| Objective:Moderate and severe mechanical trauma(MT)can cause cardiac myocyte Apoptosis,resulting in decreased cardiac function.Many studies have shown that MT can cause strong oxidative stress reaction and release of a large number of cytokines.Tumor necrosis factor-?(TNF-?)produced by excessive production of Traumatic plasma(TP)is the most important pathological medium for the apoptosis of cardiomyocytes caused by non lethal MT.The TNF-?produced by monocyte-macrophages actived by lipopolysaccharide(LPS),and LPS acts as a endotoxin,it comes from the cell wall of Gram-negative bacteria.However,in the experiment,the mechanical trauma to the SD rats was the contusion and wounding of the children.There was no open wound on the body surface,that is,there was no external infection.However,the MT that rats suffered in the experiment was a pounding contusion and no open wound on the body surface,this means that the experimental rats did not have an external infection.At the same time,the intestinal tract is the largest repository for endotoxin and bacterium in the body,and it mean these organs are first affected by bacteria under the condition of MT,so it is presumed that the internal infection may occur there.There is report for the Wenchuan earthquake that there was no perforative injuries in the patients,but there was case of intestinal tissue necrosis and bacterial translocation.The first step of our experiment is to find the location of the injury in the intestinal tract and the evidence of bacterial translocation after the MT,meanwhile,we explore how the bacteria are transferred to the circulation of the blood.The second step of our experiment is to test the protective effect of glucagon like peptide-2(GLP-2)on SD rats ater MT.Method:The cecal tissue specimens of the male SD rats suffered from MT were stained by hematoxylin-eosin(HE)staining method,by this way,we can observe the mucosa change after MT at different time points in rat cecum.Our experiment focus on the brush border,crypts and mucous layer of the overall shape,then we can know the earliest time point which shows obviously damage on mucous after MT.The cecum specimens at different time points after trauma were still observed by transmission electron microscope,we focus on the change of microvilli and cell connection on the surface of mucosa.we had detected two bacterial markers in plasma in SD rats at different time points after MT,that is LPS and D-Lactatic acid.Using Enzyme-linked immunosorbent assay(Elisa)Kit and the detection wavelength is 450nm in our experiment,thus,we can get the change trend of LPS and D-lactic acid in the plasma after MTIn order to protect the cecum mucous membrane in the process of MT,the Polypeptide medicine GLP-2 was treated on SD rats before MT three days,we still use the methods described above to observe the effects of GLP-2 on cecum mucous membrane,it include HE staining,transmission electron microscope,detection of LPS and D-lactic acid by Elisa Kit,to confirm whether the GLP-2 has a significant protective effect on the cecum after MT.After the protective efficacy of GLP-2 was confirmed,the protective effect of different concentrations of GLP-2 was further detected,and the detection molecular was D-lactic acid.There was some content in the caecum of the rat,which may be the cause of the damage of the cecum mucous membrane in the process of MT and even the main cause.Then these rats were subjected to different periods of starvation,after starvation,these rats experienced the process of MT and we observe the effect of starvation by HE staining.Main results:1.Injury location of the digestive tract by MT1.1 MT(200 rotates)can cause significant damage on cecum mucous layer of cecumin SD ratsThe results of HE staining on cecum showed that,compared with the blank control and the false trauma group,the damage of rats after MT 12h,24h,and 48h was obvious severely than the others.It can cause the brushing edge of the caecum mucosa of male SD rats to be irregular,the structure is scattered1.2 Mechanical trauma has no serious damage to the stomach and the distal ileum of SD ratsThe results of HE staining gastric mucosa showed that the surface of gastric mucosa in SD rats was slightly vacuolated in 6h time point after MT.With the prolongation of posttraumatic time,the trend of self healing gradually appeared.Compared with the control group(false trauma),there was no obvious difference between 12h,24h and 48h after MT.In the end of the ileum of SD rats,there was no obvious change at all time points after the MT.Compared with the stomach and the distal ileum,the cecum has the most obviously damage2.The earliest time point for the obvious damage of the cecum ultrastructure:20min after MTDue to ultrastructure observation,compared with the control group and the sham group,the microvilli in the mucosa of 20min after MT were thinly reduced,the cellular connections were widened,the cellular structure become loose,and the damage gradually aggravated with time increase after MT.Autophagy bodies appeared in the widened cell connections at different time points after MT.After MT 12h,bacterial microbes appeared near capillaries.3.The changes of LPS in the plasma of SD rats at different time points after MT were determinedCompared with the control group,plasma LPS of SD rats increased gradually after the trauma and reached the highest level at 20min after trauma.There is significant difference between 10min and 20min after MT in level of LPS concentration.The level of LPS in plasma keep a high statue until the 24h after MT,in the time point of 48h after MT,the level of LPS in plasma showed a obviously decrease.4.The changes of D-lactic acid level in plasma of SD rats at different time points after MT were determinedCompared with the control group,D-lactic acid levels in SD rats plasma after MT reached the highest point at 20min after trauma,then the level of D-lactic acid keep an high statue until 24h after MT,then,it showed an obviously decrease on time point of 48h after trauma.5.GLP-2 can protect the cecum of SD rats after mechanical traumaAccording to the results of HE staining cecum tissue,transmission electron microscopy results,the LPS and D-lactic acid level of plasma.SD rats treated with GLP-2 group show a obviously protective effect rather than PBS groups and control groups.The protective effect on cecum include complete cell connection and pretty mucosa microvilli on cecum.6.The effect of different concentrations of GLP-2 treateed on SD rats after MTSubcutaneous doses of GLP-2 were divided into 4 groups:2.5?g/100g/time,5?g/100g/time,10?g/100g/time,20?g/100g/time,data show that 20?g/100g/time of the concentration of GLP-2 has more beneficial effect on decreaseing level of D-lactic acid in the plasma.7.VK2(Vitamin K2)mixture may have a certain protective effect on the cecum Transmission electron microscopy results of VK2 treated groups showed that the VK2 had more complete cecum villi and a certain degree of cell connection recovery.It suggests that VK2 mixture might have a certain degree of protective effect on intestinal mucosa of SD rats after MT.8.It is determined that the cecum is damaged by MT and the food residue in the cecum is not the main reason for cecum mucous membranethe results showed that MT after starvation did not significantly reduce the mucosa damage,even aggravate the degree of injury of mucosa,the main cause of cecal mucosa layer is MT.Conclusion:The results above showed that MT can lead to obviously damage on cecum mucosa of SD rats,it can cause destruction of microvilli and widen cell junction,it only spend 20 minutes after MT,at the same time,the intestinal bacterial markers such as LPS and D-lactic acid can show an significantly increase,thus,intestinal bacterium translocation started.GLP-2 can protect the cecal mucosa after MT,and its protective effect is positively related to the concentration of GLP-2 in range of 25 and 200?g/ml. |
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