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《Scientia Agricultura Sinica》 2012-02
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Establishment of Aortic Endothelial Cells Model in Vitro from Wuzhishan of Inbred Miniature Pig and Expression of In2ammatory Cytokines Mediated by TLR2

WANG Pan1,2,TAO Xiao-li3,SHEN Zu-nan2,LI He-gang3,PAN Jie1,MU Yu-lian2,LI Kui2(1College of Life Science,Shandong Normal University,Ji'nan 250014;2Institute of Beijing Animal Science and veterinary,Chinese Academy of Agricultural Sciences,Beijing 100193;3College of Veterinary Medicine,Huazhong Agricultural University,Wuhan 430070)  
【Objective】 This study aimed to explore a protocol of isolation,culture and identification of Wuzhishan Inbred Miniature Pig aortic endothelial cells(EC),and to study the regulation of Toll-like receptor 2(TLR2) expression by LPS in EC with the expression of in2 ammatory cytokines mediated by TLR2.【Method】Thoracoabdominal aortic were isolated from 3-4 month-old Wuzhishan Inbred Miniature Pig,disassociated with 0.1% collagenase I.Aortic EC were identified with the methods of uptake of fluorescence-labeled acetylated low density lipoproteins(DiI-Ac-LDL) and detecting of CD31 expression in the cells by flow cytometry.After stimulation of aortic derived EC with 1 μg·mL-1 lipopolysaccharide(LPS) for 0,6,8 and 12 h,the level of TLR2 and TLR4 mRNA was detected by real-time PCR.Furthermore,we detected the influence of lipoteichoic acid(LTA) on IL-6,IL-8 and ICAM-1 expression in aortic EC pretreated for 12 h with 1 μg·mL-1 LPS.【Result】 Obtained Wuzhishan Inbred Miniature Pig aortic EC displayed paving stone-like pattern with high vitality.The cultured cells expressed CD31 and Ac-LDL Receptor in membrane,demonstrating that they were indeed EC.Stimulation of aortic derived EC with LPS induced a strong increase in TLR2 mRNA but not in TLR4 mRNA.LTA potentiated the increase of the in2 ammatory cytokines IL-6,IL-8 and ICAM-1 expression in EC pretreated with LPS,but not in EC without LTA.【Conclusion】The present study successfully isolated and identified Wuzhishan Inbred Miniature Pig aortic EC in vitro culture system.LPS stimuli induced a high TLR2 expression in EC,which mediated the expression of inflammatory cyokines.
【Fund】: 国家自然科学基金项目(30871775)
【CateGory Index】: S828
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