| Objectives:Sepsis is systemic inflammatory response syndrome originated from infection. Sepsis is delitescent at onset while develops quickly into severe status with high mortality. It is currently the leading cause of death for critical patients. pathogen-associated molecular patterns, the conserved structure for pathogens, can be recognized by host pattern recognition receptors to induce sepsis. As the major component of the outer membrane in G-bacteria, LPS is regarded as the most important PAMPs. When release from bacteria, LPS is quite potent in inducing sepsis. Its level is also positively correlated with patients’ pathogenetic conditions. Herenin, it is necessary to investigate the release patterns of LPS in sepsis animal models and find corresponding intervention measures to improve sepsis treatment. CLP model is widely used for sepsis study. It is also regarded as the golden standard for sepsis model, for its resemblance with sepsis in human. It will provide necessary experimental materials for clinical sepsis research that study on LPS release and its intervention to be held in CLP model.Based on the above knowledge, we will first investigate the dynamic trend of LPS release in CLP rats. Additionally, antibiotics will be included to hinder infection. The LPS level under this condition will also be detected. Then we will observe the in vivo activity of KB an anti LPS agent in CLP rats. Lastly, we will further investigate the protective effects and drug safety of KB in sepsis mice model produced via heat killed bacteria injection.Methods:1. To detect time dependent and dose dependent release of LPS induced by different kinds of antibiotics in vitro and to choose a suitable kind of antibiotics that could be introduced for CLP model.2. To detect LPS levels at each time points in CLP model as well as those treated with antibiotics selected in in vitro experiments. Then KB will be introduced foe efficacy determination, in which its effects on LPS level and mortality of CLP rats will be observed. 3. In sepsis mice challenged via heat killed bacteria injection, protective effects of KB as well as clinical drug Xuebijin and ulinastatin will be observed and compared, in order to verify the relation between anti LPS activity and protective effects in sepsis mice.Results:1. Bactericidal activity of three antibiotics is in an order decrease as ceftazidime, o imipenem and ampicillin sodium-sulbactam sodium. However, ampicillin sodium-sulbactam sodium was most potent and stable in inducing release of LPS.2. In CLP rats, blood culture was positive as early as4h after modeling, which reaches peak in8h and maintained at2000CFU/ml. LPS was positive8h after modeling and reached peak in12-16h. LPS was detectable20tO72h, although at relatively low levels. When ampicillin sodium-sulbactam sodium was introduced, fewer bacteria but more LPS will be detected. In KB group, LPS level was decreased and more rats survived.3. KB could increase model mice survival by40%and15%separately in heat killed E.coli. or Staphylococcus aureus injection model separately.Conclusions:1. Ampicillin sodium-sulbactam sodium was mostly effective in eliminating bateria and increase release of LPS. The use of ampicillin sodium-sulbactam sodium in CLP will make it more comparable with clinical sepsis;2. The LPS release dynamics in CLP model is determined and effects of KB in reducing LPS and improve survival is also verified, which means KB plays antagonistic roles against LPS in vivo.3. KB could protect mice challenged with both heat killed E.coli. and Staphylococcus aureus, which further verifies its anti LPS acitivity and means it promising future as an anti sepsis drug candidate. |
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