Objective To explore the expression of GATA-3 in pulmonary tissue of asthmatic mice and the inhibitory effect of dexamethsone(Dex)on it.Methods The Blab/c mice asthma model was induced by ovalbumin(OVA) with classic method.Twenty-four male mice were randomly divided into control group,asthmatic group and Dex treated group.The expression of GATA-3 protein was measured by immunohistochemical staining.The expression of GATA-3 mRNA was measured by reverse transcription-polymerase chain reaction(RT-PCR).The level of IL-4 in mice spleen CD4 T cell was measured by flow cytometry.The airway inflammation was evaluated by HE staining.Results The percentages of postive GATA-3,GATA-3 mRNA and IL-4 protein of asthmatic group were significantly higher than those of control group(P

Objective To investigate the transfection efficacy and expression of Ang-1 gene and proangiogenesis in vitro and vivo by ultrasound-mediated microbubble destruction.Methods 293T cells were divided into three groups:group A was given hAng-1 plasmid and microbubbles plus ultrasonic irradiation,group B was given hAng-1 plasmid and ultrasound,group C was given hAng-1 plasmid only (without ultrasound).Forty-eight hours after transfection,the transient expression rate was observed under fluorescence microscopy and flow cytometry.RT-PCR and Western blot analysis were taken to evaluate the mRNA and protein expression of Ang-1 respectively.Twenty-seven rabbit models of ligated left circumflex branch coronary artery were divided into 3 groups randomly as follow:group Ⅰ (accepted intravenous injection of SonoVue microbubble and Ang-1 plus ultrasonic irradiation),group Ⅱ (accepted intravenous injection of Ang-1 with ultrasound),group Ⅲ (control group).Myocardial contrast echocardiography (MCE) was executed on all animals before and after the treatment.Two weeks after gene delivery,RT-PCR and Western blot analysis were taken to evaluate mRNA and protein expression of Ang-1 respectively.Microvessel density (MVD) counting of infracted myocardium,observed by Factor Ⅷ immunochemical staining,was performed to value the proangiogenesis effect of Ang-1 delivered by ultrasound mediated cavitation of microbubble.Results Green fluorescence was observed in group A and B by fluorescence microscopy,which was negative in group C.The transfection expression rate was significantly improved in group A ( P ＜ 0.01).In vivo,Microbubbles could be observed in former ischemic myocardium in MCEexamination and the Ang-1 mRNA and protein could be detected in group Ⅰ.On the other hand,the contrast agent was defected obviously and none of the animals showed Ang-1 mRNA and protein expression in other two groups.The MVD counting showed significant improvement in group Ⅰ whereas other two groups didn't.ConclusionsMicrobubble-enhanced ultrasound exposure can improve the Ang-1 gene transfection expression rate observably both in vitro and in vivo.This strategy for delivering has great proangiogenesis effect in vivo.

Objective To explore the capability of Ang-1 gene delivery to acute myocardial infarction using targeted microbubbles carrying ICAM-1 antibody.Methods Thirty-seven rabbits' left circumfles branch coronary arteries were ligated for models.Three rabbits were injected with microbubbles carrying ICAM-1 to detect the ability of targeting.Thirty-four rabbit models were divided into 3 groups randomly as follow:IM group (n=12,accept direct intramuscular injection),ICAM-1 group (n=12,accept intravenous injection of targeted microbubbles and Ang-1) and control group (n=10,without any treatment).Ultrasonography were executed on all animals before and 2 weeks after the treatment.All rabbits were killed after 2 weeks and examined for Ang-1 mRNA and protein by RT-PCR and Western-Blot respectively.Microvessel density (MVD) counting of infracted myocardium,observed by factor Ⅷ immunochemical staining,was performed to value the proangiogenesis effect of Ang-1 delivered by targeted microbubbles carrying ICAM-1 antibody.The liver and the kidney in ICAM-1 group were taken to assess the systemic delivery.Results IM and ICAM-1 group showed significantly improvement in the ejection fraction (P＜0.05) while control group did not.Ang-1 mRNA and protein could be detected in IM and ICAM-1 group;however,the expression between the two groups showed no siginificant difference.None of the control animals showed Ang-1 expression.compared with ICAM-1 group,the MVD was greater in IM group.Ang-1 was not detected either in liver or in kidney in ICAM-1 group.Conclusions Targeted microbubbles carrying ICAM-1 antibody can deliver Ang-1 gene to ischemic myocardium directly.Meanwhile,it's as effective as IM injections besides the greater angiogenesis effect.This strategy improves the perfusion of acute myocardial infarction and the function of heart.

