Screening For Differentially Expressed Genes In Rat Brain Injury Induced By Repeated+Gz Exposures

Modern high-performance fighter planes can develop very high acceleration (+Gz). Under such a situation, pilots have to undertake a high-sustained G acceleration stress, which may produce loss of consciousness (G-LOC), even result in fatal flight accidents. So, it is necessary to further investigate the effects of brain damage induced by repeated +Gz acceleration stress. Research on the molecular mechanism of brain damage induced by repeated +Gz exposures has become one of the most important fields in aerospace medicine. However, most of the previous studies were only related to the changs in morphology, pathophysiology and biochemistry, and the exact molecular mechanism of G-LOC remains unknown. Therefore, it is of great importance to explore the molecular basis of brain damage induced by repeated +Gz exposures. In this study, SD rats were used to establish the animal model of brain injury induced by repeated +Gz exposure and suppression subtractive hybridization technique was adopted to screen the differentially expressed genes in rat brains of +Gz exposure group. The aim of the study was to obtain preliminary experimental data for the molecular mechanisms of the brain injury. The main results and conclusions are as follows:1. Sprague-Dawley (SD) rats were used to establish the animal model of brain injury induced by repeated +Gz exposure. In this model, it was found that the main pathological changes of the exposed brains included lyses of Nissl substance in neurons of cerebral cortex, neuronal ischemic changes, white matter swelling, capillary hyperemia or ischemia.2. A cDNA subtractive library with high subtractive efficiency of repeated +Gz exposures in rat brain was constructed with suppression subtractivehybridization (SSH). The cDNA subtractive library after amplification included 100 blue clones and 400 white clones, 75 ones of which were selected to prepare for plasmid. Identification of the clones with restriction endonuclease cleavage showed most of them had been cloned to the vector.3. Seventy positive clones were identified from SSH pools. The obtained differential display fragments were screened by differential screen technique. Sequence analysis performed for 10 positive clones showed that three clones contained unknown genes and seven clones contained genes homologous with the genes published in GenBank.4. To further confirm the expression levels of those differentially expressed genes in rat brain of +Gz exposure group, the primers for two known genes (53# and 72#) and one unknown genes (58#) were designed for semi-quantitative reverse transcription polymerase chain reation (RT-PCR), using GAPDH as the reference. The result was photographed under UV light and analyzed with a quantification system. The ratio of differentially expressed genes /GAPDH was determined to eliminate gel -to-gel variance. The results from RT-PCR of the three differentially expressed genes were significantly higher than those of control group (P<0.01)In summary, with rapidness and effectiveness of SSH technique, the screened differentially expressed genes including the known and three novel ones in this study might play an important role in the pathogenesis of rat brain injury induced by repeated +Gz exposures.