Park Young Min, Kim Chi Dae, Lee Jong Soo
Department of Ophthalmology, Pusan National University School of Medicine & Medical Research Institute, Yangsan Pusan National University Hospital, Yangsan 626-770, Korea.
Department of Pharmacology, Pusan National University College of Medicine, and MRC for Ischemic Tissue Regeneration, Yangsan 626-870, Korea.
Korean J Physiol Pharmacol. 2015 Jul;19(4):357-63. doi: 10.4196/kjpp.2015.19.4.357. Epub 2015 Jun 30.
The purpose of this study was to compare the inhibitory effect of bevacizumab on human Tenon's fibroblasts (HTFs) cultured from primary and recurrent pterygium. Cultured HTFs were exposed to 2.0, 5.0, 7.5, and 15.0 mg/mL concentration of bevacizumab for 24 hours. The 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide and lactate dehydrogenase leakage assays were then performed to assess fibroblast metabolism and viability. The matrix metalloproteinase (MMP), procollagen type I C terminal propeptide (PIP), and laminin immunoassays were performed to examine extracellular matrix production. Changes in cellular morphology were examined by phase-contrast and transmission electron microscopy. Both metabolic activity and viability of primary and recurrent pterygium HTFs were inhibited by bevacizumab in a dose-dependent manner, especially at concentrations greater than 7.5 mg/mL. Both types of HTFs had significant decreases in MMP-1, PIP, and laminin levels. Distinctly, the inhibitory effect of bevacizumab on MMP-1 level related with collagenase in primary pterygium HTFs was significantly higher than that of recurrent pterygium. Significant changes in cellular density and morphology both occurred at bevacizumab concentrations greater than 7.5 mg/mL. Only primary pterygium HTFs had a reduction in cellular density at a bevacizumab concentration of 5.0 mg/mL. Bevacizumab inhibits primary and recurrent pterygium HTFs in a dose-dependent manner, especially at concentrations greater than 7.5 mg/mL. As the primary HTFs produces larger amounts of MMP-1 compared to recurrent HTFs, significant reduction in MMP-1 level in primary pterygium HTFs after exposure to bevacizumab is likely to be related to the faster cellular density changes in primary pterygium HTFs.
本研究的目的是比较贝伐单抗对从原发性和复发性翼状胬肉培养的人结膜成纤维细胞(HTFs)的抑制作用。将培养的HTFs暴露于浓度为2.0、5.0、7.5和15.0mg/mL的贝伐单抗中24小时。然后进行3-[4,5-二甲基噻唑-2-基]-2,5-二苯基溴化四氮唑和乳酸脱氢酶泄漏试验,以评估成纤维细胞的代谢和活力。进行基质金属蛋白酶(MMP)、I型前胶原C末端前肽(PIP)和层粘连蛋白免疫测定,以检测细胞外基质的产生。通过相差显微镜和透射电子显微镜检查细胞形态的变化。贝伐单抗以剂量依赖性方式抑制原发性和复发性翼状胬肉HTFs的代谢活性和活力,尤其是在浓度大于7.5mg/mL时。两种类型的HTFs的MMP-1、PIP和层粘连蛋白水平均显著降低。明显地,贝伐单抗对原发性翼状胬肉HTFs中与胶原酶相关的MMP-1水平的抑制作用显著高于复发性翼状胬肉。在贝伐单抗浓度大于7.5mg/mL时,细胞密度和形态均发生显著变化。仅原发性翼状胬肉HTFs在贝伐单抗浓度为5.0mg/mL时细胞密度降低。贝伐单抗以剂量依赖性方式抑制原发性和复发性翼状胬肉HTFs,尤其是在浓度大于7.5mg/mL时。由于原发性HTFs比复发性HTFs产生更多的MMP-1,暴露于贝伐单抗后原发性翼状胬肉HTFs中MMP-1水平的显著降低可能与原发性翼状胬肉HTFs中更快的细胞密度变化有关。