Sodium taurocholate cotransporting polypeptide(NTCP)is identified as the functional receptor for HBV entry,which is responsible for upregulated HBV transcription in the HBV life cycle.Besides,NTCP is also implicated i...Sodium taurocholate cotransporting polypeptide(NTCP)is identified as the functional receptor for HBV entry,which is responsible for upregulated HBV transcription in the HBV life cycle.Besides,NTCP is also implicated in the progression of HBV-induced hepatocellular carcinoma(HCC).Thereby,NTCP-targeting entry inhibitors are proposed to suppress HBV infection and replication in HBV-induced hepatoma therapy.Herein,we integrated in silico screening and chemical synthesis to obtain a small-molecule NTCP inhibitor B7,which exhibited moderate anti-proliferative activities against HepG2 cells and anti-HBV activity in vitro.Additionally,CETSA assay,molecular docking,and MD simulation validated that B7 could bind to NTCP.Furthermore,western blot analysis demonstrated that B7 induced apoptosis with an increased expression of Bax and caspase 3 cleaving as well as a decreasing expression of Bcl-2 in HepG2 cells.Taken together,our study identified B7 as a novel NTCP inhibitor with anti-proliferation activities which might provide a new opportunity for HCC therapy.展开更多
Background and aims:The sodium taurocholate co-transporting polypeptide(NTCP)is a functional receptor for the hepatitis B virus(HBV),and it is critical for bile acid homeostasis.Previous studies of the association bet...Background and aims:The sodium taurocholate co-transporting polypeptide(NTCP)is a functional receptor for the hepatitis B virus(HBV),and it is critical for bile acid homeostasis.Previous studies of the association between the S267F variant and chronic hepatitis B(CHB)have generated conflicting results.This study analyzed the correlation between the NTCP S267F variant and CHB susceptibility by using a large sample of participants classified by gender and age,and this study also analyzed the relationship between this variant and the level of serum total bile acids.Methods:In total,543 patients with CHB and 429 control subjects underwent S267F variant genotyping using SNaPshot technology.Logistic regression was utilized to evaluate any association of the NTCP S267F variant with CHB susceptibility.Results:The S267F variant was inversely correlated with the risk of chronic HBV infection in both the dominant model(GG genotype vs.AG genotype:odds ratio(OR)=0.46,95%confidence interval(CI)0.30 -0.71,P<0.001)and the allele model(G allele vs.A allele:OR=0.50,95%CI 0.33-0.76,P=0.001),and this correlation was not affected by gender and age stratification.The carriers of the heterozygous NTCP variant exhibited higher total bile acids levels than the carriers of wild-type NTCP,regardless of whether they were control subjects or patients with CHB.Heterozygous carriers exhibited reduced hepatitis B e antigen(HBeAg)-positivity rates and had lower ALT,AST,and lg DNA concentrations compared with wild-type carriers in patients with CHB.Conclusions:The S267F variant of NTCP is a protective factor that reduces the risk of chronic HBV infection and exhibits a higher total bile acids level.Patients with CHB who carry this variant may have a better prognosis than those carrying wild-type NTCP.展开更多
AIM To determine the variability/conservation of the domain of hepatitis B virus(HBV) pre S1 region that interacts with sodium-taurocholate cotransporting polypeptide(hereafter, NTCP-interacting domain) and the preval...AIM To determine the variability/conservation of the domain of hepatitis B virus(HBV) pre S1 region that interacts with sodium-taurocholate cotransporting polypeptide(hereafter, NTCP-interacting domain) and the prevalence of the rs2296651 polymorphism(S267 F, NTCP variant) in a Spanish population. METHODS Serum samples from 246 individuals were included and divided into 3 groups: patients