Hepatitis B virus efficiently infects non-adherent hepatoma cells via human sodium taurocholate cotransporting polypeptide.
Hepatitis B virus efficiently infects non-adherent hepatoma cells via human sodium taurocholate cotransporting polypeptide.
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DOI：
10.1038/srep17047
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发表时间：
2015-11-23
影响因子：
4.6
通讯作者：
Moriishi K
中科院分区：
文献类型：
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作者：
Okuyama-Dobashi K;Kasai H;Tanaka T;Yamashita A;Yasumoto J;Chen W;Okamoto T;Maekawa S;Watashi K;Wakita T;Ryo A;Suzuki T;Matsuura Y;Enomoto N;Moriishi K
牛磺胆酸钠共转运多肽(NTCP)被认为是乙肝病毒(HBV)感染的功能性受体。然而，在贴壁单层细胞条件下，乙肝病毒不能有效感染表达NTCP的HepG2细胞(NTCP-HepG2细胞)。在本研究中，Ntcp主要在单层Ntcp-HepG2细胞的基侧膜区检测到，而在顶端部位未检测到。我们假设感染的非贴壁细胞条件会增强乙肝病毒的传染性。用胰酶和EDTA处理制备了非贴壁的NTCP-HepG2细胞，该细胞不降解膜组分中的NTCP。非贴壁阶段的病毒剂量比贴壁阶段低10倍，成功地感染了NTCP-HepG2细胞。观察到血液传播或细胞培养来源的乙肝病毒有效地感染非贴壁的NTCP-HepG2细胞，并且在肉豆蔻酰化的PreS1肽的存在下，这种感染显著受损。在非贴壁细胞条件下，乙肝病毒也能有效地感染HepaRG细胞。我们利用我们的培养系统筛选了几个化合物，并鉴定出Proscillaridin A是一种有效的抗乙肝病毒药物，其IC50值为7.2 NM。综上所述，非贴壁宿主细胞感染条件以一种NtCP依赖的方式增强了乙肝病毒的感染性，从而为寻找抗乙肝药物和研究乙肝病毒感染机制提供了新的策略。
Sodium taurocholate cotransporting polypeptide (NTCP) has been reported as a functional receptor for hepatitis B virus (HBV) infection. However, HBV could not efficiently infect HepG2 cells expressing NTCP (NTCP-HepG2 cells) under adherent monolayer-cell conditions. In this study, NTCP was mainly detected in the basolateral membrane region, but not the apical site, of monolayer NTCP-HepG2 cells. We hypothesized that non-adherent cell conditions of infection would enhance HBV infectivity. Non-adherent NTCP-HepG2 cells were prepared by treatment with trypsin and EDTA, which did not degrade NTCP in the membrane fraction. HBV successfully infected NTCP-HepG2 cells at a viral dose 10 times lower in non-adherent phase than in adherent phase. Efficient infection of non-adherent NTCP-HepG2 cells with blood-borne or cell-culture-derived HBV was observed and was remarkably impaired in the presence of the myristoylated preS1 peptide. HBV could also efficiently infect HepaRG cells under non-adherent cell conditions. We screened several compounds using our culture system and identified proscillaridin A as a potent anti-HBV agent with an IC50 value of 7.2 nM. In conclusion, non-adherent host cell conditions of infection augmented HBV infectivity in an NTCP-dependent manner, thus providing a novel strategy to identify anti-HBV drugs and investigate the mechanism of HBV infection.