Aging-regulated PNUTS maintains endothelial barrier function via SEMA3B suppression [PNUTS]

Age-related diseases pose great challenges to health care systems worldwide. During aging, endothelial senescence increases the risk for cardiovascular disease. Recently, it was described that Phosphatase 1 Nuclear Targeting Subunit (PNUTS) has a central role in cardiomyocyte aging and homeostasis. Here, we determined the role of PNUTS in endothelial cell aging. We confirmed that PNUTS is repressed in senescent endothelial cells (ECs). Moreover, PNUTS silencing elicits several of the hallmarks of endothelial aging: senescence, reduced angiogenesis and loss of barrier function. To validate our findings in vivo, we generated an endothelial-specific inducible PNUTS-deficient mouse line (Cdh5-CreERT2;PNUTSfl/fl), termed PNUTSEC-KO. Two weeks after PNUTS deletion, PNUTSEC-KO mice presented severe multiorgan failure and vascular leakage. Transcriptomic analysis of PNUTS-silenced HUVECs and lungs of PNUTSEC-KO mice revealed that the PNUTS-PP1 axis tightly regulates the expression of semaphorin 3B (SEMA3B). Indeed, silencing of SEMA3B completely restored barrier function after PNUTS loss-of-function. These results reveal a pivotal role for PNUTS in endothelial homeostasis through a SEMA3B downstream pathway that provides a potential target against the effects of aging in ECs. 3 mice in which Pnuts (Ppp1r10) was deleted in endothelial cells only (Cdh5-CreERT2; Pnuts fl/fl) by injection of Tamoxifen 12 days prior to termination of the experiment and 3 control mice that did receive tamoxifen but lacked the CreERT2 (Pnuts fl/fl) and therefore have a functional Pnuts (Ppp1r10) gene were used for this experiment. Mice were terminated and lungs were isolated. Then, RNA was isolated from total lung tissue.