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Objective To discuss the clinical effect of neoadjuvant chemotherapy combined tumor cells to destroy the loss in treatment of patients with advanced ovarian cancer.Methods One hundred and forty-four patients with advanced ovarian cancer were divided randomly into the control group(n=72) and research group (n=72).The patients of control group were given conventional chemotherapy(ovarian tumor remove first and then neoadjuvant chemotherapy) and the patients of research group were given neoadjuvant chemotherapy (neoadjuvant chemotherapy first and then ovarian tumor remove).The operation time, intraoperative blood loss, hospital stay, ideal reduction rate, clinical efficacy and postoperative complications between the two groups were compared.Results The operation time, intraoperative blood loss, hospital stay of the research group were obviously lower than that of the control group((124.6±21.3) min vs.(186.4±32.6) min, (382.5±62.3) ml vs.(618.5± 86.4) ml, (8.9± 1.3) d vs.(12.2± 3.4) d;t =5.623,9.646,5.257), while the ideal reduction rate of the research group were obviously higher than that of the control group(70.8％ vs.47.2％, x2 =8.735), the differences were statistically significant(P＜0.05).The clinical efficacy(87.5％ vs.52.8％, x2 =6.748) of the research group were obviously higher than that of the control group, while the postoperative incision infection (9.7％ vs.19.4％, x2 =4.452) and fever(4.2％ vs.15.3％,x2 =5.536) were obviously lower than that of the control group, the differences were statistically significant (P＜0.05).Conclusion The treatment of neoadjuvant chemotherapy can obviously increase the the clinical effect of treatment of patients with advanced ovarian cancer and decrease the postoperative complications, it is worth popularization and application.
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Objective To Investigate the value of HPV genotyping in diagnosis of cervical intraepithelial neoplasia(CIN).Methods From July 2012 to February 2013,200 women from Tianjin Medical University General Hospital and 244 women from Affiliated Hospital of the Chinese People's Armed Forces Logistics College were selected to be HPV genotyping test and thin liqid-based cytology test.Consequently,132 samples were performed colposcopy test and cervical biopsy.Results HPV prevalence was 26.4％ (117/444) in this study.The infection of one type HPV was more common.The top 5 of HPV types were HPV16,58,33,18,and 52.The top 5 of the risk for CIN Ⅱ and above followed HPV16,33,39,52 and 18.There was no significant difference between age and HPV positive rate (x2 =0.948,P ＞ 0.05).Multiple infection and cervical lesions rank correlation analysis(r =0.132,P ＞0.05).For CIN Ⅱ and above disease,cytology positive rate was 90％ (44/49),and HPV positive rate was 96％ (47/49) cytology combine HPV positive rate was 98％ (48/49,x2 =0.063,P ＞ 0.05).Conclusions HPV infection should increasing trends with age.Cytology test and HPV genotyping test had good consistency.The combination of them can improve the sensitivity for high-grade lesions.
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Objective To explore the relationship of signal transduction among human papillomavirus 18 E6 oncoprotein (HPV18E6), signal transducers and activators of transcription 1 (STAT1), protein kinase R( PKR )/α subunit of eukaryotic initiation factor 2 ( eIF2α ), nuclear factor-kappa Bp65 ( NF-κBp65 ), mitogen-activated protein kinase( MAPK)/c-Jun N-terminal kinase(JNK) ,and possible molecular mechanism. Methods Construct two lentiviral vectors which contain shRNA interfering sequence aiming at the targets of HPV18E6 oncogene and NC sequence( HPV18E6-RNAi-LV, NC-GFP-LV), based on the transduction with HPV18E6-RNAi-LV and NC-GFP-LV into HeLa cell to interfere the expression of HPV18E6 oncogene and NC sequence,the expressions of mRNA and protein( including phosphating patem)of HPV18E6, STATI, PKR, eIF2α, NF-κBp65, MAPK, JNK are measured with RT-PCR and Western blot, the difference of proliferation and sensitivity to carboplatin of HeLa cell are determined with Transwell cell methods and MTT among every groups. Results The expression of HPV18E6 oncogene can affect the expression level of mRNA and protein of NF-κBp65 and PKR genes, also affect phosphating levels of phosphating protein p-STAT1, p-PKR and p-eIF2α;the restraining rates of proliferation and sensitivity to carboplatin of HeLa cell are higher in HPV18E6-RNAi-LV group than the other groups( P＜0. 05 or P＜0.01 ). Conclusion HPV18E6 oncoprotein not only reduces the expression of PKR but dephosphorylates p-STAT1, pPKR and p-eIF2α to restrain activation of PKR/eIF2α signal transduction passage, maintain the proliferation and invading ability of HeLa cell and restrain apoptosis. The signal transduction among HPV18E6, MAPK/JNK are not clear.
