Investigation of the mechanisms of VEGF-mediated compensatory lung growth: the role of the VEGF heparin-binding domain

Abstract

Morbidity and mortality for neonates with congenital diaphragmatic hernia-associated pulmonary hypoplasia remains high. These patients may be deficient in vascular endothelial growth factor (VEGF). Our lab previously established that exogenous VEGF164 accelerates compensatory lung growth (CLG) after left pneumonectomy in a murine model. We aimed to further investigate VEGF-mediated CLG by examining the role of the heparin-binding domain (HBD). Eight-week-old, male, C57BL/6J mice underwent left pneumonectomy, followed by post-operative and daily intraperitoneal injections of equimolar VEGF164 or VEGF120, which lacks the HBD. Isovolumetric saline was used as a control. VEGF164 significantly increased lung volume, total lung capacity, and alveolarization, while VEGF120 did not. Treadmill exercise tolerance testing (TETT) demonstrated improved functional outcomes post-pneumonectomy with VEGF164 treatment. In lung protein analysis, VEGF treatment modulated downstream angiogenic signaling. Activation of epithelial growth factor receptor and pulmonary cell proliferation was also upregulated. Human microvascular lung endothelial cells (HMVEC-L) treated with VEGF demonstrated decreased potency of VEGFR2 activation with VEGF121 treatment compared to VEGF165 treatment. Taken together, these data indicate that the VEGF HBD contributes to angiogenic and proliferative signaling, is required for accelerated compensatory lung growth, and improves functional outcomes in a murine CLG model.

Direct thrombin inhibitors as alternatives to heparin to preserve lung growth and function in a murine model of compensatory lung growth

“…In this study, protein analysis of lung tissue (Fig. 6 ) confirmed decreased angiogenic signaling in heparinized mice as demonstrated by the 0.41-fold ( P = 0.02) reduction in the activation of VEGFR2; an important receptor implicated in angiogenesis 42 . Bivalirudin and argatroban did not significantly affect activation of VEGFR2.…”

Lipopolysaccharide-induced murine lung injury results in long-term pulmonary changes and downregulation of angiogenic pathways

“…5). The use of exercise tolerance testing as a functional measurement of lung capacity is well established [28][29][30] . These results suggest that isolated LPS exposure contributes to impaired long-term functional capacity.…”

“…Immunohistochemistry. Paraffin-embedded lung sections (N = 2-3 per group) from three time points (24 h, 7 days, 4 weeks) were assessed for type I collagen (COL1A1), type IV collagen (COL4A1), CD31 (endothelial marker), and proSPC (type II pneumocyte marker) expression using immunofluorescence as previously described 30,37 . After deparaffinization with xylene, sections were progressively rehydrated in decreasing concentrations of ethanol, followed by PBS.…”

Alveologenesis: What Governs Secondary Septa Formation

“…Endothelial cells, in turn, play an important role in stimulating AT2 proliferation during neo-alveolarization. VEGF accelerates compensatory lung growth after PNX in mice by increasing the number of alveolar units [211]. These changes can be mediated by both VEGFR2 and EGF-dependent mechanisms [212,213] and a paracrine mechanism through the upregulation of epithelial cell mitogen HB-EGF [214].…”