Vascular Smooth Muscle Cell Apoptosis Promotes Allograft Arteriosclerosis Through Inducing The Production Of SDF-1α

Objective-Allograft arteriosclerosis is a vascular proliferative disease characterized by diffuse intima hyperplasia and is also a leading cause of late allograft loss, limiting the long-term success of organ transplantation. Allograft medial vascular smooth muscle cell (VSMC) apoptosis is the early event after organ transplantation and is involved in allograft arteriosclerosis by recruitment of bone marrow-mesenchymal stem cells (BM-MSCs). The present study elucidates the underlying molecular mechanism and the contribution of medial VSMC apoptosis to allograft arteriosclerosis. Methods-Recombinant lentivirus vector of wild-type p53 carrying a SM-22αpromoter was constructed and transferred into primarily cultured VSMCs. Nuclei staining with DAPI and flow cytometry was used to detect the roles of overexpressing p53 in inducing VSMC apoptosis. The effect of VSMC apoptosis on expression of SDF-1α, TGF-βand PDGF-BB was assessed by Western blot and ELISA. BM-MSCs were primarily cultured in vitro and conditioned medium was prepared from VSMCs with overexpressing p53. The influence of VSMC apoptosis on BM-MSCs migration and proliferation was further evaluated by Transwell and BrdU labeling. BM-MSCs were pretreated with selective inhibitor of cell signaling pathway to explore the molecular mechanism that VSMC apoptosis promoted migration and proliferation of BM-MSCs. Rat aortic transplant models were established. Recombinant lentivirus vector of BclxL carrying a SM-22αpromoter was constructed and transferred into rat aortic allografts. TUNEL assay was performed to detect the medial VSMC apoptosis in allograft aorta. Expression of SDF-1αin allograft arteries was detected by immunostaining. Histopathology of allograft arteries was assessed by HE and Masson staining to explore the influence of BclxL gene transfer in allograft arterial neointima formation. On the other hand, the allograft recipient rats were treated with rapamycin and PD98059, the inhibitor of mTOR and MAPK/ERK signaling, to investigate their effect on allograft arterial neointima formation thereby exploring the roles of PI3K/Akt/mTOR and MAPK/ERK signaling in allograft arteriosclerosis. Results-Lentivirus-mediated p53 overexpression effectively induced VSMC apoptosis. Apoptotic VSMCs released chemokine SDF-1αand induced neighbor vital VSMCs express SDF-1αthereby elevating SDF-1αlevel in conditioned medium from VSMCs with overexpressing p53. VSMC apoptosis was able to promote BM-MSCs migration and proliferation however, neutralizing antibody of SDF-1αwas capable of abolishing this effect. Western blot showed that VSMC apoptosis activated PI3K/Akt/mTOR and MAPK/ERK signaling in BM-MSCs by SDF-1α. Pretreatment of BM-MSCs with inhibitor of PI3K/Akt/mTOR signaling or MAPK/ERK signaling, Ly294002 and rapamycin or PD98059 obviously prevented VSMC apoptosis-induced migration and proliferation of BM-MSCs. Compared with isograft arteries, medial VSMC apoptosis and expression of SDF-1αsignificantly revealed in allograft arteries. After 8 weeks transplantation, neointima with activation of PI3K/Akt/mTOR and MAPK/ERK signaling obviously formed in allograft arteries. BclxL was selectively overexpressed in medial VSMCs by lentivirus-mediated BclxL gene transfer under regulation of SM-22αpromoter. Overexpression of BclxL significantly inhibited medial VSMC apoptosis and reduced SDF-1αexpression level both in allograft arterial media and serum. Inhibition of early medial VSMC apoptosis in allograft arteries accompanied notable decrease of CD90+CD105+ double-positive cells in neointima as well as attenuation of neointima areas and luminal stenosis for 8 weeks transplantation. Likewise, both of rapamycin and PD98059 was able to suppress recruitment of bone marrow-mesenchymal stem cells (BM-MSCs) to neointima and prevent allograft arteriosclerosis. Conclusions-Allograft arterial medial VSMC apoptosis plays crucial roles in development of allograft arteriosclerosis through recruitment of BM-MSCs by inducing the production of SDF-1α. Activation of PI3K/Akt/mTOR or MAPK/ERK signaling is the molecular mechanism for VSMC apoptosis-induced migration and proliferation of BM-MSCs. VSMC apoptosis represents a relevant target of therapeutic interventions for allograft arteriosclerosis.