Telomeric Repeat-Containing RNA (TERRA) Related To Polycystic Ovary Syndrome (PCOS)

Chapter 1 Telomeric repeat-containing RNA in Leukocyte and Ovarian granulose cellObjective:To test the expression of telomeric repeat-containing RNA(TERRA)in peripheral blood leukocytes and Ovarian granulose cell.Method:Leukocyte was isolated from peripheral blood leukocyte from patients with PCOS and heath women.Ovarian granulose cell was isolated from patients with PCOS,RNA-FISH was performed to detect the expression of TERRA.Results:We performed RNA-FISH on peripheral blood leukocytes of both groups using the strand-specific PNA probe(CCCTAA)3-cy3.The results revealed that TERRA was expressed in the human peripheral blood leukocytes of the controls and PCOS patients,and it was distributed in the nucleus.The TERRA signal was abolished after culture with RNase A.Same was founded in ovarian granulose cell.Conclusion:TERRA was detected in peripheral blood leukocyte and ovarian granulose cell.This is the first report on TERRA expression in peripheral blood leukocyte.Chapter 2 Telomeric Repeat-Containing RNA(TERRA)Related To PCOSObjective:To evaluate the role of TERRA on PCOS and relationship between TERRA and peripheral blood leukocyte telomere length(LTL).Method:Real time PCR technique to measure the mean LTL and TERRA expression in PCOS and controls.Cobas 6000 and Abbott 1000 were used to detect clinical parameters.logistic regression and multiple linear regression were used to analyze the association between several related clinical parameters and the age-adjusted ratio of the telomere repeat length(T/S ratio)in PCOS and controls.Results:The body mass index(BMI),LH,TTE,DHEAS,FINS,and FGP were all significantly higher in the PCOS group than in the control group(p<0.01).There were no significant differences in the FSH,E2,P,and PRL levels between the two groups.The age-adjusted LTLs significantly decreased with age in both the groups.The age-adjusted LTLs were significantly longer in the PCOS group than in the control group(2.74±0.19 vs 2.54±0.13;p<0.01).The age-adjusted TERRA level was significantly lower in the PCOS group than in the control group(1.03±0.58 vs 1.67±0,46,p<0.01).The testosterone(TTE)was related positively to LTL and negatively to TERRA in the PCOS group(r = 0.532,p = 0.002;r =-0.477,p = 0.017).Conclusion:This is the first report on the relationship between TERRA and TTE in human peripheral blood leukocytes.These findings indicate that TERRA and TTE play an important role in the LTL regulation in PCOS.TERRA and TTE may be potential targets to regulate LTL in PCOS.Chapter 3 Role of regulation of androgens on TERRA and telomerase activityObjective:To explore the role of regulation of TEREA and testosterone on telomerase activity.Method:Ovarian granulose cells were cultured in plates in complete medium.After 24h,0.1-100nmol/L testosterone was added,cells were incubated for 48h..Human PCR-ELISAT was used to detect telomerase activity.Real time PCR was used to evaluate hTERT mRNA and TERRA levels.Westernblot was used to detect hTERT protein expression.MTT was used to detect cellular activity and Annexin V-FITC/PI was used to assess cellular apoptosis.Results:Testosterone(100nmoll/L)inhibited cellular viability of ovarian granule cells,P<0.01;Testosterone induced cellular apoptosis of ovarian granule cells,With the increase of testosterone concentration(from 0.1 to 100nmol/L),cellular apoptosis rate increased,from 9.2%±0.05 to 32.1%.± 0.02,P＜0.01.Apoptosis rate was higher in testosterone groups than in control,P<0.01.Testosterone(0.1 nmol/L)increased hTERT mRNA expression(P<0.05)and testosterone(1nmol/L,10nmol/L,100nmol/L)inhibit TERT mRNA expression.(P<0.01).hTERT protein was found as same as hTERT mRNA.Testosterone(0.1 nmol/L,lnmol/L)induced telomerase activity(P<0.05).Conclusion:Testosterone induced the protein expression of telomerase(hTERT)and hTERT mRNA.Testosterone decreased TERRA levels.Testosterone and TERRA play an important role in regulation of telomerase activity.