General Capillary Endothelial Cells Undergo Reprogramming into Arterial Endothelial Cells in Pulmonary Hypertension through HIF-2α/Notch4 Pathway.

Pulmonary arterial hypertension (PAH) is characterized by a progressive increase of pulmonary vascular resistance and obliterative pulmonary vascular remodeling that result in right heart hypertrophy, failure, and premature death. The underlying mechanisms of loss of distal capillary endothelial cells (ECs) and obliterative vascular lesion formation remain unclear. Our recent single-cell RNA sequencing, spatial transcriptomics analysis, RNASCOPE, and immunostaining analysis showed that arterial ECs accumulation and loss of capillary ECs were evident in human PAH patients and pulmonary hypertension (PH) rodents. Pseudotime trajectory analysis of the single-cell RNA sequencing data suggest that lung capillary ECs transit to arterial ECs during the development of PH. Our study also identified CXCL12 as the marker for arterial ECs in PH. Capillary EC lineage tracing approach using capillary specific-Dre;Tdtomato reporter mice demonstrated that capillary ECs gave rise to arterial ECs during PH development. Genetic deletion of HIF-2a or pharmacological inhibition of Notch4 normalized the arterial programming in PH. In conclusion, our study demonstrates that capillary endothelium transits to arterial endothelium through the HIF-2a-Notch4 pathway during the development of PAH. Thus, targeting arterial EC transition might be a novel approach for treating PAH patients.

Single-cell and Spatial Transcriptomics Identified Fatty Acid-binding Proteins Controlling Endothelial Glycolytic and Arterial Programming in Pulmonary Hypertension.

Pulmonary arterial hypertension (PAH) is a devastating disease characterized by obliterative vascular remodeling and persistent increase of vascular resistance, leading to right heart failure and premature death. Understanding the cellular and molecular mechanisms will help develop novel therapeutic approaches for PAH patients. Single-cell RNA sequencing (scRNAseq) analysis found that both FABP4 and FABP5 were highly induced in endothelial cells (ECs) of Egln1Tie2Cre (CKO) mice, which was also observed in pulmonary arterial ECs (PAECs) from idiopathic PAH (IPAH) patients, and in whole lungs of pulmonary hypertension (PH) rats. Plasma levels of FABP4/5 were upregulated in IPAH patients and directly correlated with severity of hemodynamics and biochemical parameters using plasma proteome analysis. Genetic deletion of both Fabp4 and 5 in CKO mice (Egln1Tie2Cre/Fabp4-5-/- ,TKO) caused a reduction of right ventricular systolic pressure (RVSP) and RV hypertrophy, attenuated pulmonary vascular remodeling and prevented the right heart failure assessed by echocardiography, hemodynamic and histological analysis. Employing bulk RNA-seq and scRNA-seq, and spatial transcriptomic analysis, we showed that Fabp4/5 deletion also inhibited EC glycolysis and distal arterial programming, reduced ROS and HIF-2α expression in PH lungs. Thus, PH causes aberrant expression of FABP4/5 in pulmonary ECs which leads to enhanced ECs glycolysis and distal arterial programming, contributing to the accumulation of arterial ECs and vascular remodeling and exacerbating the disease.

Hypoxia Induced Changes of Exosome Cargo and Subsequent Biological Effects.

Exosomes are small extracellular vesicles that are secreted by almost all types of cells and exist in almost all extracellular spaces. As an important mediator of intercellular communication, exosomes encapsulate the miRNA, lncRNA, cirRNA, mRNA, cytokine, enzyme, lipid, and other components from the cytoplasm into its closed single membrane structure and transfer them to recipient units in an autocrine, paracrine, or endocrine manner. Hypoxia is a state of low oxygen tension and is involved in many pathological processes. Hypoxia influences the size, quantity, and expression of exosome cargos. Exosomes derived from hypoxic tumor cells transfer genetics, proteins, and lipids to the recipient units to exert pleiotropic effects. Different donor cells produce different cargo contents, target different recipient units and lead to different biological effects. Hypoxic exosomes derived from tumor cells uptaken by normoxic tumor cells lead to promoted proliferation, migration, and invasion; uptaken by extracellular space or liver lead to promoted metastasis; uptaken by endothelial cells lead to promoted angiogenesis; uptaken by immune cells lead to promoted macrophage polarization and changed tumor immune microenvironment. In addition to various types of tumors, hypoxic exosomes also participate in the development of diseases in the cardiovascular system, neuron system, respiratory system, hematology system, endocrine system, urinary system, reproduction system, and skeletomuscular system. Understanding the special characteristics of hypoxic exosomes provide new insight into elaborating the pathogenesis of hypoxia related disease. This review summarizes hypoxia induced cargo changes and the biological effects of hypoxic exosomes in tumors and non-malignant diseases in different systems.

Hypoxia Activates Notch4 via ERK/JNK/P38 MAPK Signaling Pathways to Promote Lung Adenocarcinoma Progression and Metastasis.

Hypoxia contributes to the progression and metastasis of lung adenocarcinoma (LUAD). However, the specific underlying molecular mechanisms have not been fully elucidated. Here we report that Notch4 is upregulated in lung tissue from lung cancer patients. Functionally, Hypoxia activates the expressions of Delta-like 4 and Notch4, resulting in the excessive proliferation and migration of LUAD cells as well as apoptotic resistance. Notch4 silencing reduced ERK, JNK, and P38 activation. Meanwhile, Notch4 overexpression enhanced ERK, JNK, and P38 activation in LUAD cells. Furthermore, Notch4 exerted pro-proliferation, anti-apoptosis and pro-migration effects on LUAD cells that were partly reversed by the inhibitors of ERK, JNK, and p38. The binding interaction between Notch4 and ERK/JNK/P38 were confirmed by the co-immunoprecipitation assay. In vivo study revealed that Notch4 played a key role in the growth and metastasis of LUAD using two xenograft models. This study demonstrates that hypoxia activates Notch4-ERK/JNK/P38 MAPK signaling pathways to promote LUAD cell progression and metastasis.