Objective To compare the clinical efficacy of modified multi-side hole nasobiliary drainage(MHND)via the percuta-neous transhepatic cholangiography(PTC)route with traditional percutaneous transhepatic cholangial drainage(PTCD)for palliative treatment of patients with advanced obstructive cholangiocarcinoma.Methods A retrospective analysis was conducted on the data from 66 patients with advanced cholangiocarcinoma who underwent biliary drainage.Results Both groups normalize temperature and alleviate symptoms of acute cholangitis within 24 h post-puncture.There was no statistically significant difference in laboratory indicators such as white blood cell(WBC),total bilirubin(TBiL),alanine transaminase(ALT),aspartic transaminase(AST)at 48 h post-operation,and in the incidence of bile leakage,biliary peritonitis,and cholangitis during the postoperative hospital stay(P>0.05).However,the incidence of postoperative electrolyte disorders,gastrointestinal symptoms,and the recurrence rate of gastrointestinal symptoms during the follow-up period were significantly higher in the PTCD group compared to the MHND group,while the recur-rence rate of biliary tract infections was slightly higher in the MHND group compared to the PTCD group.The differences between the two groups were statistically significant(P<0.05).Conclusion Modified MHND shows better clinical efficacy in the treatment of patients with advanced tumor jaundice.Compared with traditional PTCD,it not only effectively reduces jaundice and relieves acute cholangitis but also significantly reduces gastrointestinal symptoms during the postoperative period,thereby improving the quality of life for patients.However,it is noteworthy that it may also increase the risk of biliary tract infections.

Enterotoxigenic Escherichia coli(ETEC)is the main cause of diarrhea in piglets,which seriously threatens the development of animal husbandry.In order to effectively prevent and con-trol piglet diarrhea,the neutralizing epitopes of the screened K88(FTDYEGASVELRKPDGGT-NK),K99(NVGNGSGGANIN),987P(LAAPAENNTSQAN),F41(VMAADWTEGQPGDII)and F18(PPNAQTYPLSSGDLK)ETEC adhesins were assembled to construct the multi-epitope fusion antigen(MEFA),whose immunogenicity was verified by in silico simulations and in vitro.The computer simulation results showed that MEFA has excellent physicochemical properties and reasonable prediction results of secondary and tertiary structures and exhibits strong interaction force when docking with toll-like receptor 3,and the kinetic simulation shows that it can be stably presented.In vitro validation results showed that anti-K88,K99,987P,F18 and F41 IgY could spe-cifically recognize MEFA proteins.After immunization of laying hens with MEFA,the titers of IgY antibodies against the five ETEC were higher than those of non-specific IgY.In addition,specific anti-MEFA IgY significantly inhibited the adhesion of K88,K99,987P,F41 and F18 ETEC strains to piglet intestinal IPEC-J2 cell lines.These results indicate that MEFA has strong immunogenicity and can be used as an ideal vaccine candidate for the prevention of K88,K99,987P,F41 and F18 ETEC infection,and provide reference and technical support for the development of multivalent broad-spectrum ETEC vaccine candidates.

