【摘要】：背景創(chuàng)傷性顱腦損傷(traumatic brain injury,TBI)在神經(jīng)外科疾病中非常常見,顱腦損傷后啟動一系列復(fù)雜的神經(jīng)化學(xué)信號變化導(dǎo)致的病理事件包括神經(jīng)元過度活躍,過多的谷氨酸釋放,炎癥,血腦屏障(blood-brain barrier,BBB)通透性增加和腦水腫。顱腦損傷后基因表達的改變和一系列神經(jīng)功能損傷對患者的預(yù)后產(chǎn)生極大的影響,帶來身體和經(jīng)濟的雙重壓力。目前,并沒有確切的藥物和治療方法來應(yīng)對對顱腦損傷后的繼發(fā)損傷。目前研究的重點主要致力于顱腦損傷后對神經(jīng)細(xì)胞的保護以及炎癥反應(yīng)對腦組織的損傷。丙戊酸(valproic acid,VPA)一種組蛋白去乙�；敢种苿┮恢北挥糜谂R床,它的神經(jīng)保護作用是多方面的。本研究利用大鼠腦損傷打擊器建立閉合性大鼠腦損傷模型,并給予丙戊酸干預(yù),通過對不同時間點的大鼠進行行為學(xué)評價及磁共振成像、乙�；M蛋白H3、NF-κB表達的變化,探討VPA的抗炎機制,為臨床應(yīng)用VPA治療TBI提供可靠依據(jù)。目的通過觀察VPA對TBI大鼠各時間點腦組織乙�；M蛋白H3、NF-κB蛋白表達的影響,探討丙戊酸對實驗性腦損傷大鼠的減輕炎性反應(yīng)的作用。方法按照隨機的原則把75只雄性SD大鼠分為3組:假手術(shù)組(C組)、損傷組(TBI組)和丙戊酸治療組(VPA組)。TBI和VPA組建立大鼠閉合性損傷模型,C組在相同部位僅切開頭皮,縫合。VPA治療組術(shù)后每天按300mg/kg腹腔注射VPA,至處死當(dāng)天;單純損傷組和假手術(shù)組在相同時間腹腔注射等體積的生理鹽水。傷后各時間點對大鼠進行行為學(xué)評價。傷后6 h、24 h、48 h和72 h每組取5只處死取材,免疫熒光雙標(biāo)法在激光共聚焦顯微鏡下觀察核因子κB(NF-κB)途徑的激活狀態(tài),免疫組化檢測乙�；M蛋白H3含量,并在傷后72 h每組取一只大鼠腹腔注射水合氯醛,待麻醉完全后于北京國家納米中心做磁共振成像。結(jié)果1.行為學(xué)評分結(jié)果:在給予VPA干預(yù)后,VPA組大鼠和TBI組大鼠相比損傷程度有所減輕,在傷后24 h內(nèi)效果不明顯,3組間差異無統(tǒng)計學(xué)意義(P0.05);VPA干預(yù)后7 d、14 d,VPA治療組大鼠的運動、感覺及反射功能均優(yōu)于TBI組,兩組間差異有統(tǒng)計學(xué)意義(P0.05)。2.7.0T小動物超導(dǎo)磁共振成像顯示:單純損傷組和正常對照組相比損傷灶顏色接近紅色,說明水腫程度較重,其中損傷灶中心為黃色,說明水腫程度最重,VPA治療組和單純損傷組相比損傷灶顏色接近藍色,說明在應(yīng)用VPA治療后大鼠損傷腦組織水腫有所減輕。從圖中還可以看出,VPA治療組損傷灶腦組織體積比相同時間點單純損傷組明顯減小,說明VPA能減少腦損傷體積。3.免疫組織化學(xué)檢測顯示:3組大鼠乙�；癄顟B(tài)組蛋白H3表達水平差異均有統(tǒng)計學(xué)意義(P0.05),VPA組和C組相比無明顯差異,TBI組明顯低于C組(P0.05)。4.免疫熒光技術(shù)檢測:C組大鼠腦組織切片NF-κB核陽性細(xì)胞各個時間點表達很少,甚至不表達,與C組相比,傷后6 h TBI組和VPA組大鼠腦組織切片NF-κB核陽性細(xì)胞數(shù)量高于C組,24 h的大鼠腦組織切片中NF-κB核陽性細(xì)胞數(shù)明顯高于C組,48 h和72 h NF-κB核陽性細(xì)胞表達開始逐漸減少,但仍比C組表達高(P0.05);VPA治療組NF-κB核陽性細(xì)胞各時間點表達量均低于同時間點TBI組(P0.05)。結(jié)論丙戊酸對大鼠腦損傷后的抗炎作用可能是通過抑制NF-κB途經(jīng)的激活來實現(xiàn)的。
[Abstract]:Background Traumatic brain injury (TBI) is very common in neurosurgical diseases. The pathological events caused by a series of complex neurochemical signal changes after brain injury include hyperactive neurons, excessive glutamate release, inflammation, blood-brain barrier (blood-brain barrier), BBB) increased permeability and brain edema. The change of gene expression after head injury and a series of nerve function damage have a great effect on the prognosis of the patient, bringing the double pressure of the body and the economy. At present, there is no definite drug and treatment method to deal with the secondary injury after brain injury. The research focuses on the protection of nerve cells after brain injury and the damage of the inflammatory response to the brain tissue. Valproic acid (VPA), a histone deethanizing enzyme inhibitor, has been used in clinical and its neuroprotective effects are various. In this study, the model of brain injury of closed rats was established by using the rat brain injury killer, and the intervention of valproic acid was given, and the anti-inflammatory mechanism of VPA was discussed by the behavior evaluation of rats at different time points and the changes of the expression of the magnetic resonance imaging, the histone H3 and NF-EMAB. Provide reliable basis for clinical application of VPA in the treatment of TBI. Objective To study the effect of VPA on the expression of histone H3 and NF-B protein in the brain tissue of TBI rats, and to study the effect of valproic acid on the inflammatory