Gene Transfer Of Calcitonin Gene-Related Peptide Suppresses The Development Of Allograft Vasculopathy

Objective To investigate construction of enhanced green fluorescent protein (EGFP)labeled recombinant adenovirus containing Calcitonin Gene-related Peptide (CGRP);To establish an experimental rat thoracic-to-abdominal aorta transplantation model for study of allograft vasculopathy; to explore gene transfer calcitonin gene-related peptide suppresses the development of allograft vasculopathy and its mechanism.Methods To identify CGRP,the recombinant adenovirus Ad5-CGRP-EGFP was constructed by us in Ad5Easy system based on the homologous recombination in bacteria. A rat allograft vasculopathy model established by thoracic to infrarenal abdominal aorta transplantation using F344 and Lewis rats.The rats were randomly divided into 5 groups: F344 group,isograft group and allograft group,gene transfer Ad5-EGFP of EGFP group and gene transfer Ad5-CGRP-EGFP of CGRP group; to study the aorta pathological morphology after operation 4 weeks,8 weeks. to view the fluorescence with freezen tissue sections under fluorescence microscope.to mensurate CGRP of tissue with RIA,to view the CGRP,VCAM-1 and iNOS intensity and position with immunohistochemistry,to mensurate the tissue apoptotic index(AI)with TUNEL. Results 1 The adenovirus Ad5-CGRP-EGFP was quickly constructed by homologous recombination bacteria using AdEasy system.2 A rat model of allograft vasculopathy was successfully established. after operation 4 weeks,8 weeks,The Lewis isograft group only appears a few cellulars infiltrate of monocytes,lymphocytes,The F344 to Lewis allograft group appears difference grades of pathological changes.3 After operation of 4 weeks,the media and adventitia of allograft vessel of CGRP group was viewed green fluorescence under fluorescence microscope by freezen tissue sections. the media isn,t evidence thickening in pathological morphology.The tissue CGRP is higher at statistically significant than other groups by to mensurate with RIA.The adventitia CGRP of CGRP group immunohistochemistry score is higher at statistically significant than other groups.The instance isn,t seen in other groups. After operation of 8 weeks,the tissue CGRP of CGRP group is only higher at statistically significant than EGFP group and allograft group.4 After operation of 4 weeks,the CGRP group is approximately normal in pathological morphology. the intima is smoothness,the media is intact,the adventitia have a little lymphocyte and inflammation cell,the vascular luminal occlusion score is lower than allograft group and EGFP group,the allograft group and EGFP group are difference occlusion score of pathological morphology. After operation of 8 weeks, the CGRP group appear difference grades changes in pathological morphology,but,the vascular luminal occlusion score is still lower than allograft group and EGFP group. After operation of 4 weeks and 8 weeks,the isograft group is approximately normal in pathological morphology,only appear a little lymphocyte encroach.5 After operation of 4 weeks,the AI of CGRP group is statistically significant lower than allograft group and EGFP group at intima,media and adventitia;After...