AIM: To get large amounts of pure antigens to raise specific antibodies and to perform quantifications.METHODS: CYP2B6 (cytochrome P) cDNA fragments was ligated into BamHI restricted PGEX-3b to generate recombinants PGEX/2B6. We identified recombinants PGEX/2B6 by EcoRI digestion. The expression of fusion proteins were induced by adding isopropyl-thiogalactoside(IPTG). Several clones showed high-level expression of fusion proteins. Insoluble proteins was isolated from the bacteria and the fusion proteins was recovered and purified from a preparative (2mm) SDS-PAGE. The polyarcrylamide gel containing the fusion proteins glutathione S-transferase(GST-2B6) were used to immunize BALB/C mice from which polyclonal ascites fluid was prepared. The purified fusion proteins GST-1A1(GST fusion protein of CYP1A1 cDNA246～386aa expressed in this library ,purified by preparative SDS-PAGE), GST-2B6 were used to test the specificity of 2B6pAb. RESULTS:Fusion proteins constructed between GST and CYP2B6 was expressed in Escherichia coli DH5?. Mouse antibodies are raised against the fusion proteins GST-2B6. 2B6pAb was fond to be specific antibody.CONCLUSION:Recombinant PGEX/2B6 were constructed and purified fusion proteins GST-2B6, and specific 2B6pAb were obtained.

Lymphocyte-activation gene 3 (LAG-3), also known as CD223, is a 498-amino-acid type I transmembrane protein encoded by LAG-3 gene, which consists of extracellular, transmembrane and intracellular regions.LAG-3 negatively regulates T lymphocyte by binding extracellular domain to ligand, thus avoiding autoimmunitycaused by T cell over-activation. Like programmed cell death 1 (PD-1) and cytotoxic T lymphocyte antigen 4 (CTLA-4), LAG-3 is an important immune checkpoint in vivo and plays a balanced regulatory role in human immune system.Tumor cells escape the surveillance of the immune system by over-expressing LAG-3 ligand. With the development in research of immune checkpoints, LAG-3 has become a new generation of immunotherapy targets after PD-1 and CTLA-4. This article reviews the structure and function of LAG-3 and the application of its inhibitors in tumor immunotherapy, in order to provide reference for the further study of LAG-3.

Objective To observe the repairing effect of the acellular nerve allografts on the sciatic nerve gap of rat. Methods The acellular nerve allografts,treated by hypotonic-chemical detergent,were put on the 10 mm gap of the sciatic nerve in the rat.The action potential of the regenerated nerves was determined by the electrophysiologic method 13 weeks after operation.The morphology of the regenerated nerves was observed under light microscope and electron microscope,and the results were analyzed statistically. Results No inflammation and rejected reaction were found in the period of 13 weeks after operation in the operated and control groups.There was no significant difference in number of the regenerated nerve fibers,diameter of the axons,and the thickness of the regenerated myelinated nerve between the experimental group and control group.Conclusion The present results indicated that the acellular nerve allografts had good biocompatibility for the host rat in vivo and might as a bridge promote the regeneration of the injured sciatic nerve.;

Objective The experiment was designed to investigate the effect of acellular nerve allografts on the functional recovery and reconstruction of the nerve-muscle structure of the sciatic nerve defect in rats. Methods Acellular nerve allograft was transferred into the defected rat sciatic nerve with 10mm long.The wet weight of tibialis anterior was weighed at 12 and 24 weeks postoperatively compared with control group.The conducted velocity of regenerated nerve and the effect of regenerated nerve on tibialis anterior were investigated by electrophysiologic test,and silver staining combined with AChE histochemical methods were used in the experiment separately. Results The wet weight of tibialis anterior and the conducted velocity of regenerated nerve in experimental group were similar to those in control group in 12 and 24 weeks after transplantation.The positive acetylcholinesterase(AChE)histochemical reaction was observed in the tibialis anterior at 12 weeks with deeper staining and located in the middle of tibialis anterior tidily at 24 weeks after operation.The regenerated nerve bundles and nerve terminals were found to grow into the motor end-plate of the tibialis anterior in silver staining combined with AChE staining in experiment group.Electromyogram showed that the regenerated nerve has innervated tibialis anterior already.Conclusion The results indicated that extracted nerve allografts as a bridge can promote the motor functional recovery and reconstruction of the nerve-muscle structure of the defected rat sciatic nerve.

