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<h3>Background</h3> Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial lung disease. A multi-centre genome-wide meta-analysis involving participants from the US, UK, and Spain identified Protein Kinase N2 (PKN2) as significantly associated with the decline in forced vital capacity in IPF patients. Although PKN2 has been linked to the proliferation of mouse embryonic fibroblasts, its role in IPF remains largely unexplored. We hypothesise that PKN2 contributes to the pro-fibrotic activity of fibroblasts, thereby promoting fibrosis. We aim to investigate how PKN2 knockdown influences TGFβ<sub>1</sub>-mediated pro-fibrotic processes in human lung myofibroblasts (HLMFs), to better understand its impact on fibroblast function and lung function decline. <h3>Methods</h3> PKN2 expression in lung tissue and in total lung cells was examined using qRT-PCR and Human Gene 2.1 ST genome-wide microarray data. The effect of PKN2 knockdown (siRNA) on HLMFs was examined by scratch wound assays, RNA-sequencing (n=5) and liquid chromatography-mass spectrometry (LC-MS) proteomics (n=3). <h3>Results</h3> PKN2 mRNA is expressed in bulk lung tissue and specific cell types such as epithelial cells, airway smooth muscle cells and HLMF cells, with affymetrix gene expression data showing significant downregulation of PKN2 expression in IPF fibroblasts versus non-IPF fibroblasts in airway (p=0.0044) and parenchymal lung fibroblasts (p=0.0011). Successful downregulation of PKN2 was achieved using siRNA (n=7, p=0.0402). RNA-sequencing shows that PKN2 knockdown in HLMFs promotes a pro-fibrotic genetic profile, with upregulation of collagens and extracellular matrix (ECM) components at gene level. Knockdown was associated with dysregulation of ECM, adhesion and migration gene pathways. LC-MS analysis confirms successful downregulation of PKN2 at protein level. PKN2 knockdown leads to dysregulation of fibrosis associated proteins and pathways including integrin-signalling and regulation of WNT signalling pathways. Functional assays showed that PKN2 knockdown does not impact HLMF proliferation (n=3, p=0.7439). PKN2 knockdown accelerates HLMF wound healing 24-hours post-injury (n=4, p=0.0061). <h3>Conclusion</h3> Our findings confirm PKN2 expression in lung and IPF-associated cells, with lower expression in IPF and upon TGFβ<sub>1</sub> stimulation. Knockdown of PKN2 enhances pro-fibrotic processes, including wound healing and disrupts the expression of fibrosis associated genes, proteins and pathways. Collectively suggesting that PKN2 plays a role in protecting against fibrosis within the lungs.