LKB1 Knockdown Triggered PDGFRβ Signaling Leading to Fibrosis and Development of Chronic Lung Allograft Dysfunction

Abstract

<h3>Purpose</h3> The platelet-derived growth factor and its receptor (PDGFRβ) signaling pathway has been recently shown to be involved in epithelial mesenchymal transition facilitating pulmonary fibrosis following lung transplantation (LTx). We have shown that LKB1 was down regulated in lung transplant recipients (LTxR) diagnosed with chronic lung allograft dysfunction (CLAD). Since LKB1, a protein kinase 1, can regulate fibrosis, the goal of this study was to investigate the role of LKB1 in the regulation of PDGFRβ leading to fibrosis resulting in CLAD following human LTx. <h3>Methods</h3> To investigate the role of LKB1 in a human airway epithelial cell line, BEAS-2B, we employed LKB1 siRNA to ablate LKB1. The effect of LKB1 knockdown on gene regulation was measured by NanoSTRING technology. Differential gene expression analysis was performed to identify genes regulated by LKB1. Downstream analysis was further confirmed knocking down PDGFRβ using siRNA and its effect was measured by western blot analysis. To validate our results, we also analyzed LKB1 and PDGFRβ expression by qRT-PCR in biopsy samples from LTxR diagnosed with bronchiolitis obliterans syndrome (BOS, n=12) and stable (n=12). <h3>Results</h3> Using NanoSTRING, we identified that LKB1 knockdown significantly induced PDGFRβ expression (2.5 fold) in BEAS-2B cells. Densitometry analysis showed that specific ablation of PDGFRβ in BEAS-2B significantly inhibited NF-κB (p=0.04), TNF-α (p=0.001), and phosphorylation of STAT3 (p=0.001). qRT-PCR analysis from biopsies LTxR with BOS samples confirmed that there is inverse correlation between PDGFRβ and LKB1 expression. PDGFRβ expression was increased (p=0.0001) whereas LKB1 expression was decreased (p=0.04) compared to stable biopsies. <h3>Conclusion</h3> With our cell line and biopsy samples assay, we have identified a novel molecular mechanism showing inverse correlation between LKB1 and PDGFRβ expression in human LTxR diagnosed with BOS. LKB1/PDGFRβ signaling pathways, can play a significant role in the pathogenesis of CLAD following human LTx which can be targeted developing therapeutics blocking LKB1/PDGFRβ signaling.