Objective To obtain the endophytic fungi which produce Huperzine A from four species in Hupziaceae and improve the culture conditions of screening endophytic fungi. Methods Plant materials were cultivated in culture medium after sterilization and their endophytic fungi were isolated. Hupzine A from metabolic was determined by HPLC, and the strains were identified by microscopic features. Results Thirty-two endophytic fungi were isolated from Huperzia serrata (Thunb. ex Murray) Trev, H. serrata (Thunb. ex Murray) Trev var. longipetiolata (Spring) H. M. Chang, H. appressa (Desv.) Love et D. Love, Phlegmariurus cryptomerianus (Maxim.) Ching ex L. B. Zhang et H. S. Kung. Two of these strains were isolated from P. cryptomerianus, HA15 (Blastomyces sp.) and HA23 (Botrytis sp.), which produced Huperzine A. Conclusions Endophytic fungi producing Hupzine A has been successfully isolated from P. cryptomerianus.

ObjectiveTo study the effect of anti-oncogene p15 and p16 on the proliferation of human cholangiocarcinoma cell line.MethodsThe cDNA of anti-oncogene p15 and p16 was constructed into pcDNA3-neo plasmid carrier. The human cholangiocarcinoma cell line QBC939 were transfected with the recombinants pcDNA3p15 and pcDNA3p16 using lipofectin, respectively. The expression products were analyzed by Western blot. Cell viability and death were measured with MTT assay. Cell cycle was determined by flow cytophotometry and the formation of cell clone was detected. Results The growth of QBC939 cells was inhibited. The flow cytophotometry verified p15 and p16 induced QBC939 cell G1 blockade. Conclusion Anti-oncogene p15 and p16 together lead to the inhibition of cell cycle.

AIM: To Establish the standard of Qirong Medicinal Wine (Radix Angelicae Sinensis, Fructus Psoraleae, Radix Polygoni Multiflori Preparata, Fructus Lycii and Flos Carthami, etc.). METHODS: Radix Angelicae Sinensis, Fructus Psoraleae, Radix Polygoni Multiflori Preparata, Fructus Lycii and Flos Carthami in the preparation were identified by TLC. Psoralen and isopsoralen were determined by HPLC. RESULTS: The average recoveries of psoralen and isopsoralen were 98.7% and 100.8%, respectively., RSD were 2.18%( n =5) and 1.67%( n =5),respectively. CONCLUSION: The methods are simple, accurate and reproducible.

Objective To study the effect of candesartan on the ERK1/2 protein expression in myocardium of epilepsy rats induced by kainic acid(KA) and its mechanism.Methods 105 male Wistar rats were divided into: control group(A);epilepsy groups(B1-5);candesartan groups(C1-5);1-5 meaned 0,0.5,2,4,6 h after epilepsy,respectively.Epilepsy rat models were made by injecting KA into amygdale under stereotactic instrument.The ERK1/2 protein expression in various groups were tested with immunohistological method.Results Compared with control group,the protein expressions of ERK1/2 increased significantly in the 0.5 h groups of B and C(P0.05).The differences in the level of ERK1/2 protein expression between B and C groups were significant(P

