Abstract:
Human inherited retinal degenerative disorders exhibit a large genetic and phenotypic heterogeneity. Retinitis Pigmentosa (RP), one of the most severe forms, denotes a group of hereditary disorders of the rod system that cause progressive retinal degeneration and may eventually lead to blindness. Achromatopsia is characterized by a total loss of function of all cone photoreceptors in the retina, leading to a severe visual impairment. So far, no cure or scientifically proven symptomatic treatment has been found for neither RP nor Achromatopsia. Because the full course of the disease is difficult to follow in patients, the availability of genetically engineered animal models is instrumental to uncover the pathophysiological mechanisms underlying the disease.
The aim of this project was to apply and further develop retinal imaging technology in order to refine in vivo structural information as a basis to better understand the mechanisms of the pathophysiology associated with inherited retinal diseases. Particularly within the scope of this thesis, major advances have been made in an early-onset form of RP called Leber’s Congenital Amaurosis (LCA) and two photoreceptor channelopathies that affect the rod and the cone system, respectively. This is complemented by the development of novel structural biomarkers for the follow-up of therapeutic strategies in both rod- and cone-centered disease classes.
The first part of the thesis introduces recent insights in the mechanisms of interaction of Crumbs proteins and the potential exchangeability of the different forms by means of respective mouse models: Crb1-/-, Crb2-/- and the Crb1Crb2 double knock-out mouse. It has been found that the lack of Crb1 resulted in a partial degeneration of the retina, whereas the lack of Crb2 induced a severe retinal degeneration similar to that of Retinitis Pigmentosa. The simultaneous ablation of both proteins mimics the characteristic retinal degeneration pattern showed by Leber Congenital Amarosis type (LCA) patients. This work has led to substantial progress in the field and several publications in a number of high-ranking journals (publication list 2,5,9,17,18).
The second part addresses the work on the restoration of function and morphology in a mouse model of Retinitis Pigmentosa or Achromatopsia, lacking the Cngb1 and Cnga3 proteins, respectively. Both approaches were performed by means of an adeno-associated viral (AAV) therapy. For the short –and long term assessment of therapeutic effects, novel morphological biomarkers based on optical coherence (OCT) data were develop. This work was documented in Koch et. al. 2012 and in Mühlfriedel et al. 2013.
The third part is devoted to the expansion of the diagnostic repertoire by improved, more informative biomarkers for therapy assessment beyond the state-of-the-art. The work is based on OCT, a novel technique for the in vivo visualization of retinal layers. Here, a quantitative method for a more detailed description of the retinal structure by means of reflectivity profiles was designed. This method was verified in three common laboratory species with differences in retinal architechture. Finally, the problem of unequal scales for the measurement of two-dimensional structures was resolved. Intraocular objects of known dimensions in the murine eye were used for the equal calibration of axes in OCT images. This work was reported in Garcia Garrido et al. 2014 and Garcia Garrido et al. 2015.
Retina
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