INHIBITION OF HIV-1 GP120-INDUCED APOPTOSIS IN NEUROBLASTOMA SK-N-SH CELLS BY AN ANTISENSE OLIGODEOXYNUCLEOTIDE AGAINST P53

Objectives: This study examines the cytotoxicity potential and the mec hanism of toxicity of the HIV-1 gp120 on human neuroblastoma cells. De sign: Previous data from our group have suggested that the HIV-1 envel ope protein gp120 promotes the secretion of tumor necrosis factor-alph a and other factors by astrocytes and microglial cells present in prim ary human brain cell cultures, thereby contributing to the injury of n eurons in these cultures. This study investigates the cytotoxicity pot ential and the mechanism of toxicity of gp120 on human neuroblastoma c ells. Methods: SK-N-SH cells were treated with HIV-1 gp120, and was fo llowed by in situ DNA fragmentation staining and small molecular weigh t DNA extraction studies to ascertain the induction of apoptosis by gp 120 in these cells. To evaluate a potential role of the growth suppres sor gene p53, gp120-treated SK-N-SH cells were subjected to reverse tr anscription polymerase chain reaction (RT-PCR) and Western blot analys es for the induction of p53. An antisense oligodeoxynucleotide against p53 was used to investigate the role of p53 in the gp120-induced apop tosis in these cells. Results: Data from T7 DNA polymerase staining an d small molecular weight DNA extraction studies demonstrated that gp12 0-induced DNA breakage in SK-N-SH cells with fragmentation patterns ch aracteristic of apoptosis. RT-PCR and Western blot analyses revealed t hat the gp120-mediated induction of apoptosis was dependent on a gp120 -induced and gp120-sustained upregulation of p53. The induction of p53 by gp120 was specific, since an antibody against gp120 prevented both the induction of p53 and subsequent apoptosis in SK-N-SH cells. The c ritical role of p53 was further illustrated by the effectiveness of a p53 antisense oligodeoxynucleotide to inhibit the gp120-induced apopto sis. As a control, the apoptosis-inducing potential of gp120 on SK-N-S H cells was not seen in the HIV-1 Gag proteins even when used at up to 5 nM. Conclusions: These results established that HIV-1 gp120 is pote ntially cytotoxic to human neuronal cells through the induction of p53 , which may eventually lead to induction of apoptosis. (C) 1998 Rapid Science Ltd.