Role of Dual-layer Detector Energy Spectral CT in Resting Myocardial Perfusion Imaging for Patients with Normal Coronary Artery / 中国医学科学院学报

Objective To investigate the role of dual-layer detector energy spectral CT in resting myocardial perfusion imaging for patients with normal coronary artery. Methods One hundred and fifty-six patients with suspected coronary heart disease underwent dual-layer detector energy spectral CT coronary angiography,and resting myocardial perfusion imaging was performed for 28 patients with normal coronary artery.According to American Heart Association's 17-segmentmodel,the iodine density and effective atomic number(Z

Prokaryotic expression and one-step purification of soluble diphtheria toxin variant CRM197 / 基础医学与临床

Objective To construct a novel prokaryotic expression system, in which cross-reacting material 197 (CRM197) can be expressed in a soluble form in Escherichia coli(E. coli) cytoplasm and purified simply by one-step Ni-NTA affinity purification. Methods The CRM197 coding sequence was cloned into the prokaryotic expres-sion vector pET-32a(+) as an fusion protein with Trx tag,the HRV3C(human rhinovirus 3C) protease recognition sequence and 6 histidine sequence were added to the N-terminal of CRM197.HRV3C protease gene was cloned into another prokaryotic expression plasmid pGArasd. Both plasmids were co-transformed into E. coli Origami B (DE3) and induced mildly at 15℃. CRM197 recombinant protein was purified by Ni-NTA affinity matrix. Results The free soluble His-tagged CRM197 protein was released by cleavage of the accompanying expressed HRV3C protease after the CRM197 fusion protein was expressed. After one-step affinity purification recombinant CRM197 protein with a purity of almost 95% was obtained. Outcoming of the final preparation incubated with DNA indicated the pu-rified CRM197 recombinant protein has deoxyribonuclease activity. Conclusions By constructing a novel double-plasmid auto-cleavage prokaryotic expression system in this study, the production process of obtaining soluble CRM197 recombinant protein in E. coli has been simplified,with expression and purification efficiency improved and the production cost reduced.

Secreted CTLA-4 fusion Plasmodium falciparum DNA vaccine combined with GM-CSF enhances immune response of mice / 基础医学与临床

Objective To study effects of secreted cytotoxic T-lymphocyte-associated protein-4(CTLA-4)fusion Plasmodium falciparum DNA vaccine combined with granulocyte-macrophage colony stimulating factor(GM-CSF) on humoral and cellular immune responses in mice.Methods The malaria antigen coding sequence fused with CTLA-4 extracellular region of mouse was constructed as eukaryotic secretory expression vector VR 1012-sES312-CTLA,recombinant protein in culture of transfected HEK 293 cells was detected by Western blot.Balb/c mice were co-administrated with VR1012-sES312-CTLA and GM-CSF expression vector.After immunization specific antibody IgG titers and cytokines IFN-γand IL-4 expression levels were evaluated by ELISA and ELISPOT respectively. Results The introduction of CTLA-4 into malaria DNA vaccine system and application of GM-CSF adjuvant signifi-cantly enhanced the specific immune response to the vaccine.Antibody titers in VR1012-sES312-CTLA and GM-CSF co-immunized mice showed a 190-fold increase compared with the simple designed VR1012-ES312 immunization(P<0.001).Conclusions Humoral and cellular immunity induced by malaria DNA vaccine are significantly en -hanced by both dendritic cell-targeting modification and the introduction of GM-CSF molecular adjuvant into the im-mune system.This result provides a new idea for effectively raising the immune response to malaria DNA vaccine.

Tunicamycin enhances TRAIL-induced apoptosis by inhibition of cyclin D1 and the subsequent downregulation of survivin

TNF-related apoptosis-inducing ligand (TRAIL) has been proposed as a promising cancer therapy that preferentially induces apoptosis in cancer cells, but not most normal tissues. However, many cancers are resistant to TRAIL by mechanisms that are poorly understood. In this study, we showed that tunicamycin, a naturally occurring antibiotic, was a potent enhancer of TRAIL-induced apoptosis through downregulation of survivin. The tunicamycin-mediated sensitization to TRAIL was efficiently reduced by forced expression of survivin, suggesting that the sensitization was mediated at least in part through inhibition of survivin expression. Tunicamycin also repressed expression of cyclin D1, a cell cycle regulator commonly overexpressed in thyroid carcinoma. Furthermore, silencing cyclin D1 by RNA interference reduced survivin expression and sensitized thyroid cancer cells to TRAIL; in contrast, forced expression of cyclin D1 attenuated tunicamycin-potentiated TRAIL-induced apoptosis via over-riding downregulation of survivin. Collectively, our results demonstrated that tunicamycin promoted TRAIL-induced apoptosis, at least in part, by inhibiting the expression of cyclin D1 and subsequent survivin. Of note, tunicamycin did not sensitize the differentiated thyroid epithelial cells to TRAIL-induced apoptosis. Thus, combined treatment with tunicamycin and TRAIL may offer an attractive strategy for safely treating resistant thyroid cancers.

Tunicamycin enhances TRAIL-induced apoptosis by inhibition of cyclin D1 and the subsequent downregulation of survivin

TNF-related apoptosis-inducing ligand (TRAIL) has been proposed as a promising cancer therapy that preferentially induces apoptosis in cancer cells, but not most normal tissues. However, many cancers are resistant to TRAIL by mechanisms that are poorly understood. In this study, we showed that tunicamycin, a naturally occurring antibiotic, was a potent enhancer of TRAIL-induced apoptosis through downregulation of survivin. The tunicamycin-mediated sensitization to TRAIL was efficiently reduced by forced expression of survivin, suggesting that the sensitization was mediated at least in part through inhibition of survivin expression. Tunicamycin also repressed expression of cyclin D1, a cell cycle regulator commonly overexpressed in thyroid carcinoma. Furthermore, silencing cyclin D1 by RNA interference reduced survivin expression and sensitized thyroid cancer cells to TRAIL; in contrast, forced expression of cyclin D1 attenuated tunicamycin-potentiated TRAIL-induced apoptosis via over-riding downregulation of survivin. Collectively, our results demonstrated that tunicamycin promoted TRAIL-induced apoptosis, at least in part, by inhibiting the expression of cyclin D1 and subsequent survivin. Of note, tunicamycin did not sensitize the differentiated thyroid epithelial cells to TRAIL-induced apoptosis. Thus, combined treatment with tunicamycin and TRAIL may offer an attractive strategy for safely treating resistant thyroid cancers.