Preliminary Studies On The Expression And Mechanism Of Nucleostemin In Prostate Cancer

Prostate cancer is the most common primary malignant tumor among malepopulation in the European and American countries. In China, the incidence andmortality rate have increased significantly due to population aging, changes in lifestyles and other causes. Prostate cancer has become a world-wide health problemwhich is not so sensitive to radiotherapy and chemotherapy, and many cases willbecome hormone-refractory prostate cancer when treated with hormone deprivation.Therefore, the gene therapy is a promising method, especially for the patients withlocal or systemic metastasis. The gene therapy for the treatment of prostate cancer,which targets the virulence genes of prostate cancer to cure this diseasefundamentally, has become the focus of research and many Scientists are nowexploring the new target genes.Recently, a p53-binding protein, named nucleostemin (NS) exclusively expressedin the stem cells and cancer cells but not in differentiated adult tissues and cells, hasbeen shown to be essential for the proliferation and self-renewal of stem and cancercells. NS may be the common controlling gene of stem and cancer cells. Tumor stemcells (TSCs) which are thought to be related to the carcinogenesis of prostate cancerhave been identified in prostate cancer. Therefore, studies on the function andpathway of NS are of great importance to the diagnosis and treatment of prostatecancer. However, it is currently unknown whether NS is expressed in prostate cancerand how this newly identified molecule involved in prostate cancer pathogenesis. Thepresent study will detect the expression level of NS in prostate cancer andinvestigate its mechanism by RNA interference and DNA microarray technology. Partâ… Detection of NS expression in prostate cancer tissuesand cell linesObjective To reveal the NS expression level in prostate cancer tissues and celllines and investigate the correlation between NS gene level and clinical variables ofprostate cancer, taking this part as a base for later study. Methods RT-PCR,western blot and immunohistochemistry were used to detect the NS mRNA andprotein level in prostate cancer tissues and PC-3, LNCap and DU 145 cells, analyzingthe correlation between NS protein level and clinical variables of prostate cancer.Results NS gene was expressed in all the detected samples and cell lines, the NSmRNA and protein level in prostate cancer tissues were significantly higher thanthose in BPH tissues. NS protein level was negatively correlated with the degree ofcell differentiation in prostate cancer tissues, the worse the differentiation, the higherthe NS protein level. Conclusion NS gene is highly expressed in prostate cancerand may be of great importance to the adverse differentiation and malignantproliferation of prostate cancer.Partâ…¡The effect on the proliferation of PC-3 cells afterdown-regulating the NS gene levelObjective To investigate the relationship between the NS gene and themalignant proliferation of prostate cancer and explore the new target gene for prostatecancer gene therapy. Methods NS-specific short-hairpin RNA (shRNA) expressionplasmid was used to transfect PC-3 cells. The changes of cell morphology, cell cycle,proliferation ability and apoptosis were studied after down-regulating the NS genelevel. Results The cells became larger and showed more pseudopodia, having atendency to differentiate, the detection of cell cycle showed a decrease of S stage andan increase of G1 stare, cell proliferation ability in vitro and tumorigenesis ability in nude mice were discounted, the final tumor volume and weight were also decreasedafter knocking down NS gene. Conclusion NS may act as an important G1/Sregulator to regulate the malignant proliferation of prostate cancer PC-3 cells. NSmay serve as an ideal therapeutic gene target for prostate cancer.Partâ…¢Gene profiling after knocking-down the expression of NSgene in PC-3 cellsObjective To screen the genes and possible signal transduction pathways withwhich NS interact and explore the mechanism of NS in pro.state cancer. MethodsOligonucleotide DNA microarray was used to screen the genome changes afterknocking down expression of NS in PC-3 cells and real-time quantitative PCR wasused to further confirm the important differentially expressed genes. Results 219differentially expressed genes were found and theses genes were involved in cellcycle, cell proliferation, signal transduction, cell apoptosis and cell differentiation.Some important differentially expressed genes were further Certified using real-timequantitative PCR. INK4 family genes (P15, P16, P18) were up-regulated and cyclinD1, HDAC1 were down-regulated, the main action points were CDK4/6-Cyclin Dand pRb-E2F1 complexes. Conclusion NS is an important G1/S checkpointregulator and its regulatory activity is certified at the gene level. Many tumor-relatedgenes are found involving in the NS-regulating pathways. However, the present studystill can not completely reveal the NS signal pathways and more studies are needed tofurther discover and confirm other mechanism.