| Summary: | 博士 === 國防醫學院 === 醫學科學研究所 === 95 === Paraplegia due to spinal cord ischemic injury remains one of serious complications after surgical repair of thoracoabdominal aortic aneurysms. Studies had shown that cyclooxygenase (COX) may contribute to ischemic neuronal damage and COX inhibitors may reduce this injury. Ketorolac inhibits both COX-1 and COX-2, and, in contrast to other NSAIDs, is water-soluble. This research was examined the protective effect of ketorolac, via intrathecal pretreatment against ischemic spinal cord injury; and tried to explore the role of COX in spinal cord ischemic injury. This research was divided into four parts: (1) examining the effects of intrathecal ketorolac pretreatment by assessing hind limbs motor function of rats and observing histopathological changes in lumbar spinal cord at 24 hours after ischemic injury; (2) evaluating the safety of ketorolac by recording the survival of rats 28 days after reperfusion; (3) investigating whether the neuroprotective effect of ketorolac was related to changes of spinal cord release of nitric oxide (NO) or excitatory amino acids (EAAs) by using spinal cord microdialysis, high performance liquid chromatography (HPLC) and NO analyzer; (4) detecting changes of several kinds of messenger RNAs (COX-2, subtypes of NO synthase (NOS), and subtypes of excitatory amino acid transporter (EAAT)) in the rat spinal cords after ischemic injury by real time polymerase chain reaction (real time PCR).
Intrathecal pretreatment with ketorolac 60 g prevented the hind limbs motor dysfunction and the histological changes in the spinal cord at 24 hours after the ischemia reperfusion, but the lower dose (30 g) had no protective effect. Therefore, 60 g ketorolac was used as the dose for the following experiments. In evaluating the safety of ketorolac, only 20% ischemic control rats survived for 4 weeks after injury. In contrast, ketorolac pretreatment, only one rat showed delayed paraplegia at 48 hours after ischemia and died one day later, and three of the other rats died within 4 weeks. The other six survived rats remained in good motor function until 28 days after reperfusion. The overall survival rate, at 28 days after surgery, was higher in rats receiving ketorolac (60%) than in control animals (20%). In fact, interruption of aortic blood flow during aortic aneurysm surgery may not only induce spinal cord ischemia, but also deleterious the heart, gut and urinary systems. Our data showed a neuroprotective effect of i.t. 60 g ketorolac in reducing the incidence of paraplegia, but it may not exert a protective effect on other organs. In spinal cord microdialysis study, a slightly, short-lasting increase of aspartate was observed at 2 h after reperfusion and returned to baseline thereafter. Glutamate level in ischemic animals dramatically increased at the first hour after reperfusion, and returned to baseline within four hours. In most rats, a second, long-lasting elevation of glutamate was observed six hours after ischemia-reperfusion, and lasted for 24 hours.
The maximum NO release was observed at 3h after ischemia-reperfusion. We found that NO and EAAs aspartate and glutamate were increased at different time points after spinal cord ischemia, and i.t. ketorolac inhibited these increases. Real time PCR showed increasing of mRNA expression of COX-2, EAAT1, EAAT2 and EAAT3, peaked at 3h after spinal cord ischemic injury. The expression of NOS subtypes showed diversity; iNOS and nNOS mRNA began their increases from 30 min after ischemia; in contrast, eNOS began its increase from 3h after ischemia. and iNOS mRNA peaked its increase at 12h after ischemia.
It is thought that the excessive increasing of glutamate concentration following neuronal ischemia increases intracellular calcium influx, consequently induces proteins expressing and initiates inflammatory processes. PGs, the major products of COX, and NO will then be increased by the glutamate stimulation. NO has been suggested as a retrograde transmitter in this signal transduction cascade. As NO and prostaglandins are formed, they may diffuse back to the presynaptic neuron where it, in turns, activates further glutamate release. The protective effects of intrathecal ketorolac pretreatment in spinal cord ischemia may come from inhibiting the production of prostaglandins, then further preventing the ongoing of “glutamate-nitric oxide-prostaglandins “
mutual potentiation. Further studies are needed to elucidate the downstream signaling pathway to beneficial future clinical application in this entity.
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