中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2012年
12期
1080-1084
,共5页
葛薇%徐哲%刘升强%王朝晖%邵彦%韩浩%李静敏
葛薇%徐哲%劉升彊%王朝暉%邵彥%韓浩%李靜敏
갈미%서철%류승강%왕조휘%소언%한호%리정민
黄芪多糖%高眼压%视网膜神经节细胞%凋亡
黃芪多糖%高眼壓%視網膜神經節細胞%凋亡
황기다당%고안압%시망막신경절세포%조망
Astragalus polysaccharides%Ocular hypertension%Retinal ganglion cell%Apoptosis
背景 青光眼患者视神经保护的问题日益引起关注.黄芪多糖(APS)是黄芪的主要活性成分,可增加再生神经蛋白的表达并促进损伤的周围神经修复,但其对视网膜神经节细胞(RGCs)再生作用的研究少见报道. 目的 探讨APS对急性高眼压状态下RGCs的保护作用.方法 采用抽签法将40只SPF级SD大鼠随机分为正常对照组、模型对照组、低剂量APS组和高剂量APS组,每组各10只.低剂量APS组和高剂量APS组大鼠自实验开始每日分别给予APS 500 mg/kg、2000 mg/kg(均溶于2.5 ml生理盐水)灌胃,模型对照组仅给予2.5ml生理盐水灌胃,正常对照组不做任何处理.用药2周后,除正常对照组外,其余3个组均抽取0.2ml房水继而单眼前房注射等体积甲基纤维素使眼压升高至22 mmHg(1 mmHg=0.133 kPa)以上制作急性高眼压模型.造模后5d,过量麻醉法处死动物,摘除该眼球制作视网膜石蜡切片,常规组织病理学观察视网膜的形态结构变化,采用免疫组织化学法检测caspase-3蛋白在大鼠视网膜中的表达,采用TUNEL染色法观察和计算各组大鼠RGCs的凋亡率.Image Pro Plus 5.1软件测量各组大鼠视网膜厚度和神经纤维层厚度. 结果 大鼠成模后5d,模型对照组、低剂量APS组和高剂量APS组大鼠的眼压均明显高于正常对照组,差异均有统计学意义(t=-8.900、-10.700、-11.300,P＜0.01).正常对照组大鼠视网膜形态正常；模型对照组大鼠视网膜水肿,细胞排列紊乱；低剂量APS组视网膜可见空泡样变性,但视网膜细胞排列较模型对照组整齐,视网膜水肿减轻；高剂量APS组视网膜水肿较明显.低剂量APS组视网膜厚度、外颗粒层及视神经纤维层厚度值均明显低于模型对照组,差异均有统计学意义(t=-23.700、-14.770、-11.640,P＜0.0l),但高剂量APS组外颗粒层及视神经纤维层厚度与模型对照组比较差异均无统计学意义(t=-0.780、-0.460,P＞0.05).低剂量APS组大鼠caspase-3蛋白阳性RGCs百分比及RGCs凋亡百分率均明显低于模型对照组(caspase-3蛋白:F=87.710,P=0.001；RGCs凋亡:F=272.840,P＜0.01),差异均有统计学意义(t=-11.700、-8.600,P＜0.01),高剂量APS组与模型对照组比较caspase-3蛋白阳性RGCs百分比及RGCs凋亡百分率的差异均有统计学意义(t=-7.900、-6.400,P＜0.05). 结论 500 mg/kg APS可有效抑制急性高眼压模型大鼠的视网膜水肿及RGCs的凋亡率,对急性高眼压大鼠的RGCs有保护作用.