Objective To investigate the effects of ultrasound and microbubbles on transient transfection of angiopoietin-1(Ang-1) into vascular endothelial CRL1730 cells and the effects of cell migration. Methods CRL1730 cells were divided into four groups:a control group (without any treatment), an Ang-1 group (with Ang-1 added only), a microbubble group (with only microbubbles added) and an ultrasound with microbubbles group (Ang-1, microbubbles and 1 MHz ultrasound irradiation for 30 s). All were cultivated for 24 h. The expression of Ang-1 mRNA and protein were measured using RT-PCR and Western blotting. The cells' migration ability was detected using the Transwell method. Results There was no difference in the expression of Ang-1 mRNA or protein among the control group, the Ang-1 group and the microbubbles group, but the expression of Ang-1 mRNA and protein was increased significantly in the ultrasound plus microbubbles group. Cell migration ability was also enhanced significantly in the ultrasound and microbubbles group compared with the other groups. Conclusion A combination of ultrasound and microbubbles might successfully transfect Ang-1 genes into vascular endothelial cells, and the migration ability of the transfected cells is also enhanced.

Objective To test the efficiency of gene transfer and expression mediated by ultrasound and microbubble strategy in 293T cell. Methods SonoVue microbubbles were mixed with pla.smid DNA encoding hAng-1 and green fluorescent protein. The mixture of the DNA and microbubbles was administer to cultured 293T cells by ultrasound exposue. The ultrasound condition was 1. 5 W/cm~2 and 30 s. Forty eight hours later, transfer rate was assessed by fluorescence microscopy and flow cytometry. Cell viability was assayed by Trypan Blue staining. RT-PCR and Western blot analysis was used to examine the expression of hAng-1 mRNA and protein. Agarose gel electrophoresis was used to evaluate the integrity of the plasmid. Results The transfection expression rate of eGFP in 293T cells was markedly increased with the additon of 20% microbubbles and 15 mg/L DNA. Fetal calf serum had no influence on the gene transfer rate. Ultrasound irradiation combined with microbubbles couldn't destroy the integrity of plasmid. Conclusions Ultrasound-mediated microbubbles destruction can increase the transfection and expression of hAng-1 gene in 293T.

Objective To certificate the effects on transfection ratio and cells viability of ultrasound (US) acoustic intensity, radiation duration, microbubbles concentration and DNA concentration in delivery human angiopioetin-1 gene (hAng-1) into 293T cells by SonoVue microbubbles and decide the optimal transfection parameters. Methods Mix 293T cells and SonoVue microbubbles linked with eGFP-C_3-hAng-1 in a different way, detect the gene transfection ratio and cells viability under the various US intensity, radiation duration, microbubbles and DNA concentrations. Results The gene expression would be increased if enhanced the intenstiy of US,radiation time,microbubbles and DNA concentrations,and the cells viability would be kept more than 90% ( P <0. 01). Whereas,if the US intensity increased over 1. 5 W/cm~2 ,the duration over 30 s and microbubbles and DNA concentrations over 20% and 15 mg/L respectively,the gene expression would not increase significantly ( P > 0. 05),whereas coupled with obviously decreased cells viability( P <0. 01). Conclusions The optimal conditions of deliver hAng-1 gene into 293T cells by SonoVue microbubbles was mixing cells and microbubbles in a cell wall-sticky way,US intensity was 1. 5 W/cm~2, duration 30 s,20% microbubbles and 15 mg/L DNA concentration.