BACKGROUND: There is a positive correlation between the serum cholesterol and osteoarthrosis incidence. However, it is still unclear whether the high-sugar and high-fat diet participates in the process of articular cartilage degeneration leading to osteoarthrosis, and the participation mechanism is unknown. OBJECTIVE: To investigate the changes in knee cartilage morphology of New Zealand white rabbits fed with high-sugar and high-fat diet and to explore the effect of high-sugar and high-fat diet on articular cartilage degeneration. DESIGN, TIME AND SETTING: The randomized controlled experiment was performed in Animal Laboratory, Xiangya Medical College from January to September 2008. MATERIALS: Forty healthy adult male New Zealand white rabbits, weighing about 2 kg, were randomly divided into 4 groups with 10 in each group. METHODS: Male New Zealand white rabbits were divided into four groups: ①normal diet group(NG), ②high-sugar diet (mixture of 37% cane sugar and 63% normal fodder) group(SG), ③high-fat diet (mixture of 20% lard and 80% normal fodder) group(FG), ④high-sugar and high-fat diet (mixture of 37% cane sugar, 10% lard and 53% normal fodder) group(SFG), MAIN OUTCOME MEASURES: Fasting blood samples were extracted every 4 weeks. The levels of blood sugar, triglyceride, cholesterol and insulin were detected, and body mass was recorded. The femoral condyle cartilage were dyed with toluidine blue, observed with transmit electron microscope after general observation of rabbit knee joints at the 28th weeks. RESULTS: ①Blood sugar, triglyceride, total cholesterol in FG and SFG were obviously higher than that in NG, the level of blood insulin in FG and SFG increased first, and then decreased significantly (P 0.05). ② By light microscope and transmit electron microscope observation, the FG and SFG demonstrated that tide line disappeared, cartilage cells were ranged in disorder, collagen fibers fractured, cartilage cells and a change in shape, such as shrinkage. These changes of morphology were not observed in SG. CONCLUSION: Long-term high-fat diet or high-sugar and high-fat diet may induce or aggravate the disorder of articular cartilage, suggested that it may take part in the development of osteoarthritis.

Objective To investigate the predictive value of neutrophil and lymphocyte ratios (NLR)for the prognosis in patients with acute cerebral infarction. Methods From January 2014 to December 2015,307 consecutive patients with acute cerebral infarction admitted to the Department of Neurology,the Fifth Affiliated Hospital of Zhengzhou University were enrolled retrospectively,including 80 females and 227 males. They were divided into ether a good prognosis group (n = 195)or a poor prognosis group (n = 112)according to the scoring criteria of the modified Rankin scale (mRS). The age,gender, past medical history,National Institutes of Health stroke scale (NIHSS)score were documented on admission. The NLR values were calculated according to the neutrophil and lymphocyte counts on admission. Logistic regression analysis was used to analyze the influencing factors of poor prognosis of acute cerebral infarction. The receiver operating characteristic curve (ROC)was used to evaluate the predictive effect of the NLR level on patients with acute cerebral infarction on admission. Results (1)Compared with the good prognosis group,the age,incidence of recurrent cerebral infarction,NIHSS score on admission, NLR levels on admission in the poor prognosis group were higher. There were significant differences between groups (69 ± 12 years vs. 62 ± 14 years,25. 0% [28 / 112]vs. 14. 4% [28 / 195],5. 00 [3. 00, 9. 00]vs. 3. 00 [1. 75,5. 00],and 3. 66 [2. 62,7. 91]vs. 2. 47 [1. 94,3. 40];all P < 0. 05). There were no significant differences in other baseline data and clinical characteristics between the groups (all P >0. 05). (2)Multivariate logistic regression analysis showed that the increase of the age,NLR level on admission,and increased NIHSS score on admission,were independent risk factor for poor prognosis (OR 1. 030,1. 148,and 1. 427,respectively,95% CI were 1. 007 -1. 053,1. 059 -1. 246,and 1. 247 -1. 634, respectively;all P < 0. 05). (3)The diagnostic cut-off value of the NLR level on admission for the poor prognosis in patients with acute cerebral infarction was 2. 84. Its sensitivity was 69. 6% and specificity was 64. 6% . Conclusion The increase of the NLR level on admission had certain reference function on the poor prognosis in patients with acute ischemic stroke.