揹景 青光眼患者視神經保護的問題日益引起關註.黃芪多糖(APS)是黃芪的主要活性成分,可增加再生神經蛋白的錶達併促進損傷的週圍神經脩複,但其對視網膜神經節細胞(RGCs)再生作用的研究少見報道. 目的 探討APS對急性高眼壓狀態下RGCs的保護作用.方法 採用抽籤法將40隻SPF級SD大鼠隨機分為正常對照組、模型對照組、低劑量APS組和高劑量APS組,每組各10隻.低劑量APS組和高劑量APS組大鼠自實驗開始每日分彆給予APS 500 mg/kg、2000 mg/kg(均溶于2.5 ml生理鹽水)灌胃,模型對照組僅給予2.5ml生理鹽水灌胃,正常對照組不做任何處理.用藥2週後,除正常對照組外,其餘3箇組均抽取0.2ml房水繼而單眼前房註射等體積甲基纖維素使眼壓升高至22 mmHg(1 mmHg=0.133 kPa)以上製作急性高眼壓模型.造模後5d,過量痳醉法處死動物,摘除該眼毬製作視網膜石蠟切片,常規組織病理學觀察視網膜的形態結構變化,採用免疫組織化學法檢測caspase-3蛋白在大鼠視網膜中的錶達,採用TUNEL染色法觀察和計算各組大鼠RGCs的凋亡率.Image Pro Plus 5.1軟件測量各組大鼠視網膜厚度和神經纖維層厚度. 結果 大鼠成模後5d,模型對照組、低劑量APS組和高劑量APS組大鼠的眼壓均明顯高于正常對照組,差異均有統計學意義(t=-8.900、-10.700、-11.300,P＜0.01).正常對照組大鼠視網膜形態正常；模型對照組大鼠視網膜水腫,細胞排列紊亂；低劑量APS組視網膜可見空泡樣變性,但視網膜細胞排列較模型對照組整齊,視網膜水腫減輕；高劑量APS組視網膜水腫較明顯.低劑量APS組視網膜厚度、外顆粒層及視神經纖維層厚度值均明顯低于模型對照組,差異均有統計學意義(t=-23.700、-14.770、-11.640,P＜0.0l),但高劑量APS組外顆粒層及視神經纖維層厚度與模型對照組比較差異均無統計學意義(t=-0.780、-0.460,P＞0.05).低劑量APS組大鼠caspase-3蛋白暘性RGCs百分比及RGCs凋亡百分率均明顯低于模型對照組(caspase-3蛋白:F=87.710,P=0.001；RGCs凋亡:F=272.840,P＜0.01),差異均有統計學意義(t=-11.700、-8.600,P＜0.01),高劑量APS組與模型對照組比較caspase-3蛋白暘性RGCs百分比及RGCs凋亡百分率的差異均有統計學意義(t=-7.900、-6.400,P＜0.05). 結論 500 mg/kg APS可有效抑製急性高眼壓模型大鼠的視網膜水腫及RGCs的凋亡率,對急性高眼壓大鼠的RGCs有保護作用.
배경 청광안환자시신경보호적문제일익인기관주.황기다당(APS)시황기적주요활성성분,가증가재생신경단백적표체병촉진손상적주위신경수복,단기대시망막신경절세포(RGCs)재생작용적연구소견보도. 목적 탐토APS대급성고안압상태하RGCs적보호작용.방법 채용추첨법장40지SPF급SD대서수궤분위정상대조조、모형대조조、저제량APS조화고제량APS조,매조각10지.저제량APS조화고제량APS조대서자실험개시매일분별급여APS 500 mg/kg、2000 mg/kg(균용우2.5 ml생리염수)관위,모형대조조부급여2.5ml생리염수관위,정상대조조불주임하처리.용약2주후,제정상대조조외,기여3개조균추취0.2ml방수계이단안전방주사등체적갑기섬유소사안압승고지22 mmHg(1 mmHg=0.133 kPa)이상제작급성고안압모형.조모후5d,과량마취법처사동물,적제해안구제작시망막석사절편,상규조직병이학관찰시망막적형태결구변화,채용면역조직화학법검측caspase-3단백재대서시망막중적표체,채용TUNEL염색법관찰화계산각조대서RGCs적조망솔.Image Pro Plus 5.1연건측량각조대서시망막후도화신경섬유층후도. 결과 대서성모후5d,모형대조조、저제량APS조화고제량APS조대서적안압균명현고우정상대조조,차이균유통계학의의(t=-8.900、-10.700、-11.300,P＜0.01).정상대조조대서시망막형태정상；모형대조조대서시망막수종,세포배렬문란；저제량APS조시망막가견공포양변성,단시망막세포배렬교모형대조조정제,시망막수종감경；고제량APS조시망막수종교명현.저제량APS조시망막후도、외과립층급시신경섬유층후도치균명현저우모형대조조,차이균유통계학의의(t=-23.700、-14.770、-11.640,P＜0.0l),단고제량APS조외과립층급시신경섬유층후도여모형대조조비교차이균무통계학의의(t=-0.780、-0.460,P＞0.05).저제량APS조대서caspase-3단백양성RGCs백분비급RGCs조망백분솔균명현저우모형대조조(caspase-3단백:F=87.710,P=0.001；RGCs조망:F=272.840,P＜0.01),차이균유통계학의의(t=-11.700、-8.600,P＜0.01),고제량APS조여모형대조조비교caspase-3단백양성RGCs백분비급RGCs조망백분솔적차이균유통계학의의(t=-7.900、-6.400,P＜0.05). 결론 500 mg/kg APS가유효억제급성고안압모형대서적시망막수종급RGCs적조망솔,대급성고안압대서적RGCs유보호작용.
