發(fā)布時間：2018-06-17 14:08

  本文選題：脊髓損傷 + 異氟醚預(yù)處理　； 參考：《第四軍醫(yī)大學(xué)》2014年碩士論文

【摘要】：脊髓損傷（SCI）是一個具有嚴(yán)重破壞性而且病理生理比較復(fù)雜的臨床疾病，引起SCI的原因很多，其中主要原因為外傷，由于中樞神經(jīng)系統(tǒng)的不可再生，SCI帶來的損傷往往也是不可逆的。有研究表明SCI發(fā)生后，缺血是引起損傷加重的一個不可忽視的角色。脊髓損傷后存在兩個缺血區(qū)域，一個區(qū)域緊靠損傷位點(diǎn)，該區(qū)域大部分神經(jīng)元正在或者已經(jīng)壞死或者凋亡，屬于已經(jīng)不可挽救區(qū)域；另一個區(qū)域離損傷位點(diǎn)比較遠(yuǎn)，緊靠第一個損傷區(qū)，該區(qū)域的神經(jīng)元尚未發(fā)生凋亡或壞死，屬于可挽救區(qū)域，因此如果能及時挽救該區(qū)域處于早期變性的神經(jīng)元，就能很大程度的緩解脊髓損傷，從而促進(jìn)神經(jīng)功能的恢復(fù)[1]。異氟醚作為吸入麻醉藥的一種已經(jīng)廣泛用運(yùn)于臨床，我們實驗室的前期實驗已經(jīng)證實了異氟醚預(yù)處理可以明顯改善大腦局部缺血損傷時導(dǎo)致的神經(jīng)功能的破壞[2]，也有部分實驗證實異氟醚預(yù)處理可以改善脊髓缺血損傷導(dǎo)致的神經(jīng)功能損傷[3-4]。在臨床實際操作中，異氟醚預(yù)處理已經(jīng)用于保護(hù)心臟手術(shù)引起的心肌缺血性損傷[5]。然而異氟醚預(yù)處理是否可以緩解脊髓損傷導(dǎo)致的缺血損傷尚未報道，因此異氟醚預(yù)處理是否可以用于減輕SCI，有必要進(jìn)一步進(jìn)行研究和探討。 異氟醚預(yù)處理產(chǎn)生神經(jīng)保護(hù)作用所涉及的機(jī)制很多，離子通道的抑制，抑制凋亡，抑制炎癥反應(yīng)等。我們實驗室的前期實驗證實異氟醚預(yù)處理減輕腦缺血再灌注損傷是通過抑制TLR4一髓樣分化因子88(MyD88)信號通路，從而抑制炎癥和免疫應(yīng)答的級聯(lián)反應(yīng)。白介素-1β（IL-1β）作為白介素-1的主要組成部分在引起炎癥反應(yīng)中發(fā)揮重要作用，而IL-1β又是通過激活核轉(zhuǎn)錄因子（NF-κB）引起多種炎癥因子的激活引起炎癥反應(yīng)。 因此，本研究擬探討異氟醚預(yù)處理對大鼠SCI能否產(chǎn)生保護(hù)作用及其可能的機(jī)制是否與IL-1信號通路有關(guān)，為以后SCI的治療及麻醉提供實驗基礎(chǔ)。 目的： 觀察異氟醚預(yù)處理對大鼠SCI神經(jīng)行為學(xué)的變化、脊髓組織病理學(xué)的改變和脊髓組織IL-1β、NF-κB表達(dá)變化，探討異氟醚預(yù)處理對大鼠SCI產(chǎn)生保護(hù)作用及其可能的機(jī)制是否與IL-1信號通路有關(guān)，為SCI的治療提供理論基礎(chǔ)。 方法： 選取健康成年雄性SD大鼠30只，隨機(jī)分為3組（n=10）。假手術(shù)組（Sham組）：只暴露脊髓組織不做打擊損傷；損傷組（SCI組）：分離脊髓并進(jìn)行打擊損傷；異氟醚預(yù)處理組（ISO組）：吸入2%異氟醚2h，預(yù)處理結(jié)束后24h時做脊髓損傷模型。術(shù)后連續(xù)七天進(jìn)行行為學(xué)評分，觀察結(jié)束后取脊髓組織進(jìn)行HE染色計數(shù)存活神經(jīng)元數(shù)量。隨后再選取SD大鼠40只進(jìn)行第二部分實驗，隨機(jī)將大鼠分為4組（n=10）：假手術(shù)組（Sham組）：只暴露脊髓組織不做打擊損傷；損傷組（SCI組）：分離脊髓并進(jìn)行打擊損傷；異氟醚預(yù)處理組（ISO組）：吸入2%異氟醚2h，預(yù)處理結(jié)束后24h時做脊髓損傷模型:；IL-1抑制劑組（IL-Ra組）：先做脊髓損傷模型，用含2g/L白介素-1抑制劑明膠海綿放于損傷處2h。免疫組化染色觀察IL-1β的表達(dá)變化。通過Western-blot法觀察脊髓組織IL-1β、NF-κB的變化。 結(jié)果： （1）BBB評分結(jié)果顯示：隨著時間的推移SCI組和ISO組大鼠后肢功能逐漸恢復(fù)，，但是異氟醚預(yù)處理組BBB評分較單純損傷組增加更為明顯。7天后ISO組神經(jīng)行為學(xué)評分比SCI組明顯增高，差異有統(tǒng)計學(xué)意義（P0.05）。 （2）HE染色結(jié)果顯示：SCI組脊髓組織水腫，細(xì)胞核固縮。ISO組脊髓組織水腫減輕，存活細(xì)胞數(shù)量較多。