based on the theory of tree structure and followed the characteristics of medical talents growth,Wuxi No.2 People's Hospital carried out hospitalTalent Tree Project innovatively,which is to trained talents corresponding tobase-trunk-crown of a tree.The project has won the 2013 Human Resource Development Excellence Award of Asian hospital management awards.We fostered talents hierarchically according to the talents training echelon structure and this provided a way of guarantee and innovation for the sustainable development of hospital.

Objective To explore the reversion effect of Gemcitabine-resistance A549 cell (A549/Gem) by silencing CXCR4.Methods A549 cell was induced by continuous stepwise exposure to Gemcitabine in order to obtain Gemcitabine-resistance A549 cell ( A549/Gem) in vitro.The CXCR4 expressions level of A549 and A549/Gem were detected by Quantitative RT-PCR ( RT-qPCR) and Western blot analyses.The CXCR4 shRNA vector was transfected into the A549/Gem cell by targeting silence CXCR4.Furthermore, MTT assay was used to explore the IC50 and RI in A549, A549/Gem and A549/Gem-CXCR4 cells.Moreover, Western blot analysis was performed to detect the expressions of phospho-JNK, phospho-p38 and phospho-ERK 1/2 in A549, A549/Gem and A549/Gem-CXCR4 cells.Results Gemcitabine-resistance A549 cell ( A549/Gem) was successful constructed by using continuous stepwise exposure to Gemcitabine in vitro.The expression level of CXCR4 was up-regulated in A549/Gem cell than in A549 cell.The CXCR4 shRNA vector could significantly decrease CXCR4 expression in A549/Gem cell.The IC50 values of Gemcitabine in A549, A549/Gem and A549/Gem-CXCR4 cell were (0.08 ±0.01)μmol/L, (14.01 ±0.21)μmol/L and (1.84 ±0.61)μmol/L, respectively.The RI value of Gemcitabine was (127.12 ±12.28) in A549/Gem cells, while the value of RI was (27.3 ±0.98) in A549/Gem-CXCR4 cells.The expression level of phospho-JNK, phospho-p38 and phospho-ERK 1/2 were also markedly inhibited in A549/Gem-CXCR4 cell than in A549/Gem cell.Conclusion CXCR4 is up-regulated in A549/Gem cell.Targeting silence CXCR4 can successfully reverse drug-resistance of Gemcitabine in A549/Gem cells, which hints CXCR4 is associated with lung cancer radiation therapy as an effective molecular target.

Objective To detect the effect of oridonin(ORI)on the gene expression of human esophageal carcinoma cell SHEEC and to screen the tumor cell apoptosis target genes.Methods The gene expression of human esophageal carcinoma cell SHEEC without and with ORI induction for 1 hours and 8 hours were detected with microarray technique,respectively.The differentially expressed genes were identified and verified with fluorecent quantitative PCR.Results A total of 1011 genes showed up or down regulation more than twice after ORI induction(including 280 genes after 1 hour and 731 genes after 8 hours induction respectively).In these genes,17 genes with the top extent of up or down regulation were identified,which were involved in the cell signal transduction,transcription regulation,and cell apoptosis.These 17 differentially expressed genes were verified with real-time PCR,and 12 genes were statistically significant.Conclusion In the 12 differentially expressed genes with statistically significance,there may have tumor cell apoptosis target genes induced by ORI through mitochondrion route.