In order to construct a prokaryotic expression vector of human receptor syt II N-fragment and to express recombinant MBP-Syt fusion protein in E.coli and to purify and identify its activity. According to codon preference of E.coli, a DNA fragment encoding human syt II N-fragment was synthesized, and then cloned into prokaryotic vector pMAL-c2x for sequencing. Then the recombinant plasmid pMAL-Syt was introduced into E.coli ER2566 by transformation for expression and the obtained engineered bacteria were induced by IPTG. The fusion protein was purified by amylose resin affinity chromatography and identified by SDS-PAGE and Western blot. The binding activity of the protein was determined by ELISA. It is concluded that MBP-Syt protein is of good binding activity.

Hemodynamic monitoring is important for critically ill patients in emergency medicine. While the conventional static hemodynamic monitoring may not accurately reflect the hemodynamic status, functional hemodynamic monitoring can dynamicly and individually monitor the hemodynamic status, and thus becomes a valuable supplementation for conventional static hemodynamic monitoring. This article reviews the limitations of conventional hemodynamic monitoring and introduces the methodology of functional hemodynamic monitoring.
Emergency Medicine ; methods ; Hemodynamics ; physiology ; Humans ; Monitoring, Physiologic ; methods

Objective To explore the relationship of post-transplant major histocompatibility complex class I chain-related gene A(MICA)antibody status and renal allograft function in clinical stable phase.Methods Fifty-seven patients accepted renal allografts followed up for at least 6 months were detected with the levels and specialties of MICA antibodies by Flow PRATM beads.Simultaneously,their serum ereatinine levels were tested as well.The impact of MICA antibody status on renal allograft function was assessed.Results Among the 57 patients,38 cases showed no HLA and MICA antibody.11 cases had HLA antibodies but not MICA antibody,8 cases had MICA antibodies and 3 cases had both MICA and HLA antibodies.There were 5 patients with MICA019 antibodies.3 patients with MICA027 antibodies,2 patients with MICA018 antibodies,while 1 patient with MICA004 and MICA017 antibodies,respectively.There were 9 patients with antibody positive score higher than 6,accounting 75%(9/12).Except age,there was no significant difference between patients with positive and negative MICA antibodies in the aspects of blood transfusion history,CDC,and cold ischemia time(P＞0.05).The average ages were(32.5±7.9)years for MICA antibodypositive patients and were(43.0±1 0.4)years for MICA antibody-negative patients(P=0.008).MICA antibody-positive patients without HLA antibody had higher serum creatinine level[(117.20±12.30)μmol/L]than MICA and HLA antibody-negative patients[(89.40±28.95)μmol/L,P＜0.05].Conclusions The measurement of MICA antibodies has prognostic value in the assessment of patients without HLA antibodies after renal transplantation.MICA antibody positive has clear association with chronic renal allograft function decline.

Objective To summarize the clinical experience of three different treatments for gallbladder and common bile duct stones.Methods The clinical data of 180 cases of gallbladder stones combined with bile duct stones undergoing surgery from May 2010 to May 2013 were retrospectively analyzed.They were divided into three groups,A group of 60 patients underwent a period of endoscopic sphincterotomy (EST),under the second phase of laparoscopic cholecystectomy (LC);Group B 60 patients underwent laparoscopic cholecystectomy (LC) + laparoscopic common bile duct exploration surgery (LCBDE) + T tube drainage;Group C 60 patients underwent conventional open cholecystectomy (OC) + bile duct exploration (OCBDE) + T tube drainage.Results A group of 53 cases completed surgery successfully,5 cases of remaining 7 patients failured in the first phase surgery,2 cases of the 5 patients did LC + LCBDE,3 of the 5 patients underwent conventional surgery.Two patients underwent the conventional surgery in the second phase surgery.B group of 57 cases completed surgery successfully,three cases convert to open surgery.All of the group C completed the surgery successfully.Group A complication was the most in the three groups (P＜0.05);group B had the shortest time of hospitalization (P＜0.05),the complication rate was lower than that in group A (P＜0.05),the complication had no significant difference between A and B.(P and group B ＞ 0.05),group B had shortest operation time (P＜0.05);no statistically significant differences were found among three groups in fasting time.Conclusion Three treatment methods have advantages as well as disadvantages,a reasonable treatment should be selected according to the specific circumstances.

