Objective:To investigate the structural alterations of mitochondria and its role in neutrophil death induced by ONO-AE-248.Methods:Human neutrophils were cultured in vitro with ONO-AE-248(5?10-5 mol/L)and medium alone. Transmission electron microscopy(TEM) was used to detect the structural alterations of mitochondria and the level of mitochondria membrane potential by flow cytometry using mitocapture dying.Results:ONO-AE-248 resulted in extremely swollen mitochondria within 6 hours. Meanwhile, a rapid loss of mitochondrial membrane potential of neutrophils occurred, especially in 3 hours and 6 hours. There were obviously differences between spontaneous apoptosis and non-apoptosis programmed cell death induced by ONO-AE-248.Conclusion:The experiment results suggest that changes of mitochondrial structure and function be typically morphological, physiological and biochemical features in this unique form of neutrophil death, and that the mitochondrial pathway might play a more important role in ONO-AE-248-induced death of neutrophils.

Objective To compare the time course of distribution and elimination of gelatin and lactated Ringer's solution (LR) by volume kinetics and mass balance analysis during hemorrhagic shock in dogs, and try to design and optimize fluid therapy in a more scientific manner. Methods Twenty dogs were randomly divided into 4 groups: CL group, CG group, BL group, and BG group. Each animal was subjected to two randomly ordered experiments that separated for at least 1 week. In the first phase, plasma volume expansion was studied in the state of anesthesia, animals received 30 mL/kg of LR (CL group) or 10 mL/kg of gelatin (CG group) over 30 min. In the second phase, plasma volume expansion was studied in the state of hemorrhagic shock, animals received 30 mL/kg of LR (BL group) or 10 mL/kg of gelatin (BG group) over 30 min. Hb concentration and Hct were measured every 5 min during and after infusion for 90 min. Hemodynamic parameters were recorded at the same time. The distribution and elimination of infused fluid were studied by volume kinetics, based on serial analysis of hemoglobin dilution in arterial blood, and by mass balance that incorporated volume calculations derived from volume kinetic analysis and measurements of urinary volumes. Results When a one-volume kinetic model was fitted to the data, the value of V and Kr in CG, BL, and BG group were significantly smaller than those in CL group (P<0.05), which could be found from the computer-generated curves.When a two-volume kinetic model was fitted to the data, the value of V1, Kr, Kt in BL group were significantly smaller than those in CL group (P<0.05). The calculations based on mass balance corresponded to the predicted based on volume kinetics. The change of central volume (CCV) in BL, BG, and CG group was significantly greater than those in CL group (P<0.05). The VEE in BG and CG group was significantly higher than that in BL and CL group. The value of VEE in BL group was significantly higher than that in CL group (P<0.05). Conclusions Both of the efficacy of lactated Ringer's solution and gelatin increased significantly in the state of hemorrhagic shock, and the former increased more.

Objective To explore the effects of PP242 on the expression of apoptosis protein Bcl?2 in human lens epithelial cells(HLECs). Meth?ods Immortal HLECs SRA01/04 were cultured and treated with different concentrations of PP242. The cell growth was examined by MTT assay at 24 h and 48 h after PP242 treatment. Real?time quantitative polymerase chain reaction(RT?PCR)and Western blot were adopted to detect the mRNA and protein expression of Bcl?2 respectively. Results The cultured HLECs SRA01/04 were collected and treated with different concentra?tions(100,250,500,750,1 000,1 500 nmol/L)of PP242. After treatment for 24 h and 48 h,the inhibitory rate in each well was determined by MTT assay. The inhibition rate was as below,24 h:7.55%,9.43%,16.98%,22.64%,26.42%,30.19%;48 h:11.11%,23.81%,36.51%,42.86%, 49.21%,63.49%. Compared with the control group,the difference was statistically significant(P<0.05). Compared with the control group(0 nmol/L),the expression of Bcl?2 protein was significantly decreased with the increasing concentrations of PP242(P<0.05),while the expression of Bcl?2 mRNA was inhibited by PP242 based on the results of RT?PCR. Compared with the control group(0 nmol/L),the RT?PCR results of Bcl?2 mRNA were 0.723±0.039,0.517±0.028,0.353±0.052,0.167±0.046,respectively(P<0.05). Conclusion PP242 could inhibit cell proliferation of HLECs in vitro with a concentration and time depended manner. Bcl?2 protein is expressed in HLECs,which could be down regulated by PP242 treatment.

