發(fā)布時(shí)間：2018-09-08 13:41

【摘要】：目的:青光眼已經(jīng)成為世界上主要的致盲眼病之一，其典型表現(xiàn)是視乳頭的凹陷性萎縮與視野的特征性缺損和縮小。目前關(guān)于青光眼視神經(jīng)萎縮的發(fā)病機(jī)制還不十分清楚，傳統(tǒng)上有兩種學(xué)說(shuō)：機(jī)械壓力學(xué)說(shuō)和血管缺血學(xué)說(shuō)。機(jī)械壓迫學(xué)說(shuō)認(rèn)為眼壓作用于篩板直接壓迫視神經(jīng)纖維，阻礙了軸漿流的運(yùn)輸，軸突蛋白的生成和移動(dòng)減少，導(dǎo)致細(xì)胞代謝受損；血管缺血學(xué)說(shuō)強(qiáng)調(diào)視神經(jīng)纖維由于缺血缺氧及失去周?chē)M織的保護(hù)而發(fā)生損害。目前認(rèn)為是機(jī)械壓迫和血供障礙共同參與了青光眼視神經(jīng)的損害，而眼壓的升高進(jìn)一步加重視網(wǎng)膜神經(jīng)血管的調(diào)節(jié)障礙，促成了視神經(jīng)的損害。阻止視神經(jīng)損害、保護(hù)視功能的方法目前主要是通過(guò)藥物和手術(shù)的方法降低眼壓以減緩壓力對(duì)視神經(jīng)的損害，，已有明顯的效果，這一途徑得到共識(shí)。然而，單純降眼壓并不能完全制止青光眼視神經(jīng)的損害，仍有一部分患者視野和視神經(jīng)在不斷受損。因此，人們?cè)絹?lái)越關(guān)注加強(qiáng)視神經(jīng)保護(hù)的治療，找到一種能保護(hù)視神經(jīng)的藥物，阻斷和/或延緩神經(jīng)節(jié)細(xì)胞原發(fā)性或繼發(fā)性損傷，已成為當(dāng)今青光眼研究的焦點(diǎn)和熱點(diǎn)。 視網(wǎng)膜神經(jīng)節(jié)細(xì)胞（retinal ganglion cells RGCS）的凋亡是一種高度調(diào)控的細(xì)胞死亡，是青光眼視神經(jīng)損傷的最終通路。本實(shí)驗(yàn)通過(guò)制作高眼壓大鼠模型，探討黃芪多糖對(duì)大鼠急性高眼壓狀態(tài)下視網(wǎng)膜神經(jīng)節(jié)細(xì)胞的保護(hù)作用，從而揭示具有活血化瘀功效的中藥制劑在改善微循環(huán)，提高機(jī)體抗氧化能力，改善青光眼視神經(jīng)軸漿流等方面具有明顯的效果，為臨床青光眼視神經(jīng)保護(hù)的治療提供實(shí)驗(yàn)基礎(chǔ)和理論依據(jù)。 方法：選用大連醫(yī)科大學(xué)實(shí)驗(yàn)動(dòng)物中心供給的SD雌性大鼠60只（其中已剔除實(shí)驗(yàn)過(guò)程中死亡3只，丟失1只），隨機(jī)分為4組，正常對(duì)照組（C組）、高眼壓模型對(duì)照組（M組）、低濃度黃芪多糖治療組（N組）、高濃度黃芪多糖治療組（H組），每組各15只。各組大鼠每日實(shí)驗(yàn)處理如下：C組和M組給予2.5ml生理鹽水灌胃，N組給予黃芪多糖500mg/kg（體重）2.5ml灌胃，H組給予黃芪多糖2000mg/kg（體重）2.5ml灌胃。用藥2周后，除正常對(duì)照組，其余三組均雙眼前房注射甲基纖維素方法制作急性高眼壓模型，模型確定成功后的第5天過(guò)量麻醉處死，取眼球做石蠟切片，行HE染色觀察視網(wǎng)膜形態(tài)結(jié)構(gòu)變化，免疫組化檢測(cè)caspase-3表達(dá)，TUNEL(TdT-mediated dUTP nick-end labeling)染色觀察節(jié)細(xì)胞凋亡。利用Image Pro Plus5.1軟件測(cè)量視網(wǎng)膜內(nèi)外顆粒層厚度、神經(jīng)纖維層厚度，計(jì)數(shù)陽(yáng)性染色的節(jié)細(xì)胞和凋亡節(jié)細(xì)胞所占節(jié)細(xì)胞總數(shù)百分比，使用SPSS16.0統(tǒng)計(jì)軟件包采用單因素方差分析(ANOVA)，組間兩兩比較采用t檢驗(yàn)進(jìn)行統(tǒng)計(jì)學(xué)分析，以P＜0.05為差異具有統(tǒng)計(jì)學(xué)意義。 結(jié)果：C組大鼠視網(wǎng)膜各層組織結(jié)構(gòu)清晰，RGCs排列密集，細(xì)胞核邊界清楚，caspase-3極少表達(dá)。M組大鼠視網(wǎng)膜全層、內(nèi)外顆粒層及神經(jīng)纖維層均高度水腫，RGCs排列疏松，細(xì)胞核腫脹，caspase-3在視網(wǎng)膜節(jié)細(xì)胞層和內(nèi)顆粒層均有大量表達(dá)。N組大鼠視網(wǎng)膜各層組織結(jié)構(gòu)清晰，節(jié)細(xì)胞整齊密集，caspase-3在視網(wǎng)膜節(jié)細(xì)胞層表達(dá)顯著減少，N組較M組差異有統(tǒng)計(jì)學(xué)意義（P＜0.001）。H組大鼠視網(wǎng)膜內(nèi)外顆粒層細(xì)胞基質(zhì)仍然水腫，神經(jīng)纖維層厚度較M組并無(wú)明顯好轉(zhuǎn)，H組較M組差異無(wú)統(tǒng)計(jì)學(xué)意義（P＞0.05），caspase-3陽(yáng)性細(xì)胞數(shù)明顯減少，RGCs凋亡率降低，H組較M組差異有統(tǒng)計(jì)學(xué)意義（P＜0.05、P＜0.001）。 結(jié)論：1、高眼壓可以導(dǎo)致視網(wǎng)膜全層水腫、RGCs排列疏松、數(shù)目減少、細(xì)胞核腫脹、神經(jīng)節(jié)細(xì)胞發(fā)生調(diào)亡。2、一定濃度的黃芪多糖對(duì)視網(wǎng)膜神經(jīng)節(jié)細(xì)胞有顯著的保護(hù)作用。3、高濃度的黃芪多糖雖然能夠部分改善視網(wǎng)膜神經(jīng)節(jié)細(xì)胞的凋亡，但應(yīng)當(dāng)注意其毒性作用的損害和影響。
[Abstract]:Objective: Glaucoma has become one of the leading blindness diseases in the world. Its typical manifestation is the concave atrophy of the optic papilla and the characteristic defect and narrowing of the visual field. The theory of vascular ischemia emphasizes that the optic nerve fibers are damaged by ischemia and hypoxia and loss of protection of the surrounding tissues. It is also involved in the damage of optic nerve in glaucoma, and the increase of intraocular pressure further aggravates the disturbance of retinal nerve and blood vessel regulation and contributes to the damage of optic nerve. However, simple hypotension does not completely stop the damage of the optic nerve in glaucoma. There are still some patients whose visual field and optic nerve are constantly damaged. Secondary injury has become the focus and hotspot of glaucoma research.
