Promotion of Astragalus membranaceus polysaccharides on intestinal epithelial cell migration via polyamine mediated Ca2+-RhoA signaling pathway / 中草药

ABSTRACT

Objective: To study the effect of Astragalus membranaceus polysaccharides (AMP) on migration, intracellular polyamines content, cytosolic free Ca2+ concentration ([Ca2+]cyto), and RhoA protein expression of intestinal epithelial (IEC-6) cells, so as to explore the repairing mechanism of Astragalus membranaceus (AM) on gastrointestinal injury. Methods: AM was extracted by water and precipitated by ethanol, AM crude polysaccharides were obtained after removing protein. AMP I, II, III, and IV were obtained by DEAE cellulose column chromatography. AMP V was further obtained by Sephadex LH-20 gel column chromatography from AMP I. Cell migration model was established by Tips scratch method; High performance liquid chromatography was used to determine the polyamines content; Flow cytometry was used to detect the [Ca2+]cyto; Western blotting analysis was used to detect RhoA protein expression. The improving effect of AMP on migration, [Ca2+]cyto, and RhoA protein expression of normal and polyamine-depleted IEC-6 cells was observed. Results: AM crude polysaccharides, AMP I, and AMP V (100 mg /L or 200 mg /L) promoted cell migration and reversed the inhibition of cell migration induced by DFMO (P < 0.01); AMP V increased the intracellular polyamines content in normal and polyamine-deficient IEC-6 cells; AMP V also enhanced [Ca2+]cyto in the process of IEC-6 cell migration and reversed the reduction of [Ca2+]cyto induced by DFMO (P < 0.01); Further study suggested that AMP V increased RhoA protein expression in normal and polyamine-deficient IEC-6 cells (P < 0.01). Conclusion: These results indicate that the repairing effect of AM on the gastrointestinal mucosa damage may be related to its role of increasing polyamines content, then improving [Ca2+]cyto and RhoA protein expression and thereby promoting cell migration.