Background and Aims:Complex choledocholithiasis often coexists with stenosis of the ampulla of Vater,which increases the difficulty and complexity of treatment.If only the stones in the bile duct are removed without addressing the ampullary stenosis,the disease is prone to recurrence.Previously,most treatments involved the use of endoscopic retrograde cholangiopancreatography(ERCP)to guide the wire for sphincterotomy and stone extraction,followed by laparoscopic cholecystectomy.However,ERCP has limitations in handling complex cases.In response,our team pioneered a new method of treating choledocholithiasis combined with stenosis of the ampulla of Vater via a transabdominal approach.This study was performed to investigate the feasibility and efficacy of this method,aiming to provide a new therapeutic option for clinical practice.Methods:A randomized controlled study was conducted with 120 patients treated at Chengdu Second People's Hospital from 2021 to 2023 for gallbladder stones and choledocholithiasis with stenosis of the ampulla of Vater.Patients were divided into an observation group and a control group,with 60 cases in each group.The observation group underwent laparoscopic cholecystectomy with choledochotomy for stone extraction,followed by retrograde guidance of duodenal papillary sphincterotomy through the opened bile duct,simultaneously treating gallbladder,bile duct stones,and stenosis of the ampulla of Vater.The control group underwent traditional ERCP approach for sphincterotomy,stone extraction,and laparoscopic cholecystectomy.Perioperative variables were collected for both groups and the surgical outcomes were compared.Results:Among the 120 patients,54 were male and 66 were female.There were no statistically significant differences between the two groups in terms of stone extraction success rate,intraoperative blood loss,postoperative 24-h total bilirubin,direct bilirubin,transaminases,white blood cell count,jaundice relief time,or incidence rates of bile leakage,retroperitoneal bleeding/infection,and severe pancreatitis(all P>0.05).The observation group had significantly shorter average operative time and postoperative hospital stay compared to the control group(98.67 min vs.110.8 min,P<0.05;3.81 d vs.5.61 d,P<0.05).Additionally,the observation group had a significantly lower incidence of postoperative hyperamylasemia and/or hyperlipasemia and mild pancreatitis(1.67%vs.25.00%,P<0.001;0 vs.10%,P=0.027).Conclusion:The novel transabdominal approach is superior to the ERCP approach in terms of reducing surgery time and hospitalization time,and it carries a lower risk of postoperative mild pancreatitis and hyperamylasemia and/or hyperlipasemia.The stone extraction success rate is comparable to that of ERCP,making it a viable alternative treatment option.

Objective To compare the clinical efficacy of modified multi-side hole nasobiliary drainage(MHND)via the percuta-neous transhepatic cholangiography(PTC)route with traditional percutaneous transhepatic cholangial drainage(PTCD)for palliative treatment of patients with advanced obstructive cholangiocarcinoma.Methods A retrospective analysis was conducted on the data from 66 patients with advanced cholangiocarcinoma who underwent biliary drainage.Results Both groups normalize temperature and alleviate symptoms of acute cholangitis within 24 h post-puncture.There was no statistically significant difference in laboratory indicators such as white blood cell(WBC),total bilirubin(TBiL),alanine transaminase(ALT),aspartic transaminase(AST)at 48 h post-operation,and in the incidence of bile leakage,biliary peritonitis,and cholangitis during the postoperative hospital stay(P>0.05).However,the incidence of postoperative electrolyte disorders,gastrointestinal symptoms,and the recurrence rate of gastrointestinal symptoms during the follow-up period were significantly higher in the PTCD group compared to the MHND group,while the recur-rence rate of biliary tract infections was slightly higher in the MHND group compared to the PTCD group.The differences between the two groups were statistically significant(P<0.05).Conclusion Modified MHND shows better clinical efficacy in the treatment of patients with advanced tumor jaundice.Compared with traditional PTCD,it not only effectively reduces jaundice and relieves acute cholangitis but also significantly reduces gastrointestinal symptoms during the postoperative period,thereby improving the quality of life for patients.However,it is noteworthy that it may also increase the risk of biliary tract infections.

Background and Aims:Complex choledocholithiasis often coexists with stenosis of the ampulla of Vater,which increases the difficulty and complexity of treatment.If only the stones in the bile duct are removed without addressing the ampullary stenosis,the disease is prone to recurrence.Previously,most treatments involved the use of endoscopic retrograde cholangiopancreatography(ERCP)to guide the wire for sphincterotomy and stone extraction,followed by laparoscopic cholecystectomy.However,ERCP has limitations in handling complex cases.In response,our team pioneered a new method of treating choledocholithiasis combined with stenosis of the ampulla of Vater via a transabdominal approach.This study was performed to investigate the feasibility and efficacy of this method,aiming to provide a new therapeutic option for clinical practice.Methods:A randomized controlled study was conducted with 120 patients treated at Chengdu Second People's Hospital from 2021 to 2023 for gallbladder stones and choledocholithiasis with stenosis of the ampulla of Vater.Patients were divided into an observation group and a control group,with 60 cases in each group.The observation group underwent laparoscopic cholecystectomy with choledochotomy for stone extraction,followed by retrograde guidance of duodenal papillary sphincterotomy through the opened bile duct,simultaneously treating gallbladder,bile duct stones,and stenosis of the ampulla of Vater.The control group underwent traditional ERCP approach for sphincterotomy,stone extraction,and laparoscopic cholecystectomy.Perioperative variables were collected for both groups and the surgical outcomes were compared.Results:Among the 120 patients,54 were male and 66 were female.There were no statistically significant differences between the two groups in terms of stone extraction success rate,intraoperative blood loss,postoperative 24-h total bilirubin,direct bilirubin,transaminases,white blood cell count,jaundice relief time,or incidence rates of bile leakage,retroperitoneal bleeding/infection,and severe pancreatitis(all P>0.05).The observation group had significantly shorter average operative time and postoperative hospital stay compared to the control group(98.67 min vs.110.8 min,P<0.05;3.81 d vs.5.61 d,P<0.05).Additionally,the observation group had a significantly lower incidence of postoperative hyperamylasemia and/or hyperlipasemia and mild pancreatitis(1.67%vs.25.00%,P<0.001;0 vs.10%,P=0.027).Conclusion:The novel transabdominal approach is superior to the ERCP approach in terms of reducing surgery time and hospitalization time,and it carries a lower risk of postoperative mild pancreatitis and hyperamylasemia and/or hyperlipasemia.The stone extraction success rate is comparable to that of ERCP,making it a viable alternative treatment option.

