Objective To discuss the prevention and treatment of bile duct injures io the gallbladder bed during laparoscopic cholecystectomy(LC).Methods A retrospective analysis was made on clinical features,treatment,and curative effects of 15 cases of bile duct injures in the gallbladder bed out of 2 032 cases of LC from January 1997 to December 2004.Results There were 5 cases of acute attack of chronic calculous cholecystitis and 10 cases of chronic atrophic calculous cholecystitis.The location of injury was at the branches of the right hepatic ducts in 8 cases and at aberrant bile ducts in 7 cases.The injury was treated with titanium clipping in 8 cases,primary suture in 5 cases,and open surgery of bile duct repair in 2 cases because the injury was relativly large and the site of injury was near the trunk of the right hepatic duct.Postoperatively,bile leakage happened in 1 case and was cured by drainage for 5 days.Follow-up surveys for 6~36 months (mean,23 months) showed free of symptoms and no jaundice or cholangitis.Conclusions Surgical dissection closely near the gallbladder wall when mobilizing the gallbladder is the key to preventing bile duct injuries in the gallbladder bed during laparoscopic cholecystectomy.Prompt detection and proper management of the injury have satisfactory curative effects.

Objective To investigate the treatment of common bile duct or common hepatic duct transverse injures(CBDTI) in cholecystectomy.Methods From January 1993 to March 2008,3362 patients received cholecystectomy in our hospital,among them 5 developed CBDTI.We reviewed the clinic feature,management,and outcomes of these patients. Results Of the 5 patients,2 had common hepatic duct transverse injuries and 3 showed common bile transverse injuries.One of the five received postenterobiliary drainage and the other four underwent choledoch end-to-end anastomosis.No biliary leakage was found after the operation.One patient developed biliary stenosis and cholangitis after the end-to-end anastomosis and thus received postenterobiliary drainage.This case was followed up for 15 years,during which no abdominal pain,fever or jaundice occurred.The patient who underwent postenterobiliary drainage showed symptoms of cholangitis for 3 times whithin 2 months after the treatment;therefore,conservative therapy was carried out.Afterwards,the patient was cured and showed no abdominal pain,fever or jaundice during a 5-year follow-up.The other 3 patients who received end-to-end anastomosis were followed up for 1,2,or 4 years,no complications were noticed during the period.Conclusions Based on personal experience and the type of injury,surgeons may chose choledoch end-to-end anastomosis or postenterobiliary drainage to treat CBDTI.

Objective:To improve the immunoblotting of immunoprecipitated proteins and decrease the interference of immunoprecipitation antibody in the interaction of endogenous proteins. Methods:Transient transfect cells with fusion protein expression vector containing the targeted S5b gene and the FLAG tag, the transfected cells or untransfected cells were harvested to study the exogenous or endogenous protein interaction. The total cell lysate was immunoprecipitated by specific antibody. Then the eluted immunocomplex was separated by SDS-PAGE, and the TrueBlot antibody or conventional antibody was used as the secondary antibody for immunoblotting detection of S5b and its partner (Rpt1 and Rpt2). Results:Clear immunoblotting bands for S5b, Rpt1 and Rpt2 were obtained. Conclusion:TrueBlot antibody prefers the immunoblot antibody to immunoprecipitation antibody, and decreases the interruption of immunoprecipitation antibody to display clear protein band.

Objective To apply a fluorescent nucleotide dye Sybr Green I in detecting telomerase activity and assess its clinical value in the diagnosis of colorectal carcinoma. Methods Telomerase activity was (measured) by telomeric repeat amplification protocol (TRAP) combined with fluorescent nucleotide dye Sybr Green I in 46 cases of colorectal carcinoma tissue specimens and 19 tumor-adjacent tissue (specimens). Results (Telomeric) repeat amplification product was displayed as clear bands in PAGE stained with Sybr Green I; the positive rate of telomoerase activity in colorectal carcinoma tissue was 89.13%, whereas no telomerase (activity) was observed in 19 tumor-adjacent tissue specimens. There was no (relationship) between telomerase (activity) and the level of cell differentiation, Dukes stages, or metastasis of lymph nodes. Conclusions The method of TRAP combined with Sybr Green I to determine telemerase (activity) is an important method for the diagnosis of colorectal carcinoma.

