Guidelines for Constructing Animal Models of Sepsis and Experimental Procedures

Overview of Sepsis and Its Research Significance

Sepsis is a systemic inflammatory response syndrome triggered by infection, characterized by organ dysfunction accompanied by the abnormal release of various cytokines and inflammatory mediators. This pathological process involves complex dysregulation of immune networks, which can further develop into septic shock or severe sepsis, ultimately leading to multiple organ dysfunction syndrome (MODS). In clinical practice, sepsis remains one of the leading causes of death among patients in intensive care units, with mortality rates as high as 30-50%.

Establishing standardized animal models is invaluable for in-depth research on the pathogenesis of sepsis and exploring new therapeutic strategies. An ideal sepsis model should be able to simulate the pathophysiological characteristics of human disease, including systemic inflammatory responses, organ dysfunctions, and immune system dysregulation. Currently used modeling methods internationally are mainly divided into two categories: endotoxin-induced acute inflammation models and progressive infection models based on live bacterial infections.

Detailed Explanation of Cecal Ligation and Puncture (CLP) Model

Model Principles and Pathological Features The cecal ligation puncture model creates a perforation at a specific site in the cecum through surgical intervention to allow intestinal contents containing various gut flora to continuously leak into the abdominal cavity, simulating common clinical polymicrobial peritonitis pathology. This model has several significant advantages: first, it reproduces the natural progression process of human sepsis, including early pro-inflammatory response stages and late immunosuppressive phases; second, by adjusting parameters such as ligation length or size/number of punctures, disease severity can be precisely controlled; finally, this model can present typical organ dysfunction associated with sepsis such as acute kidney injury (AKI), liver function impairment, and lung injury.

From a pathophysiological perspective, the CLP model fully presents the dynamic process involved in developing sepsis. Within 12 hours post-surgery, animals begin exhibiting typical signs consistent with systemic inflammatory response syndrome (SIRS), including abnormal body temperature changes or reduced activity levels. After 18-24 hours they enter multi-organ failure stages where significant capillary leakage, microcirculation disorders, and mitochondrial dysfunction become observable pathological changes. These features make CLP an ideal choice for studying mechanisms underlying septic pathology along with treatment interventions.

Standardized Operating Procedure Selecting appropriate experimental animals is crucial for successful modeling outcomes. It is recommended that male C57BL/6 mice aged between 8-12 weeks weighing between 25-30g be utilized while ensuring all animals adaptively raised under SPF conditions for at least seven days maintaining light/dark cycles over twelve hours allowing free access to food/water prior experimentation procedures wherein random allocation must occur dividing them into modeled groups versus sham-operated controls ensuring comparability across baseline traits like weight/age etc. nSurgical operations need strict adherence towards aseptic techniques throughout procedure execution involving inhalational anesthesia via Isoflurane keeping anesthetic depth sufficient enough preventing limb flexion reflexes during incision preparation exceeding two centimeters around intended cut area employing three-step disinfection using iodine solution followed up utilizing midline approach measuring approximately one point five-two centimeters opening skin fascia layer sequentially avoiding excessive tension applied onto mesentery preventing vascular damage while performing single ligature typically located distal third region secured using four-zero silk sutures appropriately placed prior executing puncturing step deemed critical success factor determining overall efficacy achieved herewith recommending eighteen-gauge needle insertion creating full-thickness hole whilst gently squeezing out minimal intestinal content thereafter closing abdomen employing layered suture technique initially stitching peritoneum/muscle layers subsequently applying interrupted closure method over dermis layer before immediate postoperative care administering pre-warmed blankets sustaining body temperature alongside subcutaneous injections delivering warmed saline solutions aimed combating hypovolemic shock risks encountered postoperatively thus enhancing survival probabilities within experimental settings following these guidelines meticulously will yield reproducible results