【作者】 王文宏；

【导师】 迟焕芳；

【作者基本信息】 青岛大学 ， 生理学， 2011， 博士

【摘要】 目的建立单眼视觉剥夺性弱视大鼠动物模型,研究枸杞子提取物(LBA)对视觉剥夺性弱视大鼠视网膜神经节细胞(RGCs)是否具有保护作用。方法新生Wistar大鼠30只,随机分成生理盐水组(10只)、左旋多巴组(10只)及LBA组(10只)。出生1周后,应用褥式缝合法缝合所有幼鼠左侧眼睑,建立单眼视觉剥夺性弱视动物模型,三组每日分别经口给予等量的NS (0.9% NaCl)、L-dopa(20mg/kg)及LBA(10mg/kg),喂养满6周。取材前5天将30g/L荧光金注射到所有大鼠双侧的上丘和外侧膝状体,通过荧光金逆行标记观察视网膜铺片中RGCs形态和数量的变化。结果①视网膜铺片中标记的RGCs为黄白色,胞体形态多样,可见荧光充盈的细胞突起。②NS组、L-dopa组及LBA组中左眼的RGCs密度明显低于右眼,差异均有显著性(P<0.01)。③与NS组相比,L-dopa组及LBA组左眼RGCs密度均明显升高(P<0.01),而L-dopa组及LBA组之间差异无显著性(P>0.05)。④L-dopa组及LBA组RGCs的存活率分别为85.17%±4.42%和84.31%±5.97%,均明显高于NS组71.35%±6.34%(P<0.01),而L-dopa组及LBA组之间差异无显著性(P>0.05)。结论①在剥夺性弱视大鼠视觉发育可塑性的关键期,LBA可有效地保护RGCs,提高RGCs存活率。②LBA对RGCs的视觉保护作用与治疗弱视的常规用药左旋多巴效果接近。目的建立单眼剥夺性弱视大鼠动物模型,研究视觉发育敏感期不同阶段枸杞子提取物(LBA)对视皮层17区Bcl-2和NGF表达的影响。方法新生Wistar大鼠60只,随机分成正常组(12只)、生理盐水组(24只)、LBA组(24只)。出生7天后,应用褥式缝合法缝合NS组和LBA组幼鼠左侧眼睑,建立单眼视觉剥夺性弱视大鼠模型。NS组及LBA组每日分别经口给予等量的0.9%NaCl及10mg/kg LBA,正常组自由进食,于生后14天、21天、28天、35天、42天、70天分别取材制片,采用Nissl染色测量视皮层17区神经元的数目和截面积,采用SABC免疫组化法检测视皮层17区相应区域Bcl-2和NGF的表达。结果①Nissl染色显示剥夺性弱视大鼠右侧视皮层分层欠清晰,神经元排列相对疏松,细胞形态及突起不规则,Nissl小体表达有所减少,可见胞核缩小、着色加深。②LBA组大鼠生后28天至70天,右侧视皮层神经元的数量和截面积明显同龄NS组大鼠的对应区域,差异有显著性(P<0.05)。③LBA组大鼠生后21天至70天右侧视皮层Bcl-2阳性细胞的数量和OD值高于同龄NS组并低于正常组大鼠的对应区域,差异有显著性(P<0.05)。④LBA组大鼠生后42天及70天,右侧视皮层17区NGF阳性细胞的数目和OD值高于同龄NS组大鼠的对应区域,差异有显著性(P<0.05)。结论①大鼠视觉发育敏感期,剥夺性弱视可导致视皮层神经元细胞数目和截面积低于正常组。②LBA对剥夺性弱视大鼠在视觉发育可塑性的关键期内具有一定保护作用,可作为弱视治疗的临床备选药物。③LBA对剥夺性弱视大鼠视觉发育的保护机制可能与上调Bcl-2及NGF的表达有关,以促进神经细胞的营养和生长。更多还原

【Abstract】 Objective:To construct the rat models of monocular visual deprived amblyopia, to investigate the possible neuroprotection of LBA on the retinal ganglion cells (RGCs) in rats with visual deprived amblyopia.Methods:A total of 30 postnatal Wistar rats were randomly divided into Normal Saline group (10), L-dopa group (10) and LBA group (10). After postnatal 1 week, the left palpebras of all rats was sewed by mattress suture in order to construct the models. Each day three groups were respectively fed with the same amount of 0.9% NaCl,20mg/kg L-dopa and 10mg/kg LBA up to 6 weeks. RGCs were retrograde labeled by injection of 30g/L fluorogold (FG) into bilateral superior colliculi and lateral geniculate body 5 days before the death in all rats, then the RGCs changes of morphology and numbers in retinal flat were observed.Results:①The labeled RGCs in retinal flats were yellow-white with various morphology, the cellular prominence with fluorescence could be seen.②RGCs density of left eyes in the three groups were obviously lower than right eyes, the differences were significant (P<0.01).③Compared with NS group, RGCs density of left eyes were obviously higher in L-dopa and LBA group (P<0.01), but there was no significance between L-dopa and LBA group (P>0.05).④The survival rates of RGCs in L-dopa group (85.17%±4.42%) and LBA group (84.31%±5.97%) were obviously higher than NS group (71.35%±6.34%, P<0.01), but there was no significance between L-dopa group and LBA group (P>0.05).Conclusion:①LBA might have neuroprotection on RGCs in rats with deprived amblyopia during the critical periods of visual plasticity and then increase the survival rates of RGCs.②The visual neuroprotection of LBA on RGCs is similar to L-dopa, which is used as conventional medicine in amblyopia therapy. Objective:To construct the rat model with monocular deprived amblyopia, to investigate the effects of LBA on Bcl-2 and NGF expression in visual cortex area 17 in the different sensitive periods of visual development.Methods:60 postnatal Wistar rats were randomly divided into normal group (12), normal saline group (24), LBA group (24). One week after left eyelid mattress sulture in NS group and LBA group, rats were respectively fed with the same amount of 0.9% NaCl and 10mg/kg LBA each day, while normal group were fed freely. In postnatal 14d,21d,28d, 35d,42d and 70d, rats were executed. Tissue sections were used to measure the neuron numbers and sectional areas in visual cortex area 17 by Nissl staining, then detect the corresponding expression of Bcl-2 and NGF by SABC immunohistochemistry.Results:①Nissl staining showed unsharp layering of the right visual cortex in deprived amblyopia rats, loose arrangement, irregular morphology and prominence, reduction of Nissl bodies, apperance of pyknosis and dark staining.②The neuron numbers and sectional areas in right visual cortex of LBA group were obviously higher than the corresponding area in the evenaged NS group from P28 to P70 (P<0.05).③The positive cell numbers of Bcl-2 and optical density (OD) in right visual cortex of LBA group were obviously higher than the evenaged NS group from P21 to P70 (P<0.05).④The positive cell numbers of NGF and OD in right visual cortex of LBA group were higher than the evenaged NS group from P14 to P28 (P<0.05).Conclusion:①In the sensitive stage of visual development, deprived amblyopia in rats might reduce the cell numbers and sectional areas in visual cortex, which were lower than normal group.②LBA might have neuroprotection during the critical period of visual plasticity in deprived amblyopia rats, and be used as an alternative drug for amblyopia treatment.③The neuroprotective mechanism of LBA on visual development in deprived amblyopia rats might be related with up-regulating the expression of Bcl-2 and NGF, which could improve nutrition and growth of the nerve cells.更多还原