bioRxiv2024
BackgroundPulmonary arterial hypertension (PAH) is a severe lung condition with unmet clinical needs, marked by endothelial damage, excessive repair, and arterial narrowing, though mechanisms remain unclear. MethodsThis study investigates Kruppel-like transcription factor 6 (KLF6), known for its role in tissue injury response and cancer onset, in PAH through functional and expression analyses in human pulmonary artery endothelial cells (HPAECs) and human and rodent PAH lung tissues. FindingsKLF6 expression increased in early experimental PAH in response to hypoxia and inflammation, while the expression of endothelial homeostasis regulators KLF2 and KLF4, previously linked to PAH, decreased. KLF6 overexpression enhanced pulmonary endothelial survival and angiogenesis through broad transcriptomic remodelling, including changes in genes governing endothelial homeostasis and arterial identity (e.g., SOX17, ERG, BMPR2) and promoted human pulmonary artery smooth muscle cells (HPASMCs) proliferation, which was inhibited by bosentan and imatinib. KLF6 functional and transcriptomic responses differed from those of KLF2 and KLF4. Comparative analysis of RNA-seq PAH databases and spatial transcriptomic analysis of human idiopathic PAH (IPAH) tissues highlighted strong association of KLF6 with vascular remodelling, especially with the formation of angioproliferative (plexiform) lesions. High KLF6 expression was observed in IPAH vascular endothelium and IPAH blood-derived endothelial progenitor cells. Single nucleus RNA-seq in PAH associated with Alveolar Capillary Dysplasia confirmed disease-related elevated KLF6 expression in arterial endothelial cells. InterpretationAccumulation and reorganization of KLF6+ endothelial cells characterize human PAH. KLF6 drives endothelial repair and an apoptosis-resistant, angioproliferative endothelial phenotype. Targeting KLF6 could be a novel therapeutic approach for PAH. RESEARCH IN CONTEXTO_ST_ABSEvidence before this studyC_ST_ABSPulmonary arterial hypertension (PAH) is a progressive and life-shortening lung disease with no cure. In PAH development, endothelial damage is believed to initiate an abnormal repair process, leading to extensive vascular remodelling and the formation of complex angio-proliferative (plexiform) lesions. We conducted a systematic search of the PubMed database to identify transcription factors potentially involved in driving endothelial repair and promoting an apoptosis-resistant, angio-proliferative vascular phenotype. Previous research has linked the loss of endothelial homeostasis in PAH to the inhibition of transcription factors KLF2 and KLF4. While KLF6 is known to play a vital role in vascular development and supports endothelial repair, its specific role in PAH remains unexplored. Added value of this studyThis study is the first to establish a connection between KLF6 and PAH pathogenesis. Our findings reveal that KLF6 activation is a key feature of an apoptosis-resistant, angio-proliferative endothelial phenotype characteristic of human PAH-associated plexogenic arteriopathy. Furthermore, we identify both overlapping and unique activation patterns and transcriptional programs regulated by KLF2, KLF4, and KLF6 in lung endothelial cells, highlighting KLF6s unique role in driving endothelial dysfunction in PAH. Implications of all the available evidenceTargeting KLF6 offers a promising therapeutic strategy to counteract excessive vascular repair and prevent the vascular remodelling in PAH. FUNDINGPhD studentship from the University of Hafr Al Batin, KSA, and the Saudi Cultural Bureau in London (UKSACB) (Rehab Alharbi). Spatial transcriptomic reagents were funded by the British Heart Foundation Centre of Research Excellence Award and Senior BHF Fellowship FS/18/52/33808 (Allan Lawrie). Human samples used in this research project were obtained from the Imperial College Healthcare Tissue Bank (ICHTB) supported by the National Institute for Health Research (NIHR) Biomedical Research Centre based at Imperial College Healthcare NHS Trust and Imperial College London.