The Research Of Dog Nerve Cell Apoptosis In The Improvement Of Cervical Syringomyelia With Spinal Shortening And Spinal Cord Hypotension

Objective:Our previous study found continuous lower dynamic state of cerebrospinal fluid circulation and apoptosis of spinal cord neural cells play an important role in the pathological processes of scoliosis with cervical syringomyelia.Our clinical studies have also confirmed that shortening of the spine and decreased tension of the spinal cord by posterior vertebral column resection can stabilize or even shrink the syrinx on the basis of improving cerebrospinal fluid circulation.But the specific mechanism involved is not yet clear.Therefore,in order to clarify the molecular biological mechanism of improved syringomyelia through spinal shortening-spinal cord hypotension,this study,based on the establishment of dog model of cervical syringomyelia,intends to dynamically detect changes of apoptotic factors expression in the cerebrospinal fluid at various time points after spinal shortening-spinal cord hypotension.Methods:1.A total of 15 beagle dogs were randomly selected to establish cervical syringomyelia model by intraspinal injection of trace Kaolin at the 7th cervical vertebra segment.MRI was used to detect the formation of syringomyelia before and after modeling 6th weeks.to evaluate neurological function by Tarlov score.2.L1 vertebra body was resected by PVCR after modeling 6th weeks to create a short space for the spine,intraoperative real-time neural evoked potential monitoring.Based on the pedicle screw rod system,the short space was reducted quantitatively after resection of Ll vertebra body.The reduction distance was 1/3-1/2(1.0-1.5 cm)of the sum of the height of the L1 vertebral body and the upper and lower adjacent intervertebral discs.3.Cerebrospinal fluid sample was collected from the Cerebellomedullary cistern at four time points,which is pre-modeling,1st week after the modeling,and 1st week and 2nd weeks after the shortening of spine by PVCR.Real-time PCR was used to detect expression changes of apoptotic factors,including BAX、Bcl-2、AKT1、Caspase-3、Caspase-9、Cytochrome C(Cyt-C)in cerebrospinal fluid.The ELISA kit was used to measure the expression level of Caspase-3 and Cyt-C protein.Results:1.After modeling,3 cases of 15 dogs were died and 12 survived:8 of them had formed cervical syringomyelia,and the success rate of the model was 66.7%(8/12)at 6th week after modeling,MRI showed that syringomyelia was mainly located in theThe canine syringomyelia is mainly located in the cervical and cervical-thoracic junction spinal cord.The Tarlov scores of neurological function at 4th and 6th after modeling were 3.88±0.13 and 3.00±0.27,respectively.2.8 dogs have formed syringomyelia 6th week after modering,and 2 cases out of 8 dogs were died and 6 were survived after the spine was shortened by PVCR surgery.The success rate of surgery was 75%(6/8).Tarlov scores of neurological function at 1st and 2nd week after PVCR surgery were improved to 3.50 ± 0.19,3.75± 0.31 respectively 3.RT-PCR resultsa.Anti-apoptosis gene AKT-1,Bcl-2:The expression of AKT-1 and Bcl-2 was lower than that pre-modeling at 1w after modeling(P<0.05);the expression of AKT-1 and Bcl-2 increased after spine was increase after spine was shorted compared witih that in 1w after modeling(P<0.05),and the increase in 2nd week is more significant higher than that at 1st week after spine was shorted by PCVR(P<0.05).b.Pro-apoptotic gene BAX and Caspase-3:The expression of BAX and Caspase-3 was higher than that pre-modeling at 1w after modeling(P<0.05);the expression of BAX and Caspase-3 at 2nd week after spine was shorted by PVCR was lower compared with that at 1w after modeling(P<0.05),decline at 2nd week is more significant lower than at 1st week after spine was shorted by PCVR(P<0.05).the expression of BAX and Caspase-3 at 1st week after spine was shorted by PCVR was slightly lower than that at 1w after modeling(P>0.05).c.Mitochondrial pro-apoptotic gene Caspase-9 and Cyt-C:the expression of Caspase-9 and Cyt-C was higher than that pre-modeling at 1w after modeling(P<0.05);the expression of them after spine was shorted by PCVR was lower compared with that at 1w after modeling(P<0.05),decline in 2nd week is more significant lower than that atlst week after spine was shorted by PCVR(P<0.05).4.ELISA detection of protein levels of pro-apoptotic proteins Cyt-C and Caspase-3The expression of Cyt-C、Caspase-3 was higher than that pre-modeling at 1w after modeling(P<0.05);the expression of them after spine was shorted by PCVR was lower compared with that at 1w after modeling(P<0.05),decline at 2nd week is more significant lower than that at 1st week after spine was shorted by PCVR(P<0.05).Conclusion:1.apoptosis-related factors BAX,Bcl-2,AKT1,Caspase-3,Caspase-9,and Cyt-C participated in the pathological process of Kaolin-induced Cervical syringomyelia formation2.After the formation of cervical syringomyelia in dog,with spinal shortening-spinal cord hypotension by PVCR and up-regulating the expression of anti-apoptosis gene and down-regulating the expression of pro-apoptotic genes,it can inhibit the apoptosis,and may inhibit the apoptosis of spinal cord neural cells through this mechanism,and gain syringomyelia improvement.3.Apoptosis-related factor AKT in cerebrospinal fluid may be used as a biomarker to reflect the pathological process of syringomyelia,providing reference for treatment and prognosis.