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Updated: May 13, 2026
Analyzing Beneficial Effects of Nutritional Supplements on Intestinal Epithelial Barrier Functions During Experimental Colitis
Published on: January 5, 2017
Myosin light chain kinase-mediated epithelial barrier dysfunction as a potential pathogenic mechanism of
Nichakorn Worakajit1, Saravut Satitsri2, Taya Kitiyakara3
1Program in Translational Medicine, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Ratchathewi, Bangkok, 10400, Thailand; Chakri Naruebodindra Medical Institute, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Bang Phli, Samut Prakarn, 10540, Thailand.
Abstract:
Diarrheas are an important adverse effect of afatinib, a tyrosine kinase inhibitor (TKI) anti-cancer drug, leading to mortality and morbidity in cancer patients with their pathophysiological mechanisms related to intestinal barrier dysfunctions being poorly understood. This study aimed to investigate the effect of afatinib on intestinal epithelial barrier integrity using a human colon-derived organoid model (colonoids). Afatinib (0.5 μM) significantly decreased the transepithelial electrical resistance (TEER) by ∼60% and increased apical-to-basolateral dextran flux by > 20 folds without causing apparent cytotoxicity in human colonoids. The delocalization of zonula occludens-1 (ZO-1) and a decrease in mRNA and protein expression of claudin-4 and ZO-1 were also observed in the afatinib-treated human colonoids. Afatinib induced nuclear translocation of nuclear factor kappa B (NF-κB) as well as mRNA and protein expression of NF-κB targets including tumor necrosis factor (TNF)-alpha, interleukin-8 (IL-8), and inducible nitric oxide synthase (iNOS) indicating the initiation of the NF-κB-mediated epithelial inflammatory responses. Interestingly, afatinib induced mRNA and protein expression of myosin light chain (MLC) kinase (MLCK) and MLC phosphorylation, a known inducer of intestinal epithelial barrier disruption. Treatment with iNOS inhibitor (1400W) or MLCK inhibitor (ML-7) reversed the effect of afatinib on mRNA expressions of ZO-1 and claudin-4, and TEER. Collectively, our results indicate that afatinib induces intestinal epithelial barrier dysfunction via mechanisms involving NF-κB-iNOS-MLCK pathways. This finding may pave the way for developing therapeutic strategies to reduce adverse effects and enhance efficacy of TKI in cancer patients.
Insights
Afatinib, an anti-cancer drug, disrupts the intestinal barrier by activating NF-κB-iNOS-MLCK pathways, leading to diarrhea. Inhibiting these pathways may mitigate this adverse effect in cancer patients.
Area of Science:
- Gastroenterology
- Oncology
- Molecular Biology
Background:
- Diarrhea is a common, severe side effect of afatinib, a tyrosine kinase inhibitor (TKI) used in cancer therapy.
- The mechanisms underlying afatinib-induced intestinal barrier dysfunction are not fully understood, contributing to patient morbidity.
Purpose of the Study:
- To investigate how afatinib affects intestinal epithelial barrier integrity.
- To elucidate the molecular pathways involved in afatinib-induced gut toxicity.
Main Methods:
- Utilized a human colon-derived organoid (colonoid) model.
- Assessed barrier integrity via transepithelial electrical resistance (TEER) and dextran flux.
- Analyzed protein and mRNA expression of tight junction proteins (ZO-1, claudin-4), NF-κB signaling, iNOS, and MLCK.
Main Results:
- Afatinib impaired barrier integrity (decreased TEER, increased flux) without cytotoxicity.
- Afatinib reduced ZO-1 and claudin-4 expression and caused ZO-1 delocalization.
- Afatinib activated NF-κB signaling, upregulating inflammatory mediators (TNF-α, IL-8, iNOS) and MLCK, leading to MLC phosphorylation.
Conclusions:
- Afatinib induces intestinal epithelial barrier dysfunction through NF-κB-iNOS-MLCK signaling.
- Inhibitors of iNOS and MLCK partially reversed afatinib's effects on barrier integrity and tight junction proteins.
- Findings suggest potential therapeutic targets to manage afatinib-induced diarrhea and improve TKI efficacy.
