Abstract
BackgroundThe role of the alternative complement pathway and its mediation by retinal microglia and macrophages, is well-established in the pathogenesis of Age-Related Macular Degeneration (AMD). However, the contribution of the classical complement pathway towards the progression of retinal degenerations is not fully understood, including the role of complement component 1q (C1q) as a critical activator molecule of the classical pathway. Here, we investigated the contribution of C1q to progressive photoreceptor loss and neuroinflammation in retinal degenerations.MethodsWild-type (WT), C1qa knockout (C1qa−/−) and mice treated with a C1q inhibitor (ANX-M1; Annexon Biosciences), were exposed to photo-oxidative damage (PD) and were observed for progressive lesion development. Retinal function was assessed by electroretinography, followed by histological analyses to assess photoreceptor degeneration. Retinal inflammation was investigated through complement activation, macrophage recruitment and inflammasome expression using western blotting, qPCR and immunofluorescence. C1q was localised in human AMD donor retinas using immunohistochemistry.ResultsPD mice had increased levels of C1qa which correlated with increasing photoreceptor cell death and macrophage recruitment. C1qa−/− mice did not show any differences in photoreceptor loss or inflammation at 7 days compared to WT, however at 14 days after the onset of damage, C1qa−/− retinas displayed less photoreceptor cell death, reduced microglia/macrophage recruitment to the photoreceptor lesion, and higher visual function. C1qa−/− mice displayed reduced inflammasome and IL-1β expression in microglia and macrophages in the degenerating retina. Retinal neutralisation of C1q, using an intravitreally-delivered anti-C1q antibody, reduced the progression of retinal degeneration following PD, while systemic delivery had no effect. Finally, retinal C1q was found to be expressed by subretinal microglia/macrophages located in the outer retina of early AMD donor eyes, and in mouse PD retinas.ConclusionsOur data implicate subretinal macrophages, C1q and the classical pathway in progressive retinal degeneration. We demonstrate a role of local C1q produced by microglia/macrophages as an instigator of inflammasome activation and inflammation. Crucially, we have shown that retinal C1q neutralisation during disease progression may slow retinal atrophy, providing a novel strategy for the treatment of complement-mediated retinal degenerations including AMD.
Highlights
The role of the alternative complement pathway and its mediation by retinal microglia and macrophages, is well-established in the pathogenesis of Age-Related Macular Degeneration (AMD)
Temporal expression of retinal Complement component 1q (C1qa) correlated with IBA1+ microglia/macrophage recruitment to the Outer nuclear layer (ONL) and subretinal space of the outer retina, peaking at 5 days photo-oxidative damage (Fig. 1g-l), with significance detected at both 5 and 7 days for both C1qa expression and IBA1+ cell recruitment (P < 0.05, Fig. 1g-l)
Double-labelling of Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) and IBA1 demonstrated that IBA1+ macrophages were in close proximity and direct contact with multiple TUNEL+ apoptotic cells in the subretinal space and ONL at 7 days of photo-oxidative damage (Fig. 1n)
Summary
The role of the alternative complement pathway and its mediation by retinal microglia and macrophages, is well-established in the pathogenesis of Age-Related Macular Degeneration (AMD). C1q binding promotes the proteolysis of C4, C2, C3 and the downstream activation of the complement cascade, propagating the effector functions of complement, including the lysis of target cells [1] In neurodegenerative diseases such as Age-Related Macular Degeneration (AMD), complement dysregulation as a consequence of environmental pressures and/ or gene mutations, is known to contribute to disease progression [5,6,7]. Histopathological investigations show a range of complement components and factors from all pathways present in drusen of AMD donor eyes (reviewed in [7]) These drusen components are associated with the accrual of activated microglia and infiltrating blood-borne macrophages in the outer retina, which is a well-established feature of AMD pathogenesis [10]
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