腺病毒介导RA538及反义c-myc在不同细胞系中作用及其机制

目的比较重组RA538,反义c-myc及LacZ腺病毒(adenovirus,AV)对不同靶细胞的转染效率、生物学特性并探讨其作用的分子机制.方法以人胃癌细胞(SGC7901)、食管癌细胞(EC109)及人胚肺二倍体细胞(2BS)系为靶细胞,采用LacZ基因转染X-gal染色、形态学观察、MTT,RT-PCR等方法,研究重组RA538,反义c-myc及LacZ AV对上述细胞的转染效率,生物学作用及其分子机制.结果 AV-LacZ进行重组腺病毒转导效率检测显示其对SGC7901,2BS细胞具有很高的转导效率,对EC109细胞转导效率较低.AV-RA538及AV-ASc-myc对SGC7901细胞能产生明显的生长抑制效应并诱导凋亡,其生长抑制率分别为76.3%和44.1%.AV-RA538及AV-ASc-myc对SGC7901细胞内源性c-myc,bcl-2基因的表达具有抑制作用.AV-RA538及AV-ASc-myc对EC109细胞及2BS细胞无明显的生长抑制及凋亡诱导作用,AV-RA538对EC109及2BS细胞中内源性c-myc,bcl-2基因的表达无调节作用.结论 AV载体转导效率很高,能实现目的基因在转导细胞中的高水平表达,但对不同靶细胞的转染效率存在差别.AV-RA538,AV-ASc-myc对SGC7901的生长抑制及凋亡诱导作用可能是通过AV的高效转导及抑制c-myc,bcl-2的表达而实现的.AV-RA538,AV-ASc-myc对食管癌、2BS细胞系无类似作用可能与其对上述细胞的转导的作用及内源性基因表达的作用有关.

Transduction efficiency, biologic effects and mechanism of recombinant RA538, antisense C-myc adenovirus on different cell lines

AIM To compare the transduction efficiency, biological and molecular mechanism of recombinant RA538, antisense c-myc adenovirus on different cell lines. METHODS Human gastric, esophageal cancer and 2BS cell line were treated with AV-RA 538, AV-ASc-myc or AV-LacZ. In order to compare their transduction efficiency, biological effects and molecular mechanism, X-gal stain, cellular morphology, MTT and RT-PCR analysis were used.RESULTS Adenovirus transduction efficiency determined by Recombinant LacZ adenovirus was found higher in SGC7901, EC109 and 2BS cell lines, but was lower in EC109 cell line. AV-RA538 and AV-ASc-myc could strongly inhibit cell growth and induce apoptosis of SGC7901 cells. The growth rates of the AV-RA538 or AV-ASc-myc-infected SGC7901 cells were inhibited by 76.3% and 44.1% respectively. The overexpression of RA538 and ASc-myc could down-regulate expression of c-myc, bcl-2 gene. AV-RA538 or AV-ASc-myc could not inhibit cell growth and induce apoptosis of EC109 and 2BS cells, and could not down-regulate expression of c-myc and bcl-2. CONCLUSION Adenovirus vector has highly transfering efficiency, and can mediate a high level of expression of interest gene in transducted cells, but the transduction efficiency of adenovirus is different on each cell line. AV-RA538 or AV-ASc-myc may inhibit growth and induce apoptosis of gastric cancer cell through high transduction efficiency of AV and inhibition of c-myc and be/-2 expression. AV-RAS38 and AV-ASc-myc can not inhibit cell growth and induce apoptosis of esophageal cancer and 2BS cell line, which may be related to transduction efficiency and regulation of expression gene on them.