存活神經(jīng)元計數(shù)，ISO組存活神經(jīng)元數(shù)量較SCI組明顯增高，差異具有統(tǒng)計學(xué)意義（P0.05）。 （3）免疫組織化學(xué)實驗結(jié)果顯示：本實驗主要觀察脊髓組織中IL-1β的表達(dá)量的變化，其中認(rèn)為胞漿、間質(zhì)被染成棕褐色為陽性。在Sham組中幾乎檢測不到IL-1β的表達(dá)。SCI組中可見IL-1β在細(xì)胞質(zhì)及胞外基質(zhì)中大量表達(dá)。而ISO組以及IL-1Ra組中細(xì)胞質(zhì)及間質(zhì)中IL-1β表達(dá)量減少。 （4）Western-blot結(jié)果顯示：分別在2h、6h、24h、48h取SCI組脊髓組織進(jìn)行IL-1β的蛋白測定，發(fā)現(xiàn)在6h時IL-1β的表達(dá)量最高。在Sham組中，IL-1β及NF-κB蛋白表達(dá)與其他組相比較少。SCI組內(nèi)IL-1β及NF-κB蛋白表達(dá)量最高，IL-1β及NF-κB蛋白表達(dá)在ISO組和IL-1Ra組表達(dá)明顯下降，其差異具有統(tǒng)計學(xué)意義（P0.05）。 結(jié)論： 異氟醚預(yù)處理能明顯降低脊髓損傷后神經(jīng)行為學(xué)障礙，增加損傷脊髓組織存活神經(jīng)元數(shù)量，說明異氟醚預(yù)處理對脊髓損傷產(chǎn)生保護(hù)作用。ISO組IL-1β及NF-κB蛋白表達(dá)量明顯降低，說明異氟醚預(yù)處理對脊髓產(chǎn)生保護(hù)作用是通過抑制IL-1信號通路產(chǎn)生作用的。
[Abstract]:Spinal cord injury ( SCI ) is a kind of clinical disease with severe damage and complicated pathological physiology , which causes many reasons . The main reason is trauma . Due to the non - regeneration of the central nervous system , the injury caused by SCI is often irreversible . There are two ischemic regions after SCI . One region is close to the injury site . Most of the neurons in the region are either already necrosis or apoptosis , which belong to the region which has not been saved ;
The other region is far from the injury site , close to the first lesion region , and the neurons in the region have not yet undergone apoptosis or necrosis , and belong to the salvage region , so that if the region in early degeneration can be saved in time , the spinal cord injury can be relieved to a large extent , thereby promoting the restoration of the neurological function . It has been proved that isoflurane preconditioning can improve the neurological deficit caused by ischemia injury in the brain , and some experiments prove that isoflurane preconditioning can improve the neurological deficit caused by ischemia injury in the brain . However , whether isoflurane preconditioning can alleviate ischemia injury caused by spinal cord injury has not been reported , and it is necessary to study and discuss whether isoflurane preconditioning can be used to alleviate SCI .