with chronic HBV infection(CHB)(n = 41, 73% Caucasians), patients with resolved HBV infection(n = 100, 100% Caucasians) and an HBV-uninfected control group(n = 105, 100% Caucasians). Variability/conservation of the amino acid(aa) sequences of the NTCPinteracting domain,(aa 2-48 in viral genotype D) and a highly conserved pre S1 domain associated with virion morphogenesis(aa 92-103 in viral genotype D) were analyzed by next-generation sequencing and compared in 18 CHB patients with viremia > 4 log IU/mL. The rs2296651 polymorphism was determined in all individuals in all 3 groups using an in-house real-time PCR melting curve analysis.RESULTS The HBV pre S1 NTCP-interacting domain showed a high degree of conservation among the examined viral genomes especially between aa 9 and 21(in the genotype D consensus sequence). As compared with the virion morphogenesis domain, the NTCPinteracting domain had a smaller proportion of HBV genotype-unrelated changes comprising > 1% of the quasispecies(25.5% vs 31.8%), but a larger proportion of genotype-associated viral polymorphisms(34% vs 27.3%), according to consensus sequences from Gen Bank patterns of HBV genotypes A to H. Variation/conservation in both domains depended on viral genotype, with genotype C being the most highly conserved and genotype E the most variable(limited finding, only 2 genotype E included). Of note, proline residues were highly conserved in both domains, and serine residues showed changes only to threonine or tyrosine in the virion morphogenesis domain. The rs2296651 polymorphism was not detected in any participant.CONCLUSION In our CHB population, the NTCP-interacting domain was highly conserved, particularly the proline residues and essential amino acids related with the NTCP interaction, and the prevalence of rs2296651 was low/null.展开更多
目的检测乙型肝炎病毒(HBV)受体蛋白在人外周血单个核细胞(PBMC)与肝细胞的表达情况,探讨HBV进入PBMC的可能途径,以期了解HBV感染PBMC的可能机制。方法用淋巴细胞分离液分离人PBMC。用RIPA提取人PBMC和肝细胞(肝脏肿瘤癌旁组织)总蛋白,...目的检测乙型肝炎病毒(HBV)受体蛋白在人外周血单个核细胞(PBMC)与肝细胞的表达情况,探讨HBV进入PBMC的可能途径,以期了解HBV感染PBMC的可能机制。方法用淋巴细胞分离液分离人PBMC。用RIPA提取人PBMC和肝细胞(肝脏肿瘤癌旁组织)总蛋白,采用Western blot以检测HBV受体蛋白在两种细胞的表达情况。结果 Western blot检测肝细胞HBV受体蛋白阳性,与hNTCP抗体反应条带位于(35~40)×103之间,与预期一致;人PBMC蛋白无此反应条带,即hNTCP检测阴性。结论 PBMC不表达肝细胞表面的HBV受体蛋白,病毒可能循非受体途径进入PBMC,或PBMC表面的HBV受体与肝细胞表面的HBV蛋白受体蛋白不同。关于HBV感染PBMC的具体机制尚需进一步研究。展开更多
基金supported by National Science and Technology Major Project of the Ministry of Science and Technology of China(No.2018ZX09735005)the National Natural Science Foundation of China(Nos.81922064,81874290,81673290,81803347 and 81903502)+1 种基金the Natural Science Foundation of Guangdong Province(No.2018A030313707)Post-Doctor Research Project,West China Hospital,Sichuan University(No.2019HXBH034)。
文摘Sodium taurocholate cotransporting polypeptide(NTCP)is identified as the functional receptor for HBV entry,which is responsible for upregulated HBV transcription in the HBV life cycle.Besides,NTCP is also implicated in the progression of HBV-induced hepatocellular carcinoma(HCC).Thereby,NTCP-targeting entry inhibitors are proposed to suppress HBV infection and replication in HBV-induced hepatoma therapy.Herein,we integrated in silico screening and chemical synthesis to obtain a small-molecule NTCP inhibitor B7,which exhibited moderate anti-proliferative activities against HepG2 cells and anti-HBV activity in vitro.Additionally,CETSA assay,molecular docking,and MD simulation validated that B7 could bind to NTCP.Furthermore,western blot analysis demonstrated that B7 induced apoptosis with an increased expression of Bax and caspase 3 cleaving as well as a decreasing expression of Bcl-2 in HepG2 cells.Taken together,our study identified B7 as a novel NTCP inhibitor with anti-proliferation activities which might provide a new opportunity for HCC therapy.
基金This study was funded by the Natural Science Foundation of Fujian Province(No.2021J011447)Ningde Science and Technology Plan Project(No.20170032).