To produce antibodies capable of neutralizing botulinum neurotoxin type B(BoNT/B),We cloned the carboxy-terminal end of Hc containing the major determinants responsible for specific toxin,induced and purifed.The heavy-chain and kappa light-chain variable region gene repertoire of immunoglobulin were amplified individually from the spleen cell mRNA by RT-PCR and joined as a single-chain Fv(scFv)DNA fragment.These fragment were cloned into the phagemid pCANTAB5E and the phage display library was constructed.Results showed that the high affinity scFv was obtained after 4 rounds of panning,with its DNA sequence conforming to that of mouse antibody.

Objective To study the influence of human leucocyte antigen(HLA) and major his-tocompatibility complex class Ⅰ chain-related gene A (MICA) specific antibodies on renal allograft function and graft rejective reaction by monitoring their changes from preoperative to postoperative pe-riods. Methods Twenty-seven patients with renal aliografts were tested with the specificity of anti-HLA antibodies (anti-HLA class Ⅰ and anti-HLA class Ⅱ) and anti-MICA antibodies and their posi-tive value changes by flow PRATM beads. The HLA genotype was integrated to distinguish donor specific antibody(DSA) and non-donor specific antibody(NDSA). Their serum creatinine levels and clinical data were analyzed simultaneously. Results Of the 27 patients, 22 cases accepted renal transplantation from dead bodies and 5 eases accepted from live donors. Except 1 failed patient, the other 26 patients had good functional renal allografts. Twenty-four survival patients were followed up on month 1, 3, 6 and 12 after transplantation. Seven out of 27 patients had pre-exist antibody before transplantation. Among them, 2 patients had anti-HLA antibody; 3 patients had anti-MICA antibody; 2 patients had both anti-HLA and anti-MICA antibody. Three patients with no anti-HLA and anti-MICA antibodies before transplantation created antibodies after transplantation from 3 to 6 months. One patient created NDSA after transplantation and appeared chronic rejection. There were 3 patients who had anti-MICA antibodies before transplantation. The expression levels of antibodies had changed from high to low, but the specific anti-MICA antibody had not changed during the follow-up on month 1, 3, 6 and 12 after transplantation. The patient with pre-transplantation low level of anti-HLA class Ⅱ antibody appeared acute rejection with fever and his CMV was positive as well. The patient's SCr levels changed from 171 μmol/L to 236 μmol/L after I to 3 months post-transplantation. Twenty-four patients were divided into positive and negative groups according to the specific antibody. There was significant difference of SCr levels between the 2 groups 1 month and 1 year after transplantation(P= 0.03, 0.05). Conclusions It is important to detect the specificity and positive value of anti-HLA antibodies and anti-MICA antibody regularly during the post transplantation follow-up. This will make an effective therapy for decreasing the occurrenee and development of acute or chronic rejection and hy-pofunction on renal allograft.

OBJECTIVETo explore the traumatic mechanism and the effectiveness of conservative and operative therapy for intra-articular fracture of calcaneus.

METHODSForty-eight patients (52 feet) with intra-articular fractures were divided into two groups named as conservative group and operative group. There were 30 patients in the conservative group, including 26 males and 4 females, ranging in age from 2 to 73 years, with an average of (49.2 +/- 8.5) years. The patients in conservative group were treated with manual reduction, external fixation and functional exercise at early time. Among 18 patients in the operative group, 14 patients were male and 4 patients were female, ranging in age from 26 to 66 years,with an average of (50.1 +/- 19.3) years. The patients in the operative group were treated with open reduction and internal fixation. The height and width of calcaneus, Böhler and Gissane angles were measured before and after treatment. The assessment in the function of ankle joints was scored according to AOFAS scoring system.

RESULTSAll the patients were followed up, and the duration ranged from 8 to 18 months,with a mean of 12 months. The time of weight-bearing ranged from 3 to 6 months, averaged 4 months. The fractures were fused completely on average of 12 months. The height and width of calcaneus, Böhler and Gissane angles had no statistically significance between two groups. According to AOFAS criteria,in the conservative group, 18 patients reached an excellent result, 6 good, 5 poor and 1 bad; in operative group, 4 patients reached an excellent result, 7 good, 6 poor and 1 bad. The therapeutic effect of conservative group was better than that of operative group.

CONCLUSIONOperative therapy in intra-articular fracture of calcaneus with steel plate can directly perceive the restoration (or roughly restoration sometimes) of the anatomical shapes of calcaneus, while conservative therapy has advantages such as restoration of the anatomical shapes of calcaneus, early functional practice, early healing, and avoiding operational complications as well.

