A Functional and Histological Analysis on Oxygen Sensitivity During the Critical Period of Retina Development

Date of Award

5-2020

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Arturo Bravo Nuevo, PhD

Second Advisor

Mindy George-Weinstein, PhD

Third Advisor

Zeus Andrea Antonello, PhD

Abstract

The death of neuronal cells is a part of the normal development of mammalian central nervous system, including the retina. In the retina, soon after the genesis of photoreceptor metabolism, the inner retinal layers go through a state of oxygen deprivation due to its consumption by the highly metabolically active photoreceptors. This metabolic stress induces apoptosis in a fraction of photoreceptors during a time known as the critical period. During the critical period, due to the lack of oxygen, levels of the pro-angiogenic factor Vascular Endothelial Growth Factor rise inducing the formation of new vessels that in turn will provide the adequate supply of oxygen for the stable retina photoreceptor population of the adult. This balanced process that involves metabolic stress, photoreceptors death, and angiogenesis can be altered by genetic factors but also simply by changes in oxygen exposure during development, as exemplified by the cases of retinopathy of prematurity.

The goal of this study was to investigate the effects of oxygen administered during the critical period on vision and retinal morphology in wildtype mice (C57BL/6J) and a mouse model of retinitis pigmentosa (C3H/HeJ). This disease is characterized by a degeneration of photoreceptors, predominantly the rod photoreceptor cells, that leads to a loss of peripheral vision and night blindness. The study was conducted with the hypothesis that modulation of oxygen could reduce or prevent photoreceptor degeneration. Additionally, we explored the effect of varying oxygen levels on the number of Myo/Nog cells in wild type mice. These cells were previously shown to aggregate in areas of stress and demonstrated to be neuroprotective in the retina. Conclusively, it was hypothesized that Myo/Nog cells would be present during retinal development and linked to areas of apoptosis. Not only that but it was also hypothesized that Myo/Nog cells would vary with modulating oxygen levels.

Animals were reared in either hyperoxia (75% O2), hypoxia (12% O2), or normoxia (21% O2) during the critical period (P7-P20). Electroretinography (ERG) and Optical Coherence Tomography (OCT) were performed on P26 and 27, to assess function and structure of the retina, respectively. Additionally, histological examination was conducted on P12, P16, and P28 to assess the effect of varying oxygen levels on cell death by staining with a marker for apoptosis.

The ERG data showed that the average A and B wave amplitudes were higher in C57 and C3H mice exposed to hyperoxia during the critical period compared to mice raised in normoxic conditions suggesting rescue of photoreceptor death in both normal and pathological retinal apoptosis. This was confirmed by OCT measurements, which indicated that the treatment of C57 and C3H mice with elevated O2 resulted in a thicker ONL compared to normoxic controls. Hypoxia induction in C57 mice increased cell death in the ONL (P16) and the GCL (P28) and resulted in a decrease in visual function due to deterioration of the retina.

Although further experimentation is required to distinguish between direct and indirect effects of oxygen on photoreceptor viability in retinitis pigmentosa (RP), it can be concluded that hyperoxia can reduce apoptosis of photoreceptors in normal retina as well as in the C3H model, suggesting that hyperoxia could be used to reduce or prevent photoreceptor degeneration in RP.

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