ObjectiveDysfunction of the intestinal epithelial tight junction (TJ) barrier is known to have an important etiologic role in the pathophysiology of heat stroke. N-3 polyunsaturated fatty acids (PUFAs), including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), play a role in maintaining and protecting the TJ structure and function. This study is aimed at investigating whether n-3 PUFAs could alleviate heat stress-induced dysfunction of intestinal tight junction.MethodsHuman intestinal epithelial Caco-2 cells were pre-incubated with EPA, DHA or arachidonic acid (AA) and then exposed to heat stress. Transepithelial electrical resistance (TEER) and Horseradish Peroxidase (HRP) permeability were measured to analyze barrier integrity. Levels of TJ proteins, including occludin, ZO-1 and claudin-2, were analyzed by Western blot and localized by immunofluorescence microscopy. Messenger RNA levels were determined by quantitative real time polymerase chain reaction (Q-PCR). TJ morphology was observed by transmission electron microscopy.ResultsEPA effectively attenuated the decrease in TEER and impairment of intestinal permeability in HRP flux induced by heat exposure. EPA significantly elevated the expression of occludin and ZO-1, while DHA was less effective and AA was not at all effective. The distortion and redistribution of TJ proteins, and disruption of morphology were also effectively prevented by pretreatment with EPA.ConclusionThis study indicates for the first time that EPA is more potent than DHA in protecting against heat-induced permeability dysfunction and epithelial barrier damage of tight junction.

Our results indicate that EPA pretreatment is more effective than DHA pretreatment in attenuating heat-induced intestinal dysfunction and preventing TJ damage. Enhanced expression of TJ proteins that support the epithelial barrier integrity may be important for maintaining a functional intestinal barrier during heatstroke.

Objective: Heatstroke can induce serious physiological dysfunction in the intestine. However, the underlying mechanisms of this condition are unknown, and therapeutic strategies are not available. In this study, we explored the role of endoplasmic reticulum (ER) stress signaling in this process and assessed whether pretreating mice with an inhibitor of ER stress could alleviate intestinal damage. Methods: A heatstroke model was established in male mice. Mice were pretreated with 4-phenylbutyrate (4-PBA) before exposure to heat stress. Intestinal morphological changes were observed by hematoxylin and eosin (H&E) staining and transmission electron microscopy. The TUNEL assay was used to detect intestinal apoptosis. The expression of the ER stress-related proteins and apoptosis-related proteins was investigated by the Western blot assay. Results: Compared with control group, mice with heatstroke exhibited evidence of intestinal injury and epithelial apoptosis, accompanied by significantly increased expression of ER stress-related proteins in the intestines. The intestinal injury score and level of intestinal epithelial apoptosis were significantly reduced after administration of 4-PBA. Furthermore, the levels of the intestinal ER stress-related proteins GRP78, PERK, p-eIF2α, ATF4, and CHOP were decreased after 4-PBA treatment. Conclusions: Our results indicate that the ER stress-mediated apoptosis pathway is activated during heat stress-induced intestinal injury. 4-PBA can inhibit heatstroke-induced intestinal ER stress and attenuate intestinal injury. We provide evidence that the beneficial effect of 4-PBA is closely related to the inhibition of ER stress-mediated apoptosis. These findings suggest that ER stress may be a novel therapeutic target in patients with heatstroke.

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