Antonini J M, Starks K, Roberts J R, Millecchia L, Yang H M, Rao K M
Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, WV 26505, USA.
In Vitr Mol Toxicol. 2000 Spring;13(1):5-16.
Chronic inhalation of hard metal (WC-Co) particles causes alveolitis and the eventual development of pulmonary fibrosis. The initial inflammatory response includes a change in the alveolar epithelial cell-capillary barrier, which has been shown to be regulated by the state of assembly and organization of the actin cytoskeletal network. The objective of this study was to evaluate the effect WC-Co particles have on F-actin organization of lung epithelial cells in an in vitro culture system. Rat lung epithelial (L2) cells were exposed to 5, 25, and 100 microg/mL of WC-Co particles, as well as the individual components (Co and WC) of the hard metal mixture particles for 24 h. The effect on F-actin organization was visualized by laser scanning confocal microscopy (LSCM) following Bodipy-Phallacidin staining. Minimal changes in the F-actin microfilaments of L2 cells were observed by LSCM after exposure to WC and WC-Co at 5 and 25 microg/mL, while at 100 microg/mL, there was a noticeable disruption in the uniform distribution of L2 cell F-actin microfilaments. After exposure to Co, a dose-dependent change in the F-actin organization of the L2 cells was observed. Little change in F-actin assembly was observed after treatment with 5 microg/mL of Co (the concentration equivalent to the 5% amount of Co commonly present in 100 microg/mL of the WC-Co sample mixture). However, at 100 microg/mL of Co, the microfilaments aggregated into homogeneous masses within the cells, and a significant loss in the organization of L2 F-actin was observed. These dramatic alterations in F-actin organization seen after exposure to the higher doses of Co were attributed to an increase in L2 cell injury as measured by lactate dehydrogenase and trypan blue exclusion. We conclude the pulmonary response evoked in the lung by inhalation of high levels of WC-Co particles is unlikely due to alterations in the F-actin microfilaments of lung-epithelial cells.
长期吸入硬质金属（WC-Co）颗粒会引发肺泡炎，并最终导致肺纤维化。最初的炎症反应包括肺泡上皮细胞-毛细血管屏障的改变，研究表明这种改变受肌动蛋白细胞骨架网络的组装和组织状态调控。本研究的目的是评估WC-Co颗粒在体外培养系统中对肺上皮细胞F-肌动蛋白组织的影响。将大鼠肺上皮（L2）细胞暴露于5、25和100微克/毫升的WC-Co颗粒以及硬质金属混合颗粒的单个成分（Co和WC）中24小时。用Bodipy-鬼笔环肽染色后，通过激光扫描共聚焦显微镜（LSCM）观察对F-肌动蛋白组织的影响。暴露于5和25微克/毫升的WC和WC-Co后，通过LSCM观察到L2细胞的F-肌动蛋白微丝变化极小，而在100微克/毫升时，L2细胞F-肌动蛋白微丝的均匀分布出现明显破坏。暴露于Co后，观察到L2细胞的F-肌动蛋白组织出现剂量依赖性变化。用5微克/毫升的Co（该浓度相当于100微克/毫升的WC-Co样品混合物中通常含有的5%的Co量）处理后，F-肌动蛋白组装变化很小。然而，在100微克/毫升的Co时，微丝在细胞内聚集成均匀的团块，并且观察到L2 F-肌动蛋白组织明显丧失。暴露于高剂量Co后F-肌动蛋白组织的这些显著改变归因于通过乳酸脱氢酶和台盼蓝排斥法测量的L2细胞损伤增加。我们得出结论，吸入高水平的WC-Co颗粒在肺部引发的肺反应不太可能是由于肺上皮细胞F-肌动蛋白微丝的改变。
