Objective To isolate the rat insulin gene enhancer binding protein 1(Islet-1)gene and construct plEGFP-C1-Islet-1 recombinant retroviral expression vector.Methods The cDNA encoding the rat Islet-1 gene was isolated by RT-PCR method,the cDNA was first cloned into PGET-1 TA vector to facilitate the sequence and then subcloned into the retroviral vector plEGFP-C1.plEGFP-C1-Islet-1 was transfected into PA317 packaging cells with lipofectamine 2000.Transformants were selected in medium containing G418.Results A 1 050 bp DNA fragment was obtained by RT-PCR;plEGFP-C1-Islet-1 recombinant retroviral expression vector was identified by restrictive enzymes digestion,PA317 cells transfected with recombinant vector expressed enhancer green fluorescent protein(EGFP).Conclusion The gene encoding the rat Islet-1 is obtained and plEGFP-C1-Islet-1 expression vector is constructed successfully.

Objective To evaluate the combined effects of vascular endothelial growth factor (VEGF) and placental growth factor (PLGF) on angiogenesis and cardiac function and compare with VEGF or PLGF only in acute myocardial infarction rats.Methods Seventy-five males Sprague-Dawley(SD) rats were randomly divided into five groups:sham group,NS group,VEGF group,PLGF group,and VEGF + PLGF group with 15 rats in each group.All the rats underwent LAD ligation and injection of NS,VEGF,PLGF,VEGF + PLGF,in the peri-infarct area,respectively,besides the sham group.Three weeks after coronary artery ligation and different agents injection,cardiac function,myocardial scar area,angiogenesis and arteriogenesis were studied.Cardiac structure and function,and infarct size were assessed by echocardiography.The number of new vessels and the number of new arterioles were evaluated by haematoxylin-eosin staining and immunohistochemistry staining.Results Three weeks after LAD ligation and different agents injection,the LVEDD and LVESD were significantly decreased (P ＜ 0.01)in NS group,VEGF group and PLGF group.While the LVEF and LVFS were higher in VEGF + PLGF group than that in other groups.Myocardial infarct size was reduced in VEGF group (P ＜ 0.05).Angiogenesis and arteriogenesis were higher in VEGF + PLGF group than that in VEGF group (P ＜ 0.01) and PLGF group (P ＜0.05).Angiogenesis and arteriogenesis were significantly higher in PLGF group than that in VEGF group (P＜0.01).The density of microvessels in VEGF group was higher than that in NS group (P ＜ 0.05),while arteriogenesis was of no statistical difference.Conclusion The combination of half VEGF and PLGF can increase angiogenesis and arteriogenesis in the ischemic marginal zone of myocardial infarction,decrease myocardial infarction area,and improve cardiac function.

Objective To evaluate the clinical application of CT perfusion imaging (CTPI) and CT subtraction angiography (CTSA) in the diagnosis of acute ischemic cerebrovascular disease (AICVD). Methods 24 cases with AICVD onset within 24 hours were examined with regular CT, CTPI and CTSA. Some of them took CTPI, MRI, MRA, DSA, SPECT by follow up examinations. Results In 24 cases 11 had regular CT negative results after onset of stroke 3～6 hours in 6 cases,6～12 hours in 3 cases,12～24 hours in 2 cases. Ten cases of them were confirmed by CTPI as having ischemic lesions, 2 cases had middle cerebral artery occlusion (MCAO), and 1 had transient ischemic attack (TIA) with CTPI negative. In 24 cases 13 had regular CT positive rseults, 9 cases had ischemic lesions larger in CTPI than in regular CT,1 case had MCAO and 1 case had internal carotid artery occlusion(ICAO). There were 4 cases with ischemic lesions on regular CT almost having the same range as that of lacunar infarction in CTPI. The peak value of time(PT), mean transit time(MTT), relative flow (RF) in all 24 cases were found obviously changed. The side of ischemic lesion as compared with the opposite side, and the core of ischemic lesion as compared with peripheral zone were found changed significantly ( P

