| Sepsis, also known as systemic inflammatory response syndrome (SIRS), is the first fatal disease in the intensive care unit. The causes for SIRS are various. For instance, infection caused by bacteria, fungi, mycoplasma and virus after severe trauma, burns, organ transplantation or major surgery. Then it developed into a systemic inflammatory response, accompanied with broken immune balance and coagulation dysfunction. Inflammatory reaction, broken immune balance, blood coagulation dysfunction can lead to the interaction of multiple organ dysfunction syndrome (MODS), and ultimately results in the death of patients. Sepsis incidence is increasing in recent years. About 19 million cases of sepsis patients appear each year, of which more than 1/4 died. Sepsis has a rapid onset and rapid progression. What’s more, its mechanism is very complex, but the effective treatment in sepsis has not yet appeared. Thus sepsis and septic shock has become the primary problem facing severe medicine.Endothelial cell glycocalyx damage is an important part in the pathological process of sepsis. After the structure of vascular endothelial glycocalyx destroyed, its function changes. First, vascular endothelial glycocalyx barrier function impaired or losed, the various signaling molecules and enzymes hidden in it will also lose; Secondly, the protection for endothelial cells of endothelial glycocalyx destroyed, the glycocalyx’s mechanical force transduction function is weakened. Then a series of pathological changes appear, including increased vascular permeability, edema, endothelial cell and leucocyte interaction patterns change, the coagulation system and antioxidant system imbalanced and angiotensin degrees lower. Endothelial cell glycocalyx component shedding also exposed the leukocyte adhesion receptor, followed by subsequent leukocyte and endothelial cell adhesion.This paper is based on heparin and its derivatives and Sulodexide(SLX),to further explore how the glycosaminoglycans (GAGs) play a role in sepsis treatment and the mechanism behind it. The main research contents include:1 Establish septic mice survival rate model and study the influence of SST0001, M402, ROH, HP and HS on 28 natural survival rate sepsis in mice. The ROH, HP, SLX could significantly increase the survival rate of sepsis in mice. Choose the three to carry out follow-up study.2 Establish the model of acute injury in septic mice. The content of HPA, MMP-9, HS, Syndecan-1 in serum was detected by Elisa method, and then described the phase change of them.2.1 HPA levels in sepsis mice serum and the effects of glycosaminoglycan. In sepsis, the serum content of HPA in mice was significantly decreased compared with control group (P< 0.05), and the serum content of HPA was decreased (P< 0.05) in 3h-12h time period. Heparin, ROH and sulodexide intervention does not make the blood serum content of HPA recovered to the normal level (P< 0.05).2.2 MMP-9 levels in sepsis mice serum and the effects of glycosaminoglycan The serum content of MMP-9 was significantly increased in 3 hours (P< 0.05), the peak value is 6 hours, and then fell back, after modeling 9h and 12 hours it had no significant difference with control group (P> 0.05). After the modeling 3hours, HP group and ROH group decreased significantly compared with the model group (P< 0.05), SLX group had no significant difference with the model group (P> 0.05); after modeling 6h, HP group, ROH group and SLX group decreased significantly compared with the model group (P< 0.05). Thus heparin, ROH and sulodexide can decrease the expression of MMP-9 in sepsis.2.3 In sepsis mice serum HS levels and the effects of glycosaminoglycan In the time of modeling 3h-12h, mice serum HS content and control group significantly increased (P< 0.05), indicating that the glycocalyx damage has occurred. After modeling 3h-12h HP, mice serum HS content in ROH and SLX groups decreased significantly compared with the model group (P< 0.05). The experimental results indicated that HP, ROH and SLX can inhibit HS fall off calyx in sepsis.2.4 Syndecan-1 levels in sepsis mice serum and the effects of glycosaminoglycan In the time of modeling 3h-12h, mice serum Syndecan-1 content rise significantly compared with the control group (P< 0.05), indicating that the glycocalyx damage has occurred. There was no significant difference in HP group between 3h and 6h compared with the model group. In 9h it significantly decreased compared with the model group, (P< 0.05), and there was no significant difference