Objective To study the effect of direct cover vitrification (DCV ) on the morphology of follicles from human o‐varian tissue .Methods Biopsies of ovarian tissue was cryopreserved by DCV and some of them was cultured for 7 days .The rates of primordial follicles ,primary follicles and secondary follicles were counted .Results Compared to the fresh group and frozen group ,the percent of primordial follicle of fresh‐cultured group ,frozen group decreased ,primary follicle significantly increased(P<0 .05) .The percent of primordial follicle ,primary follicle ,secondary follicles of fresh‐cultured group were lower than those of fresh group(P<0 .01) .In frozen group ,the rates of the primary and secondary follicles with normal morphology were lower than those of fresh cultured group(P<0 .05) .The rate of normal morphology of secondary follicles of frozen‐cultured group was demonstrably lower than that of fresh‐cultured group(P<0 .05) .Conclusion Partial follicles might be injured in cryopreserved human ovarian tissues by direct cover vitrification .A period of post‐thaw culture may enhance follicle recovery .

Objective To study the safety and efficacy of retroperitoneal laparoscopic resection of adrenal neurilemmoma.Methods The data of 7 cases with adrenal neurilemmoma undergoing retroperitoneal laparoscopic resection were analyzed.2 cases were diagnosed by ultrasound scan,3 cases had blood pressure elevation,and 2 cases had pain in waist and abdomen.All cases underwent ultrasound and CT scan.The neurilemmoma was located in left adrenal area in 2 cases and located in right adrenal area in 5 cases.Results All the 7 cases were successfully operated.No conversion or severe blood loss happened.The average tumor size was 5.0 cm,ranging from 3.0 to 7.0 cm.The average operation time was 75 min,ranging from 45 to 120 min.The estimated blood loss was 50 ml,ranging from 20 to 100 ml.The patients were discharged 7-8 day after the operation.During the 2-12 months of follow-up,no recurrence or metastasis was found.Conclusion Retroperitoneal laparoscopic resection offers an effective and better treatment for adrenal neurilemmoma,with the advantages of less blood loss,less trauma,and faster recovery.

Objective To compare the clinical efficacy of subinguinal microsurgical and single -port laparoscopic high ligation in the treatment of varicocele.Methods 218 patients with varicocele were enrolled in this study.According to the digital table,they were randomly divided into two group.148 cases were treated by subinguinal microscopic varicocelectomy(microscopic group),70 cases were treated by single -port laparoscopic high ligation varicocele(single -port laparoscopic group).Postoperative follow up was 3 -24months.The operative duration,length of hospital stay,hospitalization expense,postoperative complications and semen quality parameters were compared between the two groups.Results There were statistically significant differences in operative duration and hospitalization expense between the two groups(all P <0.05).There was no statistically significant difference in length of hospital stay (P >0.05).In the 218 followed -up patients,the sperm concentration and motility (grade a + b sperm)all significantly improved,which of the microscopic group and single -port laparoscopic group preoperation were (19.1 ± 8.2)×106 /mL,(18.2 ±7.9)×106 /mL and (22.7 ±7.8)%,(21.6 ±8.9)% respectively,which at 3 -6 months after operation were (56.2 ±10.8)×106 /mL,(45.8 ±12.9)×106 /mL and (58.8 ±9.7)%,(44.6 ±10.7)%, there were statistically significant differences compared with preoperation (t =6.227,9.579,all P <0.05 ). Conclusion The surgical methods in the treatment of varicocele can improve the quality of patients,but microscopic group is obviously better than single -port laparoscopic group in improvement of semen quality parameters,safety, patient -based compliance and economy.

Objective To summarize the clinical experience and analyze the efficacy of transurethral plasma needle electrode en bloc resection for bladder cancer.Methods From February 2015 to August 2016,a total of 26 patients,including 21 males and 5 females,with bladder cancer received transurethral plasma needle electrode en bloc resection of bladder tumor.Their age ranged from 42 to 75 years,mean (56 ± 13) years.The size of tumor ranged from 1 to 4 cm,mean (2.3 ± 1.6) cm.The solitary tumor was found in 19 cases and multiple tumors were found in 7 cases,including 2 tumors in 5 cases,3 tumors in 2 cases.In 6 cases,the tumor located in the lateral bladder wall.All the pre-operative biopsy showed the urethelial carcinoma in all cases,No bladder extravasion or upper urinary tumor was noticed in the CTU before surgery.By using the electrode needle tip inserted into the bladder mucosa,blunt release or cut the tumor bases until the deep muscularis or the serosal layer,complete removal of the tumor.The specimen was removed from the bladder and sent to the pathological examination.The operation time,the volume of blood loss during operation,surgical complications,pathological diagnosis and the wounds recovery were recorded and analyzed.Results All surgeries were undergone successfully.Totally 35 tumors were resected with diameter of 1.0-4.0 cm,mean (2.3 ± 1.6)cm.The estimated blood loss was less than 10 ml.The operative duration ranged from 20 to 50 min,mean (30 ± 16)min.The duration for removing the single tumor ranged from 5 to 25 min.No obturator nerve reflex were observed during surgery.No blood loss and complications occurred after operation.All patients received 30 mg pirarubicin bladder instillation chemotherapy immediately and no adverse reaction was noticed.Postoperative pathological stages of enrolled cases were distributed as 33 cases of T1G1 staging,2 cases of T1G3 staging.No positive margin was observed.3 months after operation,cystoscopy showed that the wound healed well.A total of 26 cases followed up for 3-10 months (mean 6.0 ± 2.3 months).No one developed recurrence.Conclusions Transurethral plasma needle electrode en bloc resection of bladder tumor would reduce the incidence of complications and obturator nerve reflex.It can provide sufficient specimens for histological diagnosis.