BACKGROUND: Matrix attachment region (MAR), a DNA sequence, is still bound to the nuclear matrices after chromatindigested with restriction endonuclease, not only affects expression of endogenous gene, but also overcames transgenic silence andimproves transcription and expression of exogenous gene. OBJECTIVE: To investigate the influence of ?-interferon MAR of CHO cells on the transgenic expression of chloramphenicolacetyltransferase (CAT). DESIGN, TIME AND SETTING: The opening experiment was performed at the Department of Biochemistry and MolecularBiology, Molecular Institute, Xinxiang Medical College from October 2006 to April 2007. MATERIALS: CHO cell lines were obtained from China Center for Type Culture Collection. The pCATG vector of CAT and G418screening markers were constructed by this laboratory. METHODS: Human ?-interferon MAR by PCR was digested with SacI/KpnI and BamHI/SalI, and was inserted into pCATGvector, which was propagated in Escherichia coli JM109, then extracted and purified followed by enzyme digestion analysis. Vectorof CAT expression cassette and human ?-interferon MAR by the two sides was successfully constructed, and christened aspCAT-MAR. Two methods were compared between CHO cells of pCATG transformation and CHO cells of pCATG-MARtransformation. After G418 selecting, genome DNA of cell lines of G418 was extracted, then primers for PCR to amplify the CATtarget gene fragment was designed. MAIN OUTCOME MEASURES: The activity of CAT was analyzed by ELISA method. It was also tested to see if thepCATG-MAR was stably integrated into genomic DNA in the transfected cells. RESULTS: CHO cells of pCATG transformation was screened to have 16 strains of positive cell, and CHO cells of pCATG-MARtransformation was screened to have 17 strains of positive cell. Human ?-interferon MAR could increase the CAT gene expressionby 2.8 fold. The coefficient of variation of CHO cells of pCATG transformation was 2.065 0, and coefficient of variation of CHOcells of pCATG-MAR transformation was 0.813 1. Genome DNA of stable transformation cell lines was amplified by a fragment of437 bp. The results confirmed the pCAT-MAR vector was stably integrated into genomic DNA. CONCLUSION: Human ?-interferon MAR can increase transgenic expression in CHO cells and decrease the transgenicexpression variation in different transfected cells.

Objective To evaluate the clinical efficacy and safety of combination treatment with clofazimine ( Cfz ) and other antituberculosis drugs for patients with multidrug-resistant tuberculosis(MDR-TB).Methods 32 cases of MDR-TB patients were treated with combination regimens that included clofaziminefrom October 2011 to September 2016 in our hospitol,according to the history of drug use and drug susceptibility test results using individualized chemotherapy,the starting dose of clofazimine was 0.1 g/day,oral,some patients with adverse reactions and tolerance adjusted to 0.05 g/day,treatment for the last 12 months for three consecutive sputum mycobacterium tuberculosis culture and sputum smear acid-fast bacilli were cured,observe the clinical efficacy and safety.Results After treatment with the combined regimen,56.2%(18/32) of patients were cured,43.8%(14/32) of patients failed treatment, there was no statistically significant difference in the number of drug-resistant patients before and after CFZ treatment,there was no statistically significant difference between the time of failure and the time of CFZ treatment,after taking CFZ combined with anti-tuberculosis program,the number of drug users was statistically significant of patients cured and failed (P<0.05).The average time of sputum culture inversion was 16w.90.6% (29/32) of patients with adverse reactions,mainly including skin color change,ichthyosis and gastrointestinal tract and other adverse reactions, through dose adjustment and symptomatic treatment to continue treatment.The average duration of treatment with clofazimine was about 13 months.ConclusionClofazimine was welltolerated,combination treatment with Clofazimine and others for patients with MDR-TBhave better efficacy .

BACKGROUND:Matrix attachment region(MAR)are DNA elements that bound to the nuclear matrices after chromatin digested with restriction endonuclease.Plenty of studies have shown that MAR considered as initiaI point of DNA replication or transcription of regulatory gene.Thereby,construction of MAR expression vector can elevate the overall level of transgene expression,enhance stability of exogenous gene.as welI as increase frequency of stable transfectant cells.OBJECTIVE:To construction pLXSN-CAT recombinant retrovirus vector that containing chloramphenicol acetyltransferase(CAT)via cloning MAR sequence of human.and to explore the influence of MAR on the gene expression.METHODS:An open experiment was performed at the Department of Biochemistry and Molecular Biology.Xinxiang Medical College from September 2007 to December 2007 The PLXSN-CAT vector of CAT was constructed by the laboratory.TaqDNA polymerase,T_4 DNA ligase,DNA Marker,restriction enzyme BamH I,agarose gel DNA purification kit,as well as plasmid purification kit were purchased fromTakara Biotechn0Iogy(Dalian)Co.,Ltd.The sequence of csp-B MAR was amplified by polymerase chain reaction(PCR)method applied to human DNA.The fragment was inserted into retrovirus vector PLXSN-CAT plasmid.The recombinant plasmid was verified by double digestion and DNA sequencing.RESULTS AND CONCLUSION:The length of specific fragment applied by PCR was 931 bp,and the recombinant plasmid PLXSN-CAT-MAR presented two bands:5.9 kb and 931 bp using respective restriction enzymes BamH I The sequence of MAR was confirmed by blasting to Genbank(serial numobr:M6271 6).It suggested that MAR had been cloned into PLXSN-CATR vector correctly.The recombinant retrovirus vector PLXSN-MAR was successfully constructed.

