Research On The Prevention Effects And Mechanism Of Hyperbaric Oxygen On Brain Injury Induced By Microwave Radiation In Rats

Part Ⅰ Research on the prevention effects of hyperbaric oxygen on brain injury induced by microwave radiation in ratsObjectiveMicrowave refers to the frequency of 300MHz-300GHz electromagnetic waves,it has already widely used in telecommunications, aviation, medical and military fields,it brings human convenience at the same time, the radiation damage also has been concerned about.The central nervous system is most sensitive to microwave radiation,but the prevention measures against microwave radiation damage is relatively rare.This study was to establish an animal model of brain injury induced by microwave radiation,to observe the protective effect of hyperbaric oxygen for microwave radiation induced brain injury,for microwave radiation-induced brain injury treated by hyperbaric oxygen to enter clinical use..Methods1.100 secondary male Wistar rats were randomly divided into 4 groups:normal control group, HBO control group,radiation group and HBO therapy group,each group of 25.The rats were exposed to microwave which average power density was 30mW/cm2 for 15 minutes for 3 days,after radiation the following day, the rats were given 1.6 ATA hyperbaric oxygen treatment,1 time a day for 14 days.2.The rats of each group were training for 3 days to detect learning ability before microwave radiation.The rats were proceeded place navigation experiment to acquisition average latency at 1d,2d,3d,7d,14d,21d after radiation,to test changes in learning and spatial memory of rats.3.Weigh weight of rats and through intraperitoneal injecte 1% pentobarbital sodium (30 mg/kg) anesthesia at 6h,7d,14d,21d after radiation.Decapitate head directly,take its brain tissue,and separating the hippocampus,With hematoxylin-eosin (HE) staining,observation on morphological structure of hippocampus in rats by light microscopy.4. Weigh weight of rats and through intraperitoneal injecte 1% pentobarbital sodium (30mg/kg) anesthesia at 7d and 14d after radiation,take 1mm3 hippocampus.By combination of uranyl acetate and lead citrate staining to observation on ultrastructure of hippocampus in rats through transmission electron microscopy.5.The data were expressed as the mean ± standard deviation of the mean. Comparison data between various groups by one-way ANOVA. P value of 0.05 was considered as statistically significant..Results1. The average escape latency(AEL) was significantly longer at 1-14d after exposure to 30 mW/cm2 microwave radiation(p<0.05). HBO therapy group AEL was significantly shorten compared with radiation group,and had no significant difference compared with normal control group at 7d-14d after radiation.There were not significant difference between HBO control group and normal control group(p>0.05).2. Changes of organizational structure in hippocampus:Hippocampal neuron cell nuclear pyknosis, hyperchromatism, Eosinophilic cytoplasm increased vascular tissue wide separations, angioedema after 30mW/cm2 microwave at 6h after radiation.Lesions aggravated at 7d after radiation,and recover at 14d after radiation,it was still not fully back to normal at 21d after radiation,part of neuron degeneration, vascular edema.Hippocampal neuron cell nuclear pyknosis, hyperchromatism, Eosinophilic cytoplasm increased in HBO therapy group at 7d after radiation,compared with radiation group,pyknosis and dark stained neurons decreased,edema reduction,hippocampus injury recovery.Hippocampu had basically returned to normal at 21d after radiation.3.Changes in ultrastructure of hippocampus:Swelling of mitochondria in hippocampal neurons and partial cavitation, cristae and dissolve, rough endoplasmic reticulum, nuclear membrane increased,broadening of nuclear envelope, secondary lysosomes in the cytoplasm, or even visible phenomena of neural and glial cells degeneration of neurons,Synaptic blur reduced presynaptic membrane vesicles, increase in postsynaptic dense at 7d after radiation.HBO therapy group has lesser lesions.The ultrastructure of hippocampus is normal in the normal control group and HBO control group.Conclusion1.30mW/cm2 microwave radiation could injure the rats,brain,which was displayed as the rats, learning and memory abilities decreasing,organizational structure and Ultrastructure of hippocampus damage.2.1.6ATA HBO therapy has a protective effect on rat brain damage induced by microwave irradiation.3.1.6ATA HBO therapy would not cause injury in rats.Part Ⅱ Research on the prevention mechanism of hyperbaric oxygen on brain injury induced by microwave radiation in ratsObjectiveThe Part I of the study found that HBO therapy has a protective effect on rat brain injury induced by microwave radiation,but its mechanism is not clear.Microwave radiation can cause cerebral energy metabolism in the