Role And Mechanisms Of High Mobility Group Box1 Protein In Intestinal ICAM-1 Expression Of Severely Burned Rats With Delayed Fluid Resuscitation

Delayed resuscitation after severe burns can cause the intestinal tissue hypoxiaischemia injury which result in cell function disorder、injury and death. And low pow-er cycle for a long time, make the circulation silt up and the formation of micro t-hrombus to induce platelet aggregation and activation, gathered in ischemia area, ch emotaxis and infiltration is also caused by pre-excitation syndrome white blood cells,causing the increased capillary permeability, and cause the intestinal tissue interstitial edema.And damage of intestinal mucosa, intestinal barrier damage and cause bacteria and toxins, easy shift, resulting in systemic infection the systemic inflammatory res ponse syndrome(systemic inflammatory response syndrome, SIRS) and multiple orga n dysfunction syndrome(multiple outraged dysfunction syndrome, MODS). So the bowel injury is an important factor that resulted in the deaths of burn patients.Intercellular adhesion molecule 1(intracellular adhesion molecule-1, ICAM-1) a-ka CD54, a member of the immunoglobulin superfamily, mediated adhesion reaction important adhesion molecule that is a single transmembrane glycoprotein, its structu re by extracellular region and across the hydrophobic membrane area and short cyto-plasmic area, is a kind of important inflammatory mediators. ICAM-1 in the mo-ve ment of cells, maintain, inflammation, immune regulation, thrombosis, and to playan important role in tumor treatment. ICAM-1 expression on vascular endothelial cells, epithelial cells and fibroblasts are strongest. In the intestinal tissue capillary endoth elial cells, the expression of ICAM-1 can destroy the integrity of the endothelial cel ls, leading to blood vessels.High mobility group box chromosomal protein 1(HMGB1), known as an abu ndant, non-histone architectural chromosomal protein, is highly conserved across different species. It was originally discovered as a DNA binding protein that facilitates DNA replication and repair. During inflammation and trauma, it can be released extr-acellularly by macrophages and other immune cells. Recent studies showed that HMGB1 can also be "passively released" into the extracellular milieu by necrotic and damaged somatic cells. HMGB1 induces endothelial cytokine expression, causes epit helial barrier dysfunction, and activates macrophages and neutrophils to release infla mmatory mediators. HMGB1 is a kind of endogenous immune adjuvant, and play a n important role on the development of various inflammatory diseases.This subject adopts the rats with 30% TBSA Ⅲ burn delay recovery in animal models, and to observe the acute intestinal injury as a result of severe burns delay recovery of intestinal tissue in dynamic changes of HMGB1, and detection of intesti nal tissue cell adhesion molecules(ICAM-1) expression and activity of p38 MAPK c-hanges, to explore the mechanism of delayed recovery of intestinal injury after sev ere burn and the relationship between HMGB1.To understand HMGB1 in severely b-urned rats gut expression, the role and mechanism of ICAM-1 for further understan ding burn caused by intestinal damage occurs to provide new ideas and methods.2. ObjectiveExplore HMGB1 delayed resuscitation in severe burns the initial mechanism in the expression of rat intestinal ICAM-13. Materials and MethodsFemale Sprague–Dawley rats weighing 200 to 250 g were purchased from the Experimental Animal Center of Anhui Medical University and housed at constant temperature for one week. All rats were randomly divided into three groups: 1)sham group(n=8); 2) burn plus Lactated Ringer’s group(n = 24); 3) burn plus Lactated Ringer’s solution and sodium butyrate-treated group(n =24). The rats were anesthetized with 3% pentobarbital sodium(30 mg/kg) by IP injection. The dorsal hair of the animals was shaved to allow direct skin contact to hot water. We developed a device for creating burn injury on the back of rats. It consists of a wooden plate with a circular opening that exposes30% TBSA. After anesthesia, the animals were placed on the device in the supine position. Each rat, along with the device, was immersed into hot water(98°C) for 12 seconds, resulting in a full-thickness skin burn injury. Sham animals were subjected to identical procedure but were immersed into water at room temperature. The burned rats were then resuscitated with an IP injection of 2 ml/kg/TBSA of Lactated Ringer’s solution at 6, 12, 36 hours postburn. Sodium butyrate was diluted 1:150 in Ringer’s lactate fluid. The rats in sodium butyrate group were treated with sodium butyrate lactated Ringer’s solution in the same manner. That is, the amount of sodium butyrate given was 400mg/kg. Subsequently, burned animals were scarified at 12 h, 24 h and 48 h postburn for checking the content of HMGB1 westernblot. The normal control group with the same anesthesia. The intestinal was harvested for the examination of expressions of p38 MAPK.Another part of immunohistochemical method to examine the expression of ICAM-1.A separate group of sham burn rats were subjected to an identical preparation except that they were immersed in room temperature water and were not given any fluid resuscitation4. Results1. Burn delayed resuscitation group of rats with the progress of the course of the d-isease, intestinal tissue of HMGB1 expression gradually increased, 12 h after injury, 24 h and 48 h were significantly higher than that of normal control group.Delay r-eco very and burns + n-butyric acid sodium group each time phase after scald burn wer-e significantly lower than the same time points delayed resuscitation group.2. Normal control group intestinal epithelial cells and endothelial cell surface has a small amount of dyeing are tan, said that the expression of ICAM-1 a mild weakly positive.Delayed resuscitation group of vascular endothelial cells and burns and part of the epithelial cells of the intestinal tissue surface has obvious tan grain, show a strong positive ICAM-1.Burns delay recovery + n-butyric acid sodium group of vasc ular endothelial cells and epithelial cells of the intestinal tissue visible on the surfac e of a small amount of tan particles, but burn delayed resuscitation group decreased significantly.3. Using Western blot method to detect the activity of intestinal p38 MAPK, p38 lig htning MAPK activity with p-p38 lightning/p38 lightning grey value than said that the activity of intestinal p38 MAPK and normal control group set to1, burns delayed resuscitation group of 24 h, the activity of p38 MAPK obvious rise, at 2.17, after HMGB1 inhibitors are sodium butyrate treatment, burns delay recovery + n-butyric acid sodium treatment group 24 h activity of rat intestinal p38 MAPK burn 24 h delay recovery group was obviously lower, at 1.03.4.Immunohistochemical method of detecting the expression of intestinal tissue p38 MAPK, intestinal tissue in rats in normal control group with scattered positive protein particles, and 24 h after severe burn injury delayed resuscitation group of intestinal tissue of epithelial cells and endothelial cells have a large number of p38 MAPK positive protein particles, is sodium butyrate in burns delayed resuscitation + 24 h group, immunohistochemical method to detect intestinal epithelial cells and endothelial cells positive protein particles decreased significantly.5. ConclusionHMGB1, through activation of p38 MAPK signaling pathways, leads to the production of ICAM-1 in intestine of severely burned rats, involving in the acute intestinal damage of severe thermal injury with delayed resuscitation.