Objective To investigate the expression of cyclooxyenase 2 (COX 2)in carcinogensis and development of endometrium carcinoma Methods Immunostainings, westernblotting and quantitve reverse transcription polymerase chain reaction (RT PCR) assay were utilized to measure levels of protein and mRNA expression of COX 2 in following five study groups: 25 cases with proliferative phase, 25 cases with secretory phase,25 cases with endometritis, 23 cases with atypical proliferative phase and 34 cases with endometrium carcinoma Results COX 2 expression of both RNA and protein in patients with endometrial carcinoma was higher significantly than patients with proliferative phase, secretary phase,endometritis,atypical proliferate phase Immunostaining score was 5 46?0 12 vs 3 20?0 18,4 78?0 12,6 10?0 25,8 70?0 93, average absorbent value was 0 75?0 23 vs 0 41?0 45, 0 56?0 31, 1 10?0 56,1 46?0 41;concentration of mRNA [(93?8) vs (65?11),(79?6),(299?11),(493?30) fpg/?g respectively] Successively the expression of COX 2 in atypical proliferation group was higher than normal endometrial and endometritis group The expression in proliferative phase group was higher significantly than secretory phase group ( P

In order to screen the genes differentially expressed in two human prostate cancer cells with different metastasis potentials, suppression subtractive hybridization (SSH) was done twice on human prostate cancer cell line with high potential of metastasis PC3M-1E8 and its synogenetic cell line PC3M-2B4 with low metastasis potential. In the first subtraction PC3M-2B4 was used as tester and PC3M-1E8 as driver and the forward subtractive library was constructed. In the second on the tester and driver were interchanged and the reverse subtractive library was constructed. The screened clones of both libraries were sequenced and Gene Bank homology search was performed. Some clones were confirmed by quantitative real-time PCR. The results showed that two subtractive libraries containing 238 positive clones were constructed. Analysis of 16 sequenced clones randomly picked from two libraries showed that 4 differentially expressed gene fragments were identified as new EST with unknown functions. It was concluded that two subtractive libraries of human prostate cancer cell lines with different metastasis potentials were constructed successfully.
Cell Line, Tumor ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Gene Library ; Neoplasm Metastasis/*genetics ; Nucleic Acid Hybridization/*methods ; Prostatic Neoplasms/*genetics ; Prostatic Neoplasms/pathology

Background and purpose:Previous studies have shown that Bubl was a critical component of the spindle checkpoint.Paclitaxel sensitivity was considered to be dependent on the functionality of this spindle checkpoint.This study investigated the effects of pEGFP-Bubl-shRNA plasmid stably transfected into the cell cycle and its sensitivity in human ovarian cancer cell line A2780.Methods:After the pEGFP-Bubl-shRNA plasmid and empty plasmids were constructed.they were transfected into A2780 cells by the Lipofectamine 2000~(TM).The nontransfected cells were the control.RT-PCR and Western blotting were used to determine the target gene and protein expression.The rate of proliferation inhibition was tested by an MTT assay,apoptosis and cell cycles were determined by flow cytometry,and the mitotic index was determined bv Hoechst33342 dye.Results:RT-PCR and Western blotting showed that pEGFP-Bubl-shRNA/A2780 group displayed a low expression of Bubl compared to the A2780 and pEGFP-Cl/A2780 group(P＜0.05).The sensitivity of the pEGFP-Bubl-shRNA/A2780 group was significantly lower than the non-transfccted and pEGFP-Cl/A2780 cells(P＜0.05).Flow cytometry revealed that the rates of G2/M phase and apoptosis were significantly lower in the pEGFP-Bubl-shRNA/A2780 group than those in the control group (P＜0.05).Conclusion:Bubl plays an important role in the paclitaxel treatment.A down-regulation of Bubl could reduce the drug sensitivity and rate of G_2/M cells in human ovarian cancer cell line A2780.

In order to investigate the roles of MTA2 in the pathogenesis of ovarian epithelial cancer, the expression of MTA2 in 4 ovarian cell lines were detected by semi-quantitative RT-PCR and Western-blot assays. MTA2 expression in normal, borderline, benign and malignant epithelial ovarian tissues was immunohistochemically examined. The expression of MTA2 mRNA and protein was detected in all of 4 cell lines of ovarian epithelial cancer. The expression of MTA2 mRNA and protein was higher in strong migration cell lines than in weak migration ones. In borderline and malignant ovarian tissues tested, MTA2 staining was dramatically stronger than in normal and benign tissues (P < 0.01). The expression levels in malignant ovarian tissues were significantly higher than that in borderline epithelial ovarian tissues (P < 0.01). The expression of MTA2 was correlated with clinical stage, histopathological grade and lymph node metastasis. It was concluded that the high expression of MTA2 was associated with more aggressive behaviors of epithelial ovarian cancer. MTA2 provides a novel indicator of ovarian cancer.