theory that has been confirmed by experiments,HBO therapy can improve cerebral microcirculation, increase partial pressure of oxygen,and promote neuronal cell energy metabolism.We speculate that,HBO may be achieved by improving cerebral microcirculation in rats after microwave irradiation to correct brain energy metabolism and function.For this reason,we detecte mitochondrial enzymes in rat hippocampal activity by enzyme-linked immunosorbent assay,detective hypoxia inducible factors (HIF-la) protein expression by immunohistochemical.Discussing the mechanism of protectice effect on brain of HBO.Methods1.100 secondary male Wistar rats were randomly divided into 4 groups:normal control group, HBO control group,radiation group and HBO therapy group,each group of 25.The rats were exposed to microwave which average power density was 30mW/cm2 for 15 min for 3 days,after radiation the following day, the rats were given 1.6 ATA hyperbaric oxygen treatment,1 times a day for 14 days.2. Weigh weight of rats and through intraperitoneal injecte 1% pentobarbital sodium (30 mg/kg) anesthesia at 6h,7d,14d,21d after radiation.Decapitate head directly,take its brain tissue,and separating the hippocampus.3. By hippocampal tissue weight (g):volume (ml)=1:9 added 9 times the volume of saline solution, ultrasonic disintegrator made of crushed 10% preparation of hippocampal tissue homogenate.4. Determine hippocampus protein content by coomassie brilliant blue staining.5. Determine hippocampus changes of activity of LDH,Na+-K+-ATPase,MAO.6. We killed the rats,took its brain tissue,and separated the hippocampus at 6h, 7d,14d,21d after radiation,maked of paraffin,observed hypoxia inducible factors (HIF-1α) protein expression by immunohistochemical.With Image-Pro Plus image analysis software,we calculated within each group the same size integrated optical density (IOD).7. The data were expressed as the mean ± standard deviation of the mean. Comparison data between various groups by one-way ANOVA. P value of 0.05 was considered as statistically significant.Results1. There were no difference between each group of LDH activity in hippocampus of rats at 6h after radiation.LDH activity was significantly elevated in radiation group.compared with other groups,there was significant difference (p<0.05),it displayed a trend of first increasing and then decreasing over time.LDH activity had a trend of increasing in HBO therapy group at 7d after radiation,but it had no significant difference compared with normal control group (p>0.05).It was significantly lower than the radiation group (p<0.05).LDH activity showed a increasing trend in HBO therapy group at 14d after radiation,it had a significant difference compared with normal control group (p<0.05),but it was lower than radiation group.LDH activity was back to normal in HBO therapy group at 21d after radiation.LDH activity was not significant increase in HBO control group in all times.there was no significant difference compared with normal control group (p>0.05).2. There were no significant difference between each group of Na+-K+-ATPase activity after radiation 6h.Na+-K+-ATPase activity significantly reduced in radiation group at 7d-14d after radiation compared with other group,and it was still lower than normal.Na+-K+-ATPase activity had no significant decline in all times,it had no significant difference compared with normal control group(p>0.05).3. MAO activity was significantly elevated in radiation group,it has significant difference compared with normal control group at all times,and it has significant difference compared with HBO therapy group at 7-21d after radiation,and it was not back to normal at 21d after radiation.MAO activity was back to normal in HBO therapy group at 7-21d after radiation,it has no significant difference compared with normal control group(p>0.05).4.Observed rat hippocampal expression of Immunohistochemistry staining HIF-1α with light microscope.positive cells were distinctly yellowish-brown or yellow-brown granules.It had no HIF-1α expression in normal control group and HBO control group in all times.There were HIF-1α expression in radiation group and HBO therapy group after radiation 6h,IOD of two groups were higher than that in the normal control group.There was a large number of HIF-1α expression in radiation group at 7d after radiation,otherwise there was a few number of HIF-1α expression in HBO therapy group.HBO therapy was back to normal at 14-21d after radiation,but radiation group still has HIF-1α expression.Conclusion1. Microwave radiation can cause mitochondrial energy metabolism in rat hippocampus, increase HIF-1α expression,cause brain damage.2. Hyperbaric oxygen may play a protective role in the brain by improving mitochondrial energy metabolism in the hippocampus and inhibiting the expression of HIF-1α caused by microwave radiation.