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Objective To assess whether end-tidal carbon dioxide partial pressure (PET CO2) can predict the fluid responsiveness in septic shock patients.Methods Septic shock patients under mechanical ventilation without spontaneous breathing and with the need of a fluid challenge test were included in this study.Heart rate,central venous pressure,pulse pressure,PErCO2,and CI before and after the fluid challenge test were conducted in all the patients.Results Of the 48 septic shock patients included,34 had preload responsiveness,14 had no responsiveness.△CI and △PET CO2 after the fluid challenge test involume responders were (0.85 ± 0.47) L · min-1 · m-2 and (3.5 ± 2.5) mmHg respectively,which were higher than those in no volume responders (P ＜ 0.05).The fluid-induced changes in PET CO2 and CI were correlated (r =0.072,P ＜ 0.05).The AUCRoc of fluid challenge-induced △PET CO2 as the predictor for volume responsiveness was 0.943,and its sensitivity was 87.9％ and specificity was 93.4％ with a critical value of 5％.The AUCRoc of △PP as the predictor for volume responsiveness was 0.801,and its sensitivity was 68.1％ and specificity was 73.2％ with a critical value of 10％.Conclusion The changes of PETCO2 induced by a fluid challenge test can predict fluid responsiveness with reliability,and have a better sensitivity and specificity than the changes of PP.

Objective To construct recobinant pmRNA IRES-hKDR and lay a primary foundation for further study of gene therapy for tumors.Methods pmRNA IRES-hKDR was obtained from pDC520 hkDR and pmRNA IRES-Luc which was reserved by our laboratory through molecular biology technology and was transfected into Hepall-6 via lipsome after identification of PCR,restriction enzyme digesting and DNA ELISA and Western blot.Results The recombiant plasmid had pmRNA IRES-hKD gene which was expressed in Hepall-6//G418 in vitro and the expressed protein was secreted out of the cells could response with specific hKDR antibody,which was identified by Western blot.Conclusion An eukaryotic expression recombiant plasmid pmRNA IRES-hKDR can be constructed successfully which provides the possibility of further researches on its anti-tumor effect.

Aim To study the effects of propofol on apoptosis of cortical astrocytes isolated from neonatal rats.Methods Pure astrocytes(AST)were obtained from the cultured cerebral cortical cells and identified by the GFAP stain technology in neonatal rats.AST cells were treated with different concentrations of propofol(0,10,30,90 μmol·L-1)for 8 hours.Cell viability was measured by MTT method,and the apoptosis was detected by Annexin V/PI double staining.Cytochrome C(cyt-C)leakage was detected by Western blot.Caspase-3 and caspase-9 activities were measured by spectrophotometry.Results AST cells were administered with different concentrations of propofol(0,10,30,90 μmol·L-1)for 8 hours.It decreased the survival rate in a concentration-dependent manner,induced the leakage of cyt-C in mitochondria,up-regulated the expression of pro-apoptotic protein caspase-3 and caspase-9,and induced AST apoptosis.Conclusion Propofol induces the apoptosis of astrocytes in neonatal rat cortex in vitro,which may be related to the activation of mitochondria apoptosis pathway induced by mitochondrial cyt-C release.

Objective To study the type variation of microglial activation in spinal dorsal horn of rats after sciatic nerve injury.Methods Healthy adult male Sprague-Dawley rats were randomly divided into the control and experimental groups, 24 rats in each group.The experimental group underwent ligation of sciatic nerve trunk to generate nerve injury in the rats.The pain behavior in the rats was measured at the 1th, 7th and 14th postoperative days, and the changes of microglial activation in the rat lumbar spinal cord dorsal horn was detected by immunofluorescence staining.qRT-PCR assay was used to validate the activation trends of M1 and M2 types of microglia cells.Results No significant changes were found in the microglial cells in the spinal cord dorsal horn of rats in the sham-operation group during 14 days after operation.In the sciatic nerve ligation group at 1 day after operation, no significant change was observed in the number of microglial cells, but the expression of marker of M1 microglia was significantly increased.At 7 and 14 days after operation, the number of microglial cells and the expression of M1 microglia marker in the spinal cord dorsal horn were increased significantly.Conclusions Microglia activation in the spinal dorsal horn starts at the first day after sciatic nerve injury, and lasts at least for two weeks after the operation.M1 microglia activation dominates during this period.