| Objective:Severe acute pancreatitis(SAP) has seriously endangered the life and health of human beings,and it is often accompanied with systemic inflammatory response syndrome(SIRS),serious infections, septic shock and multiple organ dysfunction syndrome(MODS),which lead to death of patients.With the deep study on the pathogenesis of SAP mechanism,it is demonstrated that macrophages play an important role in antigen presentation-activated immune responses of SAP patients. This paper is to explore the preparation of lipsomal clodronate and the influence of function to peritoneal macrophages(PM) in SAP ratMethods:Lipsomal clodronate is prepared by means of thin film;SAP rat model was induced by injection of 5%sodium taurocholate under the pancreatic membrane,the peritoneal macrophages were obtained from SAP rat.After exposure to different doses of lipsomal clodronate(50,100,150μL),then the PM proliferation was determined by MTT colourimetry,the function of phagocytosis was evaluated by neutral red colorimetry,appraising the function of oxidative stress by detecting the contents of NO,O2-·and H2O2.the apoptosis of PM was measured by flow cytometry and agarose gel electrophoresis,respectively.Result:The prepared lipsomal clodronate had a suitable encapsulation efficiency of clodronate(5.8%)with an average size of 200nm,the spherical shape of liposome was confirmed by transmission electron microscope.The PM had a normal shape,the purity was more than 90 percent determined by Wright's staining,trypan blue staining found that the rate of cell survival was more than 95 percent.Exposed to differents doses of lipsomal clodronate resulted in a obviously growth depression(P＜0.01),inhibition ratio was follow the action time and drug concentration;neutral red colorimetry indicated the function of phagocytosis were manifestly restrained,the inhibition ratio was 17.4%, 25.8%and 38.0%,respectively.The ability of secreting NO and O2-·by peritoneal macrophages was inhibited,the difference was obviously compared with control group(P＜0.05);The ability of secreting H2O2 by peritoneal macrophages was inhibited obviously at 100μmol/L and 150μmol/L(P＜0.05);The apoptosis rate of the PM was(10.32±0.34) %,(8.16±0.49)%and(29.87±0.35)%in three different doses,the difference was evident(P＜0.01);1.2%of agarose gel electrophoresis of DNA extracted from apoptotic macrophages induced by lipsomal clodronate shows more and more clear and characteristic ladder follow the lipsomal clodronate concentration.Conclusion:We found that it was feasible to prepare lipsomal clodronate by means of thin film,and we prepared lipsomal clodronate successfully, it had an average size confirmed by transmission electron microscope.We found that liposomal clodronate could depress the function of PM in rat,and it could to induce apoptosis of PM in SAP rat.The paper provides a new theory to treat SAP in clinic,and it was expecting.
|
PDF Full Text Request