Objective To construct double expression retroviral vectors targeting chronic myeloid leukemia (CML) b3a2-type mRNA and investigate its effect on the phenotype of K562 cells. Methods The eGFP coding sequence was inserted into the retroviral vector pMSCV-neo to construct pMSCV/GFP, then H1-RNA pol III-based transcription cassettes was subcloned into it to form pMSCV/GFP-H1-BCR/ABL40AS. Two control vectors pMSCV/GFP-H1-BCR/ABL40S & pMSCV/GFP-H1-BCR/ABL80AS were constructed in addition. All these constructions were identified by restriction enzyme analysis and DNA sequencing. After that, the recombinant vectors were transferred into retrovirus packaging cell line PT67 by using lipofectamine2000, and G418 were used to select stable virus-producing cell lines. Viral titer was determined by infection of NIH3T3 cells sequentially. The cell-growth curve was assayed, cell apoptosis was checked with Annexin V-PE/7AAD double staining and flow cytometry analysis after 24-hour infection, the PKR phosphorylation was assayed by Western blotting. Results The plasmids were successfully constructed. Four cell lines, named as PT67-MSCV/GFP, PT67-40as, PT67-40s and PT67-80as were gained by G418 selection, and virus titers were 6.2?10~ 5 , 5.6?10~ 5 , 4.6?10~ 5 and 6.0?10~ 5 CFU/ml respectively. PT67-40as suspensions could induce K562 cell apoptosis by (22.54?3.19)%, significantly different from PT67-MSCV/GFP or PT67-40s (P

To construct an eukaryotic expressing vector that expresses CH50, a recombinant CellⅠ-HepⅡ bifunctional-domain polypeptide of human fibronectin, and to investigate the chemotaxis to immune cells and the inhibitory effect on the growth of tumor by the expression of the plasmid in vivo, the plasmid was constructed by DNA recombination. Gene transfection was performed in vitro and in vivo. The expressed product was identified by Western blot. The chemotaxis after gene transfection in vivo was observed by histotomy and staining of muscle tissues. The inhibition of gene transfection on solid tumor was observed in mice. The results showed that plasmid pCH510 was constructed by the recombination of the 5′-terminal noncoding region and signal peptide coding region of human fibronectin cDNA and cDNA fragment coding CH50 polypeptide with a 3′-terminal noncoding region of human FN cDNA, and the insertion of the recombinated fragment into plasmid pcDNA3.1. After transfection with plasmid pCH510, NIH3T3 cells could produce CH50 polypeptide. The transfection of plasmid pCH510 by the injection in muscle of mouse could produce the effects of chemotaxis on immune cells and the inhibition on the growth of solid tumor. It is concluded that plasmid pCH510 can express in cells and in vivo in mouse. The expression of the plasmid in vivo has a chemotactic effect on immune cells and can inhibit the growth of solid tumor.

To construct an eukaryotic expressing vector that expresses CH50, a recombinant CellⅠ-HepⅡ bifunctional-domain polypeptide of human fibronectin, and to investigate the chemotaxis to immune cells and the inhibitory effect on the growth of tumor by the expression of the plasmid in vivo, the plasmid was constructed by DNA recombination. Gene transfection was performed in vitro and in vivo. The expressed product was identified by Western blot. The chemotaxis after gene transfection in vivo was observed by histotomy and staining of muscle tissues. The inhibition of gene transfection on solid tumor was observed in mice. The results showed that plasmid pCH510 was constructed by the recombination of the 5′-terminal noncoding region and signal peptide coding region of human fibronectin cDNA and cDNA fragment coding CH50 polypeptide with a 3′-terminal noncoding region of human FN cDNA, and the insertion of the recombinated fragment into plasmid pcDNA3.1. After transfection with plasmid pCH510, NIH3T3 cells could produce CH50 polypeptide. The transfection of plasmid pCH510 by the injection in muscle of mouse could produce the effects of chemotaxis on immune cells and the inhibition on the growth of solid tumor. It is concluded that plasmid pCH510 can express in cells and in vivo in mouse. The expression of the plasmid in vivo has a chemotactic effect on immune cells and can inhibit the growth of solid tumor.

