目的观察补肾活血方对AGEs/缺氧条件下视网膜Müller细胞血管内皮生长因子(VEGF)及色素上皮衍生因子(PEDF)的影响,并对其作用机制进行初步探讨。方法应用改进的酶消化法培养SD大鼠视网膜Müller细胞,将其分为正常对照组(NC组)...目的观察补肾活血方对AGEs/缺氧条件下视网膜Müller细胞血管内皮生长因子(VEGF)及色素上皮衍生因子(PEDF)的影响,并对其作用机制进行初步探讨。方法应用改进的酶消化法培养SD大鼠视网膜Müller细胞,将其分为正常对照组(NC组)、正常中药干预组(NCM组)、低AGEs组(LA组)、高AGEs组(HA组)、低AGEs中药干预组(LAM组)、高AGEs中药干预组(HAM组)、模拟缺氧组(SH组)、模拟缺氧中药干预组(SHM组),用酶联免疫吸附法(ELISA)对相关干预条件下体外培养视网膜Müller细胞上清液进行VEGF及PEDF含量的测定。结果(1)VEGF表达量:(1)24、48、72 h时,SH组、LA组、HA组视网膜Müller细胞VEGF表达量均高于NC组,差异均有统计学意义(P<0.05);HA组、SH组表达量均高于LA组,差异均有统计学意义(P<0.05)。(2)24 h时,SH组表达量高于HA组;48 h时,SH组表达量低于HA组,差异均有统计学意义(P<0.05)。(3)除72 h NCM组VEGF表达量与NC组相比,差异无统计学意义(P>0.05)外,其余时段有中药干预组的表达量均较对应无中药干预组降低,差异均有统计学意义(P<0.05)。(4)SH组、LA组、HA组、LAM组、HAM组、SHM组细胞48、72 h时VEGF表达量均较前一时段增高,差异均有统计学意义(P<0.05)。(2)PEDF表达量:(1)SH组、LA组、HA组各个时段视网膜Müller细胞PEDF表达量较NC组降低,差异均有统计学意义(P<0.05)。(2)24、72 h时,HA组、SH组PEDF表达量较LA组降低,差异均有统计学意义(P<0.05)。(3)24h时,SH组PEDF表达量较HA组降低,差异有统计学意义(P<0.05)。(4)24、72 h NCM组、LAM组,24、48、72 h HAM组,24 h SHM组视网膜Müller细胞PEDF表达量均较对应无中药干预组增高,差异有统计学意义(P<0.05)。(5)LA组、HA组、SH组、LAM组、HAM组、SHM组48、72 h时细胞PEDF表达量均较前一时段降低,差异均有统计学意义(P<0.05)。(3)VEGF/PEDF比值:(1)NC组、NCM组各时段其比值几乎相同,表明视网膜Müller细胞VEGF与PEDF的量处�展开更多
目的:探讨加味桃红四物汤(MTSD)对视网膜Müller细胞rMC-1缺氧损伤的保护作用。方法:用加味桃红四物汤含药血清干预缺氧条件下rMC-1细胞,随机分为正常对照组(21%O_(2))、缺氧模型组(1%O_(2))、含药血清低(1%O_(2)+5%含药血清)、中(1...目的:探讨加味桃红四物汤(MTSD)对视网膜Müller细胞rMC-1缺氧损伤的保护作用。方法:用加味桃红四物汤含药血清干预缺氧条件下rMC-1细胞,随机分为正常对照组(21%O_(2))、缺氧模型组(1%O_(2))、含药血清低(1%O_(2)+5%含药血清)、中(1%O_(2)+10%含药血清)、高剂量组(1%O_(2)+15%含药血清),CCK-8法检测细胞的活力,ELISA法检测血管内皮生长因子(VEGF)和色素上皮衍生因子(PEDF)分泌,Western blot检测磷酸化转录激活因子3(p-STAT3)、转录激活因子3(STAT3)和缺氧诱导因子-1α(HIF-1α)的蛋白表达,Real time PCR检测VEGF、PEDF、STAT3和HIF-1α的基因表达。结果:在1%O_(2)条件下培养48h,rMC-1细胞活力较正常对照组明显受到抑制(P<0.05),加味桃红四物汤含药血清低、中剂量组均可以改善rMC-1细胞缺氧48h的细胞存活率(P<0.05),而高剂量组无改善作用(P>0.05)。加味桃红四物汤含药血清低、中剂量组均可减少缺氧条件下rMC-1细胞上清液VEGF的蛋白表达量(P<0.05),但不能增加PEDF的蛋白含量(P>0.05),对p-STAT3和HIF-1α在蛋白水平均有下调作用(P<0.05),且低剂量组抑制作用优于中剂量(P<0.05)。加味桃红四物汤含药血清中剂量组对缺氧后rMC-1细胞STAT3的蛋白表达有上调作用(P<0.05)。加味桃红四物汤含药血清低、中剂量组对缺氧后rMC-1细胞VEGF基因表达均有下调作用(P<0.05),对PEDF基因表达均有上调作用(P<0.05),且低剂量组优于中剂量(P<0.05);并且加味桃红四物汤含药血清低剂量可下调缺氧后STAT3和HIF-1α的基因表达(P<0.05)。结论:加味桃红四物汤含药血清可能通过抑制STAT3/HIF-1α通路,下调缺氧诱导的视网膜Müller细胞rMC-1的VEGF蛋白和基因表达,上调PEDF基因表达,减轻该细胞的缺氧损伤。展开更多
