使用定制的 LED 暴露系统研究低强度可见光对人角质形成细胞的影响。
Investigating the effects of low intensity visible light on human keratinocytes using a customized LED exposure system.
机构信息
School of Engineering, RMIT University, Melbourne, VIC, Australia.
School of Health and Biomedical Sciences, RMIT University, Bundoora, VIC, Australia.
出版信息
Sci Rep. 2022 Nov 7;12(1):18907. doi: 10.1038/s41598-022-23751-3.
Photobiomodulation (PBM) refers to the use of light to modulate cellular processes, and has demonstrated utility in improving wound healing outcomes, and reducing pain and inflammation. Despite the potential benefits of PBM, the precise molecular mechanisms through which it influences cell behavior are not yet well understood. Inconsistent reporting of key light parameters has created uncertainty around optimal exposure profiles. In addition, very low intensities of light, < 0.1 J/cm, have not been thoroughly examined for their use in PBM. Here, we present a custom-made compact, and modular LED-based exposure system for studying the effects of very low-intensity visible light (cell proliferation, migration, ROS production, and mitochondrial membrane potential) of three different wavelengths in a parallel manner. The device allows for six repeats of three different exposure conditions plus a non-irradiated control on a single 24-well plate. The immortalised human keratinocyte cell line, HaCaT, was selected as a major cellular component of the skin epidermal barrier. Furthermore, an in vitro wound model was developed by allowing the HaCaT to form a confluent monolayer, then scratching the cells with a pipette tip to form a wound. Cells were exposed to yellow (585 nm, 0.09 mW, ~ 3.7 mJ/cm), orange (610 nm, 0.8 mW, ~ 31 mJ/cm), and red (660 nm, 0.8 mW, ~ 31 mJ/cm) light for 10 min. 48 h post-irradiation, immunohistochemistry was performed to evaluate cell viability, proliferation, ROS production, and mitochondrial membrane potential. The results demonstrate increased proliferation and decreased scratch area for all exposure conditions, however only red light increased the mitochondrial activity. Oxidative stress levels did not increase for any of the exposures. The present exposure system provides opportunities to better understand the complex cellular mechanisms driven by the irradiation of skin cells with visible light.
光生物调节(PBM)是指利用光来调节细胞过程,已证明其在改善伤口愈合效果、减轻疼痛和炎症方面具有实用价值。尽管 PBM 具有潜在的益处,但它影响细胞行为的确切分子机制尚不清楚。关键光参数的报告不一致,导致最佳暴露谱的情况存在不确定性。此外,非常低强度的光,<0.1 J/cm,其在 PBM 中的应用尚未得到彻底研究。在这里,我们提出了一种定制的紧凑型、模块化基于 LED 的暴露系统,用于以并行方式研究三种不同波长的非常低强度可见光(细胞增殖、迁移、ROS 产生和线粒体膜电位)的影响。该设备允许在单个 24 孔板上对三种不同暴露条件的六次重复加上非照射对照进行重复。永生人角质形成细胞系 HaCaT 被选为皮肤表皮屏障的主要细胞成分。此外,通过允许 HaCaT 形成致密单层,然后用移液管尖端刮擦细胞形成伤口,开发了体外伤口模型。细胞分别暴露于黄色(585nm,0.09mW,3.7mJ/cm)、橙色(610nm,0.8mW,31mJ/cm)和红色(660nm,0.8mW,~31mJ/cm)光 10min。照射后 48h,进行免疫组织化学以评估细胞活力、增殖、ROS 产生和线粒体膜电位。结果表明,所有暴露条件下的增殖都增加,划痕面积减小,但只有红光增加了线粒体活性。任何暴露都没有增加氧化应激水平。目前的暴露系统为更好地理解皮肤细胞受可见光照射驱动的复杂细胞机制提供了机会。