O-GlcNAc Transferase Regulates Angiogenesis in Idiopathic Pulmonary Arterial Hypertension

Abstract

Idiopathic pulmonary arterial hypertension (IPAH) is considered a vasculopathy characterized by elevated pulmonary vascular resistance due to vasoconstriction and/or lung remodeling such as plexiform lesions, the hallmark of the PAH, as well as cell proliferation and vascular and angiogenic dysfunction. The serine/threonine hydroxyl-linked N-Acetylglucosamine (O-GlcNAc) transferase (OGT) has been shown to drive pulmonary arterial smooth muscle cell (PASMC) proliferation in IPAH. OGT is a cellular nutrient sensor that is essential in maintaining proper cell function through the regulation of cell signaling, proliferation, and metabolism. The aim of this study was to determine the role of OGT and O-GlcNAc in vascular and angiogenic dysfunction in IPAH. Primary isolated human control and IPAH patient PASMCs and pulmonary arterial endothelial cells (PAECs) were grown in the presence or absence of OGT inhibitors and subjected to biochemical assessments in monolayer cultures and tube formation assays, in vitro vascular sprouting 3D spheroid co-culture models, and de novo vascularization models in NODSCID mice. We showed that knockdown of OGT resulted in reduced vascular endothelial growth factor (VEGF) expression in IPAH primary isolated vascular cells. In addition, specificity protein 1 (SP1), a known stimulator of VEGF expression, was shown to have higher O-GlcNAc levels in IPAH compared to control at physiological (5 mM) and high (25 mM) glucose concentrations, and knockdown resulted in decreased VEGF protein levels. Furthermore, human IPAH PAECs demonstrated a significantly higher degree of capillary tube-like structures and increased length compared to control PAECs. Addition of an OGT inhibitor, OSMI-1, significantly reduced the number of tube-like structures and tube length similar to control levels. Assessment of vascular sprouting from an in vitro 3D spheroid co-culture model using IPAH and control PAEC/PASMCs and an in vivo vascularization model using control and PAEC-embedded collagen implants demonstrated higher vascularization in IPAH compared to control. Blocking OGT activity in these experiments, however, altered the vascular sprouting and de novo vascularization in IPAH similar to control levels when compared to controls. Our findings in this report are the first to describe a role for the OGT/O-GlcNAc axis in modulating VEGF expression and vascularization in IPAH. These findings provide greater insight into the potential role that altered glucose uptake and metabolism may have on the angiogenic process and the development of plexiform lesions. Therefore, we believe that the OGT/O-GlcNAc axis may be a potential therapeutic target for treating the angiogenic dysregulation that is present in IPAH.

P53 in the impaired lungs

“…It also affects the proliferation of smooth muscle cells. The vascular wall becomes thicker, and plexiform lesions are formed [ 107 ].…”

Effects of FTMT Expression by Retinal Pigment Epithelial Cells on Features of Angiogenesis

“…The in vitro angiogenesis tube formation assay encompasses multiple features required for endothelial cells to form into vessel-like structures, including adhesion, migration, and tubule formation in a coordinated manner depending on the balance of pro-angiogenic and antiangiogenic factors [63]. This assay has been used to screen for inhibitors and to identify signaling mechanisms involved (for examples, see References [64][65][66]). We have focused mainly on VEGF and to a less extent on PEDF, as effectors of angiogenesis and angiogenesis inhibition, but a number of factors are involved to produce the end result of vessel formation.…”

Regulatory Effects of O-GlcNAcylation in Vascular Smooth Muscle Cells on Diabetic Vasculopathy

“… 68 69 70 In rat aortic smooth muscle cells, plasminogen activator inhibitor-1 (PAI-1), a well-known stimulator of VSMC migration (especially in the setting of chronic transforming growth factor [TGF]-β1 activation), is upregulated via Sp1-binding to the PAI-1 promoter region through the release of a transcriptional repressor from Sp1 complexes under high-glucose conditions. 71 Barnes et al 72 demonstrated that higher O-GlcNAcylation of Sp1 in pulmonary arterial smooth muscle cells from idiopathic pulmonary arterial hypertension facilitated cell migration via increased expression of vascular endothelial growth factor. It was also shown that the increased expression of TGF-β1 in endothelial cells cultured under high-glucose conditions was mediated by elevated O-GlcNAcylation of Sp1, 73 which may be implicated in VSMC proliferation through the synthesis of proteoglycans and extracellular matrix proteins.…”