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V. Marigo, D. Sanges, A. Comitato; Cross-Talk Between Caspase-12 and AIF During Apoptosis in Animal Models of Retinits Pigmentiosa. Invest. Ophthalmol. Vis. Sci. 2007;48(13):554.
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Molecular mechanisms underlying apoptosis in retinitis pigmentosa (RP) are still elusive and this hampers the development of a cure for this blinding disease. Changes in intracellular calcium homeostasis had been associated with retinal degeneration in an animal model for RP, the rd1 mouse. Retention of mutant protein in the endoplasmic reticulum (ER) was shown to have detrimental effects in transgenic mice expressing mutant rhodopsin. AIF and caspase-12 are activated by calpains in response to two ER stresses: protein misfolding and disruption of calcium homeostasis.
In order to correlate activation of mitochondrial AIF, caspase-12 and apoptosis, co-localization studies by triple immunofluorescence of AIF, caspase-12 and TUNEL were undertaken in sections of degenerating retinas. Similar experiments were performed in wild type and mutant retinal stem cells differentiated in vitro to rod photoreceptors. Retinal stem cells were treated with calpastatin, caspase inhibitors, siRNAs to AIF, caspase-12 and calpains followed by immunofluorescence analysis. P10 mutant mice were intravitreally injected with either calpastatin or caspase inhibitors and the outcome was evaluated by immunoflorescence and TUNEL staining.
We found that AIF and caspase-12 are activated and localize to the nucleus of the same apoptotic photoreceptors in the two murine models for RP. By reduction of AIF or caspase-12 expression we also defined that AIF plays the major role in apoptosis in the rd1 retina while caspase-12 has a reinforcing effect. By treatment with a calcium-channel blocker we established that AIF and caspase-12 activation is dependent on intracellular calcium in rd1 photoreceptors. Most interestingly, by blocking calpain activity with specific inhibitors we decreased apoptosis in the degenerating retina together with activation of AIF and caspase-12.
The option of exploiting apoptosis as a therapeutic target is complex. Nevertheless the molecular understanding of the different apoptotic factors activated during degeneration and the identification of common activators are the first step toward this goal. Our study identifies AIF and caspase-12 as two apoptotic factors triggering photoreceptor cell demise. The efficacy of calpain inhibitors in interfering with apoptosis and relocalization of AIF and caspase-12 opens new perspectives for designing therapeutic approaches to rescue photoreceptor cell death in this disease.
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