Effects of heat stress combined with lipopolysaccharides stimulation on injury to human intestinal epithelial cells

BAI Yong-cheng; PAN Zhi-guo; LI Yong-kui; ZHENG Yu-mei

Abstract：Objective To investigate the biological effects of heat stress combined with lipopolysaccharides ( LPS) stimulation on injury to human intestinal epithelial cells SW 480.Methods SW480 cells were cultured in vitro and divided into four groups .The control group was cultured in the incubator at 37℃ for 2 hours.The LPS stimulation group was treated with 1μg/mL LPS for 6 hours.The heat stress group was cultured in incubator at 43℃for 2 hours.The combined group was cultured in incubator at 43℃for 2 hours followed by treatment with 1μg/mL LPS for 6 hours.Cyto-toxic effect was detected by lactate dehydrogenase ( LDH) assay.WST-1 was used to detect cell survival rate .Flow cy-tometry was used to detect the rate of apoptosis .TEER value was used to assess cellular permeability .Enzyme marker was used to detect Caspase -3 enzyme activity .Expression of ZO-1 were determined by Western blot .The level of reactive oxygen species (ROS) was measured by DCFH-DA staining.In addition, the effect of ROS specific scavenger MnTBAP on cell injury induced by heat stress combined with LPS stimulation was also studied .Results Compared with the control group, levels of LDH in the heat stress group and LPS stimulation group were significantly elevated ( P<0.05 ) with the cell survival rates significantly reduced (P<0.05).Compared with the heat stress group and LPS stimulation group , level of LDH in the combined group was significantly elevated (P<0.05) with the cell survival rates significantly reduced (P<0.05).Compared with the control group , apoptosis rate, Caspase-3 expressions and ROS levels in the heat stress group and LPS stimulated group were significantly higher ( P<0.05 ) , while the TEER values and ZO -1 expressions signifi-cantly lower (P<0.05).Compared with the heat stress group and LPS stimulation group , apoptosis rate, Caspase-3 ex-pression and ROS level in the combined group were significantly elevated ( P<0.05) with the TEER value and ZO -1 ex-pression significantly reduced (P<0.05).In addition, the MnTBAP significantly alleviated cells injury induced by heat stress combined with LPS stimulation .Conclusion Heat stress has a synergistic effect with LPS stimulation , which medi-ates cells injury .ROS may initiate the damaging cascade as an upstream signal in the process .