ObjectiveTo explore the effects of fluoxetine on special learning and memory and serum S100B level in depressed model rats.MethodsAdult male SD rats were divided into six groups randomly according random digits table:control group ( A ),depressed model group ( B ),group of depressed model treated with single dose of fluoxetine for one day ( C ),group of depressed model treated with fluoxetine for one week (D),group of depressed model treated with fluoxetine for two weeks (E) and group of depressed model treated with fluoxetine for four weeks (F),ten rats in each group.Except control group,others were subjected to forced-swimming for four weeks,15 min a day.Fluoxetine (10 mg/kg) was given intragastric administration to group C-F before swimming everyday.Morris water maze ( MWM ) was used to measure the spatial learning and memory of rats.ELISA was used to determine the level of serum S100B.ResultsIn the hiding platform test of MWM,there was significant longer of escape latency (EL) in B group than that in A group(P ＜ 0.05 ).And the EL in all groups treated with fluoxetine became shorter with the prolonging of treatment.In the probe test,there were significant longer time in target quadrant in D,E,F than in other quadrant (F =5.162,P ＜ 0.01 ).The levels of serum S100B were lower in E,F groups ( E group ( 0.91 ± 0.23 ) ng/ml,F group ( 0.85 ± 0.21 ) ng/ml) than that in B group (( 1.26 ±0.61 )ng/ml,P＜0.05).ConclusionChronic administration of fluoxetine could improve the impairment of spatial learning and memory and reverse the increase of S100B level in serum of depressed model rats.

Purpose:To investigate the relationship between cell apoptosis and dephosphorylated RB protein in human breast cancer. Methods:In our work,human breast cell lines (MCF-7/S,the chemosensitive cell line and MCF-7/ADR,the chemoresistent cell line)were evaluated. Chemosensitivity of two cell lines was evaluated by the MTT colorimetric assay;the expressive levels of dephosphorylated RB protein were detected with immunocytochemistry. Apoptosis rates were determined by flow cytometry(FCM). Results:ADR inhibited proliferation of chemosensitive cell line MCF-7/S ,the 50% inhibition concentration (IC 50 ) was 0.128 ?g/ml;And IC 50 of MCF-7/ADR was 10.89 ?g/ml. The chemotherapeutic sensitivity of MCF-7/S was more than that of MCF-7/ADR by 86 times . Before treatment with ADR,phosphorylated RB protein was positive in two cell lines,but dephosphorylated RB protein was negative;After treatment of different concentration ADR,when the concentration of ADR was increased,expression of dephosphorylated RB protein elevated accordingly in MCF-7/S,but no significant change in MCF-7/ADR. Apoptosis and cell cycle was detected by FCM assays shows ADR induced apoptosis of MCF-7/S more than MCF-7/ADR(P0.05).

Objective To explore the feasibility,efficacy and safety of endoscopic retrograde cholangio?pancreatography ( ERCP ) drainage during peroperation of hilar cholangiocarcinoma for alleviate jaun?dice. Methods Nineteen cases patients with hilar cholangiocarcinoma who were treated with ERCP in the First Affiliated Hospital of Xi'an Jiao Tong University from January 2013 to December 2013,the drainage way,efficient rate,complication rate,and surgical situation were retrospective analyzed. Results Bilateral endoscopic drain?age was one?time achieved in all 19 patients. Among them,Eendoscopic nasobiliary drainage( ENBD) for unilat?eral bilateral drainage was 4 cases,ENBD and plastic stent for unilateral( left or right) drainage was 9 cases,EN?BD and plastic stent for bilateral drainage was 6 cases. The drainage efficiency rate was 89. 5% ( 17/19) . Serum alanine aminotransferase(ALT),total bilirubin(TBIL),direct bilirubin(DBIL),alkaline phosphatase(ALP) and Prothrombin time (PT) were significantly decreased after 7days post?ERCP((208. 4±47. 7) U/L vs. (90. 3 ±31. 57) U/L,(421. 7±85. 9) μmol/L vs. (150. 1±49. 7) μmol/L,(294. 6±30. 6) μmol/L vs. (95. 4±23. 2)μmol/L,(853. 1±133. 7) U/L vs. (600. 0±116. 4) U/L,(17. 7±1. 8) s vs. (13. 8±1. 0) s;P=0. 000,0. 001, 0. 000,0. 001,0. 004) . There were 6 cases occurred ERCP postoperative complications,including 2 cases of hy?peramylasemia, 1 case of pancreatitis, 3 cases of cholangitis. Seven cases of hilar cholangiocarcinoma patients were received hilar radical surgery by combination caudate lobectomy of the left or right hepatectomy,no postop?erative cholangitis was occurred. Conclusion ERCP biliary drainage is an important means to ensure the perio?perative safety and efficacy of hilar cholangiocarcinoma.

