Objective:To clone and express active domain of human granzyme A ( aGzmA ) and detect its biological activity.Methods:Human aGzmA gene was amplified by PCR from the full-length human granzyme A and inserted into prokaryotic ex-pression vector pET24a(+).The constructed recombinant plasmid pET24a-aGzmA was transformed to E.coli BL21(DE3) and induced with IPTG.The expressed product was identified by SDS-PAGE and Western blot.The recombinant protein was purified by the Ni2+affinity column chromatography and the enzyme activity was assayed with BLT substrate.Results: A DNA fragment of 700 bp was amplified by PCR.The recombinant plasmid pET24a-aGzmA identified by enzyme-digesting analysis and sequencing showed that aGzmA gene was inserted into vector plasmid correctly.SDS-PAGE analysis showed that there was a specific protein with a relative molecular mass of about 26 kD.Western blot analysis indicated that the protein could react with mouse anti-His monoclonal antibody specifically.The recombinant protein with high purity could be acquired from the inclusion bodies by the Ni2+ affinity column chromatography and the purified protein had good enzyme activity.Conclusion: The recombinant human granzyme A with good biological activity was prepared successfully.

0.05). There was no correlation between Bcl-2 and Fas positive cells and acetylcholine receptor antibody serum titre, disease grouping, age and sex in patients with or without hyperplastic thymic and thymoma.Conclusions In MG patients, the expression of Bcl-2 in hyperplastic thymus is upregulated. There is no correlation between the expression of apoptotic genes Bcl-2 and Fas and clinic relative factors.

Objective To evaluate prognostic factors for early bronchopleural fistula after pneumonectomy with non small cell lung cancer,and establish a validated clinical model to estimate the risk of early-BPF.Methods We reviewed the medical records of 429 patients who underwent pneumonectomy for NSCLC at our institution.We used univariate and multivariate analysis to identify potential independent risk factors for early-BPF after pneumonectomy for NSCLC.A model to estimate risk of early-BPF was developed by combining independent risk factors.Results The rate of early-BPF after pneumonectomy for NSCLC was 6.5％ (28/429).Three factors were independently associated with early-BPF:neoadjuvant therapy (HR:2.406),bleeding (HR:2.171)and diabetes (HR:1.144).A scoring system for early-BPF was developed by assigning 2 points for each major risk factor (neoadjuvant therapy and bleeding) and 1 point for each minor risk factor(diabetes).Scores were grouped as low (0-1),intermediate (2-3),and high (3),yielding the rate of early-BPF was 14％,27％,and 43％,respectively.Conclusion This clinical model is established on the basis of independent risk factors.This model can be used as a predictive tool for early-BPF after pneumonectomy for NSCLC.

Aim To explore the inhibition of Sinica Maxim′s extract( CSME) on resistant infections of burn wounds,such as the methicillin-resistant staphylococcus aureus ( MRSA ) , resistant pseudomonas aeruginosa (RPA) and resistant escherichia coli(RECO). Meth-ods The resistant strains were cultured by MH agar plates. After resistance genes of quality control strains were extracted and appraised, such as mecA, mexB, merA, qacE△1-sull, tnpU/A and mexB, etc, and then,some projects of CSME were detected,such as the antibacterial spectrum, the minimum inhibitory con-centration(MIC), different concentrations of sensitive rate and inhibition curves, etc. Finally, these results were compared with the inhibitory effects of some anti-biotics to determine the sensitivity rates of CSME. Re-sults The MIC of CSME was 62. 5 ,125 ,250 g · L-1 respectively on the MESA, RPA and RECO. The inhi-bition rates of CSME appeared concentration-dependent on these three kinds of resistant bacteria,and the inhi-bition rates of the multi-concentration CSME on RECO were significantly lower than on MRSA and RPA ( P<0. 05). While in MIC,the resistance rates of MRSA on carbenicillin, cefazolin, erythromycin were significant-ly higher than those of CSME(P<0. 05); The inhibi-tion zones of CSME were significantly smaller than those of ceftriaxone, cefepime, imipenem, but greater than those of other antibiotics( P<0. 05 ); The inhibi-tion zones of CSME on RPA were significantly smaller than those of carbenicillin, and greater than those of other antibiotics ( P <0. 05 ) . The inhibition zones of CSME on RECO were significantly smaller than those of ceftriaxone,cefepime,imipenem,ciprofloxacin,nitro-furazone,and greater than those of other antibiotics ( P<0. 05 ) . Conclusions CSME has a significant inhi-bition on burn wound infection with these three kinds of resistant bacteria,such as MRSA,RPA and RECO. It is prompted that CSME could become one of the effective drugs to control burn wound infections with multi-re-sistant strains.