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Objective:To investigate the effective method to avoid the ovarian hyperstimulation syndrome(OHSS)in infertile patients underwent in vitro fertilization-embryo transfer(IVF-ET)and whole embryo freezing and thawing 2-3 cycles after egg retrieval.Methods:The controlled ovarian hyperstimulation(COH),the number of retrieved oocytes,the number of embryo freezing,and clinical pregnancy rate were compared between 239 patients with the ET cycle(group A)and 164 patients with whole embryo freezing and thawing(group B).Results:The levels of initial dosage of gonadotropins(Gn)and total dosage of Gn were higher in group A compared with those of group B(P ＜ 0.01).The level of estradiol(E2)on day of human chorinonic gonadotrophin(hCG),the number of retrieved oocytes and the number of embryo freezing were lower in group A compared with those of group B(P＜ 0.01).There were no differences in patient age,COH,rate of OHSS and clinical pregnancy rate between group A and group B.Conclusion:Freezing the whole embryos and thawing ET 2-3 cycles after egg retrieval didn't influence the treatment results in infertile patients.
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Objective:To identify the relationship between expression of protein kinase R(PKR), phosphating PKR, EIF2α(p-PKR, p-EIF2α) in PKR→EIF2α signal transduction passage and the grades of cervical lesions, the role in generation and progression of cervical tumor and their effects to prognosis of cervical cancer patients. Methods:The expressions of PKR, p-PKR and p-EIF2α in human cervical cancer tissue of 63 cases, cervical intraepithelial neoplasia(CINⅠ-Ⅲ) of 114 cases and normal cervical epithelium of 15 cases were detected by immunohistochemical technique. Results:With the increase in grades of cervical lesions, the positive-expression rate of PKR increased and significantly correlated with the grades of cervical lesions(P < 0.05). With the increase in grades of cervical lesions, the positive-expression rates of p-PKR and p-EIF2α increased firstly, and then decreased. In cervical cancer group, the positive-expression rate of PKR was much higher than that of p-PKR(P < 0.01). The development and progression was quicker in later clinical stages of cervical cancer than that of earlier clinical stages of cervical cancer (P < 0.01). The development and progression of cervical cancer was quicker in patients with negative-expression of p-PKR and p-EIF2α than that in patients with positive-expression of p-PKR and p-EIF2α(P < 0.05). Conclusion:The positive-expression rate of PKR was correlated with the grades of cervical lesions. There are some factors which can impede PKR and EIF2α to be phosphorylated or make p-PKR and p-EIF2α dephosphorylate in high level cervical lesions, which promotes the development and progression of cervical lesions, worsens the prognosis of cervical cancer.
