High dose expression of heme oxigenase-1 induces retinal degeneration through ER stress-related DDIT3

Abstract

Background Oxidative stress is a common cause of neurodegeneration and plays a central role in retinal degenerative diseases. Heme oxygenase-1 (HMOX1) is a redox-regulated enzyme that is induced in neurodegenerative diseases and acts against oxidative stress but can also promote cell death, a phenomenon that is still unexplained in molecular terms. Here, we test whether HMOX1 has opposing effects during retinal degeneration and investigate the molecular mechanisms behind its pro-apoptotic role. Methods Basal and induced levels of HMOX1 in retinas are examined during light-induced retinal degeneration in mice. Light damage-independent HMOX1 induction at two different expression levels is achieved by intraocular injection of different doses of an adeno-associated virus vector expressing HMOX1. Activation of Müller glial cells, retinal morphology and photoreceptor cell death are examined using hematoxylin-eosin staining, TUNEL assays, immunostaining and retinal function are evaluated with electroretinograms. Downstream gene expression of HMOX1 is analyzed by RNA-seq, qPCR examination and western blotting. The role of one of these genes, the pro-apoptotic DNA damage inducible transcript 3 (Ddit3), is analyzed in a line of knockout mice. Results Light-induced retinal degeneration leads to photoreceptor degeneration and concomitant HMOX1 induction. HMOX1 expression at low levels before light exposure prevents photoreceptor degeneration but expression at high levels directly induces photoreceptor degeneration even without light stress. Photoreceptor degeneration following high level expression of HMOX1 is associated with a mislocalization of rhodopsin in photoreceptors and an increase in the expression of DDIT3. Genetic deletion of Ddit3 in knockout mice prevents photoreceptor cell degeneration normally resulting from high level HMOX1 expression. Conclusion The results reveal that the expression levels determine whether HMOX1 is protective or deleterious in the retina. Furthermore, in contrast to the protective low dose of HMOX1, the deleterious high dose is associated with induction of DDIT3 and endoplasmic reticulum stress as manifested, for instance, in rhodopsin mislocalization. Hence, future applications of HMOX1 or its regulated targets in gene therapy approaches should carefully consider expression levels in order to avoid potentially devastating effects.

Depletion of miR-96 Delays, But Does Not Arrest, Photoreceptor Development in Mice

“…All mice were handled and maintained in the animal facility of the School of Ophthalmology and Optometry at Wenzhou Medical University as authorized by the Institutional Animal Care and Use Committee of Wenzhou Medical University. 51 , 52 All animal experiments were performed in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.…”

Sensitivity of the Dorsal-Central Retinal Pigment Epithelium to Sodium Iodate-Induced Damage Is Associated With Overlying M-Cone Photoreceptors in Mice

“…Hematoxylin-eosin (H&E) staining has been described previously. 29 Briefly, a mark was made at a 12 o'clock position of the cornea, and the extracted eyeballs were incubated in a mixed fixing solution (formaldehyde, ethanol, H 2 O glacial acetic acid; 1:4:4:1) for at least one day at room temperature. Paraffin retinal sections (5 µm) were stained by a standard program.…”

“…Quantitative real time PCR was performed on a 7500 Real-Time PCR Detection System (Applied Biosystems) with Power SYBR Green PCR Master Mix as previously described. 29 All primer sequences used for real-time PCR are listed in the supplementary data ( Supplementary Table S1 ).…”

“…TUNEL staining was performed as previously described. 29 Briefly, the paraffin retinal sections were dewaxed by xylene and boiled in sodium citrate (0.01 M; Beyotime Institute of Biotechnology) for two minutes. Then, sections were incubated with the TUNEL reaction Kit (Roche, 11684795910) according to the manufacturer's protocol.…”

Cellular stress signaling and the unfolded protein response in retinal degeneration: mechanisms and therapeutic implications

“…These results suggest that maintaining a certain level of CHOP is necessary for Nrf2 activation and cell survival in the RPE and photoreceptors during stress conditions. However, excessive CHOP activation by ER stress can be detrimental to cell survival and function contributing to neurodegeneration [ 82 ].…”