Objective To study peripheral blood lymphocytic ?-adrenoceptor(?-AR) densities and ?_1-adrenoceptor gene(?_1-AR mRNA) expression levels in patients with heart failure,to investigate the influences of different ?-adrenoceptor antagonists in ?-AR densities and ?_1-AR mRNA expression levels.Methods 104 cases of patients with heart failure were randomly divided into non-?-adrenoceptor antagonist group(35 cases),metoprolol group(34 cases) and carvedilol group(35 cases),we repeatly determined the ?-AR densities and ?_1-AR mRNA expression levels after two-month therapy.Results The ?-AR densities and ?_1-AR mRNA expression levels in heart failure group reduced significantly compared to these in normal persons(P0.05).After two-month therapy,?-AR densities and ?_1-AR mRNA expression levels in metoprolol group were significantly higher than those in carvendilol group and non-?-adrenoceptor antagonist group(P0.05).Conclusions The peripheral blood lymphocytic ?-AR densities and ?_1-AR mRNA expression level down-regulates in heart failure,which correlate with the severity of heart failure but etiological factors.Metoprolol can up-regulate the ?-AR densities and ?_1-AR mRNA expression level,whereas carvedilol has no such effects.

Objective To construct the double-H1promoters siRNA expression cassettes(SECs) targeted to human telomere retrotranscriptase(hTERT),and investigate the interfering effect of its siRNA on hTERT gene expression in HepG2 cells.Methods SECs were constructed by fusing PCR,based on two different human telomerase hTERT gene fragments.When SEGs transferred into HepG2 cells respectively,the SECs were transcripted to the siRNA.The interfering effect of SECs on the telomerase activity in the cells was assessed by telomeric repeat amplification protocol(TRAP) and PCR-EIA.Results SEGs were successful constracted,and the telomerase activity was significantly inhibited when the HepG2 cells were tranfected with SECs.Conclusions The siRNA SECs display a definite RNA interference effect on the expression of telomerase.This method of SECs preparation can be applied for RNAi research in tumor inhibition.

BJECTIVE: To observe peripheral T lymphocytes changes of patients with lumbar disc herniation, and investigate the relationship between the type of herniation, signs and T lymphocyte subsets. METHODS: Blood samples from 49 patients with single.level lumbar intervertebral disc herniation were collected from General Hospital of Tianjin Medical University from January to October 2008, including 26 males and 23 females. There were 22 cases of L_(4～5) and 27 cases of L_5S_1. In the straight leg raising (SLR) test before treatment, 34 cases were positive and 15 were negative. According to the position of intervertebral disc observed in operation, the patients were divided into ruptured disc herniation (RDH, n=30) and degenerative disc herniation (DDH, n=19) groups. In addition, blood samples from 20 healthy blood donors were collected as control group. T lymphocytes subsets were detected by flow cytometry. RESULTS: The levels of CD3~+, CD4~+, and CD4~+/CD8~+ in RDH group were significantly higher than control group (P < 0.05), and the levels of CD8~+ in RDH group were significantly lower than control group (P < 0.05). The levels of CD4~+ and CD4~+/CD8~+ in positive SLR test group were significantly higher than patients with negative results, but significantly lower levels of CD8~+ was found in positive group than negative group (P < 0.05). The positive rate of SLR test in RDH group was significantly higher than that of SLR testing in DDH group. CONCLUSION: Changes of T lymphocyte subsets in peripheral blood were found after herniation of lumbar intervertebral disc. T lymphocytes mediated immune response may play an important role in the occurrence and development of signs in patients with herniated lumbar intervertebral disc. The SLR test may help to confirm that the disc herniation may cause nerve root impairment by mechanical loading or inflammatory stimulus.

Objective To reveal and compare the immunogenicity of glycoprotein from nucleus pulposus and collagen from anulus fibrosus in rabbit intervertebral disc. Methods According to the similarity principle of genome biology and the amount of nucleus pulposus in different intervertebral disc, Glycoprotein from nucleus pulposus was harvested from the intervertebral disc of 20 rabbits and placed into the psoas by anterolateral approach. Similar implantation of collagen from anulus fibrosus was performed in another 20 rabbits. The implantations of autologous muscle in rabbits serve as a control group.After one week,two weeks, four weeks, six weeks and eight weeks, the rabbits were killed, the tissue was harvested. HE staining was applied to detect the degree of vascularization and infiltration of lymphocytes. Immunohistochemistry were used for analysis of T-helper cells (CD4+) and cytotoxic T cells (CD8+) in the tissue. Results HE staining showed the infiltration of lymphocytes was found just one week after implantation and vascularization was seen at the 2nd week in the glycoprotein implanted group. However in collagen implant group,small amounts of lymphocytic and atypical vascularization were seen at the 4th week. Immunohistochemistry revealed that the proportion of activated T cells (CD4+ and CD8+) was significantly higher in the glycoproteinimplanted group at all time points, as compared with the collagen-implanted group and sham group. The proportion of activated T cells was higher in collagen-implated group than that of in sham group . Conclusion The glycoprotein in nucleus pulposus is easier to induce the differentiation and mature of T cell. The antigenicity of glycoprotein in nucleus pulposus was stronger than that of collagen in anulus fibrosus in rabbit intervertebral discs.

