30 chorinic villi in early pregnancy (8-10 weeks)and their parent blood samples were analysedusing the method of restriction fragment length polymorphisms with single locus DNA probes of PAC255 and PAC256. Based on the gene frequencies of these two loci in Chinese and the relativechance of obligatory gene in each triplet, the cumulative parternity index and cumulative relativechance of paternity were calculated. The range of the former is 56. 80- 8333. 50, and the later is 98.27 - 99. 99 %. According to the international standard of paternity test, in the 30 pedigrees, 20 cases(66. 7 % )were assured for their paternity. It is showed that the combination of PAC255 and PAC256single locus DNA probes for the paternity test of early pregnancy is very available.

The human sphincter of Oddi has been known for more than a century.The sphincter regulates periodic discharge of bile and pancreatic juice and their directional flow.Research on the sphincter is still inadequate,especially on how to accurately evaluate its function,dysfunction and effective treatment.In recent years,advances in anatomy,regulation and improvements in investigation have deepened our understanding of diagnosis and treatment of sphincter of Oddi dysfunction. This paper summarizes the new developments in this field.

Objective:To study the impact of rotary cell culture system(RCCS) on the quality and activity of fetal islets of pancreas after cryopreservation and resuscitation.Methods:The fetal islets of pancreas were assigned into three groups averagely,experiment group 1,2 and control group.In experiment group1 and 2,fetal islets of pancreas were cultivated in RCCS and common culture respectively.While in control group,fresh fetal islets of pancreas were cultivated in RCCS all the time.We resected the fetal pancreas and digested with Collagenase V and purified with ficoll solution.Then standard cryopreservation step was adopted,and after resuscitation,the islets of pancreas were cultivated continuously.And the quantity,quality and activity of the islets of pancreas,and insulin stimulation test results in all groups were detected.Results:After purification,islets got from one fetal pancreas reached 2,331.98-5,115.43 IEQ(average 3,551.27? 253.76 IEQ) .The survival rate,the insulin release and stimulation index in islets of pancreas of group 1 incubated in RCCS were higher than those in common culture.Conclusion:RCCS are good for the growth and proliferation of islets of pancreas,and the islets of pancreas cultivated in RCCS have better insulin secreting capacity.RCCS combined standard cryopreservation method may further improve the cryopreservation effect.

In the present study, 89 porcine reproductive and respiratory syndrome virus (PRRSV) isolates in China during 2007 to 2012 were randomly selected from the GenBank genetic sequence database. Evolutionary characteristics of these isolates were analyzed based on the sequences of non-struc-tural protein 2 (Nsp2) and glycoprotein 5 (GP5). The genetic variations of the isolates were also compared with six representative strains. The results showed that a high degree of genetic diversity exists among the PRRSV population in China. Highly pathogenic PRRSV isolates, with a discon-tinuous deletion of a 30 amino acid residue in the Nsp2 region, remained the most dominant virus throughout 2007-2012 in China. Owing to the extensive use of representative vaccine strains, natu-ral recombination events occurred between strains. Three isolates-HH08, DY, and YN-2011-were more closely related to vaccine strains than the other isolates. Both YN-2011 and DY were the evolu-tionary products of recombination events between strains SP and CH-1R. The results of the present study provide useful information for the epidemiology of PRRSV as well as for vaccine development.

