각막내피세포의 증식 및 손상의 회복에 대한 ROCK 억제제 Y-27632의 효과
Effect of ROCK Inhibitor on the Expansion and Wound Healing of Human Corneal Endothelial Cell
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목적: ROCK 억제제인 Y27632가 인간각막내피세포의 생체외 및 생체내 증식에 미치는 영향을 알아보았다. 대상과 방법: 인간공여각막에서 각막내피세포를 분리 및 배양한 후 Y27632 (10 μM) 투여 여부에 따른 세포증식의 정도를 유세포분석을 통해 비교하여 생체외 효과를 알아보았다. 생체내 효과의 분석을 위해, 각막경유 냉동손상으로 유도한 각막내피세포손상 토끼동물모델에서 Y27632 (10 mM) 투여 여부에 따른 각막두께 및 손상크기를 비교하였다. 결과: Ki67양성 세포는 투여군(9.1 %)이 대조군(8.0 %)에 비해 높았으며, Annexin V 양성 세포는 2.9%로 대조군(4.2%)에 비해 줄었으나 통계적 유의성은 없었다. 동물모델에서 손상크기는 대조군(45.6 mm2)이 투여군(49.3 mm2)에 비해 유의하게 작았으나(p=0.029),두 군 모두 처치 전에 비해 유의한 변화는 없었다. 세극등 및 각막두께 검사상 두 군간 유의한 차이는 없었다. 결론: Y27632가 기존의 연구와는 달리, 인간각막내피세포에서는 비생체 증식에는 유의한 효과를 관찰할 수 없었고, 토끼동물모델의손상된 각막내피세포에도 증식을 유도하지 못했다.
Abstract
Purpose: To investigate the effect of ROCK inhibitor Y27632 on the human corneal endothelial cell proliferation in vitro and in vivo. Methods: Using corneal endothelial cells isolated and cultured from human donor cornea, we compared the effect of Y27632 (10 μM ) on the proliferation in vitro by flow cytometry analysis. For the evaluation of the effect of Y27632 (10 mM)in vivo, corneal thickness and wound area were analyzed for the corneal endothelial wound rabbit model induced by transcorneal freezing. Results: Ki67 positive cells were increased in the Y27632 group (9.1 ± 4.1 %) than the control group (8.0 ± 5.9 %), whereas annexin V positive cells in the Y27632 group (2.9 ± 1.0 %) were decreased compared to the control group (4.2 ± 2.2 %). However these were not statistically significant. Wound area after Y27632 application in animal model is concerned, the control group showed significant smaller area (45.6 ± 0.6 mm2) compared to the Y27632 group (49.3 ± 0.8 mm2; p = 0.029,Mann-Whitney U test), however, these were not significantly different from the baseline. Corneal thickness was not different between the two groups. Conclusions: Different from other reports for the effect of Y27632, no significant effect on the proliferation in vitro and wound healing in vivo, regarding human corneal endothelial cell, were found in this study.
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