Experimental Study Of Mitochondria TrxR2 On The Mechanism Of Hypoxic Preconditioning Reduce Oxidative Stress On The Rats After Traumatic Brain Injury

Traumatic brain injury has become a major cause of battle-related injuries for soldiers in the modern warfare, but also the daily neurosurgery most common diseases, the rate of incidence and mortality are increased year by year. Primary brain injury directly related to the outside power, generally can not directly intervene. The pathological process of secondary brain injury are complicated and become the main direction of clinical treatment and intervention. Oxidative stress plays an important role in the pathological process of secondary brain injury, Thioredoxin reductase 2 (TrxR2) is an important anti-oxidative enzymes of mitochondrial. Study the TrxR2 expression changes and interventions may become a new target for treatment of traumatic brain injury.Part I:Experimental study of the expression changes of Mitochondria TrxR2 on rats brain cortex after traumatic brain injuryObjective: Research of the the expression changes of thioredoxin reductases2 in rats brain cortex after traumatic brain injury, and explore the role of TrxR2 in the pathological process of secondary brain injury.Methods:One hundred and eight healthy Sprague Dawley male rats were randomly divided into normal control group (Con, n=6) and traumatic brain injury group (TBI, n=96). The rats traumatic brain injury model were made by the modified Feeney’s equipment. After traumatic brain injury was made, the rats was subdivided into 8 groups, namely group 1 h,3 h,6 h,12 h,1 d,3 d,7 d,14 d, and each group had 12 rats according to sacrifice time point. Six rats in each group were selected directly decapitated the brain on ice. The content of brain tissue water, superoxide dismutase (SOD), malondialdehyde (MDA), glutathione glutathione (GSH) around 0.5 cm contusion brain cortex lesions was measured. The mitochondrial TrxR2 protein were extracted for Western-Blot detection and activity analysis. Another six rats through heart perfused with 4% paraformaldehyde and than brains immersed in 4% paraformaldehyde, dehydrated, embedded in paraffin. The expression of TrxR2 and NeuN in the brain cortex was detected by immunohistochemistry.Results: The expression of TrxR2 protein in the brian cortex enhanced at 3 h, peaked at 1 d and normalized on 14 d after traumatic brain injury. Compared with normal control group, the TrxR2 protein expression in traumatic brain injury group were significantly higher from 3 h to 7 d (P<0.05). The activity of TrxR2 reduced at 1 h, lowest at 1 d and normalized on 14 d after injury. The optical density of NeuN reduced at 1 h, lowest at 3 d and normalized on 14 d after injury. The activity of TrxR2 activity has positively correlated with the neuronal activity NeuN staining (r=0.855, P< 0.05), but negatively correlated with TrxR2 average optical density, water content and MDA content in brain tissue (r = -0.818, r = -0.824, r = -0.842, P<0.05), and positively correlated with the activity of SOD and GSH content (r-0.854, r = 0.862, P＜0.05).Conclusions:The expression of TrxR2 protein were increased after traumatic brain injury, but its activity was inhibited. The change of TrxR2 activity were reversed with the level of brain tissue water content and oxidative stress, but the same as the neuronal activity change. Therefore, focused on improving the activity of TrxR2 expression after traumatic brain injury can inhibit oxidative stress, reduce secondary brain injury and 1mprove prognosis.Part Ⅱ:Experimental study of Mitochondria TrxR2 on the Mechanism of Hypoxic Preconditioning reduce Oxidative Stress on the Rats after Traumatic brain injuryObjective: Observe the change and influence of hypoxia preconditioning on the expression of Nrf2 and TrxR2 after traumatic brain injury on the around of rats’ brain damage zone. To study Mitochondrial TrxR2 on the mechanisms of hypoxic preconditioning reduces oxidative stress on rats after traumatic brain injury.Methods: Forty eight healthy adult male Sprague Dawley rats were randomly divided into sham group (Sham), hypoxic preconditioning group (HPC), traumatic brain injury group (TBI) and hypoxic preconditioning + traumatic brain injury group (HP+TBI), each group had 12 rats. The hypoxic preconditioning model of rats were made by Hypobaric chamber for 3 days (-50 kPa、3 h/d) and the traumatic brain injury models were used the Feeney s improved equipment. The neurological functions of each experimented group were evaluated by the Modified neurological severity score (mNSS) at 24 h after injury. Six rats in each group were selected to examine the content of brain tissue water, superoxide dismutase (SOD), malondialdehyde (MDA), glutathione glutathione (GSH) around 0.5 cm contusion brain cortex lesions. The expression of Nrf2 and mitochondrial TrxR2 protein were extracted for Western-Blot detection or activity analysis. Another 6 rats were killed for immunohistochemical staining to detect the Nrf2, TrxR2 and neurons NeuN expressions.Results:Compared with Sham or HPC group, The Nrf2 and TrxR2 protein expression were significantly higher (P<0.01). However, the Modified neurological severity score, brain water content and MDA level of brain tissue in HP+TBI group were significantly lower than in TBI group (P<0.05). Whlie the TrxR2 activity, SOD activity, GSH levels and NeuN neuronal activity were significantly higher than TBI group (P<0.05).Conclusions: Hypoxic preconditioning can up-regulate the expression and activity of TrxR2, less cerebral edema and oxidative stress, improving neuronal activity, reducing secondary brain injury after traumatic brain injury.