灯盏细辛提取物对高压状态下视网膜保护作用的基因调控研究
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摘要
目的:1.观察DSX对高压培养大鼠RGCs保护作用的基因调控。2.观察DSX对持续性高眼压大鼠视网膜保护作用的基因调控。3.探讨DSX的视网膜保护机制。
方法:1.建立4小时80mmHg体外高压培养RGCs模型,利用基因芯片技术研究压力对大鼠RGCs基因表达的影响。2.在高压培养RGCs中加入DSX进行干预,利用基因芯片技术,观察DSX对RGCs基因表达的调控。3.采用Akira法,建立持续性高眼压大鼠模型,利用基因芯片技术研究持续性高眼压对大鼠视网膜基因表达的影响。4.用DSX灌胃持续性高眼压大鼠1月,利用基因芯片技术,观察DSX对高眼压大鼠视网膜基因表达的调控。
结果:1.模型组和空白组RGCs有差异表达基因108条,其中凋亡基因NGFR、PAWR、S100B、ESTs和细胞骨架蛋白基因表达上调,表达下调的基因主要涉及神经系统发育、神经冲动传导、突触传递、离子通道性能、神经生理过程、细胞间信号转导、细胞黏附与细胞外基质等功能基因.2.RGCsDSX组和模型组差异表达基因51条,其中凋亡抑制基因Bcl2a1和Spp1上调,凋亡基因NGFR下调,4条外部生理刺激调控基因上调,3条ATPase活性基因上调;3.RGCs阳性对照组和模型组差异表达基因41条,其中凋亡基因S100B和NGFR表达下调,凋亡抑制基因Hmoxl上调;4.模型组和空白组差异表达基因22条,其中凋亡基因NGFR上调,CRYBB1下调;5.DSX组和模型组差异表达基因17条,其中凋亡基因NGFR表达下调,CRYBB3、CRYGD、CRYBB2表达上调。
结论:1.高压导致谷氨酸代谢转运相关基因下调,谷氨酸代谢转运功能紊乱,造成细胞外Glu堆积,从而促进Ca~(2+)内流,细胞内Ca~(2+)超载,引发细胞信号转导系统的级联反应,上调凋亡基因的表达使RGCs凋亡。2.DSX可提高RGCs对外界刺激的反应,改善能量代谢,清除Glu,减轻Ca~(2+)内流,避免细胞内Ca~(2+)超载,阻断细胞内信号转导途径,使凋亡基因NGFR表达下调。3.DSX能上调凋亡抑制基因Bcl2a1和SPP1的表达,上调对RGCs有保护作用的晶体蛋白相关基因的表达从而保护视网膜。4.DSX对视网膜的保护作用靶点多、机制复杂,其保护作用可能是凋亡抑制基因与凋亡基因,在高眼压等病理状态下的拮抗与协同作用的结果。
Objective:1.To observe the protection of DSX on gene regulation of rat RGCs cultured in vitro with high pressure.2.To observe the protection of DSX on gene regulation of the rat retinal with chronic,moderately elevated intr aocular pressure(IOP).3.Analyze the mechanism of protection of DSX for rat retina.
Method:Setting up RGCs high pressure model cultured in vitro with the condition of 80mmHg for 4 hours,then using gene array technology to study the regulation of gene express of rat RGCs on the variation of pressure.2.Adding DSX into RGCs cultured with high pressure,then using gene array technology to observe the effect of DSX on the regulation the gene express of rat RGCs.3.Building the model of ehronicity,moderately elevated intraocular pressure by the method of Akira's,then using gene array technology to study the regulation to gene express of rat retina.4.Giving DSX orally on IOP rat for one month,then using gene array to observe the effect of DSX on the regulation on the gene express of retina.
Results:1.There was 108 genes had differential expression between the model group and the control group of RGCs.such as apoptosis gene NGFR,PAWR,S 100B and ESTs was up-regulation among,other genes hand changed including the transmission of nerve impulse, synaptic transmission,cation channel activity,cell-cell signaling,cytoskeletal protein binding and extracellular matrix.2.There was 51 genes had differential expression between the test group and the model group of RGCs.apoptosis inhibiting genes Bcl2al and Sppl were up-regulation and apoptosis gene NGFR was down-regulation among them,other response to external biotic stimulus genes(4) and ATPase activity genes(3) were up-regulation.3.There was 41 genes had differential expression between the positive control group and the model group of RGCs.apoptosis inhibiting genes Hmoxl was up-regulation and apoptosis gene S100B and NGFR was down-regulation among.4.There was 22 genes had differential expression between the model group and the blank group of retinal apoptosis genes NGFR was up-regulation and CRYBB1 was down-regulation among.5.There was 17 genes had differential expression between the test group and the model group of retinal apoptosis genes NGFR was down-regulation and CRYBB3,CRYGD and CRYBB2 were up-regulation among.
Conelusion:1.The pressure could down-regulate the genes such as glutamic acid metabolism or transporter,so made high concentration of Glu,then stimulate Ca~(2+) flowing into intra-cellular,high concentration of Ca~(2+) induced signal transduction system producing cascade reaction,apoptosis gene was up-regulated to make RGCs apoptosis.2.DSX can improve energy metabolism,clean Glu for RGCs,then lessen Ca~(2+) inflow,avoiding Ca~(2+) overloading intra-cellular,blocking up cell signal transduction pathway,making apoptosis genes NGFR down regulation.3.DSX play the role as the protectant of retina by up-regulating apoptosis inhibiting genes Bcl2a1 and Spp1 and several genes about crystalline.4.The protection of DSX for retina is multi- target,complex mechanism,and identify.
