【摘要】：研究背景 腦出血是指原發(fā)性非外傷性腦實(shí)質(zhì)出血,也稱自發(fā)性腦出血,是臨床上常見(jiàn)的血管性疾病,發(fā)病率、死亡率和致殘率均較高。除了血腫的占位效應(yīng)及對(duì)周圍組織的直接破壞之外,繼發(fā)性腦損傷也是出血后腦損傷的主要原因,包括出血后血腫擴(kuò)大壓迫腦組織,引起腦組織移位和損傷,繼發(fā)性炎性反應(yīng),產(chǎn)生的凝血素引起的凝血連鎖反應(yīng),紅細(xì)胞溶解產(chǎn)生的血紅蛋白所誘導(dǎo)的毒性反應(yīng),補(bǔ)體級(jí)聯(lián)反應(yīng)以及神經(jīng)元凋亡等方面。因此,腦出血的及時(shí)治療十分關(guān)鍵。微創(chuàng)血腫抽吸術(shù)一直是腦出血治療的研究熱點(diǎn),近年來(lái)有關(guān)干細(xì)胞移植治療腦出血的實(shí)驗(yàn)研究表明,干細(xì)胞進(jìn)入大鼠腦組織中,替代了神經(jīng)元的功能,減輕腦出血后的炎性反應(yīng),并促進(jìn)神經(jīng)功能的恢復(fù),此方法可能會(huì)成為臨床治療腦出血的新方法。目前,仍未見(jiàn)臍帶間充質(zhì)干細(xì)胞(HUC-MSCs)移植聯(lián)合微創(chuàng)血腫抽吸術(shù)治療腦出血的實(shí)驗(yàn)研究。本實(shí)驗(yàn)觀察了HUC-MSCs移植聯(lián)合微創(chuàng)血腫抽吸術(shù)對(duì)大鼠腦出血的療效,旨在為臨床進(jìn)一步應(yīng)用干細(xì)胞移植治療腦出血提供臨床實(shí)驗(yàn)基礎(chǔ)。 研究目的 本實(shí)驗(yàn)在建立大鼠腦出血模型的基礎(chǔ)上,探討HUC-MSCs移植聯(lián)合微創(chuàng)血腫抽吸術(shù)治療大鼠腦出血后腦內(nèi)P53的表達(dá)及神經(jīng)功能的恢復(fù)。 研究方法 體外分離培養(yǎng)HUC-MSCs,移植前以BrdU標(biāo)記,采用免疫細(xì)胞化學(xué)染色方法鑒定HUC-MSCs.將造模成功的120只SD大鼠隨機(jī)分為腦出血組、血腫抽吸組、細(xì)胞移植組、細(xì)胞移植聯(lián)合血腫抽吸組(聯(lián)合治療組),各組又分為1、3、7、14和30d5個(gè)時(shí)間點(diǎn)。向尾狀核注入2μL Ⅳ型膠原酶建立腦出血模型,6h后將符合標(biāo)準(zhǔn)的模型,進(jìn)行相應(yīng)的處理后,各組分別于各時(shí)間點(diǎn)處死大鼠前,用改良的大鼠神經(jīng)功能缺損評(píng)分(mNSS)評(píng)價(jià)大鼠的神經(jīng)功能狀況,后采用免疫組織化學(xué)方法檢測(cè)大鼠腦出血周圍P53基因的表達(dá)及HUC-MSCs在腦組織的分布,并進(jìn)行統(tǒng)計(jì)學(xué)處理。 研究結(jié)果 1. HUC-MSCs細(xì)胞中CD29表達(dá)陽(yáng)性,細(xì)胞漿為棕褐色；CD34表達(dá)陰性,細(xì)胞漿為藍(lán)色；CD44表達(dá)陽(yáng)性,細(xì)胞膜為棕褐色；CD90表達(dá)陽(yáng)性,細(xì)胞漿為棕褐色；CD105表達(dá)陽(yáng)性,細(xì)胞漿為棕褐色。腦出血組和血腫抽吸組未發(fā)現(xiàn)BrdU表達(dá)的陽(yáng)性細(xì)胞,細(xì)胞移植組和聯(lián)合治療組則于出血腦區(qū)、海馬及大腦皮質(zhì)等部位發(fā)現(xiàn)較多的陽(yáng)性細(xì)胞。P53在血腫側(cè)腦組織的胞核內(nèi)廣泛表達(dá),集中在血腫周圍紋狀體區(qū)、皮質(zhì)、皮質(zhì)下及海馬區(qū)等部位。2.與腦出血組比較,血腫抽吸組、細(xì)胞移植組和聯(lián)合治療組的mNSS評(píng)分、P53表達(dá)量均下降(P＜0.05),并且7d時(shí),聯(lián)合治療組的mNSS評(píng)分及P53表達(dá)量低于腦出血組和血腫抽吸組(P＜0.05),在14-30d時(shí)則低于其余3組(P＜0.05)。 研究結(jié)論 HUC-MSCs移植聯(lián)合微創(chuàng)血腫抽吸術(shù)治療可以顯著減少大鼠腦出血后神經(jīng)元的損傷,更利于神經(jīng)功能的恢復(fù)。
[Abstract]:Background intracerebral hemorrhage refers to primary non-traumatic intracerebral hemorrhage, also known as spontaneous intracerebral hemorrhage, which is a common vascular disease in clinic. The morbidity, mortality and disability rate are high. In addition to the space-occupying effect of hematoma and the direct destruction of surrounding tissues, secondary brain injury is also the main cause of brain injury after hemorrhage, including the hematoma after hemorrhage expands and compresses brain tissue, causes brain tissue displacement and injury, and secondary inflammatory reaction. The coagulation chain reaction induced by hemagglutinin, the toxic reaction induced by hemoglobin induced by erythrocyte lysis, the complement cascade reaction and neuron apoptosis. Therefore, the timely treatment of cerebral hemorrhage is very important. Minimally invasive hematoma aspiration has been a hot topic in the treatment of intracerebral hemorrhage. In recent years, experimental studies on stem cell transplantation for intracerebral hemorrhage have shown that stem cells enter the brain tissue of rats and replace the function of neurons. It may be a new method for the treatment of intracerebral hemorrhage. At present, there is no experimental study of umbilical cord mesenchymal stem cell (HUC-MSCs) transplantation combined with minimally invasive hematoma aspiration in the treatment of intracerebral hemorrhage. The effect of HUC-MSCs transplantation combined with minimally invasive hematoma aspiration on intracerebral hemorrhage in rats was observed in order to provide a clinical experimental basis for the further application of stem cell transplantation in the treatment of intracerebral hemorrhage. Objective to investigate the expression of p53 and the recovery of neural function after HUC-MSCs transplantation combined with minimally invasive hematoma aspiration in the treatment of intracerebral hemorrhage in rats. Methods HUC-MSCs, was isolated and cultured in vitro and labeled with BrdU before transplantation. HUC-MSCs. was identified by immunocytochemical staining. 120 SD rats were randomly divided into intracerebral hemorrhage group, hematoma aspiration group and cell transplantation combined with hematoma aspiration group (combined treatment group). Intracerebral hemorrhage model was established by injecting 2 渭 L type 鈪，

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