Pectin from Prunus domestica L. induces proliferation of IEC-6 cells through the alteration of cell-surface heparan sulfate on differentiated Caco-2 cells in co-culture
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Dietary fiber intake provides various physiological and metabolic effects for human health. Pectin, a water-soluble dietary fiber, induces morphological changes of the small intestine in vivo. However, the molecular mechanisms underlying pectin-derived morphological alterations have not been elucidated. Previously, we found that pectin purified from Prunus domestica L. altered the sulfated structure of cell-surface heparan sulfate (HS) on differentiated Caco-2 cells via fibronectin and α5β1 integrin. In this study, we investigated the biological significance of the effect of pectin on HS in differentiated Caco-2 cells. An in vitro intestinal epithelium model was constructed by co-culture of differentiated Caco-2 cells and rat IEC-6 cells, which were used as models of intestinal epithelium and intestinal crypt cells, respectively. We found that pectin-treated differentiated Caco-2 cells promoted growth of IEC-6 cells. Real-time RT-PCR analysis and western blotting showed that relative mRNA and protein expression levels of Wnt3a were upregulated by pectin treatment in differentiated Caco-2 cells. Analysis by surface plasmon resonance spectroscopy demonstrated that pectin-induced structural alteration of HS markedly decreased the interaction with Wnt3a. However, depression in the secretion of Wnt3a from Caco-2 cells by anti-Wnt3a antibody did not affect the proliferation of IEC-6 cells in co-culture system. These observations indicated that pectin altered the sulfated structure of cell-surface HS to promote secretion of Wnt3a from differentiated Caco-2 cells and Wnt3a indirectly stimulated the proliferation of IEC-6 cells.
KeywordsDifferentiated Caco-2 cell Heparan sulfate IEC-6 cell Pectin Wnt3a
Bovine serum albumin
Fetal bovine serum
Human HS 6-O-endosulfatase-2
Quail HS 6-O-endosulfatase-1
We thank the members of the Division of Genomics Research, Life Science Research Center, Gifu University, for their support in conducting the experiments and thank Miki Corporation (Hyogo, Japan) for providing the concentrated prune juice. This work was supported by Japan Society for the Promotion of Science (JSPS) Grant-in-Aid for Scientific Research (C) Grant Number 23580163.
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