Abstract
Epithelial mesenchymal transformation (EMT) has been proposed to play a role in chronic lung allograft dysfunction (CLAD). Liver kinase B1 (LKB1), a tumor suppressor gene, can regulate EMT. However, its role in CLAD development following lung transplantation remains unknown. Goal of this study is to determine the role of LKB1 on Rab27 mediated exosome release and immune-pathogenesis of CLAD.
We first analyzed EMT on airway epithelial cell line (BEAS-2B) with exosomes from bronchiolitis obliterans syndrome (BOS) and stable. Role for LKB1 using SiRNA mediated transfection of BEAS-2B and in a hypoxia/reperfusion model were determined. Using qRT-PCR, we analyzed LKB1 and miR-155 levels in biopsies from lung transplant recipients with BOS and stable and fibrosis in a murine lung transplant model of chronic rejection.
In BEAS-2B, knockdown of LKB1 increased exosome release (p=0.0001). Co-culture of BEAS-2B with exosomes from BOS increased vimentin (p=0.001) and α-SMA (p=0.001). LKB1 knockdown increased Rab27A (p=0.01), α-SMA (p=0.01), and vimentin (p=0.01) following hypoxia/reperfusion. qRT-PCR of biopsies from lung transplant recipients with BOS demonstrated downregulation of LKB1 (p=0.002), upregulation of Rab27A (p=0.006) and miR-155 (p=0.03) compared to stable biopsies. Using a murine lung transplant model of chronic rejection, we demonstrated correlation between development of lung fibrosis to downregulation of LKB1 (p=0.01), upregulation of vimentin (p=0.01) and α-SMA (p=0.01.
We identified a novel molecular mechanism in which exosomes released from transplanted lungs undergoing chronic rejection with inactivated tumor suppressor gene LKB1 results in EMT leading to the pathogenesis of CLAD following human lung transplantation.