Objective:To determine the roles of constitutively activated signal transduction pathway ERK1/2(p44/42 MAPK)in controlling cytotoxicity or proliferation activity of natural killer cell lines.Methods:Whole cell extracts from human NK cell lines(YT,IL 2 independent;NK 92,IL 2 dependent) was used in Western blot to detect the constitutively activated signal transduction pathway(s) in human NK cell lines. Specific inhibitor PD098059 was used to abrogate phosphorylation of ERK1/2 for further evaluation. A MTT based method was applied to analyze the cytotoxicity and proliferation capability of NK cell lines before and after specific inhibition of ERK1/2 activation by PD098059.RT PCR protocol was applied to analyze the cytotoxic related molecules possibly engaged with the ERK1/2 signal transduction pathway.Results:Western blot showed that signal transduction pathway ERK1/2,NF ?B and STAT3 was constitutively phosphorylated in two representative human NK cell lines YT and NK 92 which killed NK sensitive K562 target efficiently,other sigal pathways(STAT1,STAT6,p38 MAPK and PI 3K) were not activated.ERK1/2 inhibitor PD098059 apparently inhibited cytotoxicity of NK cell lines but did not influence their proliferation potential, RT PCR analysis revealed that the expression of lytic related molecules including IFN?,FasL,and perforin was downregulated to distince degree by PD098059 as it blocked the phosphorylation of ERK1/2 in NK cell lines.Conclusion:The constitutively phosphorylated signal pathway ERK1/2 in NK cell lines mainly involved in transducing signals controlling the cytotoxic capacity but not the proliferation potential of NK cells,ERK1/2 regulated the lytic capacity of NK cells by inducing the expression level of lytic related molecules including IFN?、FasL and perforin. [

Purpose　The aim is to obtain the cDNA sequence of encoding extramembrane human FL gene with high level expression in E.coli. Methods　The primers were designed based on the known FL cDNA sequence. The total RNA was isolated from fetal liver cells , and then RT-PCR was performed. The fragment was cloned into pUC-18T vector, and further sequenced by automatic sequence analyzer. The gene was inserted into GST fusion expression vector between BamH Ⅰ and EcoR Ⅰ sites. The recombinant plasmid was transformed into E.coli strain DH5 α and induced with 1mmol/L IPTG.Results　The 546bp DNA fragment was amplified by RT-PCR method from fetal liver cells and its sequence was identical to the published sequence encoding human FL. The expressed fusion protein, with molecular weight of about 22kD, was about 10% of the total bacteria protein by SDS-PAGE and densitometry analysis.Conclusion　cDNA was cloned successfully. This study provided a basis for the further fundamental research and clinical application of FL.

In order to investigate the application of standard score in the examination of eight-year program's clinical practice and to keep consistency of learning level among groups, we translate the original scores into standard scores and discuss the results and how to define the excellent grade so that the scores may have more comparability.

Objective To understand the cue-induced craving, physiological reactions to heroin-related cues, and to explore the factors that impact the cue-induced craving. Methods 369 abstinent heroin addicts were exposed to videotapes of heroin using and simulacrum after the self relaxation. The craving, abstinent symp-toms were measured before and after cue exposures. The physiological reactions were measured by multi-biofeed-back instrument before, during, and after cue exposure. BIS-Ⅱ questionnaire was used to collect information about their impulsive characteristic. Results After cue exposure, the self-reports scores of craving (t=11.24, P< 0.01), physiological index including heart rate((76.6±11.3) beats/min, F=19.23), skin conduct ((7.48± 4.25)μs, F=53.99), pupil size (t=11.73) had significant change in heroin addicts (P<0.01). Logistic regres-sion results showed that the scores of BIS-Ⅱ were related to cue-reduced craving (P=0.001,95% CI: 1.015～1. 065). Conclusions Exposure to drug-related cues can induces craving and physiological reactions in long term abstinence heroin addicts. High impulsive characteristic is a risk factor to induce drug-related craving. The inter-vention strategies should consider the impulsive characteristic and make a comprehensive intervention program to prevent relapse.

Objective To investigate the protective effect of RLN revealed for RLN in high -risk thyroid surgery,and the clinical value of detecting nerve,thus to provide a reference for clinical treatment.Methods 132 patients of thyroid high -risk surgery were selected,according to whether to explode recurrent laryngeal nerve,they were divided into exposed group (80 cases)and non -exploded group(52 cases).The recurrent laryngeal nerve damage rate,clinical indicators of operation time were compared between the two groups.In the explode group,80 patients according to whether implementation of the nerve monitoring,were divided into the monitored group (36 cases)and non -monitored group(44 cases).The hours to find the nerve,recurrent laryngeal nerve injury rates and other indicators were compared.Results The operative time of the exploded group was (133.66 ±21.48)min,which was significantly longer than the non -exploded group,difference was statistically significant (t =3.994,P =0.025 ).The exposed group had similar incidence of hypocalcemia compared with the other groups,there was no statistically significant difference (P >0.05).The revealing recurrent laryngeal nerve injury rate of the exposed group was 2.50%,which was significantly lower than the non -exploded group,difference was statistically significant (χ2 =5.794,P =0.008).The recurrent laryngeal nerve positioning time of the monitoring group was (4.78 ±0.94)min,which was significantly shorter than the unmonitored group,difference was statistically significant (t =7.168,P =0.001).The recurrent laryngeal nerve injury rate of the monitoring group was 2.78%,showed no significant difference (χ2 =0.911,P =0.338).Conclusion In a high risk thyroid surgery,explode recurrent laryngeal nerve can reduce the rate of recurrent laryngeal nerve damage,nerve monitoring can effectively speed up the nerve localization time,but has no significant effect on reduce the rate of recurrent laryngeal nerve injury.