Adolescent ; Adult ; Aged ; Calcaneus ; injuries ; physiopathology ; surgery ; Case-Control Studies ; Child ; Child, Preschool ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Intra-Articular Fractures ; physiopathology ; therapy ; Male ; Middle Aged

OBJECTIVETo detect potential mutation of Doublecortin (DCX) gene in a patient featuring X-linked subcortical laminar heterotopia (X-SCLH) and epilepsy.

METHODSMutation of the DCX gene was screened by PCR and direct sequencing. Pathogenicity of the mutation was analyzed with a PolyPhen-2 software.

RESULTSA de novo missense mutation c.971T>C (p.Phe324Ser) was discovered.

CONCLUSIONA diagnostic method for X-SCLH has been established, which may facilitate diagnosis and genetic counseling of patients featuring this disease.

Agenesis of Corpus Callosum ; diagnosis ; genetics ; Base Sequence ; Brain ; pathology ; Child ; Classical Lissencephalies and Subcortical Band Heterotopias ; diagnosis ; genetics ; Electroencephalography ; Epilepsy ; diagnosis ; genetics ; Exons ; Female ; Humans ; Magnetic Resonance Imaging ; Microtubule-Associated Proteins ; genetics ; Mutation ; Neuropeptides ; genetics

AIM:To investigate the expression and localization of autophagy related protein microtublule associated protein 1 light chain 3 (LC3) at various stages of follicular development and atresia in the mice.METHODS:On 0,1,2,3,4 and 5 day after intraperitoneal injection of pregnant mare serum gonadotropin (PMSG),expression and positioning situation of autophagy related protein LC3 and apoptosis related protein cleaved caspase-3 were examined by the method of immunohistochemical staining.The protein levels of cleaved caspase-3 and LC3 were determined by Western blot in cultured mouse granulosa cells after incubation under serum-free conditions in the absence or presence of FSH.LC3 subcellular localization in granulosa cells were studied by the method of immunofluorescence.RESULTS:The LC3 protein expressed in granulosa cells during all developmental stages mainly.Granulosa cells of atretic follicles that showed intense staining of cleaved caspase-3 and LC3.The protein levels of cleaved caspase-3 and LC3-Ⅱ in the granulosa cells significantly decreased at 1 d and 2 d after intraperitoneal injection of PMSG (P ＜ 0.05).The protein levels of cleaved caspase3 and LC3-Ⅱ in the granulosa cells increased in turn on 3,4 and 5 day after intraperitoneal injection of PMSG.The positive correlation between LC3-Ⅱ and cleaved caspase-3 protein levels was observed (r2 =0.8299,P ＜ 0.05).The LC3-Ⅱ protein expressed with punctuate structures in granulosa cell cytoplasm cultured under serum-free conditions in the presence of FSH.CONCLUSION:LC3 is expressed in the follicular granulosa cells with cell specificity and regional specificity.Autophagy is induced mainly in granulosa cells during folliculogenesis and shows positive correlation with apoptosis.Ovarian granulosa cell autophagy and apoptosis are gonadotropic hormone dependent.

This meta-analysis was carried out to evaluate the relationship between NM23 expression and the prognosis of patients with colorectal cancer.We searched PubMed,EMBASE and Web of Science for relevant articles.The pooled odd ratios (ORs) and corresponding 95％CI were calculated to evaluate the prognostic value of NM23 expression in patients with colorectal cancer,and the association between NM23 expression and clinicopathological factors.In total,2289 patients were pooled from 24 available studies.The incorporative OR combined by 16 studies with overall survival showed that high NM23 expression was associated with better overall survival (OR=0.67,95％CI:0.49-0.93,P=0.02,I2=56％,Ph=0.004).And a new estimate without heterogeneity was produced when only combining high-quality studies (OR=0.70,95％CI:0.56-0.86,P=0.0007,I2=46％).In disease free survival (DFS),we also obtained a good prognosis (OR=0.30,95％CI:0.14-0.68,P=0.004).Although we failed to find any significance in N status (P=0.10),elevated NM23 expression was related to well tumor differentiation (OR=0.60,95％CI:0.44-0.820,P=0.001) and Dukes' A&B (OR=0.55,95％CI:0.32-0.95,P=0.03).These results indicated that over-expressed NM23 might be an indicator of good prognosis,well tumor differentiation and Dukes' A&B of patients with colorectal cancer,but no significance was found in N status.