Showing papers by "Bärbel Rohrer published in 2020"

Immunization Against Oxidized Elastin Exacerbates Structural and Functional Damage in Mouse Model of Smoke-Induced Ocular Injury.

Abstract: Purpose Age-related macular degeneration (AMD) is the leading cause of blindness in Western populations. While an overactive complement system has been linked to pathogenesis, mechanisms contributing to its activation are largely unknown. In aged and AMD eyes, loss of the elastin layer (EL) of Bruch's membrane (BrM) has been reported. Elastin antibodies are elevated in patients with AMD, the pathogenic significance of which is unclear. Here we assess the role of elastin antibodies using a mouse model of smoke-induced ocular pathology (SIOP), which similarly demonstrates EL loss. Methods C57BL/6J mice were immunized with elastin or elastin peptide oxidatively modified by cigarette smoke (ox-elastin). Mice were then exposed to cigarette smoke or air for 6 months. Visual function was assessed by optokinetic response, retinal morphology by spectral-domain optical coherence tomography and electron microscopy, and complement activation and antibody deposition by Western blot. Results Ox-elastin IgG and IgM antibodies were elevated in ox-elastin immunized mice following 6 months of smoke, whereas elastin immunization had a smaller effect. Ox-elastin immunization exacerbated smoke-induced vision loss, with thicker BrM and more damaged retinal pigment epithelium (RPE) mitochondria compared with mice immunized with elastin or nonimmunized controls. These changes were correlated with increased levels of IgM, IgG2, IgG3, and complement activation products in RPE/choroid. Conclusions These data demonstrate that SIOP mice generate elastin-specific antibodies and that immunization with ox-elastin exacerbates ocular pathology. Elastin antibodies represented complement fixing isotypes that, together with the increased presence of complement activation seen in immunized mice, suggest that elastin antibodies exert pathogenic effects through mediating complement activation.

Encapsulated Cell Technology for the Delivery of Biologics to the Mouse Eye.

Abstract: Many current therapeutics under development for diseases of the posterior pole of the eye are biologics. These drugs need to be administered frequently, typically via intravitreal injections. Encapsulated cells expressing the biologic of choice are becoming a tool for local protein production and release (e.g., via long-term drug delivery). In addition, encapsulation systems utilize permeable materials that allow diffusion of nutrients, waste, and therapeutic factors into and out of cells. This occurs while masking the cells from the host immune response, avoiding the need for suppression of the host immune system. This protocol describes the use of alginate as a polymer in microencapsulation coupled with the electrospray method as a microencapsulation technique. ARPE-19 cells, a spontaneously arising human RPE cell line, has been used in long-term cell therapy experiments due to its lifetime functionality, and it is used here for encapsulation and delivery of the capsules to mouse eyes. The manuscript summarizes the steps for cell microencapsulation, quality control, and ocular delivery.

Systemic Inflammation by Collagen-Induced Arthritis Affects the Progression of Age-Related Macular Degeneration Differently in Two Mouse Models of the Disease.

Abstract: Purpose Age-related macular degeneration (AMD) shares similar risk factors and inflammatory responses with rheumatoid arthritis (RA). Previously, we identified increased risk for dry AMD among patients with RA compared to control subjects, using retrospective data analysis. In this current study, we investigate the role of systemic inflammation triggered in a murine model of arthritis on choroidal neovascularization and retinal pigment epithelium (RPE) degeneration mouse models. Methods Collagen-induced arthritis (CIA) was induced in C57BL/6J mice prior to laser-induced choroidal neovascularization (CNV; wet AMD model) or sodium iodate-induced retinal degeneration (NaIO3; dry AMD model). CNV lesion size and retinal thickness were quantified by optical coherence photography (OCT), visual function was analyzed using optokinetic response and electroretinography, RPE morphology was examined by immunohistochemistry, and inflammatory gene expression was analyzed by quantitative PCR. Results CIA mice demonstrated decreased spatial acuity and contrast sensitivity, whereas no difference was observed in the RPE-generated c-wave. CNV lesion size was decreased in CIA mice. NaIO3 decreased c-wave amplitude, as well as retinal thickness, which was augmented by CIA. NaIO3 treatment resulted in loss of normal RPE hexagonal shape, which was further aggravated by CIA. Increased Cxcl9 expression was observed in the presence of CIA and CIA combined with AMD. Disease severity differences were observed between sexes. Conclusions Our data suggest systemic inflammation by CIA results in increased pathology in a dry AMD model, whereas it reduces lesions in a wet AMD model. These findings highlight the need for additional investigation into the role of secondary inflammation and sex-based differences on AMD.