Enterotoxigenic Escherichia coli(ETEC)is the main cause of diarrhea in piglets,which seriously threatens the development of animal husbandry.In order to effectively prevent and con-trol piglet diarrhea,the neutralizing epitopes of the screened K88(FTDYEGASVELRKPDGGT-NK),K99(NVGNGSGGANIN),987P(LAAPAENNTSQAN),F41(VMAADWTEGQPGDII)and F18(PPNAQTYPLSSGDLK)ETEC adhesins were assembled to construct the multi-epitope fusion antigen(MEFA),whose immunogenicity was verified by in silico simulations and in vitro.The computer simulation results showed that MEFA has excellent physicochemical properties and reasonable prediction results of secondary and tertiary structures and exhibits strong interaction force when docking with toll-like receptor 3,and the kinetic simulation shows that it can be stably presented.In vitro validation results showed that anti-K88,K99,987P,F18 and F41 IgY could spe-cifically recognize MEFA proteins.After immunization of laying hens with MEFA,the titers of IgY antibodies against the five ETEC were higher than those of non-specific IgY.In addition,specific anti-MEFA IgY significantly inhibited the adhesion of K88,K99,987P,F41 and F18 ETEC strains to piglet intestinal IPEC-J2 cell lines.These results indicate that MEFA has strong immunogenicity and can be used as an ideal vaccine candidate for the prevention of K88,K99,987P,F41 and F18 ETEC infection,and provide reference and technical support for the development of multivalent broad-spectrum ETEC vaccine candidates.

Overlap extension PCR is a common method for site-directed mutagenesis. As objective gene sequence growing longer, it is often difficult to obtain the target product in the second round of PCR, and it is highly possible to introduce unexpected mutations into a long gene fragment by PCR. To circumvent these problems, we can only amplify a small gene fragment which contain the target mutation by overlap extension PCR, and then ligate it with vector to get target plasmid. If the restriction site at the end of the amplified fragment was not a single one on plasmid vector, double fragments ligation method could be used to construct target plasmid. Partial amplification, combined with double fragments ligation, could solve lots of problems in long gene mutagenesis. Taking retinoblastoma gene 1 S780E mutagenesis as an example, it is difficult to amplify whole retinoblastoma gene 1 by overlap extension PCR because of long fragment interfering the overlapping extension of second round PCR. However, it is relatively easy to amplify the F3 (1 968-2 787) fragment which contains target mutation S780E. There is a Nhe I site which can be used for ligation on 5' end of F3 fragment, but another Nhe I site on the plasmid restrained from doing so directly. In order to circumvent this obstacle, we ligated F3 fragment, combining with F2 (900-1 968) fragment which was digested from wild type plasmid, with the vector which contain F1 (1-900) fragment of the gene. That double fragments ligated with one vector at the same time, though less efficient, can recombine into a complete plasmid. The sequences of the two selected recombinant plasmids were consistent with the target mutation, which verified the feasibility of this scheme. As an improvement of overlap extension PCR, partial amplification and double fragments ligation methods could provide solutions for site directed mutagenesis of many long genes.
Base Sequence ; Cloning, Molecular ; Genetic Vectors ; genetics ; Mutagenesis, Site-Directed ; methods ; Nucleic Acid Amplification Techniques ; Plasmids ; Polymerase Chain Reaction

