RIP1 kinase mediates angiogenesis by modulating macrophages in experimental neovascularization

Abstract

Inflammation plays an important role in pathological angiogenesis. Receptor-interacting protein 1 (RIP1) is highly expressed in inflammatory cells and is known to play an important role in the regulation of apoptosis, necroptosis, and inflammation; however, a comprehensive description of its role in angiogenesis remains elusive. Here, we show that RIP1 is abundantly expressed in infiltrating macrophages during angiogenesis, and genetic or pharmacological inhibition of RIP1 kinase activity using kinase-inactive RIP1K45A/K45A mice or necrostatin-1 attenuates angiogenesis in laser-induced choroidal neovascularization, Matrigel plug angiogenesis, and alkali injury-induced corneal neovascularization in mice. The inhibitory effect on angiogenesis is mediated by caspase activation through a kinase-independent function of RIP1 and RIP3. Mechanistically, infiltrating macrophages are the key target of RIP1 kinase inhibition to attenuate pathological angiogenesis. Inhibition of RIP1 kinase activity is associated with caspase activation in infiltrating macrophages and decreased expression of proangiogenic M2-like markers but not M1-like markers. Similarly, in vitro, catalytic inhibition of RIP1 down-regulates the expression of M2-like markers in interleukin-4–activated bone marrow-derived macrophages, and this effect is blocked by simultaneous caspase inhibition. Collectively, these results demonstrate a nonnecrotic function of RIP1 kinase activity and suggest that RIP1-mediated modulation of macrophage activation may be a therapeutic target of pathological angiogenesis.

Pharmacological Inhibition of Caspase-8 Suppresses Inflammation-Induced Angiogenesis in the Cornea

“…Recently, Ueta et al demonstrated that in pathological angiogenesis mouse model, RIPK1 abundantly expresses in infiltrating macrophages and inhibition of RIPK1 alleviates angiogenesis. Furthermore, caspase inhibition can augment RIPK1 activation, which cause aggravation of pathological angiogenesis [54]. Similarly, during the study, we found that 100 µM Z-IETD-fmk treatment decreases the activity of caspase-8 but aggravates CNV comparing with the 40 µM group.…”

The P300/XBP1s/Herpud1 axis promotes macrophage M2 polarization and the development of choroidal neovascularization

“… 23 Therefore, M2 macrophage infiltration in CNV lesions is an emerging target for CNV therapy. 24 , 25 In our study, activation of the p300/XBP1s/Herpud1 axis in infiltrating M2 macrophages promoted the proliferation, migration and tube formation of choroidal endothelial cells. Furthermore, blockade of the p300/XBP1s/Herpud1 axis inhibited the M2 polarization of macrophages and alleviated the development of CNV, in consistent with previous studies.…”

“…Infiltrating macrophages interact with choroidal endothelial cells through Notch receptor 1 (Notch1) signalling during retinal sprouting angiogenesis 23 . Therefore, M2 macrophage infiltration in CNV lesions is an emerging target for CNV therapy 24,25 . In our study, activation of the p300/XBP1s/Herpud1 axis in infiltrating M2 macrophages promoted the proliferation, migration and tube formation of choroidal endothelial cells.…”

Acadesine suppresses TNF-α induced complement component 3 (C3), in retinal pigment epithelial (RPE) cells

“…Washed 4 times for 3 minutes each in TBST and then incubated with secondary HRP-conjugated antibodies accordingly for 25 minutes at room temperature before final washing step, 4 times for 3 minutes each. Membranes were developed using chemiluminescent substrate (ECL Select western blotting detection reagent, RPN 2235, GE Healthcare Life Sciences, USA) [ 87 ] and images captured by using the ChemiDoc imaging system (Bio-Rad, Hercules, CA, USA). For full list of the antibodies used see S1 Table .…”