Objective To analyze the gonadal hormone and seminal plasma of patients with abnormal semen liquefaction and investigate the influence mechanism in order to provide guidance for the diagnosis and treatment. Methods 152 men of childbearing age were divided into two groups according to the liquefaction time (cut?off point: 60 minutes). Routine semen parameters,gonadal hormone and seminal plasma were tested and compared between the above groups. T?test was applied to compare individual gland function (pH value,neutralα?glycosidase,fructopyranose,seminal plasma zinc and citric acid) and gonadal hormone (FSH,LH,PRL,T and E2). Logistic regression analysis was adopted to probe the influencing factors for abnormal semen liquefaction. Results Seminal pH value (7.47 ± 0.13 vs. 7.32 ± 0.18),citric acid(51.12 ± 12.95 vs. 83.11 ± 33.46)and FSH (4.40 ± 1.03 vs. 4.85 ± 1.50)levels were significant different between the two groups (P < 0.05),but the other indexes showed no significant difference. Correlation regression analysis showed that semen liquefaction capacity has correlative relationship with seminal plasma fructose (OR=2.644),citric acid (OR=0.922),serum T (OR=1.029) and E2,while no correlative relationship with other indexes. Conclusions Correlation between two glands (seminal vesicle and prostate) and balance in the two hormones (T and E2) influence the liquefaction time. Specific causes should be distinguished before diagnosis.

Objective To compare the effect of kidney-tonifying blood-activating recipe (KBR) and Aescuven Forte Tablets ( AFT) in improving the sperm quality of varicocele-induced male sterility, thus to optimize the therapeutic therapy for varicocele-induced male sterility. Methods A total of 102 varicocele-induced male sterility with abnormal sperm parameters after conservative treatment were randomized into KBR group (N=53) and AFT group ( N=49) . KBR group was given KBR plus natural vitamin E and AFT group was given AFT plus natural vitamin E, and the treatment lasted for 8 continuous weeks. Before and after treatment, the quality of seminal fluid was analyzed, sperm quantization parameters such as total number of sperm (TNS) , total number of progressive motility sperm ( TNPS) , total number of normal form sperm ( TNNS) and total number of nor mal form and progressive motility sperm ( TNNPS) were observed, and the improvement rate of sperm quantization parameter was compared. Results (1) Before treatment, the differences of TNS, TNPS, TNNS and TNNPS were insignificant between the two groups ( P>0.05) . After treatment, TNNS was not improved in AFT group ( P>0.05) , but TNS, TNPS, TNNPS were much improved in both groups ( P<0.01 compared with those before treatment) . The improvement of KBR group was superior to that of AFT group ( P<0.05) . ( 2) The improvement rate for TNS, TNPS, TNNS, TNNPS was 90.57%, 79.25%, 67.92%, 77.36%in KBR group, and was 75.51%, 73.47%, 28.57%, 61.22% in AFT group respectively. The improvement rate for TNS and TNNS in KBR group was superior to that in AFT group ( P<0.05 or P<0.01) . Conclusion Varicocele-induced male sterility patients usually have the syndrome of kidney deficiency and blood stasis, so KBR, which has the function of tonifying kidney and activating blood, has synergistic action on the effect of AFT in improving sperm quality of varicocele-induced male sterility patients.

Full-length coxsackie adenovirus receptor (CAR) eukaryotic expression plasmid was transfected into an ovarian cell line, SKOV3, and its effect on the change of malignant metastasis phenotype was explored. CAR mRNA and protein expression levels among 4 ovarian cancer cell lines (A2780, SKOV3, SW626, CAOV3) and the positive control 293 (a transformed human embryo kidney cell line) was detected by using semi-quantitative RT-RCR and Western blot and compared. CAR-negative SKOV3 was transfected with the eukaryotic expression plasmid containing a full-length CAR cDNA and mock-vector respectively. The positive clones were screened by G418. The biological behavior changes of positive transfected cells were gauged by colony formation in soft agar assay and cell adhesion assay. Among the cell lines, there were obviously different CAR expression levels. CAR could not be detected in SKOV3. In transfected cell group, CAR expression was enhanced obviously as compared with non-transfected or mock-transfected groups. Cell adhesion in the transfected group was promoted. The number of colony formation was reduced significantly in transfected groups (25.32 +/- 8.91) as compared with that in non-transfected group (88.75 +/- 13. 98) and mock-transfected group (82.53 +/- 19.37). Among the 4 ovarian cancer cell lines, CAR expression level was variable. Exogenous CAR expression had a potential role in inhibiting the malignant metastasis phenotype of ovary cancer cells.
Cell Line, Tumor ; Lymphatic Metastasis ; Neoplasm Invasiveness ; Ovarian Neoplasms/metabolism ; Ovarian Neoplasms/*pathology ; Phenotype ; Receptors, Virus/*biosynthesis ; Receptors, Virus/genetics ; Transfection

