Up-regulation of B-cell lymphoma 6 expression suppresses apoptosis of rat liver cells / 第二军医大学学报

ABSTRACT

Objective To clone the gene of B-cell lymphoma 6 (Bcl6), construct a green fluorescent expression vector carrying rat Bcl6 gene, and to evaluate its roles in rat liver cells. Methods Total liver RNA was isolated from rat liver samples and cDNA was synthesized by reverse transcription; the coding sequence of Bcl6 gene was obtained by nested PCR. PEGFP-Bcl6 fusion expression vector was constructed by recombination technique and confirmed by sequencing. Then the recombinant vectors were transfected into rat liver cell line BRL-3A by lipofectamine; the expression levels of Bcl6 mRNA and protein were determined by real-time PCR and Western blotting analysis, respectively. Finally, the apoptosis of BRL-3A cells transfected with recombinant vector was analyzed by flow cytometry and cell proliferation was examined by MTT assay. Results The recombinant vector pEGFP-Bcl6 was successfully constructed as determined by PCR and sequencing, and was effectively delivered to BRL-3A cells by transient transfection as confirmed by PCR and Western blotting analysis. Result of flow cytometry proved that Bcl6 up-regulation suppressed apoptosis of rat liver cells and MTT assay found that Bcl6 up-regulation promoted proliferation of liver cells. Conclusion We have successfully cloned rat Bcl6 gene and constructed its green fluorescent expression vector. Our results initially prove that Bcl6 may inhibit cell apoptosis and promote cell proliferation in rat liver cells.