Alteration And Significance Of Protein Expression Pattern In Rats Following Mechanical Brain Injury

Objective: To study the alteration of protein expression pattern in rats serum and hippocampus after closed head injury and to find biomarkers of brain injury. Methods: Male Sprague-Dawley rats were randomly divided into sham operation group and injury groups which were subjected to Marmarou's brain injury and then subdivided into 4 h,8 h,12 h,24 h and 48 h groups according to the time elapsed after injury. The neurological status of the rats were evaluated pre-injury, 1 h, 12 h, 24 h and 48 h after injury. The pathological study included H&E stain, cresyl violet stain and esterification-silver stain. The change of protein expression pattern in serum and hippocampus were monitored with weak cationic exchanger chips and immobilized metal affinity capture chips by surface-enhanced laser desorption ionization-time of flight-mass spectrometry. Results: 1. Clinical changes: All rats of injury groups had no skull fracture. Immediately after brain injury, 70% of rats generalized convulsions with apnea for 15 seconds. 9.17% of rats developed convulsions without apnea for 30 seconds. 14.3% of rats weakened quadruped motor function. 2. Neurologic evaluation: The ability of balance,reflex,raising head,walking and escaping was much weakened in rats of injury groups. The neurological severity scores (NSS) were reduced at 1 h and 12 h after injury (P<0.01 compared with pre-injury). There was no significant difference in the NSS between pre-injury and at 24 h or 48 h after injury. 3. Macroscopic findings: A massive and extensive subarachnoid hemorrhage could be found in all injured rats and covered the entire brain. There were no contusions in brain tissues. 4. Microscopic findings: Two types of the injured neurons could be identified by H.E stain, one is characterized by darkness and shrinkage of the nuclei, the other one is characterized by the swell nuclei. The pathologic changes in cresyl violet stain or esterification-silver stain were similar to those observed with H.E stain, and the injured neurons and nerve fibers were more obvious. 5. Changes of protein expression patterns: (1) A total of 436 protein peaks were detected on WCX-2 chips and a total of 229 protein peaks were detected on IMAC-Cu chips in serum. A total of 364 protein peaks were detected on WCX-2 chips and a total of 345 protein peaks were detected on IMAC-Cu chips in hippocampus. (2) Among the total of protein peaks detected, the relative intensity of 53 protein peaks was shown to be differentially altered on WCX-2 chips in serum between injury groups and sham operation group(P<0.05). The relative intensity of 16 protein peaks was shown to be differentially altered on IMAC-Cu chips in serum between injury groups andsham operation group(P<0.05). There were 53 protein peaks in serum of injury groups which couldn't be detected in sham operation group with WCX-2 chips. There were 19 protein peaks in serum of injury groups which couldn't be detected in sham operation group with IMAC-Cu chips. (3) The relative intensity of 37 protein peaks was shown to be differentially altered on WCX-2 chips in hippocampus between injury groups and sham operation group(P<0.05). The relative intensity of 12 protein peaks was shown to be differentially altered on IMAC-Cu chips in hippocampus between injury groups and sham operation group(P<0.05). There were 102 protein peaks in serum of injury groups which couldn't be detected in sham operation group with WCX-2 chips. There were 29 protein peaks in serum of injury groups which couldn't be detected in sham operation group with IMAC-Cu chips. (4) On WCX-2 chips, 13 protein peaks were detected not only in serum but also in hippocampus. On IMAC-Cu chips, 10 protein peaks were detected both in serum and in hippocampus. (5) On WCX-2 chips, 6 protein peaks were detected at two time-points in serum and 18 protein peaks were detected at more than two time-points in hippocampus. On IMAC-Cu chips, 2 protein peaks were detected at more than two time-points in hippocampus. (6) Compared with sham operation groups, there were 122 pro...