Objective To construct human canstatin gene eukaryotic expression vector and investigate the therapeutic effect of intramuscular canstatin gene delivered by electroporation on tumor growth.Methods Canstatin cDNA was amplified from total RNA extracted from fresh fetal liver by reversing transcription polymerase chain reaction (RT-PCR).The canstatin cDNA fragment was in serted into pEGFP-N1 eukaryotic expression vector.The recombination plasmid was delivered to the quadriceps of the mice with Lewis lung carcinomas by electroporation intramuscular.Fluorescence intension measured by fluorescence microscope,reverse-PCR assay,and immunohistochemistry assay were performed to detect the expression of canstatin gene in the muscle and in circulation.The tumor weight and volume were used to detect the biological effects of canstatin gene delivery.Results Recombinant eukaryotic expression vector of recombinant human canstatin was successfully constructed.The canstatin mRNA was significantly increased in the skeletal muscle and intramuscular delivery of canatatin gene by electroporation acquired the expression of enhanced green fluorescent protein (EGFP)/canstatin protein in the circulation and significantly inhibited tumor growth.The percent of the inhibition of tumor weight was 57.7 ％.Conclusions Electroporation mediated gene transfer efficiency in skeletal muscle was compared to simple plasmid injection and lasted for a long time.It was an efficient and safe,convenient and economic,gene transfer methods and might have certain clinical application value.Electroporation mediated canstatin gene transfer in skeletal muscle had obvious inhibitory effect on Lewis lung cancer in mice subcutaneous xenograft tumor growth.

AIM: To investigate the effects of TNF receptor-associated death domains (TRADD) of latent membrane protein 1 (LMP1) on the proliferation of nasopharyngeal cancer SP18 cells.METHODS: The SP18-LMP1 cells and SP18-LMP1TRADD cells, which expressed LMP1 and LMP1 TRADD proteins, respectively, were established.The proliferation of SP18 cells affected by LMP1TRADD was detected by cell counting to analyze the cell growth curve, and by colony formation assay, soft agar formation assay, and flow cytometry.Moreover, the expression profile of differential genes between SP18-LMP1 cells and SP18-LMP1TRADD cells was analyzed by gene chips.RESULTS: The cell growth curve, and the results of colony formation and soft agar formation displayed that the growth velocity and colony forming ability of SP18-LMP1 cells were stronger than those of SP18-LMP1TRADD cells (P<0.01).The results of flow cytometry analysis showed that the proliferation index of SP18-LMP1 cells was higher than that of SP18-LMP1TRADD cells (P<0.01).Sixty-three differentially expressed genes associated with cell proliferation were screened out, in which 33 genes were up-regulated and 30 were down-regulated in the SP18-LMP1TRADD cells.CONCLUSION: TRADD active region is an important functional site of LMP1 to promote the proliferation of SP18 cells.LMP1 may improve the cell proliferation index and induce the proliferation of SP18 cells through TRADD.

Objective To study a gastric carcinoma subcell line with higher invasive potential and its biologic characteristics screened by a transwell chamber. Methods Transwell chamber was used for the selection of tumor subline. The biological characteristics of the cell lines were studied with optics microscope, Westem blotting, Transwell, immunohistochemical staining and growth curve. Results A gastric carcinoma cell subcell line (named MKN-28S10) was established from its parent cell line MKN-28 with higher invasive potential. MKN-28S10 showed essentially the same morphous as MKN-28. The expression of E-cadherin and TIMP-1 decreased significantly in the screened subcell line(P<0.05). The expression of NM23-H1 in MKN-28S10 was significantly lower than that in MKN-28 (P<0.05). Compared with MKN-28 (61.75±2.06 per vision), the migrative ability of passing through the membrane Millipore(100.25±0.50 per vision) was obviously increased in MKN-28S10(P<0.05). Compared with MKN-28, the growth rate of MKN-28S10 was increased obviously. Conclusion MKN-28S10 cell has stronger invasive ability and more powerful proliferation than that of its mother line MKN-28.