Activation of JNK signaling promotes all-trans-retinal–induced photoreceptor apoptosis in mice

Abstract

Disrupted clearance of all-trans-retinal (atRAL), a component of the visual (retinoid) cycle in the retina, may cause photoreceptor atrophy in autosomal recessive Stargardt disease (STGD1) and dry age-related macular degeneration (AMD). However, the mechanisms underlying atRAL-induced photoreceptor loss remain elusive. Here, we report that atRAL activates c-Jun N-terminal kinase (JNK) signaling at least partially through reactive oxygen species production, which promoted mitochondria-mediated caspase- and DNA damage-dependent apoptosis in photoreceptor cells. Damage to mitochondria in atRAL-exposed photoreceptor cells resulted from JNK activation, leading to decreased expression of Bcl2 apoptosis regulator (Bcl2), increased Bcl2 antagonist/killer (Bak) levels, and cytochrome c (Cyt c) release into the cytosol. Cytosolic Cyt c specifically provoked caspase-9 and caspase-3 activation and thereby initiated apoptosis. Phosphorylation of JNK in atRAL-loaded photoreceptor cells induced the appearance of γH2AX, a sensitive marker for DNA damage, and was also associated with apoptosis onset. Suppression of JNK signaling protected photoreceptor cells against atRAL-induced apoptosis. Moreover, photoreceptor cells lacking Jnk1 and Jnk2 genes were more resistant to atRAL-associated cytotoxicity. The Abca4−/−Rdh8−/− mouse model displays defects in atRAL clearance that are characteristic of STGD1 and dry AMD. We found that JNK signaling was activated in the neural retina of light-exposed Abca4−/−Rdh8−/− mice. Of note, intraperitoneal administration of JNK–IN-8, which inhibits JNK signaling, effectively ameliorated photoreceptor degeneration and apoptosis in light-exposed Abca4−/−Rdh8−/− mice. We propose that pharmacological inhibition of JNK signaling may represent a therapeutic strategy for preventing photoreceptor loss in retinopathies arising from atRAL overload.

Ferroptosis drives photoreceptor degeneration in mice with defects in all-trans-retinal clearance

“…Photoinduced photoreceptor degeneration in Abca4 −/− Rdh8 −/− mice implicates ferroptotic cell death As illustrated in Figure 8A, C57BL/6J and Abca4 −/− Rdh8 −/− mice aged 4 weeks were dark adapted for 48 h, irradiated for 2 h by 10,000-lx light emitting diode (LED) light, and then raised in the dark for 5 days, respectively. Consistent with our most recent report (19), hematoxylin and eosin (H&E) staining indicated that neural retina from Abca4 −/− Rdh8 −/− mice exposed to light visibly suffered from histological injury (Fig. 8B), and thicknesses of whole neural retina, outer nuclear layer (ONL), and inner and outer segments (OS+IS) were all decreased observably (Fig.…”

“…Previous studies indicate that iron overload gives rise to ROS by the Fenton reaction and elicits ferroptosis through ROS-mediated lipid peroxidation (20). We have lately demonstrated that abundant ROS (including mitochondrial ROS) are generated in atRAL-loaded 661W photoreceptor cells (19). The results shown in Figures 2 and 3 confirmed that atRAL interrupted iron homeostasis to increase Fe 2+ and dramatically induced lipid peroxidation in 661W photoreceptor cells.…”

“…Previous studies also show that atRAL causes cell death in a murine photoreceptor cell line (661W) (18). Most recently, we introduce that activation of c-Jun N-terminal kinase (JNK) promotes photoreceptor apoptosis induced by atRAL, and blocking JNK significantly mitigates photoreceptor atrophy and apoptosis in Abca4 −/− Rdh8 −/− mice subjected to light exposure (19). These findings suggest that patients with dry AMD and STGD1 may benefit from the suppression of photoreceptor apoptosis.…”

“…5). In recent time, we disclose the ability of atRAL to induce the production of mitochondrial ROS in 661W photoreceptor cells (19), implying that ROS Figure 9. Fer-1, a selective inhibitor of ferroptosis, alleviates light-induced photoreceptor degeneration and ferroptosis in Abca4 −/− Rdh8 −/− mice.…”

“…On the basis of MTS data and morphological images, atRAL at the concentration of 5 μM was used to treat 661W photoreceptor cells for 3 or 6 h in subsequent experiments. Evidence from our recent research has indicated that 5-μM atRAL triggers apoptosis of 661W photoreceptor cells after 6 h of treatment ( 19 ). However, to further ascertain whether atRAL-loaded photoreceptor cells suffer from nonapoptotic forms of cell death, we used quantitative reverse transcription–polymerase chain (qRT-PCR) and western blotting to analyze the expression of recognized ferroptosis-related genes and proteins, respectively.…”