方法:1.建立4小时80mmHg体外高压培养RGCs模型,利用基因芯片技术研究压力对大鼠RGCs基因表达的影响。2.在高压培养RGCs中加入DSX进行干预,利用基因芯片技术,观察DSX对RGCs基因表达的调控。3.采用Akira法,建立持续性高眼压大鼠模型,利用基因芯片技术研究持续性高眼压对大鼠视网膜基因表达的影响。4.用DSX灌胃持续性高眼压大鼠1月,利用基因芯片技术,观察DSX对高眼压大鼠视网膜基因表达的调控。
结果:1.模型组和空白组RGCs有差异表达基因108条,其中凋亡基因NGFR、PAWR、S100B、ESTs和细胞骨架蛋白基因表达上调,表达下调的基因主要涉及神经系统发育、神经冲动传导、突触传递、离子通道性能、神经生理过程、细胞间信号转导、细胞黏附与细胞外基质等功能基因.2.RGCsDSX组和模型组差异表达基因51条,其中凋亡抑制基因Bcl2a1和Spp1上调,凋亡基因NGFR下调,4条外部生理刺激调控基因上调,3条ATPase活性基因上调;3.RGCs阳性对照组和模型组差异表达基因41条,其中凋亡基因S100B和NGFR表达下调,凋亡抑制基因Hmoxl上调;4.模型组和空白组差异表达基因22条,其中凋亡基因NGFR上调,CRYBB1下调;5.DSX组和模型组差异表达基因17条,其中凋亡基因NGFR表达下调,CRYBB3、CRYGD、CRYBB2表达上调。
结论:1.高压导致谷氨酸代谢转运相关基因下调,谷氨酸代谢转运功能紊乱,造成细胞外Glu堆积,从而促进Ca~(2+)内流,细胞内Ca~(2+)超载,引发细胞信号转导系统的级联反应,上调凋亡基因的表达使RGCs凋亡。2.DSX可提高RGCs对外界刺激的反应,改善能量代谢,清除Glu,减轻Ca~(2+)内流,避免细胞内Ca~(2+)超载,阻断细胞内信号转导途径,使凋亡基因NGFR表达下调。3.DSX能上调凋亡抑制基因Bcl2a1和SPP1的表达,上调对RGCs有保护作用的晶体蛋白相关基因的表达从而保护视网膜。4.DSX对视网膜的保护作用靶点多、机制复杂,其保护作用可能是凋亡抑制基因与凋亡基因,在高眼压等病理状态下的拮抗与协同作用的结果。
Objective:1.To observe the protection of DSX on gene regulation of rat RGCs cultured in vitro with high pressure.2.To observe the protection of DSX on gene regulation of the rat retinal with chronic,moderately elevated intr aocular pressure(IOP).3.Analyze the mechanism of protection of DSX for rat retina.
Method:Setting up RGCs high pressure model cultured in vitro with the condition of 80mmHg for 4 hours,then using gene array technology to study the regulation of gene express of rat RGCs on the variation of pressure.2.Adding DSX into RGCs cultured with high pressure,then using gene array technology to observe the effect of DSX on the regulation the gene express of rat RGCs.3.Building the model of ehronicity,moderately elevated intraocular pressure by the method of Akira's,then using gene array technology to study the regulation to gene express of rat retina.4.Giving DSX orally on IOP rat for one month,then using gene array to observe the effect of DSX on the regulation on the gene express of retina.
Results:1.There was 108 genes had differential expression between the model group and the control group of RGCs.such as apoptosis gene NGFR,PAWR,S 100B and ESTs was up-regulation among,other genes hand changed including the transmission of nerve impulse, synaptic transmission,cation channel activity,cell-cell signaling,cytoskeletal protein binding and extracellular matrix.2.There was 51 genes had differential expression between the test group and the model group of RGCs.apoptosis inhibiting genes Bcl2al and Sppl were up-regulation and apoptosis gene NGFR was down-regulation among them,other response to external biotic stimulus genes(4) and ATPase activity genes(3) were up-regulation.3.There was 41 genes had differential expression between the positive control group and the model group of RGCs.apoptosis inhibiting genes Hmoxl was up-regulation and apoptosis gene S100B and NGFR was down-regulation among.4.There was 22 genes had differential expression between the model group and the blank group of retinal apoptosis genes NGFR was up-regulation and CRYBB1 was down-regulation among.5.There was 17 genes had differential expression between the test group and the model group of retinal apoptosis genes NGFR was down-regulation and CRYBB3,CRYGD and CRYBB2 were up-regulation among.
Conelusion:1.The pressure could down-regulate the genes such as glutamic acid metabolism or transporter,so made high concentration of Glu,then stimulate Ca~(2+) flowing into intra-cellular,high concentration of Ca~(2+) induced signal transduction system producing cascade reaction,apoptosis gene was up-regulated to make RGCs apoptosis.2.DSX can improve energy metabolism,clean Glu for RGCs,then lessen Ca~(2+) inflow,avoiding Ca~(2+) overloading intra-cellular,blocking up cell signal transduction pathway,making apoptosis genes NGFR down regulation.3.DSX play the role as the protectant of retina by up-regulating apoptosis inhibiting genes Bcl2a1 and Spp1 and several genes about crystalline.4.The protection of DSX for retina is multi- target,complex mechanism,and identify.
引文
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