The construction and biological function assessment of HCC cell lines expressing IL-33 isoform

JIN Ziqi; LEI Lei; LIN DANDan; HU Bo; LIU Haiyan

doi:10.13431/j.cnki.immunol.j.20180119

Abstract：This study aimed to construct HCC cell lines stable expressing full-length-IL-33 (fIL-33), secreted-IL-33 (sIL-33), or nuclear-IL-33 (nIL-33), and to assess the effect of fIL-33, sIL-33, nIL-33 on tumor cell growth, apoptosis and cell cycle. Firstly, fIL-33, sIL-33, nIL-33 were cloned into lentivirus vector pRRL-Venus. Then lentivirus was generated by transfecting venus-fIL-33, venus-sIL-33, venus-nIL-33 or venus into the 293 T packaging cell lines. Hepa1-6 cells were infected with 1 ml lentiviral supernatant for 24 hours. Single cells were sorted into 96-well plates by flow cytometry 72 h later. A series of in vitro experiments were carried out to determine the biological function of fIL-33, sIL-33 and nIL-33, including cell proliferation by CCK8, apoptosis by Annexin V and 7 AAD, cell cycle by PI flow cytometry. Data showed that the fIL-33, sIL-33, and nIL-33 steady transfected cell lines were successfully established. fIL-33 promoted hapa1-6 cells growth while sIL-33 and nIL-33 inhibited hepa1-6 cells proliferation;fIL-33 inhibited both early and late apoptosis of tumor cells while nIL-33 only inhibited late apoptosis, and sIL-33 did not affect apoptosis of hepa1-6 cells. sIL-33 elevated the percentage of G1 period tumor cells significantly. We did not observe obvious effect of fIL-33 and nIL-33 on cell cycle. These results founded a solid foundation for further investigation of fIL-33, sIL-33 and nIL-33 in the development of hepatocellular carcinoma.