Changes in entero-insular axis function and its role in mice with severe burns
Objective To explore the changes in entero-insular axis function and its role in mice with severe burns.Methods This study was an experimental study.Ninety C57BL/6J male mice aged 8-10 weeks were divided into sham injury group and burn group(with 45 mice in each group)according to the random number table.A full-thickness scald(hereinafter referred to as burn)wound of 30％of the total body surface area was created on the back of mice in burn group,and the mice in sham injury group were simulated to cause a sham injury.Twenty-four hours after injury,the fasting blood glucose was measured(n=12),followed by intraperitoneal glucose tolerance test and oral glucose tolerance test;the curve of blood glucose concentration changes over time was plotted,and the area under the curve was calculated(n=6);the blood was taken from the heart before intraperitoneal injection or gavage of glucose solution and at 30,60,and 120 minutes after intraperitoneal injection or gavage of glucose solution for measuring the plasma insulin and glucagon like peptide-1(GLP-1)levels using enzyme-linked immunosorbent assay(ELISA),with a sample number of 3;the ileal tissue was taken from 3 mice in each group for detecting the GLP-1 expression and apoptosis levels of intestinal L cells by immunofluorescence staining and TdT-mediated dUTP nick-end labeling staining;the pancreatic islets were collected from 6 mice in each group for glucose-stimulated insulin secretion experiments.After incubation with low glucose(2.8 mmol/L glucose)and high glucose(16.7 mmol/L glucose),the supernatant was taken and the insulin level was detected using ELISA.Thirty-six C57BL/6J male mice aged 8-10 weeks were divided into sham injury group,burn group,and burn+exendin-4(Ex-4)group(with 12 mice in each group)according to the random number table.The mice in sham injury group and burn group were subjected to the same corresponding treatment as before.The mice in burn+Ex-4 group were injured in the same way as the burn group mice followed by treatment with GLP-1 receptor agonist Ex-4.Twenty-four hours after injury,mouse pancreatic islets were collected,the protein expressions of heavy-chain binding protein(BIP),protein kinase R-like endoplasmic reticulum kinase(PERK),phosphorylated PERK(p-PERK),eukaryotic translation initiation factor 2α(eIF2α),phosphorylated eIF2α(p-eIF2α),and CCAAT/enhancer-binding protein homologous protein(CHOP)were detected using Western blotting,and the p-PERK/PERK and p-eIF2α/eIF2α ratios were calculated(n=3),the apoptosis rate of pancreatic islet cells was detected using flow cytometry(n=3),the glucose stimulated insulin secretion experiment was conducted as before to detect insulin levels in the supernatant(n=6).Results Twenty-four hours after injury,the fasting blood glucose of mice in burn group was(7.3±1.0)mmol/L,which was significantly higher than(5.1±0.6)mmol/L in sham injury group(t=6.36,P＜0.05).Twenty-four hours after injury,in the intraperitoneal glucose tolerance test and oral glucose tolerance test,the areas under the curve of blood glucose concentration changes over time of mice in burn group were significantly larger than those in sham injury group(with t values of 4.32 and 6.03,respectively,P＜0.05);compared with those in sham injury group,the plasma insulin levels of mice before intraperitoneal injection of glucose solution and the plasma GLP-1 levels of mice before intraperitoneal injection or gavage of glucose solution in burn group were significantly decreased(P＜0.05),and the plasma levels of insulin of mice at 30,60,and 120 minutes after intraperitoneal injection or gavage of glucose solution,as well as the plasma levels of GLP-1 of mice at 30 and 60 minutes after gavage of glucose solution were significantly decreased in burn group(P＜0.05).Twenty-four hours after injury,compared with those in sham injury group,the GLP-1 expression level of intestinal L cells of mice in burn group was significantly decreased(t=7.74,P＜0.05),and the apoptosis level was significantly increased(t=14.28,P＜0.05).Twenty-four hours after injury,the insulin level in the supernatant of mice pancreatic islet incubated with high glucose in burn group was(8.5±0.4)ng/mg,which was significantly lower than(15.7±0.3)ng/mg in sham injury group(t=18.68,P＜0.05).Twenty-four hours after injury,compared with those in sham injury group,the protein expression levels of BIP,p-PERK/PERK,p-eIF2α/eIF2α,and CHOP in the pancreatic islets of mice in burn group were significantly increased(P＜0.05);compared with those in burn group,the protein expression levels of BIP,p-PERK/PERK,p-eIF2α/eIF2α,and CHOP in the pancreatic islets of mice in burn+Ex-4 group were significantly decreased(P＜0.05).Twenty-four hours after injury,the apoptosis rate of pancreatic islet cells of mice in burn group was(32.0±3.0)％,which was significantly higher than(10.3±2.5)％in sham injury group(P＜0.05);the apoptosis rate of pancreatic islet cells of mice in burn+Ex-4 group was(20.0±3.6)％,which was significantly lower than that in burn group(P＜0.05).Twenty-four hours after injury,the insulin level in the supernatant of mice pancreatic islet incubated with high glucose in burn group was significantly lower than that in sham injury group(P＜0.05),while the insulin level in the supernatant of mice pancreatic islet incubated with high glucose in burn+Ex-4 group was significantly higher than that in burn group(P＜0.05).Conclusions After severe burns,the mice display dysfunction of the entero-insular axis,increased apoptosis of intestinal L cells,decreased synthesis and secretion of GLP-1,endoplasmic reticulum stress and increased apoptosis in pancreatic islet cells and a decrease in glucose-stimulated insulin secretion.The GLP-1 receptor agonist Ex-4 can protect the function of pancreatic islet cells of mice with severe burns,reducing the apoptosis level of pancreatic islet cells and promoting insulin secretion possibly via the alleviation of endoplasmic reticulum stress.
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