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AIM: To observe the effects of miR-542-5p on the proliferation of rat small intestine crypt epithe-lial IEC-6 cells induced by sphingosine-1-phosphate (S1P).METHODS: Two IEC-6 cell lines (SphK1-IEC-C1 and SphK1-IEC-C2) were established, which expressed sphingosine kinase-1 (SphK1) stably.Radioactive tracer was used to detect SphK1 activity and S1P secretion.The cell proliferation was observed by cell counting and described by drawing growth curve, and the cell cycle analysis was carried out by flow cytometry.The level of miR-542-5p was evaluated by RT-qPCR.RESULTS: Compared with control vector cells without SphK1 cDNA, both SphK1-IEC-C1 and SphK1-IEC-C2 cell lines showed that Sphk1 was elevated, both intracellular and extracellular S1P increased dramatically, the rate of cell growth was faster, the percentage of the cells in S phase increased, and miR-542-5p expression decreased.S1P (0.5~10 μmol/L) led to the decrease in miR-542-5p expression.On the contrary, SphK1 silencing resulted in the increase in miR-542-5p expression in the IEC-6 cells.The miR-542-5p was elevated in SphK1-IEC-C1 cells and SphK1-IEC-C2 cells, which caused the decrease in the percentage of the cells in S phase.The cell growth rate in the above-mentioned 2 cell lines decreased compared with negative control group.CONCLUSION: In IEC-6 cells, S1P promotes proliferation by inhibiting miR-542-5p expression, which induces the cell cycle transferring from G1 phase to S phase.
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[ ABSTRACT] AIM:To investigate the role of protease activated receptor-2 ( PAR-2 ) in the process of tryptase mediated IEC-6 cell injury.METHODS:The rat intestinal epithelial cell line IEC-6 was treated with tryptase at different concentrations (1 μg/L, 10 μg/L, 100μg/L and 1 000μg/L) in the presence or absence of PAR-2 antagonist FSLLRY-NH2 for 12 h respectively.The cell survival rate was detected by MTT assay.The protein levels of PAR-2 and cleaved-caspase 3 were determined by Western blotting.The LDH activity was also measured.RESULTS:Compared with control group, the cell survival rates were significantly decreased in 100 μg/L and 1 000 μg/L tryptase treated groups, the LDH activities were significantly increased in 10 μg/L to 1 000 μg/L tryptase treated groups, and the protein levels of PAR-2 and cleaved caspase 3 were significantly increased in 100μg/L and 1 000μg/L tryptase treated groups (P<0.05).Com-pared with 1 000 μg/L tryptase treated group, the LDH activity and cleaved caspase 3 protein level were dramatically de-creased while the survival rate was significantly increased in the presence of PAR-2 antagonist FSLLRY-NH2 (P<0.05). CONCLUSION:Tryptase induces IEC-6 cell injury in a dose-dependent manner by activating PAR-2.
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Objective: To observe the effect of flavonoid from Glycyrrhizae Radix (FGR) on the migration of small intestinal epithelial cells (IEC-6) and intracellular polyamines content. Methods: The cell migration model was constructed by scratch method. Under the intervention of α-difluoromethylornithine (DFMO), the specific inhibitor of polyamine synthesis, or 4-aminopyridine (4-AP), the specific inhibitor of potassium channel, the effect of FGR on cell migration was observed. The intracellular polyamine content being cultured for 12 or 24 h in normal and DFMO intervention condition was detected by precolumn derivatization-HPLC. Results: FGR could reverse the inhibitory effects of DFMO and 4-AP on cell migration, increase the intracellular content of polyamines after 12 or 24 h treatment, and reserve the decrease of polyamine content caused by DFMO. Conclusion: FGR could affect the polyamine-dependent signaling pathway, which may be related to the mechanisms for FGR promoting cell migration.
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Objective To study the effects of Chinese herb extracts on cell proliferation and glucose absorption in intestinal epithelial cell line(IEC-6 cells) of rats.Methods The extracts of four Chinese herbs,including Herba Agastaches(HA),Rhizoma Atractylodis(RA), Cortex Phellodendri(CP),and Gypsum Fibrosum(GF),were made.Their suitable concentration on the cell proliferation in IEC-6 cells was determined by MTT method,and glucose absorption and the activity of Na+,K+-ATPase in IEC-6 cells were assayed.A method of real time PCR was applied to the determination of SGLT1 and GLUT2 mRNA expression in the cells.Results Chinese herb extracts treatment altered the cell proliferation in a dose-dependent manner.Moreover,RA volatile oil(50 ?g/mL) and CP alkaloid(10 ?g/mL) treatment increased glucose absorption and the activity of Na+,K+-ATPase genes(P0.05).Conclusion The three extracts treatment,including RA volatile oil,CP aklaloid,and HA volatile oil,could increase glucose absorption and the expression of glucose transport carrier genes,but their regulative mechanism are not totally the same.