Interleukin - 1尾 ( IL - 1尾 ) plays an important role in the inflammatory response , and IL - 1尾 ( IL - 1尾 ) plays an important role in inflammatory response by inhibiting the activation of NF - 魏B . IL - 1尾 ( IL - 1尾 ) is an important component of IL - 1 . IL - 1尾 ( IL - 1尾 ) is an important component of IL - 1 .

Therefore , it is proposed to investigate whether the protective effects of isoflurane preconditioning on SCI in rats and possible mechanisms are related to IL - 1 signaling pathway , which provides an experimental basis for the treatment and anesthesia of SCI .

Purpose :

The changes of SCI neural behavior in rats , the changes of spinal cord tissue pathology and the expression of IL - 1尾 and NF - 魏B in the spinal cord were observed . The protective effects of isoflurane preconditioning on SCI in rats and possible mechanisms were discussed .

Method :

30 male SD rats were randomly divided into 3 groups ( n = 10 ) , sham operation group ( Sham group ) : only exposed spinal cord tissue did not fight injury ;
Injury group ( SCI group ) : spinal cord was isolated and injured ;
The rats were randomly divided into 4 groups ( n = 10 ) : sham operation group ( Sham group ) .
Injury group ( SCI group ) : spinal cord was isolated and injured ;
isoflurane preconditioning group ( iso group ) : inhalation of 2 % isoflurane for 2 h , spinal cord injury model at 24 h after pretreatment :
IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 , IL - 1尾 ,

Results :

( 1 ) BBB scores showed that the function of hindlimb of SCI group and ISO group gradually recovered over time , but the BBB score of isoflurane preconditioning group was significantly higher than that in SCI group ( P0.05 ) .

( 2 ) The results of HE staining showed that the spinal cord tissue edema and cell nuclear fixation in SCI group were significantly decreased and the number of viable cells was more . The number of viable neurons in ISO group was significantly higher than that in SCI group ( P0.05 ) .

( 3 ) The results of immunohistochemistry showed that the expression of IL - 1尾 in the spinal cord tissue was mainly observed in this experiment . The expression of IL - 1尾 in the cytoplasm and extracellular matrix was almost undetectable in the Sham group , but the expression of IL - 1尾 in the cytoplasm and stroma of the ISO group and the IL - 1Ra group was decreased .

( 4 ) Western - blot showed that the expression of IL - 1尾 and NF - 魏B in SCI group was the highest at 2h , 6h , 24h and 48h . In Sham group , the expression of IL - 1尾 and NF - 魏B in SCI group was the highest , and the expression of IL - 1尾 and NF - 魏B in SCI group was the highest , and the expression of IL - 1尾 and NF - 魏B was significantly decreased in the ISO group and IL - 1Ra group , and the difference was statistically significant ( P0.05 ) .

Conclusion :

The effect of isoflurane preconditioning on spinal cord injury was significantly decreased , and the protective effect of isoflurane preconditioning on spinal cord was decreased by inhibiting IL - 1 signaling pathway .
【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R614

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 陳銀海;劉敏;何井華;;脊髓損傷患者流行病學(xué)調(diào)查[J];實用醫(yī)學(xué)雜志;2011年06期

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