Effect Of Electro-acupuncture On The Expression Of Cyt-c And Caspase-9 Protein In Brain Tissue Of TBI Rats

Objective:To observe the effects of electro-acupuncture on the neurological function,Brain tissue morphology,Cytochrome C(Cyt-C)and Caspase-9 protein expression in rats with traumatic brain injury(TBI),and to explore the possible mechanism of electro-acupuncture promoting neurological recovery,reducing brain cell apoptosis and treating brain injury through the expression of apoptosis proteins Cyt-C and Caspase-9.Method:The model of moderate craniocerebral injury was prepared by Feeney's free fall attack method.Seventy SD rats were Randomly divided into blank group(8 rats),sham-operation group(8 rats),model group(24 rats)and electro-acupuncture group(24 rats).Select acupuncture points in rats: Renzhong(GV26),Baihui(GV20),Neiguan(PC6),and Zusanli(ST36),all treated with electro-acupuncture.After the model was established,the rats in the electro-acupuncture group were treated once a day for 14 days every morning.The remaining groups did not have intervention.Rats in each group were assessed by Modified Neurological Severity Score(m NSS)scores at three time points on days 3,7 and 14 for brain function;Eight rats were sacrificed in the model group and the electro-acupuncture group at three time points on days 3,7,and 14 and specimens were collected.Eight rats were sacrificed on the 14 th day in the blank group and the sham-operation group,and the specimens were collected.Morphological changes of injured brain tissue in each group were observed by HE staining and Nissl staining.Td T-mediated nick end labeling(TUNEL)method was used to detect the apoptosis of injured brain tissue.Immunohistochemistry and Western blot were used to detect the expression of Cyt-C and Caspase-9 in the cortex of the injured area.Result:(1)m NSS score: The scores of the model group and the electro-acupuncture group were significantly higher than the sham-operation group,which was statistically significant(P<0.01).The scores of the model group and the electro-acupuncture group reached the peak on the 3rd day,and decreased with time.The decline trend of the electro-acupuncture group was more significant.The electro-acupuncture group scored lower on the 3rd,7th,and 14 th days than the model group,which was statistically significant(P< 0.05);(2)HE staining: In the pathological section,the cell bodies of the blank group and the sham-operation group were generally observed to be full and the nuclear membrane was clear.On the third day of the model group,a large amount of tissue necrosis around the damaged area was observed,and the cells were arranged in disorder,vacuolar-like changes,deformation,nuclear fragmentation,nuclear pyknosis,etc,but the electro-acupuncture group had a smaller necrotic range than the model group.In the 7th and 14 th day sections,both the model group and the electro-acupuncture group showed more nuclear pyknosis and new connective tissue.However,the damage and repair of the electro-acupuncture group is better than that of the model group.(3)Nissl staining: Nissl distribution in the blank group and the sham-operation group was relatively uniform,and no obvious neuron loss.After the modeling,the number of Nissl bodies in the 3d model group and the electro-acupuncture group was significantly reduced,and then gradually increased with time.The number of Nissl bodies in the electro-acupuncture group on day 3,7 and 14 was higher than that in the simultaneous model group.(4)TUNEL: There are only a few apoptotic nerve cells in the blank group and the sham-operation group.On the 3rd day after model establishment,the number of neuronal apoptosis in the model group and the electro-acupuncture group was significantly increased.The number of apoptosis in the two groups decreased from the 7th to the 14 th day.The number of neuronal apoptosis in the model group and the electro-acupuncture group was significantly higher than that in the sham-operation group or the blank group on the 14 th day(P<0.01),but the number of neuronal apoptosis in the electro-acupuncture group was significantly lower than that in the model group at the 3rd,7th,and 14 th day(P< 0.05).(5)The expression of Cyt-C and Caspase-9 protein was detected by immunohistochemistry and Western blot: In the model group and the electro-acupuncture group,their peaks were reached on the third day,and they all showed a downward trend with time,which is similar to the change of neuronal apoptosis.The expression of Cyt-C in electro-acupuncture group or the model group was higher than that in sham-operation group or blank group at 14 days after injury(P<0.01),but the expression of Cyt-C in electro-acupuncture group was less than that in the model group at 3,7 and 14 days after injury(P<0.05);The expression of Caspase-9 was consistent with that of Cyt-C.The expression of Caspase-9 in electroacupuncture group was less than that in the model group at 3,7 and 14 days after injury(P<0.05).Conclusion:(1)Electro-acupuncture can improve the neurological function of TBI rats;(2)Electro-acupuncture can reduce the level of neuronal apoptosis in damaged brain tissue of TBI rats;(3)Electro-acupuncture may reduce the expression of proapoptotic proteins Cyt-C and Caspase-9 in brain tissue,and alleviate apoptosis in brain tissue regions damaged by TBI,thus promoting the recovery of neurological function after TBI.