Inhibitory Effects Of Bioactive Compounds Isolated From Oiltea Camellia On Human Prostate Epithelial Cell Line And Prostate Cancer Cells In Vitro

The histological definition of benign prostatic hyperplasia(BPH)refers to an overgrowth and increased proliferation of the epithelial and stromal components of the transition zone,and of the peri-urethral area of the prostatic gland.The molecular mechanisms of BPH pathogenesis remain poorly understood.Previous studies indicated that Oiltea Camellia Extract(OCE)possess anticarcinogenic properties.The purpose of this study was to investigate the major bioactive compounds and anti-tumorigenic effects of OCE on benign prostate hyperplasia and prostate cancer.Bioactive compounds were isolated from polyamide fraction Fr8 of OCE,which showed the strongest 5a-reductase inhibitory activity.BPH-1 and PC-3 cell lines were used to represent non-tumorigenic hyperplasia and malignant prostate cancer,respectively.Real time fluorescence quantitative PCR assay was used to further investigate the regulatory mechanisms underlying BPH.Cell migration assay was introduced to test the migration ability of PC-3 cells 36 hours after being treated with bioactive compounds.The main results were obtained as follows:(1)Bioactive compounds was isolated from Fr8 by solid phase extraction(SPE)and preparative high-performance liquid chromatography(preparative HPLC),three of them were demonstrated to have growth inhibitory activity on BPH-1 cells.The chemical structures of those compounds were identified as 3-O-galloyl-4,6-[(S)-hexahydroxydiphenoyl]-?/?-D-glucopyranose(gemin D),3,4,6-tri-O-galloyl-?i?-D-glucose(GAG)and valoneic acid dilactone(VAD)by Mass Spectrometer(MS)and Nuclear magnetic resonance(NMR)analyses.(2)Pro-apoptotic actions of gemin D,GAG and VAD on BPH-1 cells were determined by Annexin V staining at 48 h treatment.Range from 25 to 200 ?g/mL,VAD showed stronger pro-apoptotic effect on BPH-1 cells than gemin D,and gemin D showed stronger than GAG at the same concentration.(3)Real time fluorescence quantitative PCR assay was performed to indicate how genes related in BPH pathogenesis changed treated with bioactive compounds.The expression of proto-oncogene C-MYC was significantly down-regulated indicated the inhibition of cell proliferation in gemin D,GAG,VAD treated BPH-1 cells.PTGS2(encoding the protein COX-2)and ALOX5(encoding the protein 5-LOX)were significantly down-regulated in treated BPH-1 cells by bioactive compounds,indicated that gemin D,GAG and VAD had anti-inflammatory effect on BPH-1 cells.(4)In PC-3 cells,bioactive compounds separatd from OCE,showed anti-proliferative and pro-apoptotic effects.MTT assay suggesting the IC50 value of gemin D,GAG and VAD decreased cell viability were 117.71 ?mol/L,140.69?mol/L,132.44 ?mol/L at 48 h treatment.Range from 25 to 200 ?g/mL,VAD showed stronger pro-apoptotic effect on PC-3 cells than gemin D,and gemin D showed stronger than GAG at the same concentration.At the concentration of 200 ?g/mL,pro-apoptotic effect of VAD on PC-3 cells was 35.8%.Cell migration assay indicated gemin D,GAG and VAD can significantly inhibit cell migration.This work suggested that gemin D,GAG and VAD,were potentially used as chemopreventive agents against prostate hyperplasia and prostate carcinogenesis via induction of apoptosis and down-regulation expression of genes belonging to immune response,cell proliferation pathways in BPH-1 cells.