cotreatment in a dose-dependent manner.Furthermore,EA supplementation sig-nificantly downregulated the CDDP exposure-induced protein and mRNA expression of NF-κB,IL-1β,TNF-α,and IL-6 but further upregulated the protein and mRNA expression of Nrf2 and HO-1.Molecular docking analysis revealed strong interactions between EA and the NF-κB or Keap1 proteins.In conclusion,our results revealed that EA sup-plementation could ameliorate CDDP-induced liver toxicity in mice by activating the Nrf2/HO-1 signaling pathway and inhibiting the NF-kB signaling pathway.

Black phosphorus quantum dots induce myocardial inflammatory responses and metabolic disorders in mice

As an ultrasmall derivative of black phosphorus(BP)sheets,BP quantum dots(BP-QDs)have been effectively used in many fields.Currently,information on the cardiotoxicity induced by BP-QDs remains limited.We aimed to evaluate BP-QD-induced cardiac toxicity in mice.Histopathological examination of heart tissue sections was performed.Transcriptome sequencing,real-time quantitative PCR(RT–qPCR),western blotting,and enzyme-linked immunosorbent assay(ELISA)assays were used to detect the m RNA and/or protein expression of proinfammatory cytokines,nuclear factor kappa B(NF-κB),phosphatidylinositol3 kinase-protein kinase B(PI3K-AKT),peroxisome proliferator-activated receptor gamma(PPARγ),and glucose/lipid metabolism pathway-related genes.We found that heart weight and heart/body weight index(HBI)were significantly reduced in mice after intragastric administration of 0.1 or 1 mg/kg BP-QDs for 28 days.In addition,obvious infammatory cell infiltration and increased cardiomyocyte diameter were observed in the BP-QD-treated groups.Altered expression of proinfammatory cytokines and genes related to the NF-κB signaling pathway further confirmed that BP-QD exposure induced infammatory responses.In addition,BP-QD treatment also affected the PI3K-AKT,PPARγ,thermogenesis,oxidative phosphorylation,and cardiac muscle contraction signaling pathways.The expression of genes related to glucose/lipid metabolism signaling pathways was dramatically affected by BP-QD exposure,and the effect was primarily mediated by the PPAR signaling pathway.Our study provides new insights into the toxicity of BP-QDs to human health.