Action Research Of Monocyte Chemoattractant Protein1Gene(mcp-1) And Lipocalin2Gene (lcn2) On Hepatoma Cell Strain BEL-7402

Monocyte chemoattractant protein1(mcp-1), a member of the CC chemokine family, could combinewith chemokine(C-C motif) ligand receptor2, and then induce lymphocyte activation, differentiation anddevelopment, and play a vital role in tumor growth or inhibition. Lipocalin-2(lcn2), a member of thelipocalin family, can be expressed and secreted by various types of cells, and participates in manyphysiological activities, such as cell proliferation, apoptosis, differentiation and immune inflammatoryresponse. mcp-1and lcn2genes could induces cell apoptosis by regulated apoptosis related genes, such asBax, bcl-2etc, and have potential as a biomarker of acute kidney injury. In order to clarify the impact ofmcp-1and lcn2on the hepatic cancer cell BEL-7402. Based on the successful cloning of mcp-1and lcn2and establishment of their kinds of vectors, the expression vectors pEGFP-N1-mcp-1, pEGFP-N1-lcn2andits control pEGFP-N1, interference vectors pGenesil-1.0-mcp-1(255), pGenesil-1.0-lcn2(292)and its controlpGenesil-1.0-HK were stably transfected into hepatoma cell line BEL-7402in vitro by lipofectamine, andtheir stably transfected cell lines were selected by G418. Then, the stably transfected cell lines and theircontrol were detected by using the following methods: cell morphology by HE staining; cell growth curvedrew by cell counting and MTT assay; cell proliferation detected by PCNA immunocytochemistry; cellapoptosis detected by Hoechst33258staining; cell colony formation rate detected by methyl cellulosesemi-solid medium; cell resistance to drugs detected by MTT assay.The results detected were as following. The cells stably transfected with pEGFP-N1-mcp-1were notsignificantly different in morphology compared with BEL-7402, but the ratio of uncleus and cytoplasmbecame smaller, and nucleolus was reduced. The growth curve showed that the cells withpEGFP-N1-mcp-1growed slower than those with pEGFP-N1; PCNA immunocytochemistry showed thatmcp-1inhibited cell growth; Hoechst33258staining showed that mcp-1had no significant effect oninducing cell apoptosis; The detection results of methyl cellulose solid medium showed that mcp-1inhibited the degree of malignant proliferation of BEL-7402cells. The uncleus of the cells withpEGFP-N1-lcn2became smaller and condensed, and the nucleolus were reduced; The growth curveshowed that he cells with pEGFP-N1-lcn2growed obviously slower than others; PCNAimmunocytochemistry showed that lcn2dramatically inhibited cell growth; Hoechst33258staining showedthat lcn2had significant effect on inducing cell apoptosis; The detection results of methyl cellulose solid medium showed that lcn2inhibited the degree of malignant proliferation of BEL-7402cells obviously; TheMTT assay of drug found that lcn2had no obvious resistant to amino acid derivative L-4-fluorophenylalanine (Fp) and L-4-fluoro phenylalanine (Clp).In summary, mcp-1could inhibite cell proliferation and malignant phenotypes of the BEL-7402cells,but had no significant effect on inducing the apoptosis of BEL-7402cells; Lcn2could inhibite cellproliferation and malignant phenotypes of the BEL-7402cells obviously, but could promote the apoptosisof BEL-7402cells.