Protective effect of Rheum tanguticum polysaccharide against H_2O_2 induced IEC-6 cell apoptosis and its possible mechanism / 中国药理学通报

ABSTRACT

Aim To investigate the protective effect of RTP1,one of the polysaccharide isolated from Rheum tanguticum,on H2O2 induced apoptosis in IEC-6 cells and its possible mechanism.Methods H2O2(100 mmol?L~-1)was used to induce IEC-6 cell apoptosis.Different doses(10,30,100 mg?L~-1)of RTP1 were administrated before H2O2 was added into IEC-6 cell culture.Cell viability was observed by MTT assay.Reactive oxygen species were measured with laser scanning confocal microscopy(LSCM).DNA content and percentage of apoptosis were assayed by DNA agarose gel electrophoresis,acridine orange staining and flow cytometry.The activation of Caspase-3 was detected with Western blot analysis.Results Following treatment with H2O2 for 2 h,H2O2 induced a significant decrease in cell viability,while DNA ladder was observed and apoptosis percentage was as high as 31.3%.Accumulation of intracellular ROS and increase in Caspase-3 activity were also detected.Pretreatment with RTP1 for 24 h exhibited cytoprotective effects in a dose-dependent manner.RTP1 obviously enhanced cell viability,reduced formation of DNA ladder and significantly reduced the number of cells labeled with Annexin V.The percentage of apoptosis/necrosis cells was markedly decreased to 24.4% and 21.5%,respectively.LSCM showed that RTP1 attenuated the accumulation of ROS.The significant decrease in Caspase-3 activity was detected.Conclusion RTP1 has cytoprotective capacity to antagonize H2O2-induced IEC-6 cell apoptosis and injury,and this effect may be related to decrease ROS and inhibit Caspase-3 activity