Objective To observe the effect of repetitive transcranial magnetic stimulation (rTMS) and functional magnetic stimulation over spinal cord (SC-MS) on motor function recovery after spinal cord hemisection in rats. Methods T10 spinal cord hemisection model was made. The stimulation (5×10 s bursts of 5 Hz at 35% maximal stimulator output, each burst separated by a 2 m interval) was delivered daily, 5 d per week for 2 weeks. The treatment began at 4 d after surgery for rTMS group and SC-MS group. Motor function recovery was assessed with Basso, Beattie & Bresnahan locomotor rating scale (BBB) and the Horizontal Ladder test. The tibialis anterior was surgically removed at 38 d after spinal cord injury for calcitonin gene-related peptide (CGRP) iummunohistochemical staining. Results The scores of BBB and Horizontal Ladder test were significantly more at 17 d after spinal cord hemisection in rTMS group than before treatment and in spinal cord injury group (P<0.05). In SC-MS group, the scores of BBB and Horizontal Ladder Test were significantly more 10 d after SCI than before treatment and in SCI group (P<0.05). The score of Horizontal Ladder test of SC-MS group was more 10 d after SCI than that in the rTMS gourp (P<0.05). The expression of CGRP on motor endplates of the tibialis anterior in rTMS group and SC-MS group were more than those of SCI group (P<0.01). Conclusion rTMS and SC-MS in acute stage can improve the motor function recovery and muscle plasticity after spinal cord hemisection in rats. The magnetic stimulation can facilitate the recovery of motor function after spinal cord hemisection in rats.

Objective To study the extracellular signal-regulated kinase 1/2(ERK1/2) expression of renal cells in epilepsy rats induced by kainic acid,and elucidate the mechanism of renal injury caused by epilepsy.Methods 70 male Wistar rats were randomly divided into two groups:epilepsy group and control group,35 rats in each group.Rat epilepsy model was prepared by injection of kainic acid into amygdala under three-dimensional positioning devices.At different time points(0,2,6,12 and 24 h) after epilepsy spasm,the renal tissue specimens were immediately prepared for immunohistochemistry.The ERK1/2 expression of rat renal cells was calculated and compared with control group.Results The ERK1/2 expression(gray value) of renal tubular epithelial cells in epilepsy rats began to increase gradually after epilepsy spasm.It reached peak at 6 h after epilepsy spasm.The ERK1/2 expression of renal tubular epithelial cells in epilepsy rats was significantly higher than at 0 h(P0.05).There were no major pathological changes in kidney of rats with epilepsy stained with HE.Conclusion ERK1/2 activation may have important role in the renal injury caused by epilepsy.Inhibition of the activation of ERK1/2 may lead to the renal protection.

Objective To explore the feasibility of the reconstruction of three dimensional dynamic finite element model of the knee joint based on two-dimensional CT and MRI image data. To analyze the impact of mobile-bearing versus fixed-bearing platform prostheses of the knee joint on patellofemoral stress by the finite element method. Methods A three-dimensional digital model of the knee joint including bone,cartilage,meniscus, ligaments and tendons was reconstructed through the Mimics software. The best clinical bone cutting angle and implant placement position measurement were simulated according to the standard of total knee replacement by computers in a three-dimensionalknee model. A three-dimensional dynamic finite element model of mobile-bearing and fixed-bearing total knee arthroplasties was reconstructed finally. The data was analyzed by the SPSS 19.0 software. The test standard level α was 0.05. Results There was no significant difference in the peak value of patellofemoral stresses between fixed-bearing and mobile-bearing platform posterior cruciate-substituting prostheses at 0° ,30° ,60° ,90° ,or 120° of knee flexion(P>0.05). Conclusion There is no significant difference in the peak value of patellofemoral stresses between fixed-bearing and mobile-bearing platform prostheses.