Objective To explore protective effect of Heme oxygenase-1(HO-1) on irradiation-induced endothelial cell apoptosis.Methods Human endothelial cell line EA.hy926 were administered with or without HO-1 activator Copp and/or HO-1 inhibitor Znpp,respectively.Then,cells were treated with or without 8 Gy radiation.The HO-1 protein expression of cells were assessed with Western blotting and apoptosis of cells treated with irradiation were evaluated with flow cytometry.Moreover,cytochrome C releasing into cytosol were also determined by Western blotting. Results In PBS+R group,HO-1 protein expression of EA.hy926 was low posterior to irradiation.When cells were preconditioned with Copp and/or Znpp,then recieved with 8Gy irradiation,the HO-1 protein expression of EA.hy926 increased significantly in comparision with the PBS+R group(P

Objective To explore the feasibility of the application of non fasting blood lipid in the hospitalized population.Methods Self-control study was used.608 patients(aged 20~86 years old) were enrolled from April 2015 to October 2016 in lipid center of FuWai hospital.Fasting sample and non-fasting sample(1~4 h after breakfast) were collected from every patient and lipid profile including TG (triglyceride), TC (total cholesterol), HDL-C (high density lipoprotein cholesterol) and LDL-C (low density lipoprotein cholesterol) were measured in clinical laboratory.The results of two tests were compared using the Wilcoxon signed-rank test.Results The differences between non-fasting and fasting lipid test were +0.47 mmol/l (+30%) for TG,-0.03 mmol/l (-2.8%) for HDL-C,-0.09 mmol/l (-3%) for LDL-C and-0.24 mmol/l (-8.7%) for calculated LDL-C (P<0.001 respectively).The differenceswere +0.01 mmol/l for TC and +0.02 mmol/l for non-HDL-C,therefore no statistical difference was observed.When the TG level was stratified,the level of non-fasting LDL-C using directing test method was not significantly different between TG> 4.5 mmol/L and the whole (0.07 vs.0.09),but the level of non-fasting LDL-C using formula method wassignificantly different between TG> 4.5 mmol/L and the whole (0.66 Vs.0.24),andthe drops were 34.9% vs.8.7%.Conclusion Non-fasting lipid test could be an effective routine method for lipid evaluation in the hospitalized population.

Objective To increase the transfection of EGFP-N3 plasmids into 293T cells using ultrasound-targeted microbubbles delivery(UTMD) mediated a peptide nucleic acid (PNA) binding nuclear localization signal (NLS).Methods Antibody-targeted microbubbles were used in the experiments which can specifically recognize the SV40T antigen receptor.The SV40T antigen receptors were expressed on the membranes of 293T cells.The PNA containing the NLS were inserted in the EGFP-N3 plasmid DNA,which increased nuclear localization.Ultrasound-targeted microbubble delivery (UTMD) and the PNA binding NLS were utilized to improve the cytoplasmic import of plasmids and the nuclear intake of the plasmid from the cytoplasm,respectively.The study was divided into five groups:Contrast (group A),Common microbubble + DNA (group B),Antibody-targeted microbubbles + DNA (group C),Common microbubbles + NLS-PNA-DNA (group D),Antibody-targeted microbubbles + NLS-PNA-DNA (group E).Fluorescence microscope was used to observe the fluorescent light in each group;flow cytometry to test the transfection;RT-PCR and Western blot to detect genes' mRNA and protein expression level.Results Ultrasound and antibody-targeted microbubble delivery (UTMD) significantly enhanced the cytoplasmic intake of exogenous genes and maintained high cell viability(＞80％).Fluorescent microscope showed that the quantities of green fluorescence in cells were increased successfully.The transfection results of flow cytometry were 0,(9.30 ± 0.46)％,(26.46 ± 2.01)％,(29.54 ± 0.62)％,(45.72 ± 1.86)％,respectively,and the differences were statistically significant(P ＜0.05).The relative mRNA and protein expression in group E were greater than those in group C and D respectively (P ＜0.05).Conclusions UTMD combined with antibody-targeted microbubbles and a PNA binding NLS plasmid can significantly improve transfection efficiency of exogenous genes by enhancing both cytoplasmic and nuclear DNA import.