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1.
J Neuroinflammation ; 21(1): 56, 2024 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-38388518

RESUMO

Inherited, age-related, and acute retinal diseases are often exacerbated by an aberrant or excessive activity of the complement system. Consequently, cells not directly affected by an acute event or genetic variants may degenerate, resulting in enhanced visual impairment. The therapeutic potential of supplementation of complement factor H (FH), a key regulator of the complement cascade, is therefore particularly promising in the context of retinal diseases caused by complement activation. In this study, we engineered adeno-associated viruses (AAVs) containing sequences of two truncated human FH variants. The expression of these variants was regulated by the glial fibrillary acidic protein (GFAP) promoter, which is selectively active in gliotic Müller cells. Both FH variants consisted of FH domains 19-20, which were connected to domains 1-4 and 1-7, respectively, by a polyglycine linker. These AAVs were intravitreally injected following ischemic injury of C57BL/6J mouse retinas. We observed transgene expression in gliotic Müller cells and to some extent in astrocytes. The expression correlated directly with damage severity. Interventions resulted in decreased complement activation, accelerated normalization of microglia activity and morphological improvements. Reduced levels of C3 transcripts and C3d protein in conjunction with higher transcript levels of inhibitory regulators like Cfi and Cfh, hinted at attenuated complement activity. This study demonstrates the great potential of complement regulatory gene addition therapy. With further in vivo testing it could be applied to treat a wide range of retinal diseases where no causative therapies are available.


Assuntos
Gliose , Doenças Retinianas , Camundongos , Animais , Humanos , Gliose/metabolismo , Fator H do Complemento/genética , Camundongos Endogâmicos C57BL , Retina/metabolismo
2.
Front Immunol ; 13: 895519, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35784369

RESUMO

The cellular events that dictate the initiation of the complement pathway in ocular degeneration, such as age-related macular degeneration (AMD), is poorly understood. Using gene expression analysis (single cell and bulk), mass spectrometry, and immunohistochemistry, we dissected the role of multiple retinal and choroidal cell types in determining the complement homeostasis. Our scRNA-seq data show that the cellular response to early AMD is more robust in the choroid, particularly in fibroblasts, pericytes and endothelial cells. In late AMD, complement changes were more prominent in the retina especially with the expression of the classical pathway initiators. Notably, we found a spatial preference for these differences. Overall, this study provides insights into the heterogeneity of cellular responses for complement expression and the cooperation of neighboring cells to complete the pathway in healthy and AMD eyes. Further, our findings provide new cellular targets for therapies directed at complement.


Assuntos
Células Endoteliais , Degeneração Macular , Corioide , Proteínas do Sistema Complemento , Humanos , Degeneração Macular/genética , Retina
3.
Int J Mol Sci ; 21(22)2020 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-33187113

RESUMO

Stargardt macular degeneration is an inherited retinal disease caused by mutations in the ATP-binding cassette subfamily A member 4 (ABCA4) gene. Here, we characterized the complement expression profile in ABCA4-/- retinae and aligned these findings with morphological markers of retinal degeneration. We found an enhanced retinal pigment epithelium (RPE) autofluorescence, cell loss in the inner retina of ABCA4-/- mice and demonstrated age-related differences in complement expression in various retinal cell types irrespective of the genotype. However, 24-week-old ABCA4-/- mice expressed more c3 in the RPE and fewer cfi transcripts in the microglia compared to controls. At the protein level, the decrease of complement inhibitors (complement factor I, CFI) in retinae, as well as an increased C3b/C3 ratio in the RPE/choroid and retinae of ABCA4-/-, mice was confirmed. We showed a corresponding increase of the C3d/C3 ratio in the serum of ABCA4-/- mice, while no changes were observed for CFI. Our findings suggest an overactive complement cascade in the ABCA4-/- retinae that possibly contributes to pathological alterations, including microglial activation and neurodegeneration. Overall, this underpins the importance of well-balanced complement homeostasis to maintain retinal integrity.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas do Sistema Complemento/metabolismo , Doença de Stargardt/metabolismo , Animais , Corioide/metabolismo , Ativação do Complemento/fisiologia , Modelos Animais de Doenças , Feminino , Degeneração Macular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microglia/metabolismo , Retina/metabolismo , Degeneração Retiniana/metabolismo , Epitélio Pigmentado da Retina/metabolismo
4.
Cell Rep ; 29(9): 2835-2848.e4, 2019 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-31775049

RESUMO

Complement dysregulation is a feature of many retinal diseases, yet mechanistic understanding at the cellular level is limited. Given this knowledge gap about which retinal cells express complement, we performed single-cell RNA sequencing on ∼92,000 mouse retinal cells and validated our results in five major purified retinal cell types. We found evidence for a distributed cell-type-specific complement expression across 11 cell types. Notably, Müller cells are the major contributor of complement activators c1s, c3, c4, and cfb. Retinal pigment epithelium (RPE) mainly expresses cfh and the terminal complement components, whereas cfi and cfp transcripts are most abundant in neurons. Aging enhances c1s, cfb, cfp, and cfi expression, while cfh expression decreases. Transient retinal ischemia increases complement expression in microglia, Müller cells, and RPE. In summary, we report a unique complement expression signature for murine retinal cell types suggesting a well-orchestrated regulation of local complement expression in the retinal microenvironment.


Assuntos
Proteínas do Sistema Complemento/metabolismo , Retina/fisiopatologia , Animais , Humanos , Camundongos
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