Objective To investigate the postoperative complications after laparoscopic splenectomy for patients with portal hypertension,and analyze the risk factors for postoperative complications.Methods The clinical data of 65 patients with portal hypertension who received laparoscopic splenectomy at the West China Hospital from September 2003 to September 2012 were retrospectively analyzed.The postoperative complications were analyzed using the modified Clavien classification system,and the risk factors for postoperative complications were analyzed.The measurement data and the count data were analyzed using the t test,chi-square test or the Fisher exact probability.The univariate and multivariate analysis were done using the Logistic regression model.Results Twenty-nine patients received hand-assisted laparoscopic splenectomy and 36 received total laparoscopic splenectomy (2 patients were converted to open surgery due to intraoperative bleeding).Sixteen patients had 20 complications.The incidence of postoperative complications of the hand-assisted laparoscopic group was 10.3％ (3/29),which was significantly lower than 38.2％ (13/34) of the total laparoscopic group (x2 =3.90,P ＜ 0.05).According to the modified Clavien classification system,there were 4 patients with grade Ⅰ complication,2 patients with grade Ⅱ complication,8 patients with grade Ⅲ a complication,1 with grade Ⅲ b complication,1 with grade Ⅳa complication,and no Ⅳb or Ⅴ complication was observed.The duration of postoperative hospital stay was (8.4 ± 2.9) days.For patients with grade Ⅱ,Ⅲ and Ⅳ complications,the duration of postoperative hospital stay was (10.1 ± 3.3) days,which was significantly longer than (7.7 ± 2.5) days of patients with grade Ⅰ complication or without complications (t =4.30,P ＜ 0.05).The results of univariate analysis showed that ASA classification and application of hand-associated device were risk factors for postoperative complications (x2=21.60,5.10,P ＜0.05).The results of multivariate analysis showed that ASA grade Ⅲ and non-application of hand-assisted device were independent risk factors for postoperative complications (OR =23.60,4.60,P ＜ 0.05).The incidence of postoperative complications of patients with ASA grade Ⅲ were 17.00 times higher than patients with ASA grade Ⅱ,and the incidence of postoperative complications of patients who received total laparoscopic splenectomy was 5.00 times higher than those who received hand-assisted laparoscopic splenectomy.Conclusions The incidence of postoperative complications is higher in patients with portal hypertension,while the severity of the complications is under ASA grade Ⅲ.ASA classification and application of hand-assisted device are correlated with the incidence of postoperative complications.

Objective To investigate the association between TGFβ1-509 C/T gene polymorphism with primary ANCA associated vasculitis (AAV) in Chinese Han population . Methods The blood DNA and clinical data of 88 patients were collected, TGFβ1-509 C/T genotypes were determined by PCR-RFLP, 107 healthy individuals were tested as controL Clinical and pathological data of the patients with different genotype were compared. Results No significant difference was found in neither genotype distributions nor allele frequencies between the patients and the control (P > 0. 05). Significant difference was found in uria protien level of the three groups of patients with different genotypes(P <0.05) ,but not in blood pressure, serum urea nitrogen or creatinine, vasculitic damage index, birminghan vasculitis activity score (P > 0. 05 ). Significant difference was found in med-heavier glomerular mesangial proliferation of the three groups ( P < 0.05 ) , but not in lighter glomerular mesangial proliferation, glomerular sclerosis, crescent formation and tubule-interstitial fibrosis and atrophy. Conclusions In Chinese Han population, TGFβ1-509 C/T polymorphism might have no relationship to susceptibility of primary AAV, but might relate to uria protein and med-heavier degree of mesenterium proliferation.

ObjectiveTo observe the effect of mesenchymal stem cells (MSCs) on enhancing rat islets viability and function in vitro by a pretransplant co-culture.Methods4-week-old Wistar rats were used as donors, bone mesenchymal stem cells were isolated and subcultured. Islets of Wistar rats were isolated and purified by one-step single-layer Histopaque-1077. Then islets were divided into four groups randomly, 2 groups co-cultured with mesenchymal stem cells (MSCs) (one using low-glucose medium; the other using high-glucose medium ); 2 groups were cultured alone (low-glucose medium; high-glucose medium), each group was further stratified into 3 subgroups(3, 7, 14 d); the survival and functionality of these islets were observed and evaluated. The amount of glucose stimulated secreted insulin were measured wth a rat/mouse insulin enzyme-linked immunosorbent assay (ELISA) kit and stimulation index was also calculated.ResultsCompared with those not co-cultured, islets co-cultured with MSCs demonstrated significantly higher survival rates and viability both in 3th, 7th and 14th day ( P ＜ 0. 01 ); furthermore, cocultured islets revealed higher levels of glucose stimulated insulin secretion and secretion indexes in 7th day (P＜0.01).ConclusionRat islet cells co-cultured with MSCs have longer in vitro survival and better functions.