response of rats with experimental brain injury. Methods 75 male SD rats were divided into three groups according to the random principle: sham operation group (group C), injury group (TBI group) and valproic acid treatment group (VPA group). The closed injury model of rats was established in the TBI and VPA groups. The VPA was injected intraperitoneally at 300 mg/ kg every day after the treatment of the VPA group, and the same time as that of the sham-operated group and the sham-operation group, the volume of saline was injected in the same time. The behavior of rats was evaluated at all time points after injury. At 6 h,24 h,48 h and 72 h after injury,5 rats were sacrificed, and the activation state of the nuclear factor B (NF-B) pathway was observed under the laser confocal microscope, and the content of the histone H3 was detected by immunohistochemistry. At 72 hours after injury, a rat was injected with water-chloral hydrate in each group, and the magnetic resonance imaging was performed in the national nano-center of Beijing after the anesthesia. Results 1. Results: After the VPA intervention, the degree of injury of the rats in the VPA group and the TBI group was reduced, the effect was not obvious in 24 hours after the injury, the difference between the groups was not significant (P0.05), and the rats in the treatment group of the VPA group were treated with the VPA intervention for 7 days,14 days, and the VPA treatment group. The sensory and reflective function was superior to that of the TBI group, and the difference between the two groups was statistically significant (P0.05). The degree of edema was the most, and the color of the lesion in the treatment group of the VPA and the simple injury group was close to the blue, indicating that the edema of the brain tissue of the rats after the treatment with VPA was reduced. It can also be seen from the figure that the volume of brain tissue in the damage range of the VPA treatment group is significantly reduced than that of the same time point, indicating that the VPA can reduce the volume of the brain injury. The results showed that there was no significant difference in the level of histone H3 in group 3 rats (P0.05), and there was no significant difference in the group of VPA and Group C, and the group of TBI was lower than that of group C (P0.05). The results showed that the expression of NF-B-B-positive cells in the brain tissue of the C-group was little or not even expressed, compared with that of the C group. The number of NF-and B-positive cells in the brain tissue of the rats at 6 h TBI and VPA was higher than that of the C group, and the number of NF-and B-positive cells in the brain tissue of the rats in 24 h was significantly higher than that of the C group, and the expression of NF-B-B-positive cells in the brain tissue of group C,48 h and 72 h was gradually decreased. However, the expression of NF-B-B-positive cells in the treatment group of VPA was lower than that of the TBI group at the same time point (P0.05). Conclusion The anti-inflammatory effect of valproic acid on rat brain injury may be achieved by inhibiting the activation of NF-B.
【學(xué)位授予單位】：新鄉(xiāng)醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R651.15

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