AIM: To investigate if pigment epithelium-derived factor(PEDF) has any protective effect on the retinal Müller cells of Sprague-Dawley rats suffering from diabetes mellitus.METHODS: Sixty Sprague-Dawley rats were...AIM: To investigate if pigment epithelium-derived factor(PEDF) has any protective effect on the retinal Müller cells of Sprague-Dawley rats suffering from diabetes mellitus.METHODS: Sixty Sprague-Dawley rats were randomly divided into a negative control group, a group receiving0.1 μg/μL PEDF, another group receiving 0.2 μg/μL PEDF,and a group receiving balanced salt solution(BSS). Rats in both the PEDF and BSS groups were treated intravitreally based on previously established diabetic models. After 4wk of treatment, morphological alterations of Müller cells and protein expression of glutamine synthase(GS) and glial fibrillary acidic protein(GFAP)were analyzed.RESULTS:PEDFateither0.1μg/μLor0.2μg/μLsignificantly improved the structures of both nuclei and organelles of Müller cells compared to the BSS-treated group.Expression of GS was significantly higher in the 0.2 μg/μL PEDF group than that in the BSS group(P =0.012), but expression of GFAP was significantly lower in the 0.2 μg/μL PEDF group than that in the BSS group(P =0.000);however, there were no significant differences in expression of these proteins between the 0.1 μg/μL PEDF group and the BSS group(P =0.608, P =0.152). CONCLUSION: PEDF protects the morphological ultrastructure of Müller cells, improves the expression of glutamate synthase and prevents cell gliosis.展开更多
文摘目的观察补肾活血方对AGEs/缺氧条件下视网膜Müller细胞血管内皮生长因子(VEGF)及色素上皮衍生因子(PEDF)的影响,并对其作用机制进行初步探讨。方法应用改进的酶消化法培养SD大鼠视网膜Müller细胞,将其分为正常对照组(NC组)、正常中药干预组(NCM组)、低AGEs组(LA组)、高AGEs组(HA组)、低AGEs中药干预组(LAM组)、高AGEs中药干预组(HAM组)、模拟缺氧组(SH组)、模拟缺氧中药干预组(SHM组),用酶联免疫吸附法(ELISA)对相关干预条件下体外培养视网膜Müller细胞上清液进行VEGF及PEDF含量的测定。结果(1)VEGF表达量:(1)24、48、72 h时,SH组、LA组、HA组视网膜Müller细胞VEGF表达量均高于NC组,差异均有统计学意义(P<0.05);HA组、SH组表达量均高于LA组,差异均有统计学意义(P<0.05)。(2)24 h时,SH组表达量高于HA组;48 h时,SH组表达量低于HA组,差异均有统计学意义(P<0.05)。(3)除72 h NCM组VEGF表达量与NC组相比,差异无统计学意义(P>0.05)外,其余时段有中药干预组的表达量均较对应无中药干预组降低,差异均有统计学意义(P<0.05)。(4)SH组、LA组、HA组、LAM组、HAM组、SHM组细胞48、72 h时VEGF表达量均较前一时段增高,差异均有统计学意义(P<0.05)。(2)PEDF表达量:(1)SH组、LA组、HA组各个时段视网膜Müller细胞PEDF表达量较NC组降低,差异均有统计学意义(P<0.05)。(2)24、72 h时,HA组、SH组PEDF表达量较LA组降低,差异均有统计学意义(P<0.05)。(3)24h时,SH组PEDF表达量较HA组降低,差异有统计学意义(P<0.05)。(4)24、72 h NCM组、LAM组,24、48、72 h HAM组,24 h SHM组视网膜Müller细胞PEDF表达量均较对应无中药干预组增高,差异有统计学意义(P<0.05)。(5)LA组、HA组、SH组、LAM组、HAM组、SHM组48、72 h时细胞PEDF表达量均较前一时段降低,差异均有统计学意义(P<0.05)。(3)VEGF/PEDF比值:(1)NC组、NCM组各时段其比值几乎相同,表明视网膜Müller细胞VEGF与PEDF的量处�
