Method: After HSG cells were treated with 3% of xylito, the expression of M3R and aquaporin 5(AQP5) was examined by PCR, western blotting and immunocytochemistry. To measure intracellular calcium release, cells were incubated with fluorescent Ca2+indicator Fluo-4 NW prior to agonist stimulation of cells in the presence or absence of xylitol and fluorescence intensity was detected at 485nm/535nm.
Result: Xylitol stimulated intracellular calcium release via M3R signaling pathway in HSG cells. The expression of M3R and AQP5 was also increased at mRNA and protein level by xylitol treatment. In addition, the induction of intracellular calcium release and the expression of M3R and AQP5 were inhibited by 4-DAMP, a selective M3R antagonist. Stimulation of muscarinic receptor induced by xylitol activated the internalization of M3R and subsequently trafficking of aquaporin5 to the plasma membrane.
Conclusion: Xylitol stimulated secretory process via muscarinic receptor signaling pathway and the trafficking of M3R and AQP5 in HSG cells.