in 12h (P> 0.05). ROH group in 3h was significantly higher than model group (P< 0.05), in 6h,9h,12h it had no significant difference compared with model group (P> 0.05). In SLX group,3h and 6h were significantly higher than model group (P< 0.05), and there was no significant difference between 9h and 12h in model group (P> 0.05). Experiments show that heparin ROH and sulodexide have a protective effect on endothelial glycocalyx.3 Transmission electron microscopy was used to explore the integrity of endothelial cell glycocalyx in acute cardiac injury of sepsis. The damage of Model group is obvious than the control grouply; heparin, sulodexide, ROH group have different degrees of recovery of cardiac aortic endothelial cell glycocalyx integrity. For HE staining, model group have a more serious inflammation lesions in the heart tissue than the control group.Heparin, sulodexide, ROH groups’s inflammatory conditions eased; Immunohistochemical study of acute heart injury in sepsis indicated that HPA and MMP-9 expression increased significantly in the model group compared with the control group. While heparin, sulodexide and ROH can inhibit the protein expression of the two kinds of enzyme.4 Transmission electron microscopy was used to explore the integrity of endothelial cell glycocalyx in acute injury of small intestine injury in sepsis. The damage of Model group is obvious than the control grouply; Heparin, sulodexide and ROH group have different degrees of recovery of small intestine endothelial cell glycocalyx integrity. For HE staining, model group have a more serious inflammation lesions in the small intestine tissue than the control group.Heparin, sulodexide, ROH groups’s inflammatory conditions eased; Immunohistochemical study of acute small intestine injury in sepsis indicated that HPA’s and MMP-9’s expression increased significantly in the model group compared with the control group. While heparin, sulodexide and ROH can inhibit the protein expression of the two kinds of enzyme.5 Transmission electron microscopy was used to explore the integrity of endothelial cell glycocalyx in sepsis associated encephalopathy. The damage of Model group is obvious than the control grouply; heparin, sulodexide and ROH group have different degrees of recovery of cerebral cortex endothelial cell glycocalyx integrity. For HE staining, model group hace no significant difference with control group in cerebral cortex. But model group have a more serious inflammation lesions in the hippocampal area than the control group.Heparin, sulodexide, ROH groups’s inflammatory conditions eased; Immunohistochemical study of acute heart injury in sepsis indicated that HPA and MMP-9 expression increased significantly in the model group compared with the control group. While heparin, sulodexide and ROH can inhibit the protein expression of the two kinds of enzyme.Through the experimental described above we can get the conclusion:heparin, sulodexide, ROH play a therapeutic role in sepsis by protecting the endothelial glycocalyx.The innovation of this paper lies in the investigation of endothelial glycocalyx for heart, small intestine and brain of septic mouse by transmission electron microscopy (TEM). We abandoned aorta perfusion in sample fixation thus avoiding the glycocalyx sheding during perfusion.This subject is to be further investigated in the following aspects:The serum levels of HPA in septic mice is not in conformity with the level of the organization which still need more experimental exploration to reveal the reasons behind it. ROH performed well in the 28 survival rate, especially in the creation of the 70% natural survival rate of up to second day after modeling. In the following study of heart, small intestine and brain protective effect, although the ROH’s protection effect on endothelial glycocalyx is obvious, it did not always play the best anti-inflammatory effect, indicating that the glycocalyx injury is not the only pathologic mechanism of sepsis. Why ROH can significantly improve the survival rate need more research to clarify; Due to brain injury influence greatly on survival rate, and glycocalyx have important significance on blood brain barrier (BBB), we should further investigate how glycosaminoglycans protect BBB and treat brain inflammation in sepsis Immunohistochemistry showed that glycosaminoglycan intervention can restrain sepsis mouse heart HPA and MMP-9 expression, but concrete mechanism remains to be elucidated. |
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