Integrity and wound healing of rainbow trout intestinal epithelial cell sheets at hypo-, normo-, and hyper-thermic temperatures

Abstract

How temperature influences fish physiological systems, such as the intestinal barrier, is important for understanding and alleviating the impact of global warming on fish and aquaculture. Monolayers of the rainbow trout cell line, RTgutGC, with or without linear 500μm wide gaps (wounds) were the in vitro models used to study the integrity and healing of intestinal epithelial sheets at different temperatures. Cultures at hypothermic (4°C) or hyperthermic (≥ 26°C) temperatures were compared to normothermic control cultures (18-22°C). Monolayers remained intact for at least a week at temperatures from 4 to 28°C, but had lost their integrity after 3h at 32°C as the cells pulled away from one another and from the plastic surface. F-actin appeared as prominent stress fibers in cells at 28°C and as blobs in cells at 32°C. At normothermia and at 26°C, cells migrated as sheets into the gaps and closed (healed) the gaps within 5-6 days. By contrast, wounds took 14 days to heal at 4°C. At 28°C some cells migrated into the gap in the first few days but mainly as single cells rather than collectively and wounds never healed. When monolayers with wounds were challenged at 32°C for 3h and returned to 18-22°C, cells lost their shape and actin organization and over the next 6 days detached and died. When monolayers were subjected to 26°C for 24h and challenged at 32°C for 3h prior to being placed at 18-22°C, cell shape and actin cytoskeleton were maintained, and wounds were healed over 6 days. Thus, intestinal epithelial cells become thermostabilized for shape, cytoskeleton and migration by a prior heat exposure.