Pulmonary Hypertension is a disease condition characterized by elevated pulmonary vascular resistance which is an outcome of excessive vascular remodeling in pulmonary arteries involving deregulated proliferation of cells in all three layers of the vessels; intima, media as well as adventitia. New therapeutic options targeting vascular remodeling process are being developed and some are curently under clinical trials. Current evidence strongly supports involvement of Notch signaling in vascular homeostasis and injury. Recently, the impact of Notch3 and its target gene, Hes5 on increased proliferation of PASMCs in PAH patients has been described. However, the role of other Notch receptors and ligands has not been studied. This led us to investigate the expression and potential contribution of Notch signaling pathway in pathogenesis of PH. Interestingly, we observed an increased expression of Notch1 in lungs and pulmonary arteries from patients with IPAH. In line with this finding, increased Notch1 immunoreactivity was detected in intima of pulmonary arteries in lung tissues from IPAH patients as compared to donors. Furthermore, similar upregulation was observed in lungs from Hypoxia+Su5416 rats as compared to healthy rats. We demonstrate for the first time an upregulation of Notch1 in pulmonary hypertension patients. Additionally, absence of this increase in other two well established experimental models depicting only medial hypertrophy phenotype, monocrotaline rat model and chronic hypoxia mice model, led us to hypothesize that Notch1 may contribute substantially to pathogenesis of plexiform lesions.In vasculature, Notch1 expression is observed in both smooth muscle cells and endothelial cells. We demonstrated a decrease in proliferation and increase in apoptosis of human pulmonary artery endothelial cells (hPAECs) with siRNA mediated Notch1 inhibition. In contrast, an increase in proliferation and survival of hPAECs was observed on constitutive Notch1 activation. These effects were associated with modulation of expression of cell cycle inhibitor, p21 and apoptosis inhibitor, Bcl-2. Interestingly, Notch1 inhibition or activation did not have any effect on proliferation of human pulmonary artery smooth muscle cells (hPASMCs), indicating, endothelial cell specific effect of Notch1 signaling in vasculature. The anti-proliferative and pro-apoptotic effect of Notch1 signaling inhibition, observed in our results, puts forward Notch inhibition as a therapeutic option for treatment of pulmonary hypertension. Presently, gamma secretase inhibitors (GSIs) are the only clinically available inhibitors targeting this pathway. GSIs inhibit the γ secretase enzyme, responsible for cleavage of Notch receptors. In order to examine the relevance of GSIs as a therapy in experimental PH, we used DBZ (a commercially available GSI) for our in vitro studies. Inhibition of Notch signaling by employing DBZ effectively reduced expression of Hes-1 (target gene of Notch signaling pathway) and attenuated proliferation of both, hPAECs and hPASMCs. Taken together, this study puts forward a role of Notch1 signaling in regulation of proliferation and apoptosis of endothelial cells which might contribute to formation of plexiform lesions observed in pulmonary hypertension. Nonetheless, our study needs to be further substantiated with in vivo studies using GSI in Hox+Su5416 rat model of PH.
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