Corrales Rosa M, Luo Lihui, Chang Eliseu Y, Pflugfelder Stephen C
Ocular Surface Center, Cullen Eye Institute, Department of Ophthalmology, Baylor College of Medicine, Houston, TX 77030, USA.
Cornea. 2008 Jun;27(5):574-9. doi: 10.1097/ICO.0b013e318165b19e.
Increased tear osmolarity in dry eye disease has been found to stimulate production of inflammatory cytokines and matrix metalloproteinases by ocular surface epithelial cells. Prokaryotic and mammalian organ system cells maintain normal function under hypertonic conditions by the synthesis or accumulation of osmoprotectant compounds. This study assessed the effect of osmoprotectant compounds on the activation state of mitogen-activated protein (MAP) kinases in human corneal epithelial cells incubated in hyperosmolar conditions.
Human corneal epithelial cells were incubated in media of isotonic, physiological osmolarity (300 mOsm) and in hyperosmolar media (400 mOsm), in the presence and absence of osmoprotectants, including several amino acids (L-carnitine and betaine), glycerol, and the polyol erythritol. The phosphorylation (activation) states of c-Jun N-terminal kinases (JNK) and p38 MAP kinases were monitored by Western blot and bead-based immunoassays.
Hyperosmolar conditions achieved by addition of sodium chloride or sucrose increased ratios of phosphorylated JNK and p38 to total JNK and p38. Compared with controls, 10 mM L-carnitine or 40 mM erythritol significantly lowered levels of activated MAP kinases in response to hyperosmolar stress. They also lowered ratios of phosphorylated to total kinases to barely detectable levels in cells cultured in isotonic media.
The osmoprotectants L-carnitine and erythritol, alone or in combination, were found to protect against stress activation of corneal epithelial cells cultured in hyperosmolar media.
干眼症中泪液渗透压升高可刺激眼表上皮细胞产生炎性细胞因子和基质金属蛋白酶。原核细胞和哺乳动物器官系统细胞通过合成或积累渗透保护剂化合物在高渗条件下维持正常功能。本研究评估了渗透保护剂化合物对在高渗条件下培养的人角膜上皮细胞中丝裂原活化蛋白（MAP）激酶激活状态的影响。
将人角膜上皮细胞在等渗、生理渗透压（300 mOsm）的培养基以及高渗培养基（400 mOsm）中培养，培养基中添加或不添加渗透保护剂，包括几种氨基酸（左旋肉碱和甜菜碱）、甘油和多元醇赤藓糖醇。通过蛋白质印迹法和基于磁珠的免疫测定法监测c-Jun氨基末端激酶（JNK）和p38 MAP激酶的磷酸化（激活）状态。
通过添加氯化钠或蔗糖实现的高渗条件增加了磷酸化JNK和p38与总JNK和p38的比例。与对照组相比，10 mM左旋肉碱或40 mM赤藓糖醇可显著降低高渗应激下活化MAP激酶的水平。它们还将等渗培养基中培养的细胞中磷酸化激酶与总激酶的比例降低至几乎检测不到的水平。
发现渗透保护剂左旋肉碱和赤藓糖醇单独或联合使用可防止在高渗培养基中培养的角膜上皮细胞发生应激激活。
