[Effects of antisense bcl-2 or survivin on the growth of human neuroblastoma cell line SK-N-MC].
OBJECTIVE
To study the effects of antisense bcl-2 and survivin (svv) mRNA on the growth of human neuroblastoma (NB) cell line SK-N-MC cells.


METHODS
The recombinant vectors PBabe puro-Asbcl-2 and PBabe puro-Assvv. were constructed by directed cloning of the EcoRI-BamHI fragments of bcl-2 cDNA or svv cDNA into the retroviral vector PBabe puro Human NB cell line SK-N-MC cells were transfected with PBabe puro-Asbcl-2, PBabe puro-Assvv, or blank vector PBabe puro as control by lipofectamine trade mark. The transfected cells were selected in the medium containing puromycine. The stably transfected cells were further studied for inhibition of protein expression of endogenous bcl-2 and SVV by immunohistochemical staining and Western blotting. The effect of antisense bcl-2 (Asbcl-2) and antisense svv (Assvv) mRNA on cell growth was determined by MTT method. The SK-N-MC cells transfected with the recombinant vectors were inoculated in nude mice to observe their carcinogenicity.


RESULTS
Both the expression of bcl-2 and the expression of SVV significantly decreased in the antisense gene transfected cells in comparison to that in the original cells and cells transfected with blank vector. Seven days after the transfection, the MTT absorption (A(550)) was 0.374 +/- 0.001 5 in the cells transfected with Assvv, 0.289 +/- 0.000 8 in the cells transfected with Asbcl-2, both significantly lower than those in the original cells and cells transfected with blank vector (1.102 +/- 0.002 1 and 1.175 +/- 0.000 9 respectively). The induced tumors in the nude mice were smaller in the PBabe puro-Asbcl-2 transfected group and PBabe puro-Assvv transfected group than in the original and control groups.


CONCLUSIONS
Stably expression of antisense bcl-2 and of antisense svv mRNA can effectively inhibit the expression of endogenous bcl-2 and SVV proteins. Both of them may play a role in the neoplastic formation of NB cells.
Guan
Jian
J
Department of Pathology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing 100730, China.
Chen
Jie
J
Luo
Yufeng
Y
Gao
Jie
J
Qiu
Honghui
H
D002455:Cell Division / Q000235:genetics; D016373:DNA, Antisense / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000235:genetics*; D009363:Neoplasm Proteins; D009447:Neuroblastoma / Q000235:genetics / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000077022:Survivin; D014162:Transfection; D014407:Tumor Cells, Cultured
C107489:BIRC5 protein, human; D016373:DNA, Antisense; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D000077022:Survivin