Objective To investigate the causes, diagnosis and surgical treatment of ERCP related duodenal per﹣foration. Methods Clinical data of 6 cases of surgical treatment of ERCP related duodenal perforation were retro﹣spective analyzed. All the 6 perforation patients underwent emergency surgical procedure, including 3 cases trans﹣fered from other hospital after duodenal perforation. 4 cases with a history of abdominal surgery. Preoperative con﹣firmed bravery manager stone 4 cases, 1 case of obstructive jaundice after gallbladder surgery, bile duct expansion in 1 case. Results Perforation causes include duodenum mirror improper operation related in 2 cases, duodenal papilla sphincterotomy related 3 cases (1 case of pre-dissection operation with needle knife), small endoscopic sphincteroto﹣my combined with endoscopic papillary balloon dilation lead to perforation in 1 case.4 cases of intraoperative found right kidney week pneumatosis, 2 cases of postoperative CT found after peritoneal pneumatosis, effusion. All patients with surgery including common bile duct exploration, T tube drainage, duodenal perforation repair, jejunum colostomy, among them 2 cases at the same time line of gastrointestinal anastomosis. 5 cases recovered, 1 case died. Conclusions Inappropriate duodenal papilla sphincter incision indications and Many previous abdominal surgery have higher perforated ration;Found in time, reasonable treatment is the most important;For serious typeⅠand typeⅡperforation, active surgical treatment in time, can effectively reduce serious consequences caused by the ERCP related perforation.

