| Objective:To study effects of prostaglandin Al (PGA1) and triptolide (TRI) on rat cardiac microvascular endothelial cells ( RCMEC ) and to observe protective effects and molecule mechanism of prostaglandin Al and triptolide on ischemical reperfusion injury of allograft in rat cardiac transplants by inhibitory effects on NF-KB activity.Methods:Isotype dystopy rat heart transplantation model were performed. Donor allografts were treated with Saline,PGA1 (20 40 80ng kg-1 three dosage groups),TRI ( 0. 1 0. 2 0. 5mg.kg-1 three dosage groups) and lipopolysaccharide (LPS,50ug .kg-1) ,respectively. Reperfusion injury was assessed with cardiac edema (percent of wet weight) ,neutrophil infiltration (MPO) ,and ultrastructure of myocardial tissues. Adhesion molecular expression was determinated by enzyme linked immunosorbent assay (ELISA) . NF-KB activity of myocardial tissues were estimated by electrophoretic Mobility Shift Assaysm (EMSA) .RCMECs were cultured and hypoxia-reoxgenation model of endothelial cells were performed. Experimental cells were divided into control group (N,treated with normoxia) ,hypoxia group (H,treated with hypoxia 15 hours) ,hypoxia /reoxgenation group (H/R,hypoxia 15 hours and reoxgenation 24 hours),PGA1 group (PGA1,treated with H/R and PGA1 20,40,80nmol/L,respectively),TRI group (TRI,treated with H/R and Tri 2. 5,5,10 u g/L,respectively) and PDTC group (PDTC,treated with H/RandPDTC). Neutrophil is isolated from blood,adhesion rate of endothelial cells and neutrophil were determined by adhesion model. NF- K B activity of endothelial cells were determined by EMSA. Protein expression of ICAM-1 and VCAM-1 of endothelial cells was estimated by ELISA,and mRNA expression of ICAM-1 and VCAM-1 of endothelial cells was assayed by Northern Blot. Endothelial cells forms were observed and the number was counted. The amounts of DNA fragmentation in the sediment and solution were determined and then the cells DNA fragmentation ratio (DNA amount in the solution/DNA amount in both of the sediment and solution) was calculated. Endothelial cells apoptosis was determined by terminal deoxynucleotidyl transf erase mediated d-UTP nick end labeling (TUNEL) . Bcl-2 and Bax protein expression was examined by Western blot and Bcl-2mRNA expression was examined by Northern blot.Results:In saline control,when donor undergo ischemic reperfusion injury,MPO activity and edema of myocardial tissue were markedly increased;ICAM-1 and VCAM-1 protein expression was incresed;and NF- K. B activity was high,graft survival were 6.4 + 0.6 days. The parameters of reperfusion injury in LPS group were high than these in saline group. MPO activity,NF- K B activity,ICAM-1 and VCAM-1 protein expression were increased markedly compared with saline control,graft survival Shortened to 4. 4 + 1. 1 days (P<0. 01) . After treatment with PGA1 and TRI,MPO activity was decreased markedly;myocardial tissues edema was relived,ICAM-1,VCAM-1 and NF- K B activity were downregulated compared with saline control;Graft survial time were prolonged to 15.6 +1.5 days and 17.0 +1.1 days,respectively (P<0. 01) . Inhibition of PGA1 on NF-K B was stronger than that of Tri,but graft survival in TRI group longer than that of PGA1 (P<0. 05) . (2) NF- K B activity,protein expression of ICAM-1 and VCAM-1 and adhesion rate of neutrophil and RCMECs treated with hypoxia were increased markedly,and after reoxgenation,these changes were more obvious. When endothelial cells were treated with reoxgenation supplement with PGA1,TRI and PDTC,adhesion rat of endothelial cells weredecreased markedly in dose-dependent manner;NF- K B activity were decreased apparently compared with H/R ( P<0. 01 ) ;Inhibition of PGAl on NF- K B activity was stronger than that of Tri and PDTC (P<0. 05) . Protein expression of ICAM-1,VCAM-1 and mRNA expression of ICAM-1,VCAM-1 in PGAl,Tri and PDTC groups are decreased markedly than these in H/R group (P<0. 01 ) ,but there are no difference apparently among PGAl,Tri and PDTC groups (P>0. 05) . (3) Numbers of RCMECs tre...
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