Objective To investigate the expression of exogenous gene transferred by piggyBac (PB) transposon in various gynecological malignant cell lines and reveal its potential application of gene therapy in gynecological cancer.Methods Amplified herpes simplex virus thymidine kinase (HSV-tk) gene coding region by PCR and integrated it into PB expression vector, PB[Act-RFP]DS, for reconstructing PB[Act-RFP, HSV-tk]DS (pPB/TK).By using different transfection reagents: FuGENE HD, jetPEI, lipofectamine 2000, pPB/TK together with helper plasmid Act-PBase were cotransfected into four mostly common gynecological malignant tumor cell lines HeLa, JEG-3, SKOV3 and HEC-1B.The mRFP1 report gene expressions was observed and detected by fluorescence microscope and flow cytometry to analyze transfection efficiency.The expressions of HSV-tk and mRFP1 gene were detected by reverse transcription PCR (RT-PCR).The cytotoxic effect of various concentration of pro-drug ganciclovir (GCV) for transfected cells was detected by methyl thiazole tetrazolium assay.The transfected cells were positive sorted by flow cytometry and limiting diluted to obtain the stable transfected cell line.The insertion sites of foreign gene tranferred by PB transposon in genome were analyzed by inverse PCR.Results (1) Double digests analysis and sequences test demonstrated that pPB/TK vector was reconstructed successfully.(2) Using three different transfective reagents, PB trausposon transferred HSV-tk gene and mRFP1 gene into HeLa, HEC-1 B, SKOV3 and JEG-3 cell efficiently, and the transfection efficiency of pPB/TK for the same cell was different by using different transfective reagents; in Hela cell, the transfection efficiency of FuGENE HD [(78.7 ± 9.2) %]was higher than that of lipofectamine 2000 [(54.1 ± 11.4) %]and jetPEI [(46.5 ± 7.4) %, all P < 0.05] ; using the same transfective reagent, the transfection efficiency of pPB/TK was also different on various cell lines, using FuGENE HD, the transfeetion efficiency of pPB/TK on HeLa, JEG-3 and SKOV3 cell was (78.7 ± 9.2) %, (74.4 ± 8.9) % and (83.2 ± 9.7) % respectively, which all were higher than that on HEC-1B [(39.5 ± 8.7) %, P < 0.05] .(3) RT-PCR showed that there were the mRNA expression of HSV-tk and mRFP1 in all cell lines.(4) 50% inhibitory concentration of GCV for transfected cells, HeLa, JEG-3, SKOV3 and HEC-1B, was 1.29, 3.35, 0.09 and 13.28 μg/ml respectively.Inhibitory effect of GCV (10 μg/ml) on SKOV3 transfected with pPB/TK was (86 ± 9) %, which was superior to that transfected with pORF-HSVtk alone [(52 ± 12)%, P < 0.05] .(5) The insertion sites of PB transposon in the target cells genome were located at TTAA sites, mRFP1 expression still could be detected in three months after transfected.Conclusions PB transposon could transfer exogenous gene into various gynecological malignant cells, which could integrated into genome and obtain a long-term and stable expression.It is expected that PB transposon may supply a more efficient and safer transgene technology platform for gene therapy in gynecological cancer.

Objective To investigate the relationship between TGFβ1-509 C/T, TCRCα-575 A/G SNPs and primary AAV using a transmission disequilibrium theory based pedigree analysis Methods Genotypes of 264 individuals from 88 AAV families include patients, their parents, brothers and sisters were determined by PCR-RFLP and direct sequencing. Transmission disequilibrium test(TDT) and HRR were employed for the data analysis to observe the transmission disequilibrium of TGF31-509 C/T and TCRCα -575 A/G polymorphisms. Results No transmission disequilibrium from heterozygous parents onto the patients was found in the trios analyzed by TDT for either TGFβ1-509 C/T (observed C/T = 36/28, expected C/T =33. 5/30. 5, x2 =0.51, P＞0.05) or TCRCo-575 A/G ( observed A/G = 29/39, expected A/G = 33.5/34. 5, x2 = 1. 59, P ＞ 0. 05 ). The genotype-based HRR and haplotype-based HRR showed there was no increased risk of AAV in the observed trios for either -509 C/T polymorphism of TGFβ1 (transmitted genotype CC/CT/TT =12/20/6, allele C/T = 44/32; nontransmitted genotype CC/CT/TT = 10/19/9,allele C/T =39/37, genotype-based HRR x2 =0.81, P ＞0. 05, haplotype-based HRR x2 =0. 66, P＞0. 05,HRR = 1.30) or -575 A/G polymorphism of TCRCα ( transmitted genotype AA/AG/GG = 9/18/12, allel A/G = 36/42; nontransmitted genotype AA/AG/GG = 15/15/9, allel A/G = 35/33, genotype-based HRR x2=2. 20, P ＞0. 05. Haplotype-based HRR x2 =0. 41, P ＞0. 05, HRR =0. 81 ). The deviation of HRR coefficient was not excessive(1.00). Conclusion TGFβ1-509 C/T and TCRCo-575 A/G polymorphism may not be associated with the genetic susceptibility of primary AAV in Guangxi Han population.

