The Telomerase Activity, Expression And Modulation Of HTERT Gene, C-myc Gene In Human Ovarian Tumor Cells

Telomeres are specialized structures of the distal end of chromosomes and function in chromosomes protection positioning, and replication .In human , telomeres consist of tandem repeats of the sequence TTAGGG that spans up to 15 K bases and associated proteins. Shortening of telomeres occurs with each cell division and eventually results in cell death. Telomerase is a ribonucleoprotion that synthesizes telomeric DNA onto chromosomal ends using a segment of its RNA component as template. The telomerase , therefor counteracts the mitotic clock of telomere shortening. Telomerase activity can be measured in vitro by a primer extension assay in which telomerase synthesize telomeric repeats onto olignucleotide primers. Using this method ,telomerase activity has been measured in a variety of tissues and cells. To date ,most cancer cells, immortal cells have been shown telomerase activity. The telomerase activity of female ovarian tumors tissus has not been investigate well. We hence studied the telomerase activity and expression of hTERT gene and c-myc gene for human ovarian malignant tumors.MATERIAL AND METHODSA total of 73 tissue samples and HO8910 cell, COQ cell were studied. Lesions were collected from in 2000-2001. These tissues were detect telomerase activity by PCR-ELISA underlines TRAP ( teromeric repeated sequences amplification protocol ) including 23 malignant tumors and its adjacent tissues, 10 borderline or low malignant potential (LMP), 28 benign ovarian tumor tissues, 12 normal ovarian. Absorbance values are reported as the A45onm reading against blank(reference wavelength A655nm). RT-PCR was used to detect all the lesion for the expression of the hTERT gene mRNA and the c-myc gene mRNA. We analyzed the relationship between all of them in ovarian malignant tumors. Detect the telomerase activity expression of hTERT mRNA and c-myc mRNA in the cells cultured with CDDP treatmentRESULTS1) It was higher in 21 of 23 malignant (91.3%), 4 of 23 adjacent (17.3%), 2 of 10 borderline (20%), 2 of 28 benign tissues(7.1%).None of normal ovarian was detect telomerase activity. The levels of telomease activity in serious subtypes, poorly differentiated , FIGO stage III and IV tend to be higher than those in others tumors(P<0.05).There were no significant correlation among the general positive rate of telomerase activity ( P>0.05 ) in histological subtypes and histological grades.2) The expression of hTERT mRNA and c-myc mRNA has analyzed by semi-quantitative amplification . In total , 20 of 73 samples examined showed detectable levels of hTERT, and 12 Of 73 samples showed detectable levels of c-myc. Both the hTERT positive and the c-myc positive lesion showed telomerase activation ,which is higher than negative ones (P<0.05).3)Detect the telomerase activity ,expression of hTERT mRNA and c-myc mRNA in the cells cultured with CDDP treatment . 24h later, both the hTERT mRNA and c-myc mRNA start decreasing .After 96h the hTERT message was almost completely disappeared. Before this observation , the c-myc gene mRNA has been disappeared at 72h. But the telomerase did not decrease untill 36h, and disappeared in 120h.ConclusionThe expression of telomerase may play a major role in ovarian malignant tumors. The detection of telomerase activity is useful in predicting prognosis of patients with malignant ovarian tumors.. Expression of hTERT gene is rate-limiting on the activation of telomerase which attribute the progression of ovarian malignant tumors. Telomerase activation is regulated by the hTERT gene, and c-myc gene may cooperate on the regulation.