In this article, the status of spindle assembly checkpoint and the alteration of its major component, Mad2 protein level were examined in A2780 and SKOV3 ovarian cancer cell lines. Recombinant eukaryotic expression plasmid pEGFP-Mad2 was transfected into paclitaxel-resistant SKOV3 cells and Mad2 protein was knocked down by Mad2-specific siRNA in paclitaxel-sensitive A2780 cells. Then the expression level of Mad2 gene was detected by Western blotting. Flow cytometry revealed that SKOV3 cells were not fully arrested in G(2)/M phase in contrast to A2780 cells in the presence of paclitaxel. However, paclitaxel sensitivity assay showed that sensitivity to paclitaxel was reversed after the transfection in both cell lines in terms of number of cells arrested at G(2)/M phase and the expression of Bcl-2 was significantly changed. These results suggest that weakened spindle checkpoint with reduced expression of Mad2 is associated with resistance to paclitaxel in ovarian cells and Bcl-2 may be involved in this process.

Chemotherapy is the preferred therapeutic approach for advanced ovarian cancer, but a successful long-term treatment is prevented by the development of drug resistance. Recent works have underlined the involvement of non-coding RNAs, microRNAs (miRNAs) in cancer development, with several conjectures regarding their possible involvement in the evolution of drug resistance. This study is to investigate the promoting effects and mechanism of miR-125b involved in the development of chemoresistance in ovarian cancer. The different expression of miR-125b in cisplatin-sensitive ovarian cancer cell line (OV2008) and its resistant variant (C13*) was identified by real-time PCR. An in vitro cytotoxicity assay and apoptosis assay using CCK-8 assay and flow cytometry, were carried out to detect the effect of miR-125b and Bak1 on cisplatin resistance of cells. Real-time PCR, Western blotting and luciferase reporter assay were used to detect whether Bak1 is a target of miR-125b. As compared with OV2008 cells, the expression levels of miR-125b in C13* cells were increased. It was found that the up-regulation of microRNA-125b caused a marked inhibition of cisplatin-induced cytotoxicity and apoptosis and a subsequent increase in the resistance to cisplatin in OV2008 and C13* cells. Moreover, Bak1 was a direct target of miR-125b, and down-regulation of Bak1 suppressed cisplatin-induced apoptosis and led to an increased resistance to cisplatin. Our study indicates that miR-125b has a significantly promoting effect on chemoresistance of C13* cells and up-regulation of miR-125b expression contributes to cisplatin resistance through suppression of Bak1 expression. This finding has important implications in the development of targeted therapeutics for overcoming cisplatin resistance in ovarian cancer.

In order to investigate the role of the PTEN expression in carcinogenesis and development of endometrial carcinoma and clarify whether and how PTEN and PI3K/Akt pathway relate to endometrial carcinoma, the expression of PTEN and phospho-Akt was detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) methods and Western-blot from 24 cases of endometrial carcinoma, 10 cases of endometrial atypical hyperplasia, 10 cases of endometrial hyperplasia, and 10 cases of normal endometrium. SP immunohistochemical methods were used to measure levels of PTEN protein expression in following 5 study groups: 31 cases of endometrium in proliferative phase, 30 cases of endometrium in secretory phase, 71 cases of endometrial hyperplasia, 25 cases of atypical hyperplasia and 73 cases of endometrial carcinoma. Immunostaining score of PTEN was 3.39+/-0.15 in proliferative phase, 1.90+/-0.21 in secretory phase, 3.34+/-0.29 in endometrial hyperplasia, 0.62+/-0.11 in atypical hyperplasia, and 0.74+/-0.19 in endometrial carcinoma, respectively. PTEN mRNA relative value in normal endometrium, endometrial hyperplasia, endometrial atypical hyperplasia, and endometrial carcinoma was 2.45+/-0.51, 2.32+/-0.32, 0.46+/-0.11, and 0.35+/-0.13 respectively. The expression levels of PTEN mRNA and protein in patients with endometrial carcinoma and atypical hyperplasia were significantly lower than in those of proliferative phase and with endometrial hyperplasia. The level of PTEN expression in patients with endometrial carcinoma was significantly related to tissue type (P<0.005), differentiation (P<0.05) and clinical stage (P<0.05), but not to depth of myometrium invasion (P>0.05). Western blot analysis revealed that Phospho-Akt level in PTEN negative cases was significantly higher, and there was a negative correlation between PTEN and phospho-Akt (r=-0.8973, P<0.0001). It was suggested that loss of PTEN expression was an early event in endometrial tumorigenesis. The phosphorylation of Akt induced by the loss of PTEN took part in the tumorigenesis and development of endometrial carcinoma.