The use of Matrigel combined with encapsulated cell technology to deliver a complement inhibitor in a mouse model of choroidal neovascularization.

Abstract: Purpose Risk for age-related macular degeneration (AMD), a slowly progressing, complex disease, is tied to an overactive complement system. Efforts are under way to develop an anticomplement-based treatment to be delivered locally or systemically. We developed an alternative pathway (AP) inhibitor fusion protein consisting of a complement receptor-2 fragment linked to the inhibitory domain of factor H (CR2-fH), which reduces the size of mouse choroidal neovascularization (CNV) when delivered locally or systemically. Specifically, we confirmed that ARPE-19 cells genetically engineered to produce CR2-fH reduce CNV lesion size when encapsulated and placed intravitreally. We extend this observation by delivering the encapsulated cells systemically in Matrigel. Methods ARPE-19 cells were generated to stably express CR2 or CR2-fH, microencapsulated using sodium alginate, and injected subcutaneously in Matrigel into 2-month-old C57BL/6J mice. Four weeks after implantation, CNV was induced using argon laser photocoagulation. Progression of CNV was analyzed using optical coherence tomography. Bioavailability of CR2-fH was evaluated in Matrigel plugs with immunohistochemistry, as well as in ocular tissue with dot blots. Efficacy as an AP inhibitor was confirmed with protein chemistry. Results An efficacious number of implanted capsules to reduce CNV was identified. Expression of the fusion protein systemically did not elicit an immune response. Bioavailability studies showed that CR2-fH was present in the RPE/choroid fractions of the treated mice, and reduced CNV-associated ocular complement activation. Conclusions These findings indicate that systemic production of the AP inhibitor CR2-fH can reduce CNV in the mouse model.

Motor Protein MYO1C is Critical for Photoreceptor Opsin Trafficking and Vision

Abstract: Unconventional myosins linked to deafness are also proposed to play a role in retinal physiology. However, their direct role in photoreceptor structure and function remains unclear. Herein, we demonstrate that systemic loss of the unconventional myosin MYO1C in mice affected opsin trafficking and photoreceptor survival, leading to vision loss. Electroretinogram analysis of Myo1c knockout (Myo1c-KO) mice showed a progressive loss of rod and cone function. Immunohistochemistry and binding assays demonstrated MYO1C localization to photoreceptor inner and outer segments (OS) and identified a direct interaction of rhodopsin with the MYO1C cargo domain. In Myo1c-KO retinas, rhodopsin mislocalized to rod inner segments and the cell bodies, while cone opsins in OS showed punctate staining. In aged mice, the histological and ultrastructural examination of Myo1c-KO retinas showed a progressively degenerating photoreceptor OS phenotype. These results demonstrate that MYO1C is critical for opsin trafficking to the photoreceptor OS and normal visual function.

Sex Related Differences in Retinal Pigment Epithelium and Retinal Disease

Abstract: Sex-related differences have been identified in various ophthalmic disorders. Sex hormone receptors, such as estrogen receptors, have been found throughout the eye including retina and retinal pigment epithelium (RPE), indicating the importance of hormone regulation in these tissues. In this chapter we will discuss sex differences within the eye and how they relate to retina and RPE health and function. As many autoimmune diseases occur more often in women, we will also examine autoimmune diseases and the secondary effects that may arise within the eye. In addition, the role estrogen plays in systemic inflammation may help in understanding the role of estrogen in the eye. By further understanding the differences between males and females in ocular health, we can provide more tailored treatments for disease and design preventative care aimed at regaining hormonal balance.