Construction of experimental course group in molecular medicine based on improving medical students' comprehensive ability / 中华医学教育探索杂志

In order to adapt to the requirements of modem medical knowledge and skills for higher medical workers,and to cultivate medical students' molecular medicine quality and comprehensive ability,Weifang Medical University broke the boundaries of disciplines and constructed experimental course group in molecular medicine based on the ideas of curriculum group construction.Molecular medical knowledge was integrated into the teaching process of experimental course group,experimental teaching content system was reasonably integrated and optimized,high quality teaching team was set up,multi-level experimental teaching platform was built and student-centered teaching mode was implemented to explore the experimental teaching approach which helped medical students to form systematic molecular medicine knowledge structure and ability structure.

Signal transduction pathway ERK1/2 involved in cytotoxicity of NK cell lines / 中国免疫学杂志

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. [

Hepatocarcinoma specific IL-1? anti-sense RNA inhibits implanted hepatocarcinoma in mice / 中国肿瘤生物治疗杂志

Objective :To construct hepatocarcinoma specific IL-1? anti-sense RNA expression vector and to explore its effect on the growth of implanted hepatocarcinoma H22 cells in mice and the possible mechanism. Methods:Murine IL-1? anti-sense RNA expression vectors pafpIRES2-antiIL-1?1 and pafpIRES2-antiIL-1?2 under the regulation of minimal alpha-feto protein (AFP) promoter and CMV enhancer were constructed,and further verified by PCR,restriction endonuclease analysis and DNA sequencing. H22 cells transfected with pafpIRES2-antiIL-1? 1 or pafpIRES2-antiIL-1? 2 were divided into 3 groups:H22/mock,H22/antiIL-1?1 and H22/antiIL-1?2 group. Expression of IL-1? was detected by RT-PCR. Transfected H22 cells were subcutaneously injected into mice to establish tumor implanted mouse model. Tumor volume was measured; the cytotocixity of spleen NK against H22 cells was detected by MTT. Results:Hepatocarcinoma specific IL-1? anti-sense RNA expression vectors pafpIRES2-antiIL-1?1 and pafpIRES2-antiIL-1?2 were successfully constructed and were verified by PCR,restriction endonuclease analysis and DNA sequencing. IL-1? expression in H22 cells was down-regulated after transfected with IL-1? anti-sense RNA expression vectors,especially with the pafpIRES2-antiIL-1?2 vector. Hepatocarcinoma cells implanted mouse model was successfully established. Tumor volume and growth of tumor in H22/antiIL-1?2 mice was obviously smaller than that in H22/mock mice,and the cytotocixity of spleen NK against H22 cells in H22/antiIL-1?1 and H22/antiIL-1?2 mice was also greatly enhanced. Conclusion:Hepatocarcinoma specific IL-1? anti-sense RNA expression vector pafpIRES2-antiIL-1? was successfully constructed. It effectively inhibits the growth of implanted hepatocarcinoma in mice probably through specifically blocking expression of IL-1? and increasing cytotocixity of spleen NK.

Construction and identification of a screening vector using enhanced green fluorescent protein(EGFP) as an indicator / 中国海洋药物

Objective To construct and identify the screening vector pUC18-EGFP,using EGFP as an indicator. Methods The EGFP gene was prepared by PCR and cloned into pUC18 resulting the vector pUC18-EGFP. Then DNA fragment was inserted into the MCS of pUC18-EGFP to test its practicability based on green fluorescence. Results The pUC18-EGFP was confirmed correctly by restriction enzyme analyses. The pUC18-EGFP was used to select recombinants. The green strains on the plate were confirmed by restriction enzyme and DNA analyses,which were E.coli harboring recombinants. Conclusion The screening vector PUCl8-EGFP was constructed successfully. Thus,we can select the positive clones on plates based on the green fluorescence of EGFP.