Objective To investigate the safety of the laparoscopic pancreaticoduodenectomy (LPD) in 70 years of age or older patients.Methods The retrospective cohort study was conducted.The clinicopathological data of 40 patients (age ≥70 years old) who underwent pancreaticoduodenectomy in the West China Hospital of Sichuan University between January 2012 and December 2016 were collected.Twenty patients undergoing LPD were allocated into the LPD group,and 20 receiving open pancreaticoduodenectomy (OPD) who were selected by random number table during the same period were allocated into the OPD group.Observation indicators included:(1) intraoperative situations;(2) postoperative situations;(3) follow-up situation.Follow-up using outpatient examination and telephone interview was performed to detect the patients' survival after discharge and tumor recurrence and metastasis up to March 2017.Measurement data with normal distribution was represented as x±s,and comparison between groups were evaluated with the t test.Measurement data with skewed distribution were described as median (range) and comparison between groups was analyzed using the nonparametric test.Comparison of count data was analyzed using the chi-square test.Comparison of ranked data was analyzed by non parametric test.Results (1) Intraoperative situations:1 patient in the LPD group was converted to open surgery,with a conversive rate of 5.0％ (1/20).Operative time and volume of intraoperative blood loss were (463 ± 10) minutes,210.5 mL (152.5-300.0 mL) in the LPD group and (332± 25) minutes,420.0 mL (350.1-493.8 mL) in the OPD group,showing statistically significant differences between the 2 groups (t =5.48,Z =-3.98,P＜0.05).Cases with intraoperative blood transfusion and pylorus preservation were respectively 4,14 in the LPD group and 6,10 in the OPD group,showing no statistically significant difference between the 2 groups (x2=0.53,1.67,P＞0.05).The results of intraoperative rapid frozen pathological examination showed negative margin of the 40 patients.(2) Postoperative situations:cases in ICU,cases with postoperative analgesia,time for out-of-bed activity,time to anal exsufflation and time for intake were 17,7,(2.2±0.7)days,(4.2± 0.9)days,(4.8±0.7)days in the LPD group and 6,15,(3.6±0.8)days,(5.7±0.9)days,(7.1 ± 2.7)days in the OPD group,showing statistically significant differences between the 2 groups (x2 =12.34,6.47,t=-6.18,-6.55,-3.65,P＜0.05).Pancreatic fistula,delayed gastric emptying (Grade B),postoperative bleeding (Grade B),biliary fistula,pulmonary infection,intestinal obstruction,wound infection,reoperation and major complication were respectively detected in 2,3,1,1,3,1,0,2,3 patients of the LPD group and 2,4,1,1,4,1,2,3,4 in patients of the OPD group,showing no statistically significant difference between the 2 groups (x2 =0.00,0.17,0.00,0.00,0.17,0.00,2.11,0.23,0.17,P＞0.05).Results of postoperative pathological examination showed that duodenal adenocarcinoma,ampullary carcinoma,lower bile duct carcinoma,pancreatic ductal adenocarcinoma and pancreatic cystic tumor were respectively detected in 8,2,5,3,2 patients of the LPD group and 10,2,4,2,2 patients of the OPD group,showing no statistically significant difference between the 2 groups (x2 =0.53,P＞0.05).Duration of postoperative hospital stay in the LPD and OPD groups were (19± 13) days and (15±7) days,respectively,showing no statistically significant difference between the 2 groups (t =1.28,P＞ 0.05).Results of postoperative oncology showed that tumor diameter,number of lymph node dissected,number of positive lymph nodes,cases with negative margin,cases in T1N0M0,T2N0M0,T3N0M0,T3N1M0,T4N0M0,T4N1M0 of TNM staging were respectively (2.4±0.7)cm,15.4±2.3,2,20,2,7,8,2,1,0 in the LPD group and (2.8±0.9)cm,14.4±2.5,3,20,1,8,5,2,3,1 in the OPD group,with no statistically significant difference between the 2 groups (t =-1.64,1.32,x2 =0.23,0.00,Z =-0.69,P＞ 0.05).(3) Follow-up situation:1 patient died respectively in both groups within the postoperative 30 days.Thirty-eight patients were followed up for 1-26 months,with a median time of 14 months.During follow-up,2 patients had tumor recurrence and 1 died of myocardial infarction in the LPD group;3 had tumor recurrence and 1 died of tumor recurrence in the OPD group.Conclusion LPD in 70 years of age or older patients is not only safe and feasible,but also significantly reduce volume of intraoperative blood loss and demand of analgesia,as well as quickly resume normal diet and activities.