Background More efforts have been made in the functional protection of the glaucoma ganglion cells (RGCs) nowadays.As main ingredient,astragalus polysaccharides (APS) enhances neuron regeneration protein expression and promotes peripheral nerve recovery.But whether APS has a protecting effect on RGCs is incompletely clear.Objective The purpose of this study was to evaluate the neuroprotective effect of APS on the RGCs in a rat model of experimental glaucoma.Methods Forty-four SPF SD rats were divided into 4 groups randomly as follows:normal control group,negative control group,low dose APS group and high dose APS group,with 10 rats for each group.APS of 500 mg/kg or 2000 mg/kg (2.5 ml) was administered by gavage feeding once daily for 2 weeks in low dose or high dose of APS group,respectively,and the same volume of normal saline solution was applied instead of APS in the model control group.Two weeks later,aspirate 0.2 ml aqueous followed by methylcellulose injected into the anterior chamber to create the acute ocular hypertension model in the three groups above.No any intervention was performed in the normal control group.The rats were sacrificed on the fifth day after model established to take a retinal section.Ocular hypertension-induced damage was evaluated by regular retina histopathologic examination.Immunolhistochemistry for caspase-3 and TUNEL kits were used to determine the expression of caspase-3 protein in retina and apoptosis rate of RGCs.Retinal cross-sections were analyzed by Image Pro Plus 5.1 software to determine the thickness of various retinal layers and the positive staining cell density in the retinal ganglion cell layer (RGCL).Results On the fifth day after establishment of models,intraocular pressure (IOP) was significantly elevated in the model control group,low dose APS group and high dose APS group in comparison with the normal control group (t=-8.900,-10.700,-11.300,P＜0.01).Retinal morphology was normal in the rats of the normal control group,but in the model control group,rat retina was significantly thickened from severe retinal edema and cell arrangement disorder.Mild retinal abnormality was seen in the low dose APS group;while obvious retina edema was in high dose APS group.The entire retinal thickness,outer nuclear layer thickness and retinal nerve fiber thickness values were lower in the low dose APS group than those of model control group (t =-23.700,-14.770,-11.640,P＜0.01).However,no difference was found in outer nuclear layer thickness and retinal nerve fiber thickness values between high dose APS group and normal control group (t =-0.780,-0.460,P ＞ 0.05).Percentage of positive RGCs for caspase-3 protein and rate of apoptotic RGCs were significantly reduced in low dose APS group compared with model control group (caspase-3:F=87.710,P=0.001;RGCs apoptosis:F=272.840,P＜0.01).Conclusions 500 mg/kg APS can protect retina and RGCs against ocular hypertension-induced damage.The protection of APS is non-dosedependent.
