ABSTRACT
Increasing the production and secretion of endogenous opioid peptide by immune cell can significantly induce myocardial protective effects against ischemia-reperfusion injury. Gene therapy is promising to increase endogenous enkephalin (ENK). However, classical viral and plasmid vectors for gene delivery are hampered by immunogenicity, gene recombination, oncogene activation, the production of antibacterial antibody and changes in physiological gene expression. Minimalistic immunologically defined gene expression (MIDGE) can overcome all the deficients of viral and plasmid vectors. The exon of rat's preproenkephalin (PPENK) gene was amplified by PCR and the fragments were cloned into pEGFP-N1 plasmids. The recombined plasmids were digested with enzymes to obtain a linear vector contained promoter, preproenkephalin gene, RNA stable sequences and oligodesoxy nucleotides (ODNs) added to both ends of the gene vector to protect gene vector from exonuclease degradation. A nuclear localization sequence (NLS) was attached to an ODN to ensure the effective transport to the nucleus and transgene expression. Flow cytometry, laser confocal microscopy and Western blotting demonstrated that PPENK-MIDGE-NLS can transfect leukocyte of rat in vivo, increase the expression of proenkephalin (PENK) in tissue, and the transfection efficiency depends on gene vector's dosage. These results indicate that PPENK-MIDGE-NLS could be an innovative method to protect and treatment of myocardial ischemia-reperfusion injury.
Subject(s)
Animals , Rats , Cloning, Molecular , Enkephalins , Genetics , Gene Expression , Genetic Therapy , Genetic Vectors , Leukocytes , Plasmids , Promoter Regions, Genetic , Protein Precursors , Genetics , Transfection , TransgenesABSTRACT
OBJECTIVE:To establish the quality standard of Belamcandin standard substance.METHODS:Different indices of standard Belamcandin were examined with quantitative and quantitative analyses according to the corresponding measures stipulated in the Appendix of pharmacopoeia of people's republic of China(part Ⅱ)published in 2005.RESULTS:The specific rotatory power of 3 batches of standard Belamcandin was —28?～—30?([?]25 D,c0.11,Pyridine);the E1% 1 cmwas 790～830;the melting point was 252～254 ℃ and the ash content was less than 0.5%.The content of Belamcandin in the products was above 99.5% as revealed by HPLC.CONCLUSION:All of the indices were shown to meet the standards for standard substances and the established standard can be used for the quality control of standard Belamcandin.
ABSTRACT
Objective To study the role of IL-2 in regulating allograft rejection mediated by alloantigen-specific CD8~+ T cell.Methods T cell proliferation in vivo at a single cell level was examined using the CFSE dilution assay. IL-2 expression by activated CD4~+ versus CD8~+ T cells was determined by intracellular cytokine staining. The ability of alloantigen-specific CD8~+T cells in mediating allograft rejection was studied using the islet transplantation model.Results CD8~+ T cells divided vigorously in vivo in the allogeneic hosts regardless the presence or absence of CD4~+ T cells. CD4~+ T cells, but not CD8~+ T cells, were the primary source of IL-2 when both subsets were present. However, CD8~+ T cells could express high levels of IL-2 in the complete absence of CD4~+ T cells. In 2C TCR transgenic (Tg) mice in which the 2C TCR transgene was selectively expressed on the CD8~+ T cells that specifically recognized alloantigen (Ld) of Balb/c origin, islet allografts from Balb/c mice was promptly rejected by the 2CTg recipients with mean survival time of only 8 days. In contrast, in 2CTg mice with a genetic deletion of the IL-2 gene (2CTg-IL-2KO mice), the alloantigen specific CD8~+ T cells failed to mediate the islet allograft rejection and all the Balb/c islets survived for more than 50 days.Conclusions CD8~+ T cells appear to be very plastic in producing and utilizing IL-2. In the presence or absence of CD4~+ T cells, CD8~+ T cells can use CD4~+ derived or self derived IL-2 for proliferation and effector function respectively. In an alloantigen specific TCR transgenic model, the effector function of CD8~+ T cells is strictly IL-2 dependent. Thus, in situations where graft rejection is mediated solely by the CD8~+T cells, blocking IL-2/IL-2R pathway may be critically important in preventing transplant rejection.
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Objective To investigate the saponins from the Chinese materia medica Huluba (the seeds of Trigonella foenum-graecum L.). Methods To isolate and purify the single saponin by the column chromatography and dry column chromatography of silica gel H. The structural elucidation was carried on by spectroscopic evidence of (()~(13)C-NMR), FAB-MS, DEPT and the results of the fraction-hydrolysis of acquiring the secondary glucosides. Results The newly isolated saponin D was consisted by six molecules of sugar with diosgenin. The chemical structure of saponin D is: diosgenin-3-O-?-L-rhamnopyranosyl (1→3)-?-D-glucopyranosyl (1→4)-?-L-rhamnopyranosyl [(1→3)-?-L-rhamnopyranosyl] (1→4)-?-D-glucopyranosyl (1→4)-?-D-glucopyranoside. Conclusion Saponin D is a new one consisted of six molecules of su-(gar) with diosgenin.
ABSTRACT
Object To do detail investigation of the saponins from the Chinese materia medica Huluba (the seeds of Trigonella foenum graecum L ) Methods The pure saponins from the total saponins were isolated by employing the column chromatography and dry column chromatography of silica gel H Their chemical structures were elucidated by 13 C NMR , FAB MS, DEPT spectroscopic evidence and the results of the fraction hydrolysis of acquiring their secondary glucosides were obtained Results Two new saponins B and C were isolated and both were the glucosides consisted by four molecules of sugar with diosgenin The chemical structure of B is: diosgenin 3 O ? L rhamnopyranosyl (1→3) ? L rhamnopyranosyl (1→4) ? D glucopyranosyl (1→4) ? D glucopyranoside And saponin C is: diosgenin 3 O ? D glucopyranosyl (1→4) ? L rhamnopyranosyl (1→4) ? D glucopyranosyl (1→4) ? D glucopyranoside Conclusion Saponins B and C are two new ones with four molecules of sugar respectively
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Object To investigate the saponin from the seeds of Trigonella foenum graecum Linn (STFG) Methods The total saponin from STFG was extracted and purified by using the absorptive resin, the single saponin was isolated by using the column chromatography as well as dry column chromatography of silica gel H The chemical structure was elucidated by 13 CNMR , FAB MS, DEPT spectroscopic evidence and the results of fraction hydrolysis of acquiring their secondary glucosides Results A new saponin A from the total saponin has been obtained, the fraction hydrolysis carried out and the secondary glucoside Ⅰ and Ⅱ identified by determining the structure of saponin A The chemical structure of saponin A is: diosgenin 3 O ? L rhamnopyranosyl(1→4) ? D glucopyranosyl(1→4) ? D glucopyranoside The secondary glucoside Ⅰ is: diosgenin 3 O ? D glucopyranoside; Ⅱ is: diosgenin 3 O ? D glucopyranosyl(1→4) ? D glucopyranoside Conclusion Glucoside A is a new saponin with three molecules of sugar