The elucidation of molecular mechanism in delayed neruonal death and its control in rat retina
大鼠视网膜迟发性神经元死亡的分子机制及其调控的阐明
基本信息
- 批准号:10307042
- 负责人:
- 金额:$ 18.88万
- 依托单位:
- 依托单位国家:日本
- 项目类别:Grant-in-Aid for Scientific Research (A)
- 财政年份:1998
- 资助国家:日本
- 起止时间:1998 至 1999
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
We evaluated the involvement of apoptosis and contributions of Bcl-2 family genes in delayed neruonal death after retinal ischemia in Sprague-Dawley rat. Retinal ischemia was induced by elevating IOP to 130 mmHg for 45 min. Histological specimens were obtained at various time after reperfusion and examined by TUNEL staining. A significant number of TUNEL positive cells were observed in the ganglion cell layer (GCL) and inner nuclear layer (INL) starting at 6 to 48 hr after transient ischemia and reached a maximum 24 hr after ischemia. DNA was extracted from the-ischemic retina at 24 and 48 hr after reperfusion and the contralateral non-ischemic retina and electrophoresed. DNA laddering was observed on agarose gel electrophoresis in retinas 24 and 48 hr after ischemia but not in normal retina. RT-PCR analysis was carried out with the retina at various time after transient ischemia using specific primers for Bcl-2 and Bax. It demonstrated that Bax gene expression gradually increased as e … More arly as 6 hr, reached a peak at 24 hr, then decreased to near the baseline level at 168 hr, while Bcl-2 gene expression did not show any obvious change at any time point after transient ischemia. Immunohistochemical study using a specific antibody for Bax was performed using retina 24 hr after transient ischemia and fingings were compared with those of the contralateral non-ischemic retina. The number of TUNEL-positive cells after transient ischemia were measured using retinas pretreated by intra-vitreous injection of a Bax antisense oligodeoxynucleotide (ODN) or a randomly sequenced ODN. intense Bax protein immunoreactivity was detected in the GCL and INL of the post-ischemic retina 24 hr after reperfusion, while little immunoreactivity was present in the non-ichemic retina. The inhibition of Bax protein expression by antisense ODNs reduced the number of TUNEL-positive cells. We conclude some of the neuronal death caused by transient retinal ischemia involves an active cell death process of apoptosis induced by the upregulaion of Bax.Furthermore, we examine histological changes after transient retinal ischemia in transgenic mice in which neurons overexpress Bcl-2, in comparison with those of wild-type littermates. Histological sections were obtained 7 days after ischemic insult. Morphometric analysis were performed to quantify the ischemic injury in transgenic mice overexpressing Bcl-2 (NSE73a/ab) vs wild type littermates (C57BL/6). The number of cells in the GCL and the thickness of the inner plexiform layer (IPL), INL, and outer nuclear layer (ONL) were quantified. For each animal, these parameters in the post-ischemic eye were normalized to those in the intact contralateral eye and shown as a percentage. The GCL cell number was approximately 60% greater in transgenic mice than that in wild-type littermates. The IPL thickness was 33.7±1.8 μm in wild-type and 43.8±6.7 μm in transgenic mice. In wild-type mice, ischemia reduced the GCL cell number and IPL thickness to 64.6±7.5, and 42.3±0.6% of the control, respectively, whereas ischemia reduced the GCL cell number and IPL thickness to 81.9±5.6, and 82.4±19.6 in transgenic mice. These results suggest that retinal neurons overexpressing Bcl-2 become torelant to ischemic injury. Less
我们评估了Bcl-2家族基因在Sprague-Dawley大鼠视网膜缺血后延迟性神经元死亡中的凋亡参与和贡献。将眼压升高至130 mmHg 45 min,诱导视网膜缺血。再灌注后各时间取组织学标本,TUNEL染色检查。瞬时缺血后6 ~ 48小时,在神经节细胞层(GCL)和内核层(INL)中观察到大量TUNEL阳性细胞,缺血后24小时达到最大值。再灌注后24和48小时提取缺血视网膜DNA,对侧非缺血视网膜进行电泳。琼脂糖凝胶电泳在缺血后24和48小时观察到DNA阶梯,而在正常视网膜则没有。利用Bcl-2和Bax特异性引物对短暂性缺血后不同时间的视网膜进行RT-PCR分析。结果表明,Bax基因表达在缺血后6小时逐渐升高,24小时达到峰值,168小时降至接近基线水平,而Bcl-2基因表达在短暂性缺血后各时间点均无明显变化。短暂性缺血24小时后,用Bax特异性抗体对视网膜进行免疫组化研究,并与对侧非缺血视网膜进行比较。用玻璃体内注射Bax反义寡脱氧核苷酸(ODN)或随机测序的ODN预处理的视网膜,测量短暂缺血后tunel阳性细胞的数量。缺血后视网膜GCL和INL在再灌注24小时后检测到强烈的Bax蛋白免疫反应性,而非缺血视网膜的Bax蛋白免疫反应性不明显。反义odn对Bax蛋白表达的抑制减少了tunel阳性细胞的数量。我们认为,短暂性视网膜缺血引起的神经元死亡与Bax上调诱导的细胞凋亡的主动死亡过程有关。此外,我们研究了神经元过表达Bcl-2的转基因小鼠与野生型小鼠相比,在短暂性视网膜缺血后的组织学变化。缺血损伤后7天取组织学切片。形态学分析量化过表达Bcl-2 (NSE73a/ab)转基因小鼠与野生型同窝小鼠(C57BL/6)的缺血性损伤。定量测定GCL细胞数及内丛状层(IPL)、内丛状层(INL)、外核层(ONL)厚度。对于每只动物,缺血后眼的这些参数与完整对侧眼的参数归一化,并以百分比显示。转基因小鼠的GCL细胞数量比野生型小鼠高约60%。野生型小鼠的IPL厚度为33.7±1.8 μm,转基因小鼠为43.8±6.7 μm。野生型小鼠GCL细胞数量和IPL厚度分别为对照组的64.6±7.5和42.3±0.6%,转基因小鼠GCL细胞数量和IPL厚度分别为81.9±5.6和82.4±19.6。这些结果提示过表达Bcl-2的视网膜神经元与缺血性损伤相关。少
项目成果
期刊论文数量(17)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Zhang S: "Involvement of NMDA-receptor in kainate-induced neurotoxicity in cultured fetal retinal neurons"Graefe's Arch Clin Exp Ophthalmol. (in press).
