Ophthalmic Laboratories and Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, People's Republic of China.
Invest Ophthalmol Vis Sci. 2010 Jul;51(7):3544-53. doi: 10.1167/iovs.09-5049. Epub 2010 Feb 24.
PURPOSE. To evaluate the potential neuroprotective effects of C-type natriuretic peptide (CNP) on rat retinal ganglion cells (RGCs). METHODS. Cultured adult rat retinal cells were treated with vehicle, CNP, or atrial natriuretic peptide (ANP), followed by cytotoxic insults (glutamate, TNFalpha, or withdrawal of trophic factor). RGC survival was analyzed by counting Thy-1-positive cells in each well. For in vivo evaluation, N-methyl-d-aspartate (NMDA) with or without CNP was injected intravitreally into rat eyes. At various time points after injection, retinal cross-sections were analyzed for thickness changes in the retinal layers, and retinal flat mounts were assessed by counting cresyl violet-labeled or TUNEL-positive cells. Expressions of natriuretic peptide receptor-B (NPRB) and apoptosis-related genes in retina, including Bcl-xL, BAX, and micro-calpain, were analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS. At 50 and 500 nM, CNP, but not ANP, significantly (P < 0.05) protected against glutamate-insult and trophic factor withdrawal-induced RGC death in vitro. Neither peptide significantly affected TNFalpha-induced cytotoxicity. Intravitreal injection of NMDA (20 nanomoles) significantly (P < 0.05) decreased the thickness of the inner plexiform layer (IPL), induced cell loss, increased the number of TUNEL-positive cells in the RGC layer, and upregulated the expression of Bcl-xL, BAX, and micro-calpain. All these effects were significantly (P < 0.05) alleviated by concomitant injection of CNP (4.5 nmol, 10 microg). The neuroprotective effects of CNP were maintained up to 14 days after CNP injection. CONCLUSIONS. CNP protects rat RGCs against the apoptotic damage induced by insults such as excitatory amino acid, both in vitro and in vivo.
目的。评估 C 型利钠肽 (CNP) 对大鼠视网膜神经节细胞 (RGC) 的潜在神经保护作用。
方法。用 vehicle、CNP 或心钠素 (ANP) 处理培养的成年大鼠视网膜细胞，然后进行细胞毒性刺激（谷氨酸、TNFalpha 或营养因子剥夺）。通过计数每个孔中的 Thy-1 阳性细胞来分析 RGC 的存活率。在体内评估中，将 N-甲基-D-天冬氨酸 (NMDA) 与或不与 CNP 一起眼内注射到大鼠眼中。在注射后的不同时间点，分析视网膜切片中视网膜各层的厚度变化，并通过计数 Cresyl Violet 标记或 TUNEL 阳性细胞评估视网膜平面安装。通过定量逆转录聚合酶链反应 (qRT-PCR) 分析视网膜中利钠肽受体-B (NPRB) 和凋亡相关基因的表达，包括 Bcl-xL、BAX 和微钙蛋白酶。
结果。在 50 和 500 nM 时，CNP 但不是 ANP，显著（P < 0.05）保护体外谷氨酸刺激和营养因子剥夺诱导的 RGC 死亡。两种肽都没有显著影响 TNFalpha 诱导的细胞毒性。眼内注射 NMDA（20 毫摩尔）显著（P < 0.05）降低内丛状层（IPL）的厚度，诱导细胞丢失，增加 RGC 层中 TUNEL 阳性细胞的数量，并上调 Bcl-xL、BAX 和微钙蛋白酶的表达。所有这些影响都显著（P < 0.05）被同时注射 CNP（4.5 nmol，10 微克）缓解。CNP 的神经保护作用在 CNP 注射后 14 天内仍保持。
结论。CNP 保护大鼠 RGC 免受兴奋性氨基酸等刺激引起的凋亡损伤，无论是在体外还是在体内。
