Molecular Pharmacology, Department of Biofunctional Evaluation, Gifu Pharmaceutical University, 1-25-4 Daigaku-nishi, Gifu 501-1196, Japan.
Eur J Pharmacol. 2012 Sep 5;690(1-3):84-9. doi: 10.1016/j.ejphar.2012.06.035. Epub 2012 Jul 1.
Crocetin, an aglycone of crocin, is found in stigmas of the saffron crocus (Crocus starus L.) and has been used in traditional medicine. We investigated the effects of oral administration of crocetin on damage induced by N-methyl-D-aspartate (NMDA) in the murine retina. Crocetin was orally administered before and after intravitreal injection of NMDA. A histological analysis was conducted by counting the cell number of ganglion cell layer (GCL). Cell apoptosis was assessed by counting cells positive for terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL). Retinal functions were measured in terms of a- and b-wave amplitudes using an electroretinogram (ERG). Activation of caspase-3/7 and cleaved caspase-3 expression were assayed. Calpain activity was evaluated by immunoblotting assays for proteolysis of α-spectrin. NMDA injection decreased the cell number in the GCL, and crocetin at a dose of 100 mg/kg inhibited this reduction. TUNEL-positive cells were observed in both GCL and inner nuclear layer (INL) after NMDA injection, and crocetin inhibited the increase in number of TUNEL-positive cells. ERG analysis showed that both a- and b-wave amplitudes were decreased by NMDA injection. Crocetin inhibited the reduction in the b-wave amplitude, but not in the a-wave. NMDA injection activated caspase-3/7 and increased expression of cleaved caspsase-3 in the GCL and INL, but both of these processes were inhibited by crocetin. NMDA injection also induced cleavage of α-spectrin, but crocetin did not affect this process. These findings indicate that oral administration of crocetin prevented NMDA-induced retinal damage via inhibition of the caspase pathway.
西红花酸苷，藏红花素的苷元，存在于番红花柱头（藏红花）中，并在传统医学中使用。我们研究了口服西红花酸苷对 N-甲基-D-天冬氨酸（NMDA）诱导的鼠视网膜损伤的影响。西红花酸苷在 NMDA 眼内注射前后口服给予。通过计数节细胞层（GCL）的细胞数进行组织学分析。通过计数末端脱氧核苷酸转移酶 dUTP 缺口末端标记（TUNEL）阳性细胞来评估细胞凋亡。视网膜功能通过视网膜电图（ERG）测量 a-和 b-波幅度来衡量。测定半胱天冬酶-3/7 的活性和裂解半胱天冬酶-3 的表达。通过免疫印迹法评估钙蛋白酶活性以测定 α- spectrin 的蛋白水解。NMDA 注射减少 GCL 中的细胞数，而 100mg/kg 的西红花酸苷抑制这种减少。NMDA 注射后，GCL 和内核层（INL）中均可观察到 TUNEL 阳性细胞，而西红花酸苷抑制 TUNEL 阳性细胞数量的增加。ERG 分析表明，NMDA 注射均降低了 a-和 b-波的幅度。西红花酸苷抑制 b-波幅度的降低，但不抑制 a-波。NMDA 注射激活半胱天冬酶-3/7 并增加 GCL 和 INL 中裂解的半胱天冬酶-3 的表达，但这些过程均被西红花酸苷抑制。NMDA 注射还诱导 α- spectrin 的裂解，但西红花酸苷不影响此过程。这些发现表明，口服西红花酸苷通过抑制半胱天冬酶途径防止 NMDA 诱导的视网膜损伤。
