The Role Of Calcium Sensitive Receptor In Neuron Apoptosis After Traumatic Brain Injury

Objective: calcium sensitive receptor(calcium-sensingreceptor,CaSR)is a key protein in calcium regulatory pathway and involved in traumatic craniocerebral injury(Traumaticbraininjury,TBI),but its relationship with neuronal apoptosis after TBI has not been fully elucidated.Therefore,this study will establish TBI models,determine the expression of key molecules in CaSR and apoptosis pathway and analyze their correlation,and further explore the relationship between CaSR expression and neuronal apoptosis,so as to provide new ideas and targets for neuronal apoptosis after TBI.Methods: six groups of animal models were established: blank control group,sham operation group,TBI group,TBI+NS group,TBI+NPS2143 group(CaSR specific blocker),TBI+NPSR568 group(CaSR agonist),HE staining was used to identify the success of TBI model,improved neurobehavioral score was used to score the neurobehavioral score of the above six groups,and EB staining was used to detect the damage degree of blood-brain barrier in the above six groups.The degree of brain edema in the above six groups was detected by dry and wet weight method,and the expression of CaSR and key molecules of apoptotic pathway in apoptotic neurons in the above six groups were detected by RT-PCR and Westernblot.Results: 1.After successful modeling,naked eye observation showed that the morphology and structure of brain tissue in the blank control group were intact,while a large number of fresh hemorrhage and obvious contusion and laceration of brain tissue could be seen in the TBI group.After modeling,HE staining showed that the brain tissue structure was intact and the cells were arranged neatly in the blank control group;sparse brain tissue,subarachnoid hemorrhage,inflammatory cell infiltration,obvious vacuoles around the cells,nuclear pyknosis and deep staining could be seen in the TBI group;sparse brain tissue,subarachnoid hemorrhage,inflammatory cell infiltration,obvious vacuoles around the cells,nuclear pyknosis and deep staining could be seen in the TBI+NPS2143 group,but the above phenomena were improved compared with the TBI group.two.The neurobehavioral score of TBI group and TBI+NS group was higher than that of blank control group and sham operation group.The neurobehavioral score of TBI+NPSR568(agonist)group was higher than that of TBI group,while the neurobehavioral score of TBI+NPS2143(inhibitor)group was lower than that of TBI group.3.Compared with the blank control group,there was no significant change in the blood-brain barrier permeability in the sham operation group,but the blood-brain barrier permeability increased in TBI group,TBI+NS group,TBI+NPS2143 group and TBI+NPSR568 group,and the brain water content in 4.TBI group and TBI+NS group was higher than that in the blank control group and sham operation group.The brain water content in TBI+NPSR568 group was higher than that in TBI group,while the brain water content in TBI+NPS2143 group was lower than that in TBI group.The gene and protein expression levels of CaSR and Caspase-3 were detected by RT-PCR and Westernblotting.The results showed that the expression of CaSR and Caspase-3 in TBI group and TBI+NS group was higher than that in blank control group,the expression of CaSR and Caspase-3 in TBI+NPS2143 model group was lower than that in TBI group,and the expression of CaSR and Caspase-3 in TBI+NPSR568 model group was higher than that in TBI group,and the difference was statistically significant.6.The gene and protein expression levels of endoplasmic reticulum stress pathway related protein Caspase-12 were detected by RT-PCR and Westernblotting.The results showed that the expression of Caspase-12 in TBI model group was higher than that in blank control group,the difference was statistically significant;the expression of Caspase-12 in TBI+NPS2143 model group was lower than that in TBI group,and the expression of Caspase-12 in TBI+NPSR568 model group was higher than that in TBI group,and the difference was statistically significant.7.The expression levels of Caspase-8 and Caspase-10 genes and proteins related to death receptor pathway were detected by RT-PCR and Westernblotting.The results showed that the expression of Caspase-8 and Caspase-10 in the death receptor pathway in the TBI model group was significantly higher than that in the blank control group,while the expression of Caspase-8 and Caspase-10 in the TBI+NPS2143 model group was significantly lower than that in the TBI group.The expression of Caspase-8 and Caspase-10,which are the key molecules in the death receptor pathway in the TBI+NPSR568 model group,was significantly higher than that in the TBI group.8.The gene and protein expression levels of Cyt C,Caspase-9 and BCL-2 in mitochondrial pathway were detected by RT-PCR and Westernblotting.The results showed that the expression of Cyt C and Caspase-9,which are the key molecules in the mitochondrial pathway in the TBI model group,was significantly higher than that in the blank control group,while the expression of BCL-2 was lower than that in the blank control group,and there was statistical significance between the two groups.The expression of Cyt C and Caspase-9 in the mitochondrial pathway in the TBI+NPS2143 model group was significantly lower than that in the TBI group,while the expression of BCL-2 was higher than that in the TBI group,and there was statistical significance between the two groups;the expression of the key molecules in the mitochondrial pathway in the TBI+NPSR568 model group was significantly higher than that in the TBI group,while the expression of BCL-2 was lower than that in the TBI group,and there was statistical significance between the two groups.Conclusion: CaSR regulates neuronal apoptosis secondary to traumatic brain injury through three Caspase-related apoptosis pathways(mitochondrial pathway,death receptor pathway and endoplasmic reticulum pathway).