Expression of inducible nitric oxide synthase in the brain tissue of rats at preoxygenation, hypoxia/reoxygenation

BACKGROUND: The disorder of the respiratory and circulation system during the anesthesia and operation often can lead to severe hypoxia. Preoxygenation is a conventional therapy for treatment of hypoxia. OBJECTIVE: To observe the change in the expression of inducible nitric oxide synthase(iNOS), i.e. brain tissue injury degree, in cerebral cortex of rats after hypoxia/reoxygenation under the condition of different levels of preoxygenation. DESIGN: Randomized controlled animal experiment. SETTING: Department of Anesthesiology and Central Laboratory, Shengjing Hospital Affiliated to China Medical University. MATERIALS: This experiment was carried out in the laboratory of Shengjing Hospital Affiliated to China Medical University from March 2003 to March 2004. Seventy-two male Wistar rats, weighing from 250 to 300 g, were provided by the Animal Experimental Room of Shengjing Hospital of China Medical University [License No. SYXK (Liao) 2003-0019]. Seventy-two rats were randomized into 3 groups: preoxygenation group, hypoxia group and reoxygentation group. Each group was divided into 4 subgroups, 6 rats in each subgroup. METHODS: 0.21, 0.50, 0.75 and 0.98 volume fraction of oxygen was given to 4 subgroups of preoxygenation group respectively. The rats in each subgroup of hypoxia group inhaled oxygen for 30 minutes according to the method of preoxygenation. Then, nitrogen gas replaced oxygen and was pumped into the cabin, and the volume fraction of oxygen was decreased to be 0.05 within 20 minutes. The rats in each subgroup of reoxygenation group were treated with the experimental methods of preoxygenation and hypoxia separately, then they were moved into the wide mouthed bottle with two-air-duct rubber stopper. Finally, oxygen was pumped and the volume fraction of oxygen reached over 0.98 within 20 minutes. After 24 hours, all the surviving rats were killed by decapitation. Cerebral tissue was sliced and stained by haematoxylin-eosin and then enveloped. Brain tissue injury was observed by pathohistological method. The change of iNOS expression was observed with immunohistochemial method. The cells with yellow-brown particle deposition in cytoplasm were judged to be positive. The percentage of positive cells was calculated: < 25% negative, 25% to 49% positive and ≥ 50% strong positive. The higher the percentage was, the severer the brain tissue injury was. MAIN OUTCOME MEASURES: iNOS expression and brain tissue injury degree in rats. RESULTS: All the 72 rats were involved in the result analysis, without deletion. ① Comparison of the percentage of positive cells stained by iNOS in the brain tissue of rats: in the preoxygenation group, the percentage in the subgroups of 0.50, 0.75 and 0.98 volume fraction was significantly higher than that in the subgroup of 0.21 volume fraction [(3.83±3.15)%, (22.07±9.57)%, (22.93±6.58)%, (1.93±1.08)%, P < 0.01].Compared with preoxygenation group, the percentage was significantly increased in the hypoxia group (P < 0.01) and further increased in the reoxygenation group (P < 0.01). In the hypoxia group and reoxygenation group, the percentage in the subgroups of 0.75 and 0.98 volume fraction was higher than that of corresponding subgroup of 0.21 volume fraction [(39.17±9.84)%, (42.93±3.95)%, (29.80±7.39)%, P < 0.01; (47.37±9.66)%, (54.53±9.05)%, (41.43±12.07)%, P < 0.01], while the percentage in the subgroup of 0.50 volume fraction [(19.43±7.92)%, (25.77±4.63)%] was lower than that of corresponding subgroup of 0.21 (P < 0.01). ② Pathohistological results of brain tissue of rats in each group: After hypoxia, the neuronal injury in each group was aggravated, and the injury in the subgroup of 0.50 volume fraction was milder than that in the subgroup of 0.21 volume fraction. After reoxygenation, the injury in each group was further aggravated and the injury in the subgroup of 0.98 volume fraction was the severest. CONCLUSION: ①Low concentration of preoxygenation can better raise the cerebral antioxidant ability , and high concentration preoxygenation is on the contrary. It suggests that the change of iNOS expression may be one of mechanisms that different concentrations of preoxygenation cause different anti-oxidative abilities of brain cells. ② iNOS has neurotoxic effect in ischemia/reoxygenation brain injury.