Prevention of allograft vasculopathy by transfection of calcitonin gene-related peptide

Abstract

To explore suppression of allograft vasculopathy (AV) by transfer of the calcitonin gene-related peptide (CGRP). The descending thoracic aortas from 39 Lewis rats were grafted to the abdominal aortas of 39 F344 rats, and then the recipient rats were randomized into 3 groups: Group A transfected with Ad5-CGRP-EGFP, a gene construct containing sequences from the adenoviral oncoprotein, CGRP, and enhanced green fluorescent protein, Group B, transfected with Ad5-EGFP containing sequences from the adenoviral oncoprotein and enhanced green fluorescent protein, and Group C without transfection. Four and 8 weeks later the abdominal aortas of 6 recipient rats from each were collected respectively. The degree of vascular obstruction was observed by microscopy. Frozen tissue sections were made and the inverted phase contrast fluorescence microscopy was used to observe the green fluorescence showing the expression of CGRP and enhanced green fluorescent protein (EGFP). RT-PCR was used to detect the expression of CGRP. Immunohistochemistry was used to examine the expression of inducible nitric oxide synthase (iNOS) and vascular cell adhesion molecule-1 (VCAM-1). The apoptosis index (AI) was detected by TUNEL method. CGRP expression was positive in both Group A and B 4 weeks later and negative in both groups 8 weeks later. Group A is approximately normal in pathological morphology 4 weeks later. The vascular luminal occlusion score of Group A was lower than Groups B and C 4 weeks later, and significantly higher than that I the same group (P < 0.05), however, still lower than those of Groups B and C 8 weeks later. The AI 4 weeks later of Group A was significantly lower than those of Groups B and group C, however, there was no significance in AI among the 3 groups. Four weeks later the VCAM-1 expression levels in the tunicae intima, media, and adventitia of Group A were all significantly lower than those of Group B C 4 weeks later, however, the iNOS expression level of Group A was only significantly lower than those of Groups B and C in the tunica intima 4 weeks later. The expression of CGRP effectively suppresses the development of AV 4 weeks after the operation.