The role of retinal microglial cells (MCs) in age-related macular degeneration (AMD) is unclear. Here we demonstrated that all retinal MCs express CX3C chemokine receptor 1 (CX3CR1) and that homozygosity for the CX3CR1 M280 allele, which is associated with impaired cell migration, increases the risk of AMD. In humans with AMD, MCs accumulated in the subretinal space at sites of retinal degeneration and choroidal neovascularization (CNV). In CX3CR1-deficient mice, MCs accumulated subretinally with age and albino background and after laser impact preceding retinal degeneration. Raising the albino mice in the dark prevented both events. The appearance of lipid-bloated subretinal MCs was drusen-like on funduscopy of senescent mice, and CX3CR1-dependent MC accumulation was associated with an exacerbation of experimental CNV. These results show that CX3CR1-dependent accumulation of subretinal MCs evokes cardinal features of AMD. These findings reveal what we believe to be a novel pathogenic process with important implications for the development of new therapies for AMD.
Christophe Combadière, Charles Feumi, William Raoul, Nicole Keller, Mathieu Rodéro, Adeline Pézard, Sophie Lavalette, Marianne Houssier, Laurent Jonet, Emilie Picard, Patrice Debré, Mirna Sirinyan, Philippe Deterre, Tania Ferroukhi, Salomon-Yves Cohen, Dominique Chauvaud, Jean-Claude Jeanny, Sylvain Chemtob, Francine Behar-Cohen, Florian Sennlaub
Sphingosine 1–phosphate (S1P), a multifunctional lipid mediator that signals via the S1P family of G protein–coupled receptors (S1PR), regulates vascular maturation, permeability, and angiogenesis. In this study, we explored the role of S1P 2 receptor (S1P2R) in normal vascularization and hypoxia-triggered pathological angiogenesis of the mouse retina. S1P2R is strongly induced in ECs during hypoxic stress. When neonatal mice were subjected to ischemia-driven retinopathy, pathologic neovascularization in the vitreous chamber was suppressed in S1p2–/– mice concomitant with reduction in endothelial gaps and inflammatory cell infiltration. In addition, EC patterning and normal revascularization into the avascular zones of the retina were augmented. Reduced expression of the proinflammatory enzyme cyclooxygenase-2 (COX-2) and increased expression of eNOS were observed in the S1p2–/– mouse retina. S1P2R activation in ECs induced COX-2 expression and suppressed the expression of eNOS. These data identify the S1P2R-driven inflammatory process as an important molecular event in pathological retinal angiogenesis. We propose that antagonism of the S1P2R may be a novel therapeutic approach for the prevention and/or treatment of pathologic ocular neovascularization.
Athanasia Skoura, Teresa Sanchez, Kevin Claffey, Suzanne M. Mandala, Richard L. Proia, Timothy Hla
Glaucoma, a progressive optic neuropathy due to retinal ganglion cell (RGC) degeneration, is one of the leading causes of irreversible blindness. Although glaucoma is often associated with elevated intraocular pressure (IOP), IOP elevation is not detected in a significant subset of glaucomas, such as normal tension glaucoma (NTG). Moreover, in some glaucoma patients, significant IOP reduction does not prevent progression of the disease. Thus, understanding IOP-independent mechanisms of RGC loss is important. Here, we show that mice deficient in the glutamate transporters GLAST or EAAC1 demonstrate spontaneous RGC and optic nerve degeneration without elevated IOP. In GLAST-deficient mice, the glutathione level in Müller glia was decreased; administration of glutamate receptor blocker prevented RGC loss. In EAAC1-deficient mice, RGCs were more vulnerable to oxidative stress. These findings suggest that glutamate transporters are necessary both to prevent excitotoxic retinal damage and to synthesize glutathione, a major cellular antioxidant and tripeptide of glutamate, cysteine, and glycine. We believe these mice are the first animal models of NTG that offer a powerful system for investigating mechanisms of neurodegeneration in NTG and developing therapies directed at IOP-independent mechanisms of RGC loss.
