Therapeutic effect of lentivirus-mediated miR-21 on chemotherapy-induced premature ovarian failure in rats / 解放军医学杂志

Objective To explore the therapeutic potential of miR-21 in rat model of chemotherapy-induced premature ovarian failure (POF).Methods Lentivirus-mediated miR-21 (LV-miR-21) was constructed successfully in vitro with molecular biology methods.Rats were divided into 4 groups named control group,model group,blank vector group and miR-21 group.Rat models of chem0therapy-induced POF were established in the latter 3 groups by intraperitoneal injection ofcytoxan (CTX).Bilateral ovaries of rats in miR-21 group were injected with LV-miR-21,of rats in blank vector group were injected with lentivirus vector,and rats in model group received no treatment.At 1,15,30,45 and 60 days after the last injection,blood sample was collected,the rats were then sacrificed and the ovaries were removed.The estrous cycle was observed by vaginal smears.The E2 level was detected by chemiluminescent immunoassay,and the follicle stimulating hormone (FSH) level was detected by homologous double antibody radiation immunoassay.Ovary weights were measured,and the follicle count was conducted through observing paraffin section under microscope.The apoptosis of ovarian granulosa cells was analyzed by TUNEL assay.Results During 15-30 days,30-45 days and 45-60 days after the last injection,regular estrous cycle was recovered respectively in 8,5 and 3 rats in miR-21 group.At the 15th,30th,45th and 60th day after the last injection,the E2 level was higher in miR-21 group than in model group and blank vector group,but the FSH level showed the opposite trend (P=0.000).At the 45th and 60th day after the last injection,the follicle numbers at all stages increased markedly in miR-21 group than in model group and blank vector group (P=0.000).At the 30th,45th and 60th day after the last injection,the ovary weights were higher in miR-21 group than in model group and blank vector group.At the 15th,30th,45th and 60th day after the last injection,the apoptosis rate of ovarian granulosa cell were significantly lower in miR-21 group than in model group and blank vector group (P=0.000).Conclusion Up-regulation of miR-21 expression may partly recover the ovarian structure and function damaged by CTX.

Inhibitory effect of short hairpin RNA targeting survivin gene on human ectopic endometrial cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct a lentiviral expression vector for short hairpin RNA (shRNA) of human survivin gene, and assess its gene silencing effect in human ectopic endometrial cells.</p><p><b>METHODS</b>Human survivin gene shRNA sequence was designed using a software available on-line. The synthesized shRNA sequence was cloned into the pGCL-GFP vector to construct LV-survivin shRNA, which was confirmed by PCR and DNA sequence analysis. The packaging 293T cells were cotransfected with LV-survivin shRNA, pHelper 1.0 and pHelper 2.0, and the titer of the lentivirus was determined. The recombinant lentivirus was injected into human ectopic endometrial cells and the survivin mRNA expression was examined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in comparison with that in the non-transfected and blank vector-transfected human ectopic endometrial cells.</p><p><b>RESULTS</b>PCR analysis and DNA sequencing confirmed correct insertion of the shRNA sequence into the lentiviral vector. The titer of virus after packaging was 8x10(8) U/ml. Survivin mRNA expression in human ectopic endometrial cells transfected by LV-survivin shRNA was significantly inhibited compared with those in the non-transfected and empty vector transfected human ectopic endometrial cells (P<0.01), and no significant difference was found between the latter two groups.</p><p><b>CONCLUSION</b>The lentiviral shRNA vector of survivin gene constructed can effectively inhibit the expression of survivin gene in human ectopic endometrial cells in vitro. This vector provides a tool for investigating the role of survivin gene in the occurrence and progression of endometriosis and for searching new therapeutic targets.</p>

Expression of hypoxia-inducible factor-1alpha in endometriosis / 南方医科大学学报

<p><b>OBJECTIVE</b>To detect the expression of hypoxia-inducible factor-1alpha(HIF-1alpha) in endometriosis and explore the possible role of HIF-1alpha in the pathogenesis of endometriosis.</p><p><b>METHODS</b>Immunohistochemistry was performed to examine the expression of HIF-1alpha in 20 normal endometrium, 20 ectopic endometrium and 68 eutopic endometrium specimens from 68 endometriosis patients, and the results were analyzed statistically.</p><p><b>RESULTS</b>The expression of HIF-1alpha was significantly increased in ectopic endometrium than in normal endometrium (P<0.01), and the expression did not undergo changes with the normal menstrual cycle in the three types of endometrium.</p><p><b>CONCLUSION</b>HIF-1alpha expression increases in ectopic endometrium, suggesting that HIF-1alpha plays an important role in the pathogenesis of endometriosis.</p>

Evaluation of tumor necrosis factor-alpha and tumor necrosis factor-beta in serum of patients with endometriosis / 中南大学学报(医学版)

OBJECTIVE@#To evaluate the levels of tumor necrosis factor-alpha (TNF-alpha) and tumor necrosis factor-beta (TNF-beta) before and after conservative laparoscopic surgery in patients with endometriosis.@*METHODS@#The levels of TNF-alpha and TNF-beta in the serum of both 82 patients with EMS and 68 controls were determined by ELISA.@*RESULTS@#The levels of TNF-alpha and TNF-beta in the serum of patients with EMS were significantly higher than those of the controls (P < 0.01), and they increased with the clinical terms ( P < 0.05). After clearance of endometrosis foci with laparoscopic conservative surgery, the TNF-alpha levels decreased significantly in EMS III - IV, and TNF-beta levels decreased significantly in EMS I - IV.@*CONCLUSION@#Measuring TNF-alpha and TNF-beta levels in the serum of patients with EMS may have important value in postoperative follow-up, surveillance and evaluation of the effectiveness of the surgery.

Association between gene mutation of cytochrome P450 1A1 in exon 7 A4889G locus and susceptibility to endometriosis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To assess the possible association between gene mutation of cytochrome P450 1A1(CYP1A1) in exon 7 A4889G locus and the susceptibility to endometriosis (EM).</p><p><b>METHODS</b>Allele specific-polymerase chain reaction method was used to analyze gene mutation in exon 7 A4889G locus of CYP1A1 in 76 patients with endometriosis and 80 healthy controls.</p><p><b>RESULTS</b>The frequency of allele G on A4889G locus of CYP1A1 gene showed a significant difference between the study cohort and the control group (Chi2=7.498, P<0.01), with an odds ratio of 1.957. Statistically significant difference in the frequencies of genotypes AA, AG and GG was observed between the two groups (Chi2=6.915, P<0.05). Individuals with homozygotes for G allele were at higher risk of suffering from EM when compared against those with homozygotes for A allele, the odds ratio being 3.437 (Chi2=5.430, P<0.05).</p><p><b>CONCLUSION</b>The above results suggest that gene mutation of CYP1A1 in exon 7 A4889G locus might be a genetic susceptible factor of endometriosis. The mutation allele of CYP1A1 gene appears to increase the risk of endometriosis.</p>