Objective To investigate the possibility of establishing a Modic changes (MCs) animal model,and explore the pathogenesis of MCs through imaging,histology and molecular biology experiments.Methods Fifty four New Zealand rabbits (weight 2.5-3.0 kg,half male and half female) were randomly divided into 3 groups:sham group (n=l8),muscle embedment group (n=18) and NP embedment group (n=18).In NP embedment group,the L4-5 and L5-6 discs were exposed by the lumbar anterolateral surgical approach.A 16 G needle was used to puncture the L5-6 vertebral body close to the epiphyseal plate.The depth of the drilling was approximately 3 mm.A 5 ml syringe was then put into the L4-5 intervertebral disc and extracted the NP,which was injected into the drilled hole of the vertebral body.The muscle embedment group and sham group shared the same operating procedures and drilling methods with the NP embedment group.Some pieces of muscle acquired from paraspinal muscles were put into the drilled hole in muscle embedment group,while nothing was put into the drilled hole in sham group.After that,the bleeding stopping,tissue washing and suture were done in all groups.12 weeks,16 weeks and 20 weeks after the surgery,MRI scan was applied to each group.All the specimens were tested by HE staining,real-time fluorescence quantitative PCR and Western blot to observe the expression of inflammatory cytokines.Results After modeling for 12 weeks,16 weeks and 20 weeks,MRI showed low signal changes on T1WI and mixed high signal in the context of low signal changes on T2WI in the NP embedment group.However,the muscle embedment and sham group showed no significant signal changes.Gross observation and HE staining confirmed that there was abnormal tissue proliferation in the imbed site of the NP embedment group.RT-PCR and Western blot showed high expression of IL-4,IL-17 and IFN-γin the NP embedment group,which were positively correlated with the length of the postoperative period.There was no significant difference between the muscle embedment group and sham group.Conclusion The puncturing of vertebral body close to endplate and putting nucleus into it can create an animal model of MCs.Autoimmune factors may play an important role in MCs.

Objective To observe the effect of the nutrition management according to the JCI(Joint Commission on Accreditation of Healthcare Organizations)management standard for critically ill pediatric patients with mechanical ventilation.Methods A total of 330 cases with mechanical ventilation were en-rolled in the control group from Hunan Province Children's Hospital PICU between Jan.2012 and Dec.2012, and these pediatric patients were managed with the conventional nutrient management.A total of 359 pediatric patients with mechanical ventilation were admitted to the experimental group from Jan.2013 to Dec.2013 and these patients were managed with nutrient management based on JCI standards.The length of mechanical venti-lation,stay in ICU,stay in hospital were compared between two groups,and the incidence of ventilator-associ-ated pneumonia,abandonment rate and mortality were also compared between experimental group and control group.Results The length of mechanical ventilation in control group[(8.39 ±1.34)days]was longer than that of experimental group[(5.69 ±1.12)days].The length of stay in PICU for control group[(12.32 ± 1.37)days]was more than that in experimental group[(9.42 ±1.53)days].The length of stay in hospital for control group [(15.37 ±2.16 )days]was higher than that of experimental group [(12.63 ±2.29 ) days].The incidence of ventilator-associated pneumonia(9.7%)in control group was higher than that of ex-perimental group,and the giving up or mortality rate in control group(8.48%)was higher than that of exper-imental group(4.35%).And there were significant differences by statistical analysis(P ﹤0.01 ,respective-ly).Conclusion According to the nutritional management in JCI standard,the length of mechanical ventila-tion,stay in PICU and stay in hospital time,the incidence of ventilator-associated pneumonia,and the aban-doned or mortality rate were reduced for critically ill pediatric patients with mechanical ventilation.