OBJECTIVE： To determine and compare the contents of catalpol and aucubin in different parts （root， stem， leaf and flower） of wild Centranthera grandiflora， and to provide reference for the selection of medicinal parts and source development. METHODS： HPLC method was used to determine the contents of catalpol and aucubin in root， stem， leaf and flower of wild C. grandiflora， and the contents of different parts were analyzed comparatively. The determination of catalpol was performed on Agilent TC-C18 column with mobile phase consisted of methanol-0.1％ phosphoric acid （1 ∶ 99， V/V） at the flow rate of 1 mL/min； the detection wavelength was set at 210 nm， and sample size was 20 μL. The column temperature was 35 ℃； the determination of aucubin was performed on SPHERI-5RP-C18 column with mobile phase consisted of acetonitrile-water （3 ∶ 97， V/V） at the flow rate of 1 mL/min； the detection wavelength was set at 205 nm， and sample size was 20 μL； the column temperature was 25 ℃. RESULTS： The linear range of catalpol and aucubin were 0.061 5-3.321 and 0.000 36-0.216 mg/mL （all r＝0.999 9）. The limits of detection were 0.016 and 0.007 μg/mL. The limits of quantitation were 0.052 and 0.023 μg/mL. RSDs of precision， stability （24 h） and reproducibility tests were all lower than 2.00％ （n＝6）. The average recoveries were 99.34％ and 99.61％， and RSDs were 1.06％ and 1.12％， respectively （n＝6）. The average content of catalpol in root， stem， leaf and flower wild C. grandiflora were 1.609， 3.030， 11.095 and 1.921 mg/g， respectively. The contents of aucubin in different parts were 0.441， 0.020， 0.005 and 0.006 mg/g，respectively. CONCLUSIONS：The established HPLC method meets the requirements of quantitative analysis. Catalpol is mainly distributed in the leaves of wild C. grandiflora， and aucubin is mainly distributed in the roots of wild C. grandiflora. The experimental conclusion provides a reference for the reasonable selection of different medicinal parts as raw materials to develop medicine with different efficacy.

Objective To evaluate laparoscopic balloon nasobiliary biliary drainage (LBNBD),vs ureteral catheter drainage in one stage laparoscopy,choledochoscopy and duodenoscopy choledocholithotomy and primary closure of the small calibered common bile duct (diameter 0.3-0.8 cm).Methods During the period of Apr 2010 to Nov 2016 102 cases were enrolled including 50 cases receiving LBNBD and 52 cases using ureteral catheter drainage.Results Between the two groups,LBNBD was superior to ureteral catheter drainage in all the following parameters:the operation time,intraoperative blood loss,postoperative liver function,blood amylase and other laboratory indicators,gastrointestinal function recovery time,gastrointestinal symptoms and electrolyte imbalance,postoperative hospital stay,and bile duct drainage time with all differences statistically significant (P ＜ 0.05).Bile drainage differences during the postoperative first 3 days (averagely 200-400 ml a day) were not statistically different (P ＞ 0.05).Postoperative pancreatitis,bile leakage,and hemobilia were not statistically different (P ＞ 0.05).Conclusions The use of LBNBD is safe and effective in endoscopic choledocholithotomy in cases of small calibered common bile duct.

Objective To explore the application of failure mode and effect analysis in management of precision instrument in operating room of ophthalmology department.Methods We designed the handover process between operating room of ophthalmology department and supply room,the process of using precision instrument,the workflow of instrument management in nurses. And then,we assessed and analyzed the potential failure mode in the management of precision instrument with the method of failure mode and effect analysis.Besides,failure mode was also evaluated and improved.Results There were failure modes with high risk in the management of precision instrument including that the handover process between operating room of ophthalmology department and supply room was not precise enough (risk priority number,RPN=448);the staff of handover process was not fixed (RPN=360);there was no full-time cleaning staff in supply room (RPN=288);use registers in nurses and the check in precision instrument were all in a lack of standardization (RPN=210, 180) and so on. After adopting targeted corrective actions,the corresponding scores of RPN fell to 0,0,40,70, 90.Conclusions The application of failure mode and effect analysis in management of precision instrument in operating room of ophthalmology department has good effects with no related adverse event on precision instrument.