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In the adult bone marrow (BM), endothelial cells (ECs) are an integral component of the hematopoietic stem cell (HSC)-supportive niche, which modulates HSC activity by producing secreted and membrane-bound paracrine signals. Within the BM, distinct vascular arteriole, transitional, and sinusoidal EC subtypes display unique paracrine expression profiles and create anatomically-discrete microenvironments. However, the relative contributions of vascular endothelial subtypes in supporting hematopoiesis is unclear. Moreover, constitutive expression and off-target activity of currently available endothelial-specific and endothelial-subtype-specific murine cre lines potentially confound data analysis and interpretation. To address this, we describe two tamoxifen-inducible cre-expressing lines, Vegfr3-creERT2 and Cx40-creERT2, that efficiently label sinusoidal/transitional and arteriole endothelium respectively in adult marrow, without off-target activity in hematopoietic or perivascular cells. Utilizing an established mouse model in which cre-dependent recombination constitutively-activates MAPK signaling within adult endothelium, we identify arteriole ECs as the driver of MAPK-mediated hematopoietic dysfunction. These results define complementary tamoxifen-inducible creERT2-expressing mouse lines that label functionally-discrete and non-overlapping sinusoidal/transitional and arteriole EC populations in the adult BM, providing a robust toolset to investigate the differential contributions of vascular subtypes in maintaining hematopoietic homeostasis. [ABSTRACT FROM AUTHOR]

SLIT2 is a secreted polypeptide that guides migration of cells expressing Roundabout 1 and 2 (ROBO1 and ROBO2) receptors. Herein, we investigated SLIT2/ROBO signaling effects in gliomas. In patients with glioblastoma (GBM), SLIT2 expression increased with malignant progression and correlated with poor survival and immunosuppression. Knockdown of SLIT2 in mouse glioma cells and patient-derived GBM xenografts reduced tumor growth and rendered tumors sensitive to immunotherapy. Tumor cell SLIT2 knockdown inhibited macrophage invasion and promoted a cytotoxic gene expression profile, which improved tumor vessel function and enhanced efficacy of chemotherapy and immunotherapy. Mechanistically, SLIT2 promoted microglia/macrophage chemotaxis and tumor-supportive polarization via ROBO1- and ROBO2-mediated PI3K-[gamma] activation. Macrophage Robo1 and Robo2 deletion and systemic SLIT2 trap delivery mimicked SLIT2 knockdown effects on tumor growth and the tumor microenvironment (TME), revealing SLIT2 signaling through macrophage ROBOs as a potentially novel regulator of the GBM microenvironment and immunotherapeutic target for brain tumors., Introduction Malignant gliomas are the most common primary brain tumors (1, 2). Among those, glioblastoma (GBM; WHO grade IV glioma) is the most frequent and aggressive tumor, accounting for more [...]

Ocular neovascular diseases are a leading cause of blindness. Vascular endothelial growth factor (VEGF) blockade improves vision, but not all individuals respond to anti-VEGF treatment, making additional means to prevent neovascularization necessary. Slit-family proteins (Slits) are ligands of Roundabout (Robo) receptors that repel developing axons in the nervous system. Robo1 expression is altered in ocular neovascular diseases, and previous in vitro studies have reported both pro- and anti-angiogenic effects of Slits. However, genetic evidence supporting a role for Slits in ocular neovascularization is lacking. Here we generated conditional knockout mice deficient in various Slit and Robo proteins and found that Slit2 potently and selectively promoted angiogenesis via Robo1 and Robo2 in mouse postnatal retina and in a model of ocular neovascular disease. Mechanistically, Slit2 acting through Robo1 and Robo2 promoted the migration of endothelial cells. These receptors are required for both Slit2- and VEGF-induced Rac1 activation and lamellipodia formation. Thus, Slit2 blockade could potentially be used therapeutically to inhibit angiogenesis in individuals with ocular neovascular disease., Ocular neovascular diseases are the most common cause of vision loss, affecting millions of people in the industrialized world. Such diseases include the wet form of age-related macular degeneration (AMD), [...]

Autonomic sympathetic nerves innervate peripheral resistance arteries, thereby regulating vascular tone and controlling blood supply to organs. Despite the fundamental importance of blood flow control, how sympathetic arterial innervation develops remains largely unknown. Here, we identified the axon guidance cue netrin-1 as an essential factor required for development of arterial innervation in mice. Netrin-1 was produced by arterial smooth muscle cells (SMCs) at the onset of innervation, and arterial innervation required the interaction of netrin-1 with its receptor, deleted in colorectal cancer (DCC), on sympathetic growth cones. Function-blocking approaches, including cell type-specific deletion of the genes encoding Ntnl in SMCs and Dee in sympathetic neurons, led to severe and selective reduction of sympathetic innervation and to defective vasoconstriction in resistance arteries. These findings indicate that netrin-1 and DCC are critical for the control of arterial innervation and blood flow regulation in peripheral organs., Introduction Arterial blood vessels and peripheral nerves both form highly branched networks that ramify together throughout the organism (1, 2). To orchestrate the formation of their exquisitely wired networks, nerves [...]