R28세포에서 고포도당이 반응성산소종의 생성에 미치는 영향
Effect of High Glucose on the Production of Reactive Oxygen Species in R28 Cells
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목적: 망막전구세포주인 R28세포에서 고포도당이 반응성산소종의 생성에 미치는 영향을 알아보고자 하였다. 대상과 방법: R28 수립세포주를 계대배양한 후 고농도(25 mM)와 저농도(5 mM)의 포도당이 포함된 배지에 각각 2일간 노출시켰다. 이때 항산화제인 50 μM N-acetyl cysteine 또는 일산화질소 생성의 기질인 100 μM L-arginine에 동시에 노출시켰다. 세포배양 과정에서 생성되는 일산화질소의 농도는 Griess assay로 측정하였으며, DCFH-DA와 modified cytochrome c assay로 반응성산소종과superoxide의 생성량을 측정하였다. 결과: 고포도당은 저농도의 포도당에 비해 일산화질소, 반응성산소종, 그리고 superoxide의 생성을 유의하게 증가시켰으며, N-acetyl cysteine과 L-arginine은 고포도당에 의한 반응성산소종의 생성증가를 감소시켰다. 결론: 고포도당은 R28세포에서 반응성산소종의 생성을 증가시켰으며 항산화제에 의해 반응성산소종의 생성이 억제되었다. 따라서 고포도당은 산화스트레스를 유발하여 망막신경절세포의 손상과 기능을 저하시킬 수 있을 것으로 생각된다.
Abstract
Purpose: To investigate the effect of high glucose (HG) on the production of reactive oxygen species (ROS) in retinal precursor R28 cells. Methods: R28 cells were incubated with low glucose (5 mM) or HG (25 mM) for two days. Additionally, the cells were co-exposed to 50 μM N-acetyl cysteine or 100 μM L-arginine. Production of nitric oxide (NO), ROS, and superoxide were assessed by Griess assay, DCFH-DA assay, and modified cytochrome c assay, respectively. Results: HG increased the production of NO, ROS, and superoxide, which were abolished by antioxidants NAC and L-arginine (a substrate for NO production). Conclusions: HG increased ROS production in R28 cells. Thus, HG may cause cellular dysfunction and damage by inducing oxidative stress in retinal ganglion cells.
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