Objective To explore the application value of plastic biliary stent and fully covered self-expandable metallic stent (FCSEMS) in endoscopic retrograde cholangio pancreatography (ERCP) for treatment of benign biliary stricture after liver transplantation.Methods The retrospective cross-sectional study was conducted.The clinical data of 54 patients with benign biliary stricture after liver transplantation undergoing ERCP treatment who were admitted to the First Affiliated Hospital of Xi'an Jiaotong University between January 2010 and August 2016 were collected.Among 54 patients,44 had simple anastomotic stricture and 10 had non-anastomotic stricture.All the patients underwent stent implantation by ERCP.Patients with stricture within 1 month postoperatively initially selected single plastic stent or endoscopic nasobiliary drainage (ENBD),and then changed into multiple plastic stents at the second stent replacement.Patients with stricture after 1 month postoperatively selected multiple plastic stents,multiple plastic stents after balloon dilation or FCSEMS.Observation indicators:ERCP situations,stent implantation,time of stent indwelling,postoperative complications,stent dislocation,treatment outcome and follow-up situations.Patients were followed up by outpatient examination and telephone interview up to November 2016.Clinical symptoms of patients were observed within 1 month postoperatively and liver function and abdominal ultrasound were retested.Liver function and remission degree of biliary stricture were monitored regularly once every 3 months.Measurement data were described as average (range).Results All the patients underwent successful ERCP,of which 53 completed the process of ERCP and 1 rejected treatment due to economic problems.All the 54 patients received 140 times ERCPs with an average of 2.59 times per person,21 times ENBDs,11 times FCSEMSs and 108 times plastic stent implantations (including 35 times single stent implantations,46 times double stents implantations,23 times 3-stents implantations and 4 times 4-stents implantations).All the 54 patients were followed up for 3-143 months,with an average time of 73 months.Of 44 with anastomotic stricture,34 received plastic stent implantation and 98 times ERCPs,with an average number of stent implantation of 2 (range,1-4) and an average time of stent indwelling of 10.7 months (range,9.0-13.0months);the postoperative acute pancreatitis,biliary infection,hyperamylasemia and adverse stent implantation or dislocation were detected in 4 persons every time,7 persons every time,10 persons every time and 3 persons every time,respectively;26 patients were cured and 5 were improved,with an effective rate of 91.2％ (31/34);3 patients with noneffective treatment continued to undergo ERCP and 3 patients had recurrence of anastomotic stricture.Among 10 patients with initial FCSEMS implantation,12 times ERCPs were performed,with an average time of stent indwelling of 7.6 months (range,6.0-12.0 months);postoperative biliary infection,hyperamylasemia and stent dislocation were detected in 1 person every time,1 person every time and 1 person every time,respectively;8 patients were cured,with an effective rate of 8/10;of 2 patients with persistent stricture,1 patient received contrast examination after stent removal,showing a comparative stricture in level 1 branch of intrahepatic duct and considering combined ischaemia,and then underwent the second implantation using multiple plastic stents;the other patient had elevated level of jaundice at 3 months after stents removal and received ERCP,showing anastomotic inflammatory polyp,and then underwent FCSEMS implantation again.Ten patients with non-anastomotic stricture received plastic stent implantation and 30 times ERCPs,with an average number of stent implantation of 3 (range,2-4) and an average time of stent indwelling of 11.3 months (range,10.0-14.0 months);the postoperative acute pancreatitis,biliary infection,hyperamylasemia and adverse stent implantation or dislocation were detected in 2 persons every time,5 persons every time,2 persons every time and 1 person every time,respectively;3 patients were cured and 3 were improved,with an effective rate of 6/10;of 4 patients with noneffective treatment,2 died of gradually deteriorating liver function and 2 underwent the second liver transplantation.Conclusions Stent implantation in ERCP is safe and effective for treatment of benign biliary stricture after liver transplantation,single plastic stent should be used in the early period (within 1 month) and multiple plastic stents should be used in the later period.Although FCSEMS has a higher displacement rate,it should be recommended due to a better clinical effect,lower incidence of complications and simple operation.For patients with non-anastomotic stricture,plastic stent should be used for extrahepatic biliary stricture,with a good clinical effect,and there is worse effect in stent implantation through ERCP for multiple intrahepatic biliary strictures.

Objective To construct the recombinant adeno-associated virus vector carrying cancerous inhibitor of PP2A (CIP2A)short hairpin RNA (shRNA)for preparation of high-titer viruses.Methods The small hairpin RNA of CIP2A (CIP2A shRNA)was designed,synthesized and cloned into pDC31 6-EGFP-U6 plasmid which was double digested by Bam HⅠ and Hin dⅢ.The resultant plasmid pDC31 6-EGFP-shRNA was confirmed and served as template to appraise primers.EGFP-CIP2A shRNA sequence was amplified by PCR,double digested with Eco RⅠand Sal Ⅰ and ligated to pSNAV2.0 plasmid digested with the same enzyme pair.pSNAV2.0-EGFP-CIP2A shRNA plasmid DNA was prepared,purified,identified and transfected into BHK-21 cells.BHK-21 cells expressing CIP2A shRNA (BHK-21/CIP2A-shRNA ) were obtained and subsequently infected with VGTC’s proprietary AAV packaging system to package the rAAV2-CIP2A shRNA.After purification,the functional and infectious virus was obtained and the titer of virus was detected.Real-time PCR and Western blot methods were used to detect the expression of CIP2A after infection with HepG2 cells,and the empty viral vector rAAV2-EGFP was used as control. Results A recombinant adeno-associated virus-2 vector carrying CIP2A shRNA was constructed successfully.The presence of the target sequence in the vector was confirmed by double enzyme digestion and sequencing.By transfecting the pSNAV2.0-EGFP-CIP2A shRNA plasmid into BHK-21 cells,BHK-21/CIP2A shRNA cells were infected with helper virus HSV1-rc/ΔUL2 to package the rAAV2-CIP2A shRNA to obtain a functional and infectious virus.The titer of the recombinant virus was 0.25×10 1 2 v.g./mL.The expression of CIP2A mRNA and screening value of 1×10 5 MOI effected HepG2 cells.Conclusion A high-titer recombinant adeno-associated virus-2 vector carrying CIP2A shRNA has been constructed successfully.