Proposed pharmacological countermeasures against apoptotic cell death in experimental models mimicking space environment damage

Space environment damage-mimicking experimental models. A number of damaging agents, acting by different pathogenetic mechanisms, could undermine human health during interplanetary exploration. In the last years it has emerged that large part of these mechanisms lead to death by apoptosis of damaged tissue cells. On the basis of these considerations, our aim is to engineer countermeasures aimed to inhibit cell apoptosis consequent to different pathogenetic mechanisms. Focusing on space environment-related eye tissue lesions, our studies have been carried out in cultured cells of cornea and retina treated with the following apoptotic stimuli: a pulse of X-ray irradiations, a several-day exposure to either 3H-labeled tymidine or to the genotoxic drug doxorubicin, or to the membrane damage apoptosis lipid mediator cerammide. Hypoxia and glucose/growth factor starvation (Locke medium) are also used as apoptotic stimuli. All these damaging agents trigger one of two alternative apoptotic pathways, namely the death receptor-dependent pathway and the mitochondrion-dependent pathway (see our review Tempestini et al. Eur J Ophthalmol. 13 Suppl 3:S11-18, 2003 Review). Our proposed countermeasures. Here we evaluate two antiapoptotic molecules. One is free radical scavenger coenzyme Q10 (CoQ10), proved to inhibit also the mitochondrial Permeability Transition Pore (mPTP) opening and thereby the mitochondrion-dependent apoptotic pathway in neuronal cells. Unexpectedly, CoQ10 also inhibited apoptosis in the response to cerammide, which trigger the receptor-dependent apoptotic pathway. Another proposed countermeasure are peculiar antisense oligoribonucleotides (ORNs) we have targeted to a cis-acting destabilizing element of bcl-2 mRNA and then patented for their ability to prevent apoptosis by preventing bcl-2 gene down-regulation in response to apoptotic stimuli. Parameters of effectiveness. We evaluated apoptosis rate on the basis of different parameters, which are examination by light-, electron-microscopy and confocal microscopy, cumulative apoptotic events scored by Time Lapse Videomicroscopy, ROS and ATP levels, mitochondrial membrane charge, activities of cytoplasmic cytochrome C, caspases and PARP, and DNA fragmentation as previously reported (Papucci et al., J Biol Chem. 278:28220-28220, 2003). Results and conclusions. Our preliminary results clearly indicate that CoQ10, as well as bcl-2 down-regulation preventing ORNs, significantly counteract apoptosis in response to different DNA damaging agents in cultured eye cells. This supports the possibility that they could be optimal countermeasures against ophthalmologic lesions during human space explorations. Furthermore, our previous observation that CoQ10 administered as eye drops (collirio) reaches the area coroide-retina (Capaccioli et al., congress communication) strongly suggests that it could be simply administered in this form. Further experiments are programmed.