[Prevention of cell death in LNCaP cells by overexpression of clusterin].
OBJECTIVE
To evaluate the effect of clusterin with and without leader sequence on overexpression preventing apoptosis in human prostate LNCaP cells.


METHODS
The plasmid pIRES2-EGFP was used to generate the clusterin expression constructs with full-length or without the leader sequence (designated as pIRES2-EGFP/cluac, pIRES2-EGFP/clubc, respectively). Western blot analysis was employed to compare clusterin expression levels in the lysis and supernatant fluid of clusterin transfected LNCaP cells in vitro. The distribution of different functional domains of clusterin in cells was detected with Immunocytochemical staining. The clusterin's protective role of Na2SeO3-induced apoptosis in LNCaP cells was examined by flow cytometry (FCM) and fluorescence microscope.


RESULTS
Clusterin expression was detected in the lysis and supernatant fluid of pIRES2-EGFP/cluac transfected LNCaP cells, while clusterin was found only in lysis liquid of pIRES2-EGFP/clubc transfected LNCaP cells, but not found in their supernatant fluid. The distribution of cluserin in the plasm of pIRES2-EGFP/cluac transfected cells was aggregative, and on the other hand, clusterin distributed dispersedly in pIRES2-EGFP/clubc transfected cells. Its anti-apoptotic property in LNCaP cells was proved by FCM and fluorescence microscope.


CONCLUSIONS
It is apparent that clusterin plays an important role in preventing apoptosis in prostate cancer, and the presence of the leader sequence is necessary for clusterin's anti-apoptotic function.
Xin
Dian-qi
DQ
Department of Urology, Peking University First Hospital, Beijing 100034, China. xindianqi@hotmail.com
Zhu
Xu-hui
XH
Ai
Jun-kui
JK
Na
Yan-qun
YQ
Guo
Ying-lu
YL
D017209:Apoptosis; D015153:Blotting, Western; D045744:Cell Line, Tumor; D051152:Clusterin; D006023:Glycoproteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008297:Male; D018832:Molecular Chaperones / Q000235:genetics / Q000502:physiology*; D011471:Prostatic Neoplasms / Q000473:pathology*; D014162:Transfection
C495242:CLU protein, human; D051152:Clusterin; D006023:Glycoproteins; D018832:Molecular Chaperones