文摘Background and aims:The sodium taurocholate co-transporting polypeptide(NTCP)is a functional receptor for the hepatitis B virus(HBV),and it is critical for bile acid homeostasis.Previous studies of the association between the S267F variant and chronic hepatitis B(CHB)have generated conflicting results.This study analyzed the correlation between the NTCP S267F variant and CHB susceptibility by using a large sample of participants classified by gender and age,and this study also analyzed the relationship between this variant and the level of serum total bile acids.Methods:In total,543 patients with CHB and 429 control subjects underwent S267F variant genotyping using SNaPshot technology.Logistic regression was utilized to evaluate any association of the NTCP S267F variant with CHB susceptibility.Results:The S267F variant was inversely correlated with the risk of chronic HBV infection in both the dominant model(GG genotype vs.AG genotype:odds ratio(OR)=0.46,95%confidence interval(CI)0.30 -0.71,P<0.001)and the allele model(G allele vs.A allele:OR=0.50,95%CI 0.33-0.76,P=0.001),and this correlation was not affected by gender and age stratification.The carriers of the heterozygous NTCP variant exhibited higher total bile acids levels than the carriers of wild-type NTCP,regardless of whether they were control subjects or patients with CHB.Heterozygous carriers exhibited reduced hepatitis B e antigen(HBeAg)-positivity rates and had lower ALT,AST,and lg DNA concentrations compared with wild-type carriers in patients with CHB.Conclusions:The S267F variant of NTCP is a protective factor that reduces the risk of chronic HBV infection and exhibits a higher total bile acids level.Patients with CHB who carry this variant may have a better prognosis than those carrying wild-type NTCP.
基金Supported by Instituto de Salud Carlos Ⅲ,No.PI14/01416 and No.PI15/00856cofinanced by the European Regional Development Fund(ERDF)the Gilead Fellowship Program,No.GLD14-00296
文摘AIM To determine the variability/conservation of the domain of hepatitis B virus(HBV) pre S1 region that interacts with sodium-taurocholate cotransporting polypeptide(hereafter, NTCP-interacting domain) and the prevalence of the rs2296651 polymorphism(S267 F, NTCP variant) in a Spanish population. METHODS Serum samples from 246 individuals were included and divided into 3 groups: patients with chronic HBV infection(CHB)(n = 41, 73% Caucasians), patients with resolved HBV infection(n = 100, 100% Caucasians) and an HBV-uninfected control group(n = 105, 100% Caucasians). Variability/conservation of the amino acid(aa) sequences of the NTCPinteracting domain,(aa 2-48 in viral genotype D) and a highly conserved pre S1 domain associated with virion morphogenesis(aa 92-103 in viral genotype D) were analyzed by next-generation sequencing and compared in 18 CHB patients with viremia > 4 log IU/mL. The rs2296651 polymorphism was determined in all individuals in all 3 groups using an in-house real-time PCR melting curve analysis.RESULTS The HBV pre S1 NTCP-interacting domain showed a high degree of conservation among the examined viral genomes especially between aa 9 and 21(in the genotype D consensus sequence). As compared with the virion morphogenesis domain, the NTCPinteracting domain had a smaller proportion of HBV genotype-unrelated changes comprising > 1% of the quasispecies(25.5% vs 31.8%), but a larger proportion of genotype-associated viral polymorphisms(34% vs 27.3%), according to consensus sequences from Gen Bank patterns of HBV genotypes A to H. Variation/conservation in both domains depended on viral genotype, with genotype C being the most highly conserved and genotype E the most variable(limited finding, only 2 genotype E included). Of note, proline residues were highly conserved in both domains, and serine residues showed changes only to threonine or tyrosine in the virion morphogenesis domain. The rs2296651 polymorphism was not detected in any participant.CONCLUSION In our CHB population, the NTCP-interacting domain was highly conserved, particularly the proline residues and essential amino acids related with the NTCP interaction, and the prevalence of rs2296651 was low/null.
文摘目的检测乙型肝炎病毒(HBV)受体蛋白在人外周血单个核细胞(PBMC)与肝细胞的表达情况,探讨HBV进入PBMC的可能途径,以期了解HBV感染PBMC的可能机制。方法用淋巴细胞分离液分离人PBMC。用RIPA提取人PBMC和肝细胞(肝脏肿瘤癌旁组织)总蛋白,采用Western blot以检测HBV受体蛋白在两种细胞的表达情况。结果 Western blot检测肝细胞HBV受体蛋白阳性,与hNTCP抗体反应条带位于(35~40)×103之间,与预期一致;人PBMC蛋白无此反应条带,即hNTCP检测阴性。结论 PBMC不表达肝细胞表面的HBV受体蛋白,病毒可能循非受体途径进入PBMC,或PBMC表面的HBV受体与肝细胞表面的HBV蛋白受体蛋白不同。关于HBV感染PBMC的具体机制尚需进一步研究。