Takayuki Harada, Chikako Harada, Kazuaki Nakamura, Hun-Meng A. Quah, Akinori Okumura, Kazuhiko Namekata, Tadashiro Saeki, Makoto Aihara, Hiroshi Yoshida, Akira Mitani, Kohichi Tanaka
Protein misfolding and aggregation are thought to underlie the pathogenesis of many amyloid diseases, such as Alzheimer and Parkinson diseases, whereby a stepwise protein misfolding process begins with the conversion of soluble protein monomers to prefibrillar oligomers and progresses to the formation of insoluble amyloid fibrils. Drusen are extracellular deposits found in aging eyes and in eyes afflicted with age-related macular degeneration (AMD). Recent characterizations of drusen have revealed protein components that are shared with amyloid deposits. However, characteristic amyloid fibrils have thus far not been identified in drusen. In this study, we tested the hypothesis that nonfibrillar oligomers may be a common link in amyloid diseases. Oligomers consisting of distinct amyloidogenic proteins and peptides can be detected by a recently developed antibody that is thought to recognize a common structure. Notably, oligomers exhibit cellular toxicity, which suggests that they play a role in the pathogenesis of neurodegenerative diseases. Through use of the anti-oligomer antibody, we came to observe the presence of nonfibrillar, toxic oligomers in drusen. Conversely, no reactivity was observed in age-matched control eyes without drusen. These results suggest that amyloid oligomers may be involved in drusen biogenesis and that similar protein misfolding processes may occur in AMD and amyloid diseases.
Volker Luibl, Jose M. Isas, Rakez Kayed, Charles G. Glabe, Ralf Langen, Jeannie Chen
In response to hypoxia, hypoxia-inducible factors act as the primary proangiogenic triggers by regulating transcription levels of target genes, including VEGF. However, little is known about the specific factors that control other components of the angiogenic process, particularly formation of matrix scaffolds that promote adhesion and migration of endothelial cells. We show that in the postnatal mouse retina, the orphan nuclear receptor tailless (Tlx) is strongly expressed in the proangiogenic astrocytes, which secrete VEGF and fibronectin. Tlx expression by retinal astrocytes is controlled by oxygen concentration and rapidly downregulated upon contact with blood vessels. In mice null for Tlx, retinal astrocytes maintain VEGF expression; however, the extracellular assembly of fibronectin matrices by astrocytes is severely impaired, leading to defective scaffold formation and a complete failure of normal retinal vascular development. This work identifies Tlx as an essential component of the molecular network involved in the hypoxia-inducible proangiogenic switch in retinal astrocytes.
Akiyoshi Uemura, Sentaro Kusuhara, Stanley J. Wiegand, Ruth T. Yu, Shin-Ichi Nishikawa
VEGF-A promotes angiogenesis in many tissues. Here we report that choroidal neovascularization (CNV) incited by injury was increased by excess VEGF-A before injury but was suppressed by VEGF-A after injury. This unorthodox antiangiogenic effect was mediated via VEGFR-1 activation and VEGFR-2 deactivation, the latter via Src homology domain 2–containing (SH2-containing) tyrosine phosphatase-1 (SHP-1). The VEGFR-1–specific ligand placental growth factor-1 (PlGF-1), but not VEGF-E, which selectively binds VEGFR-2, mimicked these responses. Excess VEGF-A increased CNV before injury because VEGFR-1 activation was silenced by secreted protein, acidic and rich in cysteine (SPARC). The transient decline of SPARC after injury revealed a temporal window in which VEGF-A signaling was routed principally through VEGFR-1. These observations indicate that therapeutic design of VEGF-A inhibition should include consideration of the level and activity of SPARC.
Miho Nozaki, Eiji Sakurai, Brian J. Raisler, Judit Z. Baffi, Jassir Witta, Yuichiro Ogura, Rolf A. Brekken, E. Helene Sage, Balamurali K. Ambati, Jayakrishna Ambati
Choroideremia (CHM) is an X-linked degeneration of the retinal pigment epithelium (RPE), photoreceptors, and choroid, caused by loss of function of the CHM/REP1 gene. REP1 is involved in lipid modification (prenylation) of Rab GTPases, key regulators of intracellular vesicular transport and organelle dynamics. To study the pathogenesis of CHM and to develop a model for assessing gene therapy, we have created a conditional mouse knockout of the Chm gene. Heterozygous-null females exhibit characteristic hallmarks of CHM: progressive degeneration of the photoreceptors, patchy depigmentation of the RPE, and Rab prenylation defects. Using tamoxifen-inducible and tissue-specific Cre expression in combination with floxed Chm alleles, we show that CHM pathogenesis involves independently triggered degeneration of photoreceptors and the RPE, associated with different subsets of defective Rabs.
Tanya Tolmachova, Ross Anders, Magnus Abrink, Laurence Bugeon, Margaret J. Dallman, Clare E. Futter, José S. Ramalho, Felix Tonagel, Naoyuki Tanimoto, Mathias W. Seeliger, Clare Huxley, Miguel C. Seabra