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Objective To study the therapeutic effect of recombinant adeno-associated virus carrying human endostatin gene therapy on endometriosis in mice model. Methods Recombinant adeno-associated virus vector carrying human endostatin gene and enhanced green fluorescent proteins gene (rAAV2-endostatin-EGFP) was constructed. Endometrium was from 12 patients with leiomyoma undergoing hysterectomy in Second Hospital, Tianjin Medical University between November and December 2008. Endometriosis models of nude mice were established by transplanting human endometrial fragments intooperitoneal surface. After 1 week, those 60 mice were divided into 3 groups: treatment group including 20 mice injected with rAAV2-endostatin-EGFP to ectopic lesion, control group including 20 mice injected with rAAV2-EGFP to ectopic lesion and blank control group including 20 mice injected with phosphate buffered saline (PBS) to the ectopic lesion. At 1, 2 and 3 weeks after treatment, those mice underwent laparotemy to observe the location and size of ectopic lesion in abdominal cavity. The expression of endostain protein, number of gland, microvessel density (MVD) and vascular endothelial growth factor (VEGF) were measured in ectopic lesions. The serum level of estradiol and progesterone were detected in nude mice among every groups. Results (1) All endometriosis of nude mice models were established successfully through peritoneum transplanting. After 1 week's treatment, flat lesion nodes, decreased gland number and narrow and atrophy glandular cavity were observed by light microscope. (2) The endostatin gene was transferred into nude mice successfully and expressed effectively. It was observed that endostatin protein expression was shown with enhanced green fluorescent proteins in ectopic lesion. (3) Glands number of ectopic lesion in rAAV2-endostatin-EGFP group(7.8±1.9,7.0±1.5 and 5.5±1.7) were significantly less than 10.1± 1.7, 10.2±2.0 and 9.8±2.4 in rAAV2-EGFP control group and 10.2±2.2,10.0±2.0 and 9.7±2.2 in PBS control group at 1,2 and 3 weeks after treatment(all P<0.05). Glands number of ectopic lesion in rAAV2-endostatin-EGFP group at 3 weeks was significantly less than those at 1 and 2 weeks after treatment (P<0.05). (4) MVD of ectopic lesion in rAAV2-endostatin-EGFP group (12.2±1.5,11.4±2.1 and 9.0±1.4) was significantly less than those at rAAV2-EGFP control group (16.5±1.7,16.5±1.9 and 16.9±1.9) and PBS control group (16.2±1.6,16.0±1.6 and 16.3±1.7) at 1,2 and 3 weeks after treatment (all P<0.05) . MVD of ectopic lesion in rAAV2-endostatin-EGFP group at 3 weeks was significantly less than those at 1 and 2 weeks after treatment(P<0.05). (5) The rate and density of VEGF expression at ectopic lesion in rAAV2-endostatin-EGFP group (35%, 30%, 25% and 1.60±0. 43,1.33± 0. 30,1.03±0.36) were significantly less than those at rAAV2-EGFP control group (80% ,75% ,85% and 2.43±0.53,2.43±0.29,2.66±0.45) and PBS control group (85% ,90% ,90% and 2.36±0.53,2.64± 0.57,2.53±0.52) at one 1, 2 and 3 ,weeks after treatment (all P<0.05). The expression of VEGF at ectopic lesion in rAAV2-endostatin-EGFP group at 3 weeks was significantly less than those at 1 and 2 weeks after treatment (P<0.05). (6) The level of estradial and progesterone in serum of nude mice of rAAV2-endostatin-EGFP group [ E_(2)> : (48±7 ) pmol/L, P: (61±8 ) nmol/L ] did not reach statistical difference when compared with those at rAAV2-EGFP control group [ E_(2): (50±9) pmol/L, P: (60±10) nmol/L] and PBS control group [E_(2):(48±7)pmol/L,P: (58±10)nmol/L,P>0.05]. Conclusions The recombinant adeno-asseciated virus carrying human endostatin gene therapy could inhibit angiogenesis at endometriotic lesions and not influence steroid level. The antiangiogenic gene therapy might become a novel option for endometriosis.
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Objective: TO explore the effect of VEGF-C gene transfection on the expression of VEGF-C in human cervical carcinoma HeLa cells and the mechanisms of its anti-apoptosis effect. Methods: The con-structed pcDNA3.1(+)NEGF-C vector was transformed into human cervical cancer HeLa cells and was select-ed by G418. The changes in the expression level of VEGF-C mRNA and protein were determined by semi-quantitive RT-PCR and ELISA. HeLa cells with overexpression of VEGF-C were named as HeLa/S1. The expression level of NF-KB and bcl-2 mRNA was determined by RT-PCR in transfected cells. Results: After transfection by liposome, the VEGF-C mRNA level and the expression of VEGF-C protein in transfected cells were higher than those in the control groups. HeLa/S1 cell line was successfully established. In HeLa/S1 cells, the expression of NF-κB (2.06±0.09 vs 1.35±0.02 vs 1.38±0.02 P<0.05) and bcl-2 gene mRNA (2.02± 0.67 vs 0.41±0.06 vs 0.37±0.06, P<0.05) level were higher than those in the control groups. Conclusion: VEGF-C gene transfection by liposome can increase the expression of VEGF-C in human cervical cancer HeLa cells. NF-κB is stimulated and induces the overexpression of bcl-2 gene in HeLa/S1 cells.
