| Spinal cord injury(SCI)resulting in sensory loss,motor weakness,and autonomic dysfunction.In autonomic dysfunction,intestinal dysfunction seriously affect the quality of life of the patients,is considered to be one of the most needed priority and urgent to solve.Previous studies have shown that intestinal dysfunction afte spinal cord injury are related to the changes of substance P,however,whether there is a change of its receptor is not clear.In addition,interstitial cells of Cajal(ICC)as a gastrointestinal slow wave pacemaker cells,closely related to gastrointestinal electrical activity spread and regulation mediated neurotransmission,but whether it is involved in intestinal dysfunction afte spinal cord injury is no report.Therefore,through animal experiments to study the changes of the substance P and its receptor NK1 R and ICC in intestinal dysfunction afte spinal cord injury is necessary,based on the successful establishment of spinal cord injury model,observe the pathological changes of spinal cord and colon tissue in rats with spinal cord injury,study the change of substance P and its receptor in spinal cord and colon tissues after spinal cord injury.Detection of colonic bioelectricity after spinal cord injury and the changes of ICC,Further discussion the mechanism of intestinal dysfunction after spinal cord injury.Methods:1.Establishment of spinal cord injury model.We established the model by modified Allen device,with 10g·25cm strength against T10 Spinal dura mater.2.Behavior detection.BBB scores were used to observe the rats’ behavioral changes,the rats with BBB score of "0-1" were selected to experiment.3.General observation.Compared with sham-operated group,the changes of the colon after spinal cord injury were observed with naked eye.4.Pathological study.Compared with normal group and sham-operated group,EC staining were performed to spinal cord and HE staining were performed to proximal colon tissue to observe the pathological changes in the injury group.5.Detection of Bioelectric activity.Compared with the normal group and sham-operated group,recording the chang of proximal colon bio-electrical with bio-electric recorder in the injury group at 1、2、4week.6.Detection of ICC.Compared with the normal group and sham-operated group,the changes of ICC in colonic tissues were detected by immunofluorescence and Western Blot in the injury group at 1、2、4week.7.Detection of substance P and its receptor NK1 R.Compared with the normal group and sham-operated group,By immunofluorescence,Western Blot was used to detect the change of substance P and its receptor NK1 R in spinal cord and colon tissue in the injury group at 1、2、4week.Main results1.Behavioral detection:BBB score: the BBB score of rats after spinal cord injury was 0-1,which indicated that both lower limbs were completely paralyzed after spinal cord injury.2.General observation.The Colon tissue was hyperemia and edema,lots of dry feces in intestinal cavity after spinal cord injury.3.Pathological observation:(1)EC staining:In normal and sham-operation group,medullary sheath of white matter in spinal cord was evenly distributed and arranged closely;In the injury group,medullary sheath of white matter in spinal cord was obviously demyelination and the local cavity was formed.(2)HE staining:In normal and sham-operation group,the proximal colon epithelial cells arranged neatly;In the injury group,villus epithelial cells shedding,lodging,mucous layer and muscular layer thickness increased.4.Detection of Bioelectric activity.In the normal group and sham-operated group,Bio-electrical recording indicated that the biological activity was regular;Compared with controls,bio-electrical recording disorder,amplitude variation is larger,and the waveform is not stable in the proximal colon of the injury group.5.Detection of ICC.The number of ICC positive cell in the colon tissue was 57.38±4.69 in the injury group at 1 week,less than the normal group and sham-operated group(P < 0.01);The number of ICC positive cell in the colon tissue was 32.26±3.62 in the injury group at 2 week,compared with the normal group and sham operated group,it was significantly reduced(P < 0.01);The number of ICC positive cell in the colon tissue was 25.07±3.05 in the injury group at 4 week,significantly less than the normal group and sham-operated group(P < 0.01).The expression of c-kit protein in the colon tissue was decreased in the injury group,Among them,the expression of c-kit protein was 0.409±0.117 in the injury group at 4 week,compared with the normal group and sham operated group,it was significantly reduced(P < 0.01).6.Detection of substance P.The content of substance P in the anterior horn of upper segment of spinal cord injury was 0.158±0.060 in the injury group at 1 week,compared with the normal group,sham operated group,it have on obvious difference(P>0.05);The content of substance P in the anterior horn of upper segment of spinal cord injury was 0.323±0.098 in the injury group at 2 week,increased significantly than the normal group,sham operated group(P<0.01);The content of substance P in the anterior horn of upper segment of spinal cord injury was 0.199±0.048,compared with the normal group,sham operated group,it have on obvious difference(P>0.05).The content of P in the anterior horn of lower segment of spinal cord injury was 0.090±0.009 in the injury group at 1 week,compared with the normal group,sham operated group,the content of P was decreased(P < 0.05);The content of P in the anterior horn of lower segment of spinal cord injury was 0.280 ±0.027 in the injury group at 2 week,significantly higher than the normal group,sham-operated group(P < 0.01);The content of P in the anterior horn of spinal cord injury was 0.129±0.022 in the injury group at 4 week,compared with the normal group,sham operated group,it have on obvious difference(P>0.05).The content of P in the colon tissue was 0.239±0.091 in the injury group at 1 week,compared with the normal group,sham operated group,the content of P was decreased(P < 0.05);The content of P in the colon tissue was 0.233±0.056 in the injury group at 2 week,compared with the normal group and sham operated group,it was significantly reduced(P < 0.01);The content of P in the colon tissue was 0.206±0.041 in the injury group at 4 week,significantly less than the normal group and sham-operated group(P < 0.01).7.Detection of NK1 R.After spinal cord injury,the changes of NK1 R expression in spinal cord showed a trend of decreasing and then increasing in the injury group.Among them,the expression of NK1 R was lowest in the injury group at 1 day,it was 0.978±0.414;the expression of NK1 R was 1.771±0.685 in the injury group at 4 week,compared with the normal group,sham operated group,the difference was statistically significant(P<0.05).Conclusions1.Lots of dry feces in intestinal cavity in spinal cord injured rats;2.Medullary sheath of white matter in spinal cord was obviously demyelination and the local cavity was formed in spinal cord injured rats;3.Villus epithelial cells shedding,lodging,mucous layer and muscular layer thickness increased in spinal cord injured rats;4.Bio-electrical were changed in colon of rats with spinal cord injury;5.The number of ICC and the expression of c-kit protein were decreased in spinal cord injured rats;6.Substance P and its receptor NK1 R in the anterior horn of the spinal cord,the dorsal horn and the colon were changed obviously in spinal cord injured rats. |
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