This study was carried out to study the protective effect of carboxytmethylpachymaran (CMP) on TNF-alpha-induced intestinal epithelial barrier dysfunction in Caco-2 cell monolayers and the underlying mechanism. The Caco-2 cell monolayers were pretreated with 50, 100 or 150 mu g/mL CMP for 72 h, and then they were exposed to 100 ng/mL tumor necrosis factor alpha (TNF-alpha) for 72 h. The results showed that CMP alleviated the drop of trans-epithelial electrical resistance (TEER) and the increase of phenol red flux induced by TNF-alpha. CMP also ameliorated TNF-alpha-induced decrease of mRNA/protein expression and distribution of Occludin and ZO-3 which were chosen as makers of tight junction (TJ). Additionally, the increased protein expressions of MLCK, phosphorylation level of myosin light chain (p-MLC), NF-kappa B p-P65 and p-IKB alpha induced by TNF-alpha were significantly inhibited by CMP. This study demonstrates the protective effect of CMP on TNF-alpha-induced damage of intestinal epithelial barrier in Caco-2 monolayers and discovers that the suppression of MLCK-p-MLC signaling regulated by NF-kappa B might be one of the mechanisms underlying the protective effect of CMP. (C) 2016 Elsevier B.V. All rights reserved.