The apoptosis of retinal ganglion cells (RGCS) is a highly regulated cell death and the ultimate pathway of glaucoma optic nerve injury. In this study, the protective effect of Astragalus Polysaccharide on retinal ganglion cells in rats with acute ocular hypertension was investigated by making a rat model of ocular hypertension. The traditional Chinese medicine preparation of blood stasis removing effect has obvious effect on improving microcirculation, antioxidant capacity and axoplasmic flow of optic nerve in glaucoma, which provides experimental and theoretical basis for the treatment of clinical glaucoma optic nerve protection.
Methods: Sixty female SD rats supplied by Experimental Animal Center of Dalian Medical University were randomly divided into 4 groups: normal control group (group C), high intraocular pressure model control group (group M), low concentration astragalus polysaccharide treatment group (group N) and high concentration astragalus polysaccharide treatment group (group H). Rats in group C and group M were given 2.5 ml normal saline, group N 500 mg/kg (body weight) 2.5 ml Astragalus polysaccharide, group H 2000 mg/kg (body weight) 2.5 ml Astragalus polysaccharide. After 2 weeks of administration, except for the normal control group, the other three groups were injected methylcellulose into the anterior chamber of both eyes to make acute ocular hypertension model. On the 5th day after the operation, the rats were sacrificed under excessive anesthesia, and the eyeballs were taken for paraffin section. The morphological and structural changes of retina were observed by HE staining, the expression of Caspase-3 was detected by immunohistochemistry, and the apoptosis of ganglion cells was observed by TUNEL staining. The positive staining ganglion cells and apoptotic ganglion cells accounted for hundreds of fractions of ganglion cells. SPSS16.0 statistical software package was used for single factor analysis of variance (ANOVA).
Results: In group C, the structure of retinal layers was clear, RGCs were densely arranged, the nuclear boundaries were clear, and caspase-3 was rarely expressed. In group M, the retinal layers were highly edematous, the RGCs were loosely arranged, the nuclei were swollen, and caspase-3 was highly expressed in ganglion cells and inner granular layers. The expression of Caspase-3 in ganglion cell layer of retina was significantly decreased in group N compared with group M (P The number of Caspase-3 positive cells and the apoptosis rate of RGCs in group H were significantly lower than those in group M (P < 0.05, P < 0.001).
Conclusion: 1. High intraocular pressure can lead to retinal edema, loose RGCs arrangement, reduced number, nuclear swelling, ganglion cell apoptosis. 2. A certain concentration of Astragalus polysaccharide has a significant protective effect on retinal ganglion cells. 3. High concentration of Astragalus polysaccharide can partly improve the apoptosis of retinal ganglion cells, but should be. Attention should be paid to the damage and effects of its toxic effects.
【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R774.1;R775

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相關(guān)期刊論文 前10條

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本文編號(hào)：2230681