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Objective: To analyze the influence of semen processing methods on the proportion of the aneuploid sperm by detecting the sperm's chromosome X, Y, 18 using fluorescence in situ hybridization. Methods: Ten patients with mild ohgoasthenosperia, who were received ICSI treatment, were included in this study. Five semen samples of the patients were randomly selected to detect using Swim-up method (A group) and 5 using sperm-grad double-density centrifugation method (B group).Another 5 patients with mild oligo-asthenosperia were as control (C group). The CEPX / Y and CEP18 probe was used to detect the sperm of these 15 patients by fluorescence in situ hybridization. The proportion of aneuploid sperm was compared in three groups. Results: The sex chromosome aneuploid rates were (4.21±2.49)%, (3.24～1.49) % and (2.62±0.89) % in control, A and B groups. The rates of aneuploid chromosome 18 were (3.00±1.22)%, (2.00～1.22)% and (2.00±1.22)% in control, A and B groups. There were no significant differences in three groups (P＞0.05). Conclusion: The results showed that the methods of Swim-up and Sperm-Grad double-density gradient centrifugation could select sperms in motility potential and teratospermia,but not in normal chromosome sperms.
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Objective: To construct the packaging plasmid pSNAV-hEndostatin-CMV-EGFP of adeno-associated virus vector (AAV) encoding human Endostatin (hEndostatin) cDNA. Methods: The hEndostatin eDNA obtained from plasmid pCD-sEndostatin was subcloned into the packaging plasmid pSNAV of AAV by molecular clone ways. The recombinant plasmid pSNAV-hEndostatin-CMV-EGFP was identified by PCR analysis, restriction enzymes analysis and sequencing analysis.Results: The recombinant pSNAV-hEndostatin-CMV-EGFP was correctly constructed and confirmed by PCR analysis, restriction enzymes analysis and sequencing analysis. Conclusion: The constructed AAV-hEndostatin packaging plasmid pSNAV-hEndostatin-CMV-EGFP could be the packaging plasmid of rAAV-hEndostatin-EGFP.
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Objective To study the changes of DNA repair genes and enhanced anti-tumor effect of cisplatin induced by mifepristone in human ovarian cancer drug resistance cells.Methods The alterations of cisplatin concentration producing 50％inhibition(IC50)in the COC1/DDP cell lines were examined by methyl thiazolyl tetrazolium(MTT)assay.RT-PCR and flow cytometry were used to analyze the changes of the mRNA of ERCC1,BRCA1,hMLH1 genes and cell cycle and apoptosis.Subcutaneous implantation of COC1/DDP was established in nude mice and the enhanced anti-tumor effect of cisplatin by mifepristone was observed in vivo.ResultsCisplatin IC50 values of COC1/DDP cell were decreased from(3.71±0.38)μg/ml to(3.18±0.46),(1.95±0.14),(0.64±0,18)μg/ml respectively when treated with 2.5,5.0,10.0 μmol/L mifepristone.Mifepristone could down-regulate the mRNA levels of ERCC1,BRCA1,hMLH1 genes and enhance G0/G1 phase block effect pf cisplatin,and 2.5,5.0,10.0 μmol/L mifepristone combined with cisplatin increased rate of cell apoptosis from 0.08％to 5.11％,9.13％and 12.24％ respectively.The percentage of inhibition of xenograft tumor volume in combined treatment group was 70.1％,which was significantly different(P＜0.05).Conclusion By down-regulating ERCC1,BRCA1,hMLH1 genes,blocking G0/G1 phase,and increasing apoptosis rate,mifepristone could enhance anti-tumor effect of cisplatin.
