hNTCP‑expressing primary pig hepatocytes are a valuable tool for investigating hepatitis B virus infection and antiviral drugs

Abstract

Primary human hepatocytes (PHHs) are the ‘gold standard’ for investigating hepatitis B virus (HBV) infection and antiviral drugs. However, poor availability, variation between batches and ethical issues regarding PHHs limit their applications. The discovery of human sodium taurocholate co-transporting polypeptide (hNTCP) as a functional HBV receptor has enabled the development of a surrogate model to supplement the use of PHHs. In the present study, the evolutionary distance of seven species was assessed based on single-copy homologous genes. Based on the evolutionary distance and availability, PHHs and primary rabbit hepatocytes (PRHs) were isolated and infected with hNTCP-recombinant lentivirus, and susceptibility to HBV infection in the two cell types was tested and compared. In addition, HBV infection efficiency of hNTCP-expressing PPHs with pooled HBV-positive serum and purified particles was determined. The potential use of HBV-infected hNTCP-expressing PPHs for drug screening was assessed. The results demonstrated that pigs and rabbits are closer to humans in the divergence tree compared with mice and rats, indicating that pigs and rabbits were more likely to facilitate the HBV post-entry lifecycle. Following hNTCP complementation and HBV infection, PPHs and Huh7D human hepatocellular carcinoma cells, but not PRHs, exhibited increased hepatitis B surface antigen and hepatitis B e-antigen secretion, covalently closed circular DNA formation and infectious particle secretion. hNTCP-expressing PPHs were susceptible to infection with HBV particles purified from pooled HBV-positive sera, but were poisoned by raw HBV-positive sera. The use of HBV-infected hNTCP-expressing PPHs for viral entry inhibitor screening was revealed to be applicable and reproducible. In conclusion, hNTCP-expressing PPHs may be valuable tool for investigating HBV infection and antiviral drugs.

Aborted infection of HBV in human sodium taurocholate cotransporting polypeptide (hNTCP) expressed woodchuck cells

“…Based on the evolutionary distance, pigs and macaque were chosen. Macaque hepatocytes [11,19] and primary pig hepatocytes [11,18] expressing hNTCP exhibited increased expression levels of HBsAg and HBeAg, as well as increased cccDNA formation after HBV infection. Therefore we speculate that, the restriction of HBV infection is not only related to hNTCPdependent steps but also caused by lacking some additional species-speci c factors.…”

“…The discovery of the HBV receptor, hNTCP, led to the successful establishment of the cell line that susceptible for HBV and HDV. Zhou et al found that primary pig hepatocytes infected with hNTCP recombinant lentivirus were susceptible to HBV infection [18]. Burwitz et al showed macaque hepatocytes could support HBV replication in vitro following transduction with viral vectors encoding hNTCP, and increased amounts of HBV DNA in serum could also be detected over time in vivo [19].…”

Aborted infection of HBV in human sodium taurocholate cotransporting polypeptide (hNTCP) expressed woodchuck cells

“…Based on the evolutionary distance, pigs and macaque were chosen. Macaque hepatocytes (Burwitz et al 2017Lempp et al 2017) and primary pig (Lempp et al 2017Zhou et al 2019) hepatocytes expressing hNTCP exhibited increased expression levels of HBsAg and HBeAg, as well as increased cccDNA formation after HBV infection (Lempp et al 2016). Therefore we speculate that, the restriction of HBV infection is not only related to hNTCP-dependent steps but also caused by lacking some additional species-speci c factors.…”

“…The discovery of the HBV receptor, hNTCP, led to the successful establishment of the cell line that susceptible for HBV and HDV. Zhou et al found that primary pig hepatocytes infected with hNTCP recombinant lentivirus were susceptible to HBV infection (Zhou et al 2019). Burwitz et al showed macaque hepatocytes could support HBV replication in vitro following transduction with viral vectors encoding hNTCP, and increased amounts of HBV DNA in serum could also be detected over time in vivo (Burwitz et al 2017).…”

ERK Activation-Mediated Autophagy Induction Resists Licochalcone A-Induced Anticancer Activities in Lung Cancer Cells in vitro

“…Recently its antitumor activities have attracted great interest of researchers. It has been demonstrated that LA inhibits the proliferation of different types of cancers, including gastric cancer BGC-823, AGS, SGC-7091 and MKN-45 cells, 12,13 bladder cancer T24 and 5367, 14 hepatocellular carcinoma HepG2 cells, 15 breast cancer MDA-MB-231 cells, 16 glioblastoma U87 cells, 17 nasopharyngeal carcinoma HONE-1, NPC-39 and NPC-BM cells, 18 cervical cancer SiHa and Hela cells, 19 osteosarcoma HOS and MG-63 cells, 20 lung cancer A549, H1299, H292 and H460, [21][22][23] and so on. In vivo studies also reveal that LA is a potent anticancer agent in gastric cancer, cervical cancer and colon cancer.…”