Construction of a recombinant adenovirus overexpressing rat TIPE2 gene / 中国组织工程研究

BACKGROUND:Tumor necrosis factor-α-induced protein-8 like-2 (TIPE2), an anti-inflammatory protein, through the T cel receptor (TCR) and TOLL-like receptor signaling pathway, implements negative regulation of adaptive immunity and innate immunity, and thus effectively maintains the stable internal environment of the body. OBJECTIVE:To construct a recombinant adenovirus that can overexpress rat TIPE2 gene. METHODS:TIPE2 cDNA target gene was amplified from rat’s lymphocytes using RT-PCR, cloned into shuttle plasmid pShuttle-clontech, and then subcloned into artificial adenovirus vector AdC68. Hereafter, HEK 293 cel s were transfected to generate a recombinant adenovirus. HEK293A cel s were infected using this recombinant adenovirus, and then TIPE2 gene level was tested by western blot method. RESULTS AND CONCLUSION:Based on results of PCR, digestion identification and sequencing, the obtained cDNA was the coding sequence region of TIPE2. Western blot findings showed that the recombinant adenovirus could overexpress TIPE2 gene. These findings indicate that the recombinant adenovirus is constructed successful y and can express TIPE2 gene stably.

The in vitro and in vivo effect of adeno-associated virus-mediated combined p53,p16 and p21 gene transfection on human hepatic carcinoma cell lines / 中国普通外科杂志

ObjectiveTo evaluate the effect of combined gene transfection with p53,p16 and p21 on the growth of human liver cancer cell lines.MethodsAfter transducing adeno associated virus(AAV) mediated p53, p16 or/and p21 genes to human hepatic carcinoma cell lines HLE,HepG2,QGY 7701,QGY 7703,BEL 7402,SMMC 7721, gene expression and tumor inhibition were studied in vitro and in BALB/c mouse model.ResultsAdeno associated virus mediated p53, p16 or p21 encoding gene could express in BEL 7402 cell line. Each individual type of recombinant AAV effected a significant induction of tumor cell apoptosis both in vitro and in vivo, the rate of apoptotic cells in vitro is about 30% and that of tumor growth inhibition is about 30～44%. And the apoptosis inducing efficiency was the highest after the tumor cell line was transfected by three recombinant AAV simultaneously, with a rate of 56% (in vitro) and 65% (in vivo).ConclusionNot only could all of the exogenous wild type p53, p16 and p21 genes mediated by AAV inhibit the growth of liver cancer cell lines, but also can the efficiency be significantly elevated by combined gene transfection.