Objective To investigate the protective mechanism of Interleukin 33 (IL-33) in preventing myocardial ischemia and reperfusion injury in rats. Methods A rat model with myocardial I/R with 32the adult male SD rats , which were randomly divided into 4 groups: SO group , I/R group , IL-33 + I/R group , SB230580 + IL-33 + I/R group. The levels of LDH, CK, TNF-α, IL-6, HMGB1, Bcl-2, total caspase-3, cleaved caspase-3 and P-P38 were detected. Results After reperfusion, IL-33 significantly decreased the levels of serum LDH and CK and the expression of TNF-α, IL-6 , cleaved caspase-3 , but significantly increased the expressions of Bcl-2, p-P38 (P < 0.05). SB230580 attenuated the protective role of IL-33 on myocardial I/R in a certain degree. Conclusions IL-33 may prevent myocardial I/R injury via inhibiting inflammation and cardiocyteapoptosis by way of P38 MAPK signaling pathway.

Purpose To observe therapeutic effect of electroacupuncture Neixiyan (Ex-LE 4 ) and Dubi (ST 35) in treating osteoarthritis of knee joint. Method All the 120 cases were randomly divided into electroacupuncture and control groups, 60 cases in each group,and they were given electroacupuncture and Ritalin slowreleased tablet respectively, and pain, mobility and swelling degree of knee joint were observed before and after treatments. Results In treatment group, the average score increased by 18, while in control group, it increased by 12.33 ( P < 0. 05 ) after treatment. Conclusion Therapeutic effect of electroacupuncture Neixiyan (Ex-LE 4) and Dubi (ST 35) in treating osteoarthritis of knee joint was better than that of administration of Ritalin slow-released tablet.

Objective To observe pathological features of peripheral vessel injury caused by ex-plosion shock wave so as to provide theoretical basis for emergency treatment, prevention and complication reduction of war extremity injuries. Methods A total of 18 rabbits were randomly divided into three groups (six rabbits in each group) and placed respectively at 1, 2 and 3 m away from the explosion cen-ter. The animal model with blast injury was made by using fluid dynamite that electrically exploded at 60 cm above the ground. The physical parameters of blast wave were recorded using pressure transducers (PCB, UAS). After explosion, the femoral arteries were examined grossly, histologically and immunohis-tochemically. Results The results showed that vascular endothelial cells were denudated, the spaces of contractile fiber cells increased and appeared puff, the vassular elastic fibers ruptured, flexed and de-formed visibly. Some parts of the vessel wall ruptured completely or partly, leading to bleeding. TUNEL staining and fluorescence microscope found large number of apoptotic cells in endothelium layer, smooth muscle layer and membrana adventitia layer of the blood vessels. Conclusion Explosion shock wave can result in severe large blood vessel injury, which should be paid much attention during treatment of ex-plosion shock injury.

