Composition Alteration Of The Intestinal Flora In A Rat Model Of Cecal Ligation And Puncture

The Sepsis is systemic inflammatory response caused by infection, which is one of the most severe complication in patients of trauma、burn、shock and infection, once be induced to Septic Shock and multiple organ dysfunction syndrome(MODS), then the prognosis would be badly and it is hard to cure patients.In the treat of Sepsis, about 30% patients can not find original source of infection in body. After all kinds of anti-infective treat, the disease still out of control, and some even be worse to MODS.Advanced investigation found that there are obvious gut flora transversion in this patients and similar animal models, which could induce and/or promote "Second Strike" in the severe trauma、burn、shock and infection. So we could conclude that the gut flora play an important role in the occurrence and development of Sepsis、SIRS and MODS.The intestinal flora is one of major microorganism colonies. A large number and various kinds of bacterium exist in gut cavity. They are significant parts of intestinal barrier system, and they also participate physiological process such as Nutrition and Metabolism、immune regulation、maintain intestinal homeostasis. Recently, several molecular biology methods has been used to investigate the relationship between intestinal bacterium and diseases, for example,Crohn disease、intestinal transplantation and ulcerative colitis. Based on the basilic role of intestinal colony in Sepsis, for research the dynamic alternation of gut microbiota in sepsis, we founded animal models and analysis the changes of gut microbiota in relation to Sepsis to offering more reference about the clinical therapeutics of sepsis.Study Ⅰ. Composition alteration of the ileal flora in rat sepsis modelObjective:To study the composition alternation in ileal microbiota after sepsis and to detect characteristic bacterial species.Methods:The rat septic model was induced by cecal ligation and puncture(CLP).Forty Sprague-Dawley(SD) rats were randomly divided into five groups. A group of rats were killed just after anesthesia as the control group,while other four groups underwent CLP and were euthanized after 6、12、18、24hours.Luminal bacterial DNA was extracted from contents of ileal near the ileocecus,then made the pathological section.The hypervariable V3 region of the 16S ribosomal RNA gene was amplified by polymerase chain reaction (PCR) and examined by denaturing gradient gel electrophoresis (DGGE). Samples from different time points after CLP were compared with the control group. Characteristic bands of DGGE gels were cut out, sequenced and matched.Results:From the DGGE picture we could find that composition of intestinal flora changed after CLP,and recovered later.The ileal flora were most notably characteristic from the control groups at 18 hours. At CLP 12 hours, Escherichia coli proliferated, meanwhile,Lachnospiraceae and Lactobacillus decreased.Conclusions:The ileal bacterium colony shifts and recovers in septic rats longitudinally. Escherichia coli overgrowth, at the same time,Lachnospiraceae and Lactobacillus diminution.Study Ⅱ. Composition alteration of the colon flora in rat sepsis modelObjective:To study the composition alternation in colon microbiota after sepsis and to detect characteristic bacterial species.Methods:The rat septic model was induced by cecal ligation and puncture(CLP).Forty Sprague-Dawley(SD) rats were randomly divided into five groups. A group of rats were killed just after anesthesia as the control group,while other four groups underwent CLP and were euthanized after 6、12、18、24hours.Luminal bacterial DNA was get from contents of colon near the ileocecus,then made the pathological section.The hypervariable V3 region of the 16S ribosomal RNA gene was amplified by polymerase chain reaction (PCR) and examined by denaturing gradient gel electrophoresis (DGGE). Samples from different time points after CLP were compared with the control group. Characteristic bands of DGGE gels were cut out, sequenced and matched.Results:From the DGGE picture we could find that colon flora pattern altered early, and recovered later after CLP. The colon flora were most notably characteristic from the control groups at 12 hours. Escherichia coli growth after CLP 6 hours, Prevotella oralis overgrowth from CLP 12 hours, while Lactobacillus decreased early and increased later.Conclusions:The research told us that the colonic flora pattern changed after sepsis, in this situation.In this time Escherichia coli and Prevotella oralis overgrowth, while Lactobacillus decreased early and increased later.