Apoptosis and autophagy in photoreceptors exposed to oxidative stress.
Studies on human and animal models of retinal dystrophy have suggested that apoptosis may be the common pathway of photoreceptor cell death. Autophagy, the major cellular degradation process in animal cells, is important in normal development and tissue remodeling, as well as under pathological conditions. Previously we provided evidence that genes, whose products are involved in apoptosis and autophagy, may be coexpressed in photoreceptors undergoing degeneration. Here, we investigated autophagy in oxidative stress-mediated cell death in photoreceptors, analyzing the light-damage mouse model and 661W photoreceptor cells challenged with H(2)O(2). In the in vivo model, we demonstrated a time-dependent increase in the number of TUNEL-positive cells, concomitant with the formation of autophagosomes. In vitro, oxidative stress increased mRNA levels of apoptotic and autophagic marker genes. H(2)O(2) treatment resulted in the accumulation of TUNEL-positive cells, the majority of which contain autophagosomes. To determine whether autophagy and apoptosis might precede each other or co-occur, we performed inhibitor studies. The autophagy inhibitor 3-methyladenine (3-MA), silencing RNA (siRNA) against two genes whose products are required for autophagy (autophagy-related (ATG) gene 5 and beclin 1), as well as the pan-caspase-3 inhibitor, Zvad-fmk, were both found to partially block cell death. Blocking autophagy also significantly decreased caspase-3 activity, whereas blocking apoptosis increased the formation of autophagosomes. The survival effects of 3?MA and zVAD-fmk were not additive; rather treatment with both inhibitors lead to increased cell death by necrosis. In summary, the study first suggests that autophagy participates in photoreceptor cell death possibly by initiating apoptosis. Second, it confirms that cells that normally die by apoptosis will execute cell death by necrosis if the normal pathway is blocked. And third, these results argue that the up-stream regulators of autophagy need to be identified as potential therapeutic targets in photoreceptor degeneration.
Kunchithapautham
Kannan
K
Department of Neurosciences, Division of Research, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Rohrer
Bärbel
B
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D001343:Autophagy / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D000071187:Autophagy-Related Protein 5; D001483:Base Sequence; D000071186:Beclin-1; D053148:Caspase 3 / Q000502:physiology; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D004195:Disease Models, Animal; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D008027:Light / Q000009:adverse effects; D051379:Mice; D008807:Mice, Inbred BALB C; D008869:Microtubule-Associated Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology; D009336:Necrosis; D018384:Oxidative Stress; D020419:Photoreceptor Cells, Vertebrate / Q000187:drug effects / Q000502:physiology* / Q000528:radiation effects; D011506:Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics; D012162:Retinal Degeneration / Q000209:etiology / Q000235:genetics / Q000473:pathology / Q000503:physiopathology
D000590:Amino Acid Chloromethyl Ketones; D051017:Apoptosis Regulatory Proteins; C495773:Atg5 protein, mouse; D000071187:Autophagy-Related Protein 5; D000071186:Beclin-1; C115010:Becn1 protein, mouse; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D017931:DNA Primers; D008869:Microtubule-Associated Proteins; D011506:Proteins; D034741:RNA, Small Interfering; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C025946:3-methyladenine; D006861:Hydrogen Peroxide; C505409:Casp3 protein, mouse; D053148:Caspase 3; D000225:Adenine