A Study Of The Relationship Between Intron 9 In CD44 Gene Transcripts And Ovarian Tumor

PARTⅠA STUDY OF THE RELATIONSHIP BETWEEN INTRON 9 IN CD44 GENE TRANSCRIPTS AND OVARIAN TUMOROBJECTIVE:To investigate the relationship between intron 9 in CD44 gene transcripts and ovarian tumor and its clinical significance.METHODS:Semi-quantitatively reverse transcription-polymerase chain reaction(RT-PCR) was used to analyze the retention rate and relative content of intron 9 in CD44 gene transcripts in 54 cases of malignant ovarian cancer,8 of ovarian borderline tumors,41 of benign ovarian tumors and 20 of normal ovary.RESULTS:The retention of intron 9 in CD44 gene transcripts was not found in normal ovarian tissue.The retention rates and relative contents of intron 9 in ovarian benign,borderline,and malignant tumor were 19.51%,7/8,92.59%and 7.86±2.35%, 88.98±3.21%,91.54±6.25%,respectively.The retention of intron 9 in ovarian borderline and malignant tumors were significantly higher than that in benign tumors (P＜0.01).There was similar retention of intron 9 in different histological types and clinical stages of ovarian tumors.No correlation was found between the clinical stages or histological types of ovarian tumors and the retention of intron 9(P＞0.05).CONCLUSION:The retention of intron 9 is usual in borderline and malignant ovarian tumors.Detection of retention of intron 9 may be a molecular diagnostic tool for ovarian cancer. PARTⅡINHIBITION OF THE EXPRESSION OF CD44 GENE BY SMALL INTERFERENCE RNA IN OVARIAN CANCER CELL LINE OVCAR-3OBJECTIVE:Since ovarian cancer cells express CD44 that cause very strong cell adhesion to peritoneal mesothelium and cause an unfavorable prognosis,we designed siRNA targeting CD44 gene to analyze the functional consequences of this inhibition in human ovarian cancer.METHODS:One pair of 21bp reverse repeated motifs of CD44 target sequence with 9 spacer were synthesized and inserted into plasmid Psilencer2.1 to generate siRNA eukaryotic expression vector.After stable transfection into OVCAR-3 cells, the relative contents of intron 9,the CD44v6 mRNA and protein expression inhibition was detected on mRNA level was detected by RT-PCR and on protein level was by immunohistochemical method.RESULTS:The recombinant plasmid P Silencer(+)2.1 CD44 was successfully constructed and transfected into OVCAR-3 cell.By use of RT-PCR and immunohistochemical method,the relative contents of intron 9,CD44v6 mRNA and protein expression were significantly inhibited(All P＜0.01).CONCOLUSIONS:The recombinant plasmid P Silencer(+)2.1 CD44 was successfully constructed and transfected into OVCAR-3.By use of RT-PCR and immunohistochemical method,the relative contents of intron 9,CD44v6 mRNA and protein expression were significantly inhibited(All P＜0.01).