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Objective To construct adenovirus vector with agiostatinK1-5 gene and to investigate the function of suppression to proliferation and migration for vascular endothelial cells.Methods With the use of gene recombination and clone technology, we constructed the adenovirus vector with the gene of angiostatin K1-5. In vitro vascular endothelial eclls proliferation assay and migration activity were performed through direct infection,MTT and transwell chemotaxis assay. Results 50% TCID indicated that the condence of resultant viruses was 1.5?109PFU/mL. It was purified by CsCL banding,final yield were generally 1.1?1010 PFU/mL plaquing-forming units. Through indirect infect assay and MTT, we found angiostatin K1-5 inhibited human vascular endothelial cells proliferation. We utilized human vascular endothelial cells to study the effect angiostatin K1-5 on cell migration,the result showed that adenoviruse vector with angiostatin K1-5 significantly inhibited HUVEC migration.Conclusion We successfully constructed adenoviruse vector with angiostatin K1-5 and demonstrated its inhibitory effect on proliferation andmigration of HUVEC.
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Objective To investigate the roles of luteinizing hormone releasing hormone (LHRH), follicle stimulating hormone (FSH), human chorionic gonadotropin (hCG) and ovary steroids in regulation of human endometrial stroma cell (ESC) decidualization Methods Stroma cells derived from human endometrium during the proliferative phase were cultured in vitro and treated with physiological doses of estradiol (E 2), testosterone (T),progesterone(P) or LHRH, FSH and hCG Their morphologic changes and prolactin (PRL) production in the media were examined and compared Results Addition of E 2 or T or P stimulated ESC proliferation, resulting in increase of the saturation density The fibroblast morphologic changes to polygonal shape and began to express PRL simutaneously after treatment of P or T P in presence of E 2 or T significantly enhanced PRL production( P
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Objective To study the effects of antisense oligodeoxynucleotides(ODN) of hEST2 (AODN) on telomerase activity and proliferation in ovarian cancer cell lines SKOV3 and COC1 Methods Antisense and sense human telomerse catalytic sub unit (hEST2) phosphorothioate (SODN)and random ODN were designed, synthesized and transfected into SKOV3 and COC1 cells by lipofectamine The expression of hEST2 mRNA and telomerase activity in SKOV3 and COC1 were tested by reverse transcription polymerase chain reaction and telomeric repeat amplification protocol before and after transfection The proliferation and growth in SKOV3 and COC1 were also investigated by methyl thiazolyl tetrazolium and growth curve before and after transfection Results AODN could down regulate the expression of hEST2 mRNA, inhibit telomerase activity and proliferation of ovarian cell lines The efficiency depends on dose and period of administration At 48 h, 30 ?mol/L AODN had the highest activity The expression of hEST2 mRNA were declined 54 6% and 44 6% in SKOV3 and COC1 respectively And also the inhibition of telomerase activity were 47 9% and 42 7% respectively in the two cell lines Conclusions AODN of hEST2 clearly inhibited the proliferation of ovarian cancer cell lines hEST2 may thus be a new target of gene therapy in ovarian carcinoma
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Objective To study changes of cisplatin sensitivity by RNA interfering the excision repair cross-complementing (ERCC) 1 gene in ovarian cancer cell lines. Methods The small interference RNA (siRNA) targeting ERCC1 gene was designed and synthesized by transcription in vitro, and transfected to ovarian cancer cell line ES-2. The mRNA and protein of ERCC1 were evaluated by means of RT-PCR, western blot and immunocytochemistry. The changes of cisplatin sensitivity after interference were examined by methyl thiazolyl tetrazolium (MTT) assay. Results In ES-2 cell, the mRNA and protein levels of ERCC1 were dramatically decreased 24, 48 and 72 hours after transfection. The sensitivity to cisplatin of ES-2 cell line was increased by 53.88 times after disturbing the ERCC1 gene. Conclusion The sensitivity to cisplatin of ovarian cancer cell lines ES-2 could be enhanced by RNA interfering ERCC1 gene.