Study Of Androgen Receptor And It's Isoforms In The Progression Of Human Prostate Cancer

Objectives: To establish androgen-independent human LNCaP prostate cancer cell model and to investigate the role of androgen receptors and its' isoforms in the progression of human LNCaP prostate cancer. The apoptosis of androgen-independent LNCaP prostate cancer cells was also analyzed.Methods: 1. Androgen-dependent parental LNCaP prostate cancer cells were maintained continuously in a regular RPMI 1640 cell-culture medium. According to the number of passage, three sublines of LNCaP C-33, C-51 and C-81 were established consecutively. The growth characteristics and androgen-dependence of the three LNCaP sublines were determined by flutamide and DHT in vivo and in vitro. 2. AR mRNA and AR protein in three LNCaP sublines were studied by RT-PCR and Western Blot methods. 3. PSA secretion in different LNCaP sublines was analyzed by radio-immunity method. 4. Effects of DHT on PSA secretion in different LNCaP sublines were detected by Western Blot method. 5. Vitalities of different LNCaP cells were studied by MTT method. 6. Apoptosis in different LNCaP sublines was determined by DNA ladder and Annexin V-PI FCM analysis.Results: 1. Three sublines of LNCaP C-33, C-51 and C-81 were established and the three LNCaP sublines demonstrated different growth characteristics and androgen -dependence. LNCaP C-81 subline displayed androgen-independent characteristics and the LNCaP cell model of androgen-independence was set up successfully. 2. There was no difference in expressions of AR mRNA and total AR protein in different LNCaP cells by RT-PCR and Western Blot methods. LNCaP C-81 cell exhibited characteristic AR expressions of increase in AR-A isoform and decrease in AR-B isoform. 3. There was difference in PSA secretion in the three LNCaP sublines and LNCaP C-81 cell had the highest rate of PSA secretion by radio-immunity method. 4. DHT manifested stronger stimulation effect on PSA secretion in LNCaP C-33 cell than that in LNCaP C-81 cell. 5. LNCaP C-81 cell showed higher vitality than that in LNCaP C-33 cell and C-51 cell by MTT method. 6. The anti-apoptosis ability in LNCaP C-81 cell was stronger than that in LNCaP C-33 cell and C-51 cell.Conclusions: 1. Studies in vivo and in vitro demonstrated that when androgen-dependent parental LNCaP prostate cancer cells were maintained continuously in a regular RPMI1640 cell-culture medium, the biological behaviors of the LNCaP cells could be altered. The androgen-dependence of these LNCaP cells decreased gradually as the passage number increasing, which displayed the process of LNCaP cell progression from androgen-dependence to androgen-independence. The LNCaP cell model established in the study was an androgen-independent cell model and the cell model can simulate progression of human prostate cancer from androgen-dependent to the hormone -refractory state without androgen intervention in clinic. This cell model may provide the opportunity to understand the molecular mechanisms underlying the acquisition of androgen independence during human prostate cancer progression. 2. AR protein was always expressed during progression of LNCaP prostate cancer, and its total expression level was not altered. This phenomena indicates that activation of AR or AR pathways is a key step in progression of human LNCaP prostate cancer. Because different AR isoforms have different function and structure, differential expression of AR isoforms in different LNCaP cells may lead to inappropriate activation of AR or AR pathways in LNCaP prostate cancer. 3. Gradual increase in PSA secretion and gradual decrease in stimulation effects of DHT on PSA secretion during progression of LNCaP prostate cancer in the experiment confirm that serum PSA level can monitor progression and relapse of human prostate cancer as an credible indicator, and can be used as an credible indicator to reflect therapeutic effect of all types of treatments in prostate cancer. 4. The acquisition of anti-apoptosis in prostate cancer cell may be an important mechanism associated with progression of human LNCaP prostate cancer, a...