Protective effect and mechanism of trehalose on ionizing radiation-induced intestinal damage in mice
Objective To investigate the protective effect and molecular mechanism of trehalose in ionizing radiation(IR)-induced small intestinal damage.Methods Eight-week-old male C57 BL/6 J mice were randomly divided into four groups,with each group containing six mice:control(no treatment given),trehalose-treated(consumption of 2 g/100 ml seaweed sugar water),IR(whole abdominal irradiation group treated with 13 Gy IR),and trehalose+IR groups(trehalose+total abdominal irradiation treatment with 13 Gy IR).Daily monitoring of body weight was performed after 13 Gy irradiation and 2 g/100 ml trehalose drinking water treatment.After 14 days,the mice were euthanized,and samples from the blood,small intestine,and colon were collected.Intestinal injury was assessed via hematoxylin-eosin(HE)and periodic acid-schiff(PAS)staining.The levels of tumor necrosis factor(TNF)-α and interleukin(IL)-6 in the serum and small intestine tissues were quantified through enzyme-linked immunosorbent assay(ELISA).Experiments on mouse small-intestine epithelial cell proliferation,cell clone formation,cell apoptosis rate,reactive oxygen species(ROS)determination,and DNA double-strand breaks(DSBs)detection were conducted to detect cell viability,clone formation ability,apoptosis,ROS levels,and the number of DSBs,respectively.The expression levels of autophagy-related genes and proteins were determined via quantitative real-time polymerase chain reaction(qRT-PCR)method and Western blot method.Inter group comparison was achieved through two independent sample t-tests.Results The animal study results demonstrated that compared with the irradiation group,trehalose significantly alleviated the weight loss,facilitated the recovery of irradiated mice,and promoted its recovery(t=4.064,P＜0.05),the trehalose+IR group exhibited milder colon shortening degree of mice,and statistically significant difference(t=4.044,P＜0.05).In addition,the HE and PAS staining evaluation results indicated that trehalose can alleviate intestinal mucosal damage in mice after IR.Compared with the irradiation group,the trehalose+irradiation group presented longer villi,deeper crypts,and more goblet cells.The two groups exhibited a statistically significant difference(t=4.373,8.414,6.515,all P＜0.05).ELISA demonstrated that trehalose substantially decreased the TNF-α and IL-6 levels in mouse serum and small intestinal tissues after IR,and the differences were statistically significant(t=4.475,8.686,3.993,6.007,all P＜0.05).In vitro,the results of cell proliferation and cloning experiments revealed that trehalose promoted the activity and clone formation ability of mouse small-intestine MODE-K cells after IR[(0.885±0.127)vs.(0.644±0.151),(97.330±5.937)vs.(64.000±7.324)],with the differences showing statistical significance(t=2.566,4.411,all P＜0.05).The findings of the apoptosis detection experiment indicated that trehalose inhibited the apoptosis rate of mouse small intestine MODE-K cells after IR treatment[(15.270±0.647)％vs.(9.334±0.854)％],and the difference was statistically significant(t=8.315,P＜0.05).According to ROS assay and DSB detection experiments,trehalose reduced intracellular ROS levels and DSBs after IR(t=8.884,4.802,both P＜0.05).The findings of qRT-PCR and Western blot analysis showed that compared with the IR group,the trehalose+IR group showed significantly increased expressions of autophagy-related genes and proteins transcription factor EB(TFEB)and microtubule-associated proteins 1 light chain 3 beta in the small-intestine tissue and MODE-K mouse cells[(208.210±26.143)vs.(8.986±5.362),(13.970±4.587)vs.(5.815±1.873)],and the differences were statistically significant(t=8.875,8.461,all P＜0.05).Conclusion Trehalose alleviates IR-induced intestinal damage in mice through the regulation of TFEB-mediated autophagy activation.
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