Zhou Xiaopeng, Fang Gengjing, Zhang Liping
The Molecular Neuropharmacology Laboratory and the Eye-Brain Research Center, The State Key Laboratory of Ophthalmology, Optometry and Vision Science, School of Optometry and Ophthalmology, Wenzhou Medical University, Wenzhou 325000, China.
MethodsX. 2022 Apr 16;9:101701. doi: 10.1016/j.mex.2022.101701. eCollection 2022.
The retinal mitochondrial injury model in rat has been developed using the mitochondrial oxidative phosphorylation uncoupler, carbonylcyanide m-chlorophenyl hydrazine (CCCP). However, the CCCP-induced murine retinal mitochondrial injury model has not been reported. Here, the optimized conditions for the murine retinal mitochondrial injury model were established by intravitreal injection of different doses of CCCP (0, 2.5, 5, 7.5, 10, 12.5, 15 μg). Indeed, it has been reported that CCCP induces Opa1 cleavage and phosphorylation of ERK in cultured cells and rat retinas. Thus, we measured phosphorylated (p) -Erk and L/S-Opa1 following CCCP-induced retinal injury. Meanwhile, KW6002 (A receptor antagonist) pretreatment inhibited retinal injury induced by CCCP at 10 and 15 μg doses differently. Intravitreal injection of 10 μg doses of CCCP can induce apoptosis of retinal ganglion cells and decrease of retinal thickness, but intravitreal injection of 15 μg doses of CCCP is the appropriate dose to study the protective effect of A receptor. (1) Dose dependent effects of intravitreal injection of CCCP on the levels of L/S-Opa1 and p-Erk; (2) A receptor antagonist (KW6002) only inhibited the apoptosis of ganglion cells, but did not affect the thickness of retina with 10µg dosage of CCCP intravitreal injection; (3) A receptor antagonist (KW6002) inhibited the apoptosis of ganglion cells and increased the thickness of retina with 15µg dosage of CCCP intravitreal injection.
已使用线粒体氧化磷酸化解偶联剂间氯苯腙（CCCP）建立了大鼠视网膜线粒体损伤模型。然而，尚未见关于CCCP诱导的小鼠视网膜线粒体损伤模型的报道。在此，通过玻璃体内注射不同剂量的CCCP（0、2.5、5、7.5、10、12.5、15μg）建立了小鼠视网膜线粒体损伤模型的优化条件。事实上，已有报道称CCCP可诱导培养细胞和大鼠视网膜中Opa1裂解和ERK磷酸化。因此，我们在CCCP诱导的视网膜损伤后测量了磷酸化（p）-Erk和L/S-Opa1。同时，KW6002（一种受体拮抗剂）预处理对10μg和15μg剂量CCCP诱导的视网膜损伤有不同程度的抑制作用。玻璃体内注射10μg剂量的CCCP可诱导视网膜神经节细胞凋亡并使视网膜厚度降低，但玻璃体内注射15μg剂量的CCCP是研究A受体保护作用的合适剂量。（1）玻璃体内注射CCCP对L/S-Opa和p-Erk水平的剂量依赖性影响；（2）A受体拮抗剂（KW6002）仅抑制神经节细胞凋亡，但在玻璃体内注射10μg剂量CCCP时不影响视网膜厚度；（3）A受体拮抗剂（KW6002）在玻璃体内注射15μg剂量CCCP时抑制神经节细胞凋亡并增加视网膜厚度。
