Kitaoka Yasushi, Kumai Toshio, Kitaoka Yuka, Lam Tim T, Munemasa Yasunari, Isenoumi Kazuyuki, Motoki Masamitsu, Kuribayashi Kohei, Kogo Jiro, Kobayashi Shinichi, Ueno Satoki
Department of Ophthalmology, St Marianna University School of Medicine, Kawasaki-shi, Kanagawa, Japan.
Brain Res Mol Brain Res. 2004 Nov 24;131(1-2):8-16. doi: 10.1016/j.molbrainres.2004.07.021.
Transcription factors of the nuclear factor-kappa B (NF-kappaB)/Rel family may be involved in neuronal cell death or survival. We examined the role of NF-kappaB p65 in N-methyl-D-aspartate (NMDA)-induced neurotoxicity in the rat retina. Western blot analysis showed that elevated levels of retinal NF-kappaB p65 protein at days 1 and 5 after intravitreal NMDA injection. Immunohistochemistry localized increased NF-kappaB p65 immunoreactivity in the ganglion cell layer (GCL) and the inner nuclear layer (INL) after NMDA injection especially in retinal ganglion cells (RGCs), displaced amacrine cells, and amacrine cells. Concomitant with the early increase in NF-kappaB p65 protein levels, there was an increase in NF-kappaB DNA binding activity after NMDA injection as shown by electrophoretic mobility shift assay (EMSA). These increases in NF-kappaB p65 protein levels and NF-kappaB DNA binding activity were totally abolished by simultaneous injection of NF-kappaB p65 antisense oligodeoxynucleotide (AS ODN). A partial but significant protective effect on the inner retina was noted when the AS ODN was given together with NMDA as shown by morphological analysis, morphometry of cells in the GCL and morphometry of inner plexiform layer thickness as well as quantitative real-time PCR of Thy-1 mRNA levels. These results suggest that activated NF-kappaB p65 may participate in NMDA-induced retinal neuronal cell death and that inhibition of NF-kappaB activation such as the use of AS ODN may be a viable neuroprotective strategy for protective RGCs and other inner retinal neurons.
核因子-κB（NF-κB）/Rel家族的转录因子可能参与神经元细胞的死亡或存活。我们研究了NF-κB p65在N-甲基-D-天冬氨酸（NMDA）诱导的大鼠视网膜神经毒性中的作用。蛋白质免疫印迹分析显示，玻璃体内注射NMDA后第1天和第5天，视网膜NF-κB p65蛋白水平升高。免疫组织化学显示，NMDA注射后，神经节细胞层（GCL）和内核层（INL）中NF-κB p65免疫反应性增加，尤其是在视网膜神经节细胞（RGC）、移位无长突细胞和无长突细胞中。与NF-κB p65蛋白水平的早期增加相一致，如电泳迁移率变动分析（EMSA）所示，NMDA注射后NF-κB DNA结合活性增加。同时注射NF-κB p65反义寡脱氧核苷酸（AS ODN）可完全消除NF-κB p65蛋白水平和NF-κB DNA结合活性的这些增加。形态学分析、GCL中细胞的形态计量学、内网状层厚度的形态计量学以及Thy-1 mRNA水平的定量实时PCR显示，当AS ODN与NMDA一起给药时，对内视网膜有部分但显著的保护作用。这些结果表明，活化的NF-κB p65可能参与NMDA诱导的视网膜神经元细胞死亡，抑制NF-κB活化，如使用AS ODN，可能是保护RGC和其他视网膜内层神经元的一种可行的神经保护策略。
