The Effect of lactoferrin and DNAJC15 domains for viral replication and cell viability in HBx-induced human hepatoma Hep3B cell line

• Main Authors: Yu-Sen Hwang, 黃裕森
• Other Authors: Chuan-Mu Chen
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/75673048182951384599

碩士 === 國立中興大學 === 生命科學系所 === 104 === Hepatocellular carcinoma (HCC) is one of the common cancers ranking secondary beside to lung cancer in Taiwan’s 2015. Currently, 55 – 60% of HCC cases worldwide are caused by hepatitis B virus (HBV) infection, most occur in Asia. Previous reports indicate that the inflammation and self-repair mechanism of hepatocytes caused by chronic HBV infection will increase the occurrence of HCC. Therefore, it has been reported that anti-HBV therapy can help reduce the risk of HCC. Lactoferrin exists in animal milks and has iron-chelating function and has been shown to have anti-inflammatory, anti-cancer, anti-microbial, anti-oxidant and immune-enhancing effects in different tissues. Some recent studies also confirmed that the treatment of lactoferrin can inhibit HBV infection. In our the previous study indicated that the overexpression of DNAJC15 in Hep3B human hepatocellular carcinoma cells, which lacks DNAJC15 and carries the HBV genome in the cells, can inhibit cancer cell invasion and migration, DNAJC15 was speculated as a tumor suppressor gene in hepatic tumorigenesis. To further investigate insight of the functional domains of DNAJC15 and the synergistic effect of lactoferrin in inhibiting cancer cell growth, a variety of the truncated DNAJC15 were constructed and subsequently transfected into Hep3B cells for stable expression and for the following treatments with lactoferrin at different concentrations. DNAJC15 is the protein integrated in the inner membrane of mitochondria, present results exhibited that the C-terminal J domain, but not N-terminal domain, is the anti-tumor functional domain of DNAJC15 due to its deletion can block the growth-inhibiting effects of DNAJC15 in Hep3B cells. However, free J domain or free C-terminal domain also cannot show the normal function of DNAJC15. Therefore, the function of DNAJC15 relies on the completeness of its C-terminal region or its localization in mitochondria membrane. Furthermore, the treatment with lactoferrin decreased the viabilities of anchored J domain transfected Hep3B cells at the dose-dependent manner. On the other hand, the results from flow cytometry indicated that DNAJC15 and lactoferrin can induce apoptosis and cause cell cycle arrest at G0/G1 phase. In summary, present study demonstrates that the J domain plays an important function of DNAJC15, but depends on its anchorage on mitochondria membrane, and lactoferrin shows the synergistic activity on the tumor suppressing effects of DNAJC15.