张S:“培养的胎儿视网膜神经元中NMDA受体参与红藻氨酸诱导的神经毒性”Graefes Arch Clin Exp Ophthalmol。
- DOI:
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- 影响因子:0
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- 通讯作者:
Suzuma I: "Contribution of E-selectin to cellular infiltration during endotoxin-induced uveitis." Invest Ophthalmol Vis Sci. 39. 1620-1630 (1998)
Suzuma I:“E-选择素在内毒素诱导的葡萄膜炎期间对细胞浸润的贡献。”
- DOI:
- 发表时间:
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- 影响因子:0
- 作者:
- 通讯作者:
Adachi K: "Mechanism of the pathogenesis of glutamate neurotoxicity in retinal ischemia."Graefe's Arch Clin Exp Ophthalmol. 236. 766-774 (1998)
Adachi K:“视网膜缺血中谷氨酸神经毒性的发病机制。”Graefes Arch Clin Exp Ophthalmol。
- DOI:
- 发表时间:
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- 影响因子:0
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Miyamoto K: "In vivo neutralization of P-selectin inhibits leukocyte-endothelial interactions in retinal microcirculation during ocular inflammation." Microvascular Research.55. 230-240 (1998)
Miyamoto K:“P-选择素的体内中和可抑制眼部炎症期间视网膜微循环中白细胞与内皮细胞的相互作用。”
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Yasuyoshi H: "Protective effect of bradykinin aainst glutamate neurotoxicity in cultured rat retinal neurons"Invest Ophthalmol Vis Sci. (in press).
Yasuyoshi H:“缓激肽对培养的大鼠视网膜神经元中谷氨酸神经毒性的保护作用”Invest Ophasemol Vis Sci。
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HONDA Yoshihito其他文献
HONDA Yoshihito的其他文献
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{{ truncateString('HONDA Yoshihito', 18)}}的其他基金
The investigation into an intreretinal pathology and a molecular biological mechanism of diabetic retinopathy
糖尿病视网膜病变的视网膜内病理学和分子生物学机制的研究
- 批准号:
13307049 - 财政年份:2001
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for Scientific Research (A)
Elucidation of the regulatory mechanism for vascular endothelial growth factor (VEGF) expression in diabetic retinopathy
阐明糖尿病视网膜病变中血管内皮生长因子(VEGF)表达的调节机制
- 批准号:
11694267 - 财政年份:1999
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for Scientific Research (A).
Targeted drug delivery using water-soluble polymer in the treatment of choroidal neovascularization.
使用水溶性聚合物靶向给药治疗脉络膜新生血管。
- 批准号:
10557154 - 财政年份:1998
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for Scientific Research (B).
The study of the intracellular mechanism and its modulatory factor in the ischemic retina
缺血性视网膜细胞内机制及其调节因子的研究
- 批准号:
08407055 - 财政年份:1996
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for Scientific Research (A)
The study of the intracellular mechanism of retinal neuronal death caused by ischemia
缺血引起视网膜神经元死亡的细胞内机制研究
- 批准号:
06404062 - 财政年份:1994
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for General Scientific Research (A)
Development of a New System for Evaluation of Retinal Microcirculation with the Use of Heat-sensitive Liposomes
开发利用热敏脂质体评估视网膜微循环的新系统
- 批准号:
05557074 - 财政年份:1993
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for Developmental Scientific Research (B)
Studies on the electrolyte transport in corneal cells.
角膜细胞电解质转运的研究。
- 批准号:
03044087 - 财政年份:1992
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for international Scientific Research
Pathophysiobgical Study of Retinal Ischenia-with Ion-seledive Microelectrode
离子选择性微电极视网膜缺血的病理生理学研究
- 批准号:
03404051 - 财政年份:1991
- 资助金额:
$ 18.88万 - 项目类别:
Grant-in-Aid for General Scientific Research (A)
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食品衍生成分通过胱氨酸/谷氨酸转运蛋白调节铁死亡的调节机制
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23KK0109 - 财政年份:2023
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代谢型谷氨酸受体异聚化对信号传导和药理学的影响
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Molecular basis of glutamate receptor trafficking in neuronal plasticity
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Dual-function glutamate transporter/chloride channels in brain physiology and neurological diseases.
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480399 - 财政年份:2023
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