Objective To investigate the clinical therapeutic effect of sigmoid neobladder after laparoscopic radical cystectomy in the treatment of invasive bladder cancer .Methods 21 patients with invasive bladder cancer accepted sigmoid neobladder after laparoscopic radical cystectomy and pelvic lymphadenectomy .The postoperative complications and clinical therapeutic effect were analyzed .Results All surgeries were completed successfully .The operation time was (245 ±125) min,blood lost was (98 ±45) mL.All the 21 patients were followed up for 3-24 months,mean of 12 months.Both side mild hydronephrosis in 6 cases (28.6%) and unilateral hydronephrosis in 1 case(0.5%).Average bladder reservoir was (548.8 ±221.6)mL,the maximum bladder pressure during the new bladder urine storage period was (2.26 ±0.24) kPa,the maximum bladder pressure during voiding period was (6.18 ±0.29) kPa,volume of residual urine was (87 ±52) mL.The mean maximum uroflow rate was (83.2 ± 5.6)mL/s.The continence of micturition was 90.4% in day-time and 76.1% in night-time.1 case had urethral stricture and got better after the urethral dilatation .Conclusion The sigmoid neobladder after laparoscopic radical cystectomy has more reliable results,without serious postoperative complications and patients with high quality of life after operation.This operation may be applied in clinical practice .

In this study,the effects of pirrolidine dithiocarbamate (PDTC) plus leflunomide (Lef) models were investigated.NIH mice and Wistar rats served as donors and recipients respectively.Mouse-to-rat cardiac xenotransplantation was performed.The recipients were divided into 5 groups:group A (the control group),group B (PDTC group),group C (PDTC plus CsA group),group D (PDTC plus Lef group) and group E (PDTC plus Lef and CsA group).The expressions of IKKα/β,by immunohistochemistry and Western blot as well as electrophoretic mobility shift assay (EMSA).The median survival time of cardiac xenografts in the control group,PDTC group,PDTC plus CsA group,PDTC plus Lef group and PDTC plus Lef and CsA group was (2.17±0.41),(2.33±0.52),(4.67±1.21),(7.00±1.79) and (9.00±1.41) days respectively.The survival time of xenografts in the PDTC plus Lef and CsA group was significantly longer than that in other four groups (P<0.05 for all),that in the PDTC plus Lef group longer than that in the control group,PDTC group and PDTC plus CsA group (P<0.05 for all),that in PDTC plus CsA group longer than the control group and PDTC binding activity were notably increased in mouse-to-rat cardiac xenografts.The expressions were decreased in the control group,PDTC group,PDTC plus CsA group,PDTC plus Lef and PDTC plus Lef and CsA group in turn.It was concluded that PDTC plus Lef and CsA can significantly suppress prolonging the survival of the cardiac xenografts.

Objective To investigate the expression of proline rich tyrosine kinase 2 (Pyk2) in non-muscle invasive bladder cancer,and analyze its correlation to clinicopathologic features and prognosis of non-muscle invasive bladder cancer.Methods 114 surgical specimens and 50 normal bladder mucosa specimens were collected from 114 non-muscle invasive bladder cancer patients who underwent TURBT at our hospital,from June 2013 to March 2018.Of the 114 patients,63 were male and 51 were female,aged 42-87 years,average age of (63.6 ± 13.8) years,73 cases of tumor ＜3 cm,41 cases of tumor ≥3 cm,83 cases were single and 31 cases were multiple tumor,53 cases were high grade and 61 cases were low grade,59 cases were Ta and 55 cases were T1 stage.Pyk2 protein expression was detected by immunohistochemistry and western blot.The correlation of the expression of Pyk2 with clinicopathologic features,including gender,age,tumor size,the number of tumors,histological grade and clinical stage were analyzed.Survival analysis was calculated by using the Kaplan-Meier method,and the difference in survival curve was analyzed by using the log-rank test.Association of Pyk2 expression with prognosis of non-muscle invasive bladder cancer analyzed by using the Cox proportional hazards regression model.Results Compared with normal bladder tissues,expression of Pyk2 protein was increased in bladder cancer tissue significantly(0.571 ±0.230 vs.0.253 ± 0.152,P ＜0.01).The expression of Pyk2 protein was closely related to clinical stage(P =0.027) and grade(P =0.010),rather than gender (P =0.275),age (P =0.419),tumor size (P =0.317),and tumor number(P =0.208).The recurrence rate in the Pyk2 positive group and negative group were 46.1％ (35/76)and 28.9％ (11/38)respectively.The progression rate in the Pyk2 positive group and negative group were 35.5％ (27/76) and 10.5 ％ (4/38) respectively.Survival analysis suggested expression of Pyk2 in non-muscle invasive bladder cancer had a significant relation to recurrence-free survival rate(P ＜0.001) and progression-free survival rate(P =0.003).In the multivariable Cox analysis,we found that Pyk2 protein was an independent predictor of recurrence-free survival rate(HR 0.245,95％ CI 0.078-0.768,P =0.016) and progression-free survival rate (HR 0.095,95％ CI 0.012-0.764,P =0.027).Conclusions The expression of Pyk2 in non-muscle invasive bladder cancer was significantly increased.The expression of Pyk2 has a significant relation to recurrence and progression of non-muscle invasive bladder cancer.High Pyk2 expression is an independent prognostic factor in non-muscle invasive bladder cancer.