Respuesta de las proteínas mal plegadas (UPR) a la isquemia global tras 48 horas de reperfusión en el cerebro de ratael efecto de la edad y del meloxicam

Abstract

The endoplasmic reticulum (RE) is a multifunctional organelle where proteins are properly folded, lipids are biosynthesized and calcium is stored. Nutrient depletion, starvation, impairment in calcium homeostasis or redox state impair the normal RE function. This impairment also occurs after the oxygen and glucose deprivation followed by the return to normoxic conditions that happens in stroke. The malfunction in the RE leads to a protein misfolding which accumulate and activate the signaling pathway called the unfolded protein response (UPR). UPR activation triggers an extensive transcriptional response, which adjusts the folding capacity of the RE protein. This study aims to analyze the role of age in the UPR induced by cerebral ischemia, and the effect of the anti-inflammatory agent meloxicam. The study was carried out on the CA1 and CA3 hippocampal areas, dentate gyrus and cerebral cortex of 3 and 18 month-old rats, using a model of 15 minutes of global cerebral ischemia followed by 48 hours of reperfusion. Typical UPR markers were used, such as the molecular chaperones GRP78/BIP and GRP94, the protein disulfide isomerase (PDI) as well as the transcription factor CHOP, a widely used death marker. The expression of these genes was measured by quantifying PDI, CHOP, GRP78 and GRP94 mRNA levels by Real Time PCR (qPCR) and PDI and GRP78 levels by Western blot. The study shows how the mRNA levels of these genes were similar in 3 and 18 month-old sham-operated animals, while the ischemic insult elicited a noticeable increase in the expression of these genes in young animals that was scarcely appreciable in older animals. The striking increase in the mRNA levels ofSummary 13 these genes in 3 month-old animals was abolished or even reverted by the treatment with meloxicam. Western blot assays showed that the UPR is still detectable 48 hours after ischemia in some of the studied areas and provided evidence that the UPR is different between younger and older animals. Western blot assays carried out in young animals also showed that meloxicam elicited different effects on the levels of PDI and GRP78/BIP in the cerebral cortex and hippocampus. We conclude that the UPR response to ischemic/reperfusion insult is dependent on age and inflammation and could play an important role in ischemic vulnerability. The UPR appears to be strongly reduced in aged animals, suggesting a reduced ability for cell survival.