Objective Severe bronchial stricture due to endobronchial tuberculosis is often accompanied by complex complication,such as obstructiv pneumonia,destroyed lung and bronchiectasis.Its treatment is very diffucult.The present report is to investigate and analyze the indication and efficacy of surgical treatment of bronchial stricture due to severe endobronchial tuberculosis.Methods Reviewed the clinico-pathological records documenting the surgical outcomes in 81 bronchial stricture due to severe endobronchial tuberculosis who underwent lobectomy or pneumonectomy enrolled in our hospital between January 1990 and December 2010.There were 29 male and 52 female.Mean age was(36 ± 12) years (ranged 16-66 years).The three most common reasons of surgery were bronchial stricture accompanied by pulmonary atelectasis,destroyed lung and bronchiectasis(76 cases,93.8％).79 cases had elective operation,whereas one patients required emergency surgery.Pueumonectomy in 51,lobectomy in 16,sleeve resection in 11,segmental resection in 2,and exploratory thoracotomy in 1.If frozen pathological examination showed that endobronchial tuberculosis remained in the bronchial stump,it was covered with muscle flaps,including intercostal muscle flap in 6 cases,latissimus dorsi muscle flap in 5 cases,serratus anterior muscle flap in 5 cases.The mean operative time was 3.2 h (range between 2 h and 5.5 h) and the blood loss averaged 546 ml (range between 100ml and 4 000 ml).The post operative hospital stay averaged(12 ±8)days.Results No intraoperative or early postoperative death occurred.Nine patients developed complications,including BPF in 2,pulmonary infection in 2,empyema in 1,hemorragic shock in 1,hemothorax in 1,incision infection in 1,chylothorax in 1.All 9 cases recovered well after treatment.Pathological examination showed that tuberculosis bronchial remained in the brinchial stump in 13 cases.Neither BPF nor empyema occurred in all the 13 cases.Multivariate analysis revealed that destroyed lung was significant risk factor of postoperative complication.There were 3 late deaths.Five year survival rate was 96.2％.Conclusion Surgical treatment is still the recommeded treatment modatity for bronchial stricture caused by endobronchial tuberculosis due to its excellent results.It should be performed in time when the drug and intraluninal treatment were no effect for avoiding of being progeressed into destroyed lung.

BACKGROUND:Vascular endothelial growth factor is a potent angiogenesis and permeability inducible factor. Vascular endothelial growth factor 165 and vascular endothelial growth factor 121 are mainly expressed in vivo, with a strong role of angiogenesis.
OBJECTIVE:To observe the feasibility of vascular endothelial growth factor 165 gene transfected bone marrow mesenchymal stem cel s to differentiate into vascular endothelial cel s.
METHODS:Bone marrow derived mesenchymal stem cel s were isolated and col ected from 50 g Sprague-Dawley rats,and identified by flow cytometry. The plasmid pGLV-EF1a carrying a vascular endothelial growth factor 165 gene was transfected to the mesenchymal stem cel s using lentiviral. Expression of green fluorescent protein was observed under a fluorescence microscope.
RESULTS AND CONCLUSION:After 12 hours of transfection, expression of green fluorescent protein was observed, increased at 48 hours, peaked at 72 hours and gradual y declined thereafter. Results prove that vascular endothelial growth factor 165 gene transfected bone marrow mesenchymal stem cel s have the expression of green fluorescent protein, indicating successful transfection. It is feasible to induce bone marrow mesenchymal stem cel s to differentiate into vascular endothelial cel s.

Objective To explore whether dithiothreitol(DTT)is helpful for PreScission Protease to cut off the GST from GST?CT3 protein. Meth?ods The pGEX?6P?3/CT3 recombinant plasmid was transfected into Escherichia coli BL21,and the GST?CT3 fusion protein was purified by B?PER method. PreScission Protease was applied with 10 mmol/L DTT to cut off the GST,then the SDS?PAGE was performed for identification of the CT3 protein. Results Without DTT,it was very difficult for PreScission Protease to cut off the GST from GST?CT3 protein. However,in the pres?ence of 10 mmol/L DTT,PreScission Protease could cut off the GST easily as identified by SDS?PAGE. Conclusion 10 mmol/L DTT can help Pre?Scission Protease to cut GST from GST?CT3 protein,so as to achieve high concentration of CT3.