Objective:To study the effect of microRNA (miR)-330-3p on hepatic ischemia-reperfusion injury (IRI) in mice, meanwhile, and to determine potential molecular mechanism.Methods:Eighty male C57BL/6 mice, aged 7-8 weeks, 23-25 g, specific pathogen free, were randomly divided into 8 groups (10 mice in each group) using random number table: reperfusion 2 h group, 6 h group, 12 h group, 24 h group, sham group, miR-330-3p agomir group (preoperative injection of agonist), miR-330-3p antagomir group (preoperative injection of inhibitor) and miRNA-NC group. Except for the sham group, the hepatic IRI model were established in mice. Polymerase chain reaction (PCR), Western blot and immunohistochemistry were used to detect the expression of miR-330-3p and phosphoglycerate mutase family member 5 (PGAM5), cleave caspase-1 and GSDMD. Luciferase reporter assay was performed to investigate whether miR-330-3p directly targets PGAM5. At the same time, AML12 cells were also treated with miR-330-3p mimics/inhibitor or PGAM5 siRNA, then the expression of PGAM5, NLRP3, cleave caspase-1 and GSDMD were detected by Western blot analysis.Results:Level of miR-330-3p gradually decreased after reperfusion, however, mRNA level of PGAM5 was increased thereafter ( P<0.05) as compared with the sham group. Serum level of AST and ALT were decreased in miR-330-3p agomir group while that of were increased in miR-330-3p antagomir group as a function of time following reperfusion, and the differences were statistically significant (all P<0.05). Cleave caspase-1 expression was decreased in miR-330-3p agomir group but was increased in miR-330-5p antagomir group ( P<0.05). Luciferase reporter assay was performed to determine PGAM5 was a target gene of miR-330-3p. SiRNA-mediated knockdown of PGAM5 decreased level of GAM5 (0.24±0.09), NLRP3(0.12±0.07), cleave caspase-1 (0.15±0.07) and GSDMD (1.08±0.08) as compared with the siRNA-NC group (1.17±0.14), (0.36±0.09), (0.68±0.09), (1.36±0.08), and the differences were statistically significant (all P<0.05). Conclusion:MiR-330-3p can alleviate hepatic IRI in mice, which may be related to inhibition of PGAM5-induced pyroptosis.

BACKGROUND: Facial nerve injury causes facial nerve paralysis (or facial palsy) and even results in psychosocial disturbances of the patients. Repair the injured facial nerve and reconstruction of the nerve function as early as possible have been the primary concern in clinical studies.OBJECTIVE: To investigate the timing and surgical approaches for repairing facial paralysis in order to provides evidences for its therapeutic and prognostic evaluation.DESIGN: Case analysis based on patients.SETTING: Hospital of Plastic Surgery of Chinese Academy of Medical Sciences.PARTICIPANTS: Nine patients with traumatic facial paralysis hospitalized in the Hospital of Plastic Surgery of Chinese Academy of Medical Sciences from December 1993 to November 2001.METHODS: Facial nerve anastomosis was performed microsurgically along with the implantation of the sural nerve graft into orbicular muscle of the eye 3 or 4 months after nerve injury in the 9 patients. The clinical data of the patients were retrospectively reviewed.MAIN OUTCOME MEASURES: The facial nerve function was evaluated with House-Brackmann scale and photographs of the patients' faces before and after surgical treatment were taken.RESULTS: In the follow-up of these patients lasting 6 months to 2 years,all the patients attained satisfactory outcome.CONCLUSION: Early operation is crucial for the treatment of traumatic facial paralysis,with facial nerve anastomosis as the primary choice. The implantation of the nerve graft into muscle is also indicated for repairing traumatic facial paralysis in some cases. The importance of individualized treatment choice is reiterated on the basis of cicatrectomy and the extent and specific features of the injury.

Objective To investigate the prognosis of liver retransplantation in patients with transplanted liver function failure caused by viral hepatitis recurrence.Methods From January 20th 2003 to November 20th 2012,the clinical data of 215 patients with liver retransplantation were retrospectively analyzed.The survival of transplanted liver of 18 cases with liver retransplantation because of hepatitis recurrence (eight cases of hepatitis C and 10 cases of hepatitis B) was compared with that of 115 cases with liver retransplantation for biliary complications.The dysfunction of transplanted liver after first transplantation and the survival after second liver retransplantation of patients with hepatitis C recurrence were compared with those of patients with hepatitis B recurrence.The prognosis analysis was compared by survival curves made by Kaplan-Meier method.Results Biliary complications were the most common reason in 215 patients with second liver retransplantation and which accounted for 115 cases (53.5 ％).Eighteen cases were hepatitis recurrence (8.4 ％).There was no significant difference in survival rate of the second transplanted liver between patients with hepatitis recurrence and biliary complication (P =0.543).The dysfunction of transplanted liver occurred at early stage (in three months) after first liver transplantation in part of patients with hepatitis C recurrence.The dysfunction of transplanted liver almost all occurred two years after first liver transplantation in patients with hepatitis B recurrence.Among eight patients with hepatitis C recurrence,the second transplanted liver of five cases survived more than one year.All the second transplanted liver of 10 patients with hepatitis B recurrence survived more than one year.There was no significant difference between them (P =0.060).Conclusions The prognosis of liver retransplantation in patients with hepatitis recurrence is similar with that of patients with biliary complications.The prognosis of liver retransplantation in patients with hepatitis B recurrence is good.

The authors of this paper confirm the goal, factors and principles in arrangement of clinical practice, design a transplacement algorithm to solve the model, and after estimating the capacity to interns of affiliated hospital, make the schedule by that algorithm so as to relieve the relative short supply of good clinical education resourses.