文摘目的:探讨加味桃红四物汤(MTSD)对视网膜Müller细胞rMC-1缺氧损伤的保护作用。方法:用加味桃红四物汤含药血清干预缺氧条件下rMC-1细胞,随机分为正常对照组(21%O_(2))、缺氧模型组(1%O_(2))、含药血清低(1%O_(2)+5%含药血清)、中(1%O_(2)+10%含药血清)、高剂量组(1%O_(2)+15%含药血清),CCK-8法检测细胞的活力,ELISA法检测血管内皮生长因子(VEGF)和色素上皮衍生因子(PEDF)分泌,Western blot检测磷酸化转录激活因子3(p-STAT3)、转录激活因子3(STAT3)和缺氧诱导因子-1α(HIF-1α)的蛋白表达,Real time PCR检测VEGF、PEDF、STAT3和HIF-1α的基因表达。结果:在1%O_(2)条件下培养48h,rMC-1细胞活力较正常对照组明显受到抑制(P<0.05),加味桃红四物汤含药血清低、中剂量组均可以改善rMC-1细胞缺氧48h的细胞存活率(P<0.05),而高剂量组无改善作用(P>0.05)。加味桃红四物汤含药血清低、中剂量组均可减少缺氧条件下rMC-1细胞上清液VEGF的蛋白表达量(P<0.05),但不能增加PEDF的蛋白含量(P>0.05),对p-STAT3和HIF-1α在蛋白水平均有下调作用(P<0.05),且低剂量组抑制作用优于中剂量(P<0.05)。加味桃红四物汤含药血清中剂量组对缺氧后rMC-1细胞STAT3的蛋白表达有上调作用(P<0.05)。加味桃红四物汤含药血清低、中剂量组对缺氧后rMC-1细胞VEGF基因表达均有下调作用(P<0.05),对PEDF基因表达均有上调作用(P<0.05),且低剂量组优于中剂量(P<0.05);并且加味桃红四物汤含药血清低剂量可下调缺氧后STAT3和HIF-1α的基因表达(P<0.05)。结论:加味桃红四物汤含药血清可能通过抑制STAT3/HIF-1α通路,下调缺氧诱导的视网膜Müller细胞rMC-1的VEGF蛋白和基因表达,上调PEDF基因表达,减轻该细胞的缺氧损伤。
基金Supported by Shaanxi Province Science and Technology Research and Development Program (No. 2012K16-06-05)
文摘AIM: To investigate if pigment epithelium-derived factor(PEDF) has any protective effect on the retinal Müller cells of Sprague-Dawley rats suffering from diabetes mellitus.METHODS: Sixty Sprague-Dawley rats were randomly divided into a negative control group, a group receiving0.1 μg/μL PEDF, another group receiving 0.2 μg/μL PEDF,and a group receiving balanced salt solution(BSS). Rats in both the PEDF and BSS groups were treated intravitreally based on previously established diabetic models. After 4wk of treatment, morphological alterations of Müller cells and protein expression of glutamine synthase(GS) and glial fibrillary acidic protein(GFAP)were analyzed.RESULTS:PEDFateither0.1μg/μLor0.2μg/μLsignificantly improved the structures of both nuclei and organelles of Müller cells compared to the BSS-treated group.Expression of GS was significantly higher in the 0.2 μg/μL PEDF group than that in the BSS group(P =0.012), but expression of GFAP was significantly lower in the 0.2 μg/μL PEDF group than that in the BSS group(P =0.000);however, there were no significant differences in expression of these proteins between the 0.1 μg/μL PEDF group and the BSS group(P =0.608, P =0.152). CONCLUSION: PEDF protects the morphological ultrastructure of Müller cells, improves the expression of glutamate synthase and prevents cell gliosis.
