| The incidence of traumatic brain injury was increased yearly with the development of the economy and transportation. Its mortality rate and mutilation rate were also enhanced. The statistics showed, in Chinese cities, patients dying from craniocerebral injury had amounted to 60% of all dead from the trauma. Recently, traumatic brain injury occurred more and more frequently, and has become the first one of most factors that lead to the death in the less than 45-aged individuals. Up to now, surgical management is most pivotal to craniocerebral injury, and, however medication still is very important. Therefore, any kind of effective remedy is very necessary.Recent researches showed that apoptosis was found and involved in secondary cell death in the neurodegenerative diseases, Alzheimer's deasease(AD), epilepticus, cerebral ischemia, spinal cord injury and traumatic brain injury. A crucial role for caspase in neuronal apoptosis has been documented through the use of specific inhibitors of cystein proteases in vivo or in vitro. Thus apoptosis may represent a therapeutic target for recovery after those central nerve diseases.Caspase-3, also named as cysteine protease P32(CPP32), act as most importantprotease in the programmed cell death. Post-translational activation of caspase-3 requires proteolytic cleavage of the precursor protein, and generated into two subunits(pl7 and p11). This process leads to enzyme(poly(ADP-ribose) polymerase(PARP), et al) activation and initialization of apoptosis.In this experiment, we evaluated the effect of a specific caspase-3 inhibitor, z-DEVD-fmk, by intracerebroventricular microinjection on the extent of apoptotic cell analysed by FCM, brain water and nitro oxygen content, following traumatic brain injury, a protocol of modified Feeney free dropping objective mode.Materials and methods36 Sprague-Dawley rats were divided randomly into sham operated group, NS treated group and Caspase-3 inhibitor treated group, each 12. There was no significant difference among the average values of body weight in 3 groups by ANOVA test (P>0.005).Animals were anesthetized with intraperitoneal injection of ketam ine(40mg/kg) and placed in a stereotaxic fram. The scalp was listerized with iodine and 75% alcohol and incised on the midline. An atraumatic craniectomy was performed by removing the right parietal bone posterior to bregma, 2mm lateral to the sagittal, and 2mm posterior to the coronary suture, to from a window about 4mm in diameter. An stainless needle connected with a micropump was penetrated into the right lateral ventricle at the coordinate: Ap 0.8mm, Lat 1.5mm, Dep 4.5mm in the same craniectomy window. 30 minutes before trauma, the caspase-3 inhibitor treated mice were applied with z-DEVD-fmk (a specific caspase-3 inhibitor, 5ug/10ul in 5 minutes), NS treated mice with 0.9%NaCl (10ul in 5 minutes), infused into the right ventricle through the needle. After infusion, the needle stayed for 10 minutes. A piston which was 3 mm in diameter, and had excursion of 3 mm was then placed over the craniectomy window. A 50g weight was droped from a height of 16cm on tothe piston. 30 minutes after trauma, the mice were applied with z-DEVD-fmk or 0.9%NaCl again at the same coordinate in the same way. The sham operated mice were only dealt with craniotomy.72 houres after the trauma, the rat's brain was removed and sectioned into coronal slices(2mm), under ice-cold circumstance. A slice contain injury field was fixed by 10% formalin, and sent to HE stain. A mass of brain cortex tissue obtained beyond 1mm from the injuried field, about 1mm3, was fixed by 2.5% glutaraldehyde, and sent to electron microscope examination. Another brain tissue about 15mg taken beyond the injured field, was disaggregated into single-cell suspension, and apoptosis was proved by FCM. One brain slice was stored in a refrigeratory(-80 ) TO investigate the brain NO(nitro oxygen) content after traumatic brain injury.Measurement of brain water contents after traumatic brain injury. After wet weighted,... |
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