The Gene Expression of Caveolin-1 in Human Androgen-independent Prostate Cancer Cells and the Intervention Study of Simvastatin on PC-3 Cells
|School||Central South University|
|Keywords||Caveolin-1 Prostate cancer cells Tumor metastasis Simvastatin Apoptosis|
In western countries, prostate cancer is the most common malignant tumor, with the first incidence and second mortality rates in all male tumors. In recent years, the incidence rate of prostate cancer in China is going up, and the reported incidence rate is up to third in malignant tumor of urinary system. Now a universal viewpoint think that the development of prostate cancer is going through prostatic intraepithelial neoplasia (PIN), local, invasive and metastatic prostate cancer from normal prostate to tumorigenesis. Howerver, almost 2/3 patients with prostate cancer have been in terminal stages of cancer when they go to see a doctor, and combined therapy with androgen blockade is the only strategy to relieve the symptom. But 1/3 cases may recur and change into androgen-independent prostate cancer after 12～18 months’ therapy. At the present, the mortality rate of androgen-independent prostate cancer patients is very high because of lacking of effective therapy. Thus, the therapy of androgen-independent prostate cancer has come to clinical attention. It has been reported that many factors are involved in the change process from androgen-dependent to androgen independent of the prostatic cancer cell. Recent studies has shown that caveolin-1 play a pivotal role in the regulation of cholesterol balance, signal transduction, vesicular transport and tumorigenesis progression. It has been reported that caveolin-1 is expressed at high levels in human prostate cancer, and plays an important role in the development of prostate cancer。Statins, the hydroxymethyglutaryl coenzyme A (HMG-CoA) reductase inhibitors, are widely used for treatment of atherosclerosis. Recent studies have demonstrated that statins inhibit the cell growth and induce apoptosis in various cancer cells such as HL-60, melanoma, fibroma, breast cancer. It is still unclear whether statins can inhibit the cell growth and induce apoptosis of prostate cancer cells, whether the effects of statins is associated with caveoline-1.In the present study, the relationship between the expression of caveolin-1 and the highly invasion and metastasis of prostate cancer was investigated by measurement of the expression of caveolin-1 in PC-3 and PC-3m cells using semiquantitative reverse transcription polymerase chain reaction (RT-PCR) and western blot analysis. And the molecular mechanisms of simvastatin on prostate cancer was also explored by analyzing proliferation, apoptosis, activity of caspase-3 and expression of caveolin-1 in PC-3 cells. ChapterⅠThe gene expression of caveolin-1 in PC-3 and PC-3m cells and its clinical valueObjectives: To study the expression of caveolin-1 in PC-3 and PC-3m cells, analyze the relationship between caveolin-1 and prostate carcinogenesis, progression, and explore the role of overexpression of caveolin-1 in PC-3 cells with higher invasion and metastasis in order to search for a novel target of gene therapy of androgen-independent prostate cancer.Methods: To measure the expression of caveolin-1 in PC-3 and PC-3m cells by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) and western blot analysis,Results: (1) The mRNA expression of caveolin-1 was detected in PC-3 and PC-3m cells, and the expression of caveolin-1 in PC-3m cells was more higher than that in PC-3 cells (P＜0.05).(2) The protein expression of caveolin-1 was detected in PC-3 and PC-3m cells, and the expression of caveolin-1 in PC-3m cells was higher than that in PC-3 cells (P＜0.05).Conclusions: (1) The mRNA and protein expression of caveolin-1 are detected in PC-3 and PC-3m cells.(2) Overexpression of caveolin-1 may be related to the invasion and metastasis of human prostate cancer.(3) Caveolin-1 may be a target of gene therapy of prostate cancer. ChapterⅡThe effects of simvastatin on human androgen-independent prostate cancer PC-3 cell line in vitroObjectives: To observe the effects of simvastatin on cell growth, and apoptosis in PC-3 cells, and explore the molecular mechanisms of simvastatin on human prostate cancer。Methods: The proliferation of PC-3 cells was measured by MTT assay. The morphological changes and percentage of apoptotic nuclei in PC-3 cells treated with simvastatin were assayed by Hoechst 33258 staining, and apoptosis was determined by Annexin V/PI double staining technique and flow cytometry. The activities of caspase-9, 8 and 3 were analyzed by Caspase Colorimetric Assay Kit. The expression of caveolin-1 was detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis.Results: (1) Simvastatin significantly inhibited the proliferation of PC-3 cells in a concentration-and time-dependent manner (P＜0.05).(2) A morphological change was observed in PC-3 cells. After 24 or 48h treatment with simvastatin (20/μmol/L), condensation of chromatin occurred, and blebbing nuclei and granular apoptotic bodies appeared. The results of FACS showed that the percentage of apoptotic cells in the PC-3 cells treated with simvastatin (20/μmol/L) for 12, 24, or 48 h were (14.81±1.08)%, (23.42±2.33)% or (40.77±4.06)%, respectively. After exposure to different concentrations of simvastatin (10, 20, or 40/μmol/L) for 48 hours, the PC-3 cells occured a lot of apoptosis cells, and percentage of apoptotic nuclei respectively were (11.20±3.33)%, (31.20±3.08)% or (56.24±4.50)%. Similarly, the results of FACS also indicated a dramatic increased in apoptosis cells, and the percentage of apoptotic cells were (17.26±1.44)%, (32.45±2.56)% or (60.47±3.98)%, respectively. The activation of Caspase-9, 8 or 3 were markedly increased in PC-3 cells treated with simvastatin (20 or 40μmol/L) at indicated periods.(3) The mRNA and protein expression of caveolin-1 in PC-3 cells treated with simvastatin (10, 20 or 40μmol/L) for 48 hours were significantly decreased by RT-PCR and western blot.Conclusions: (1) Simvastatin can significantly inhibit the proliferation of PC-3 cells.(2) Simvastatin can induce apoptosis of PC-3 cells.(3) The effects of Simvastatin on inhibition of proliferation and induction of apoptosis may be related to reduction of the expression of caveolin-1 in PC-3 cells..(4) Inhibitors of the enzyme HMG-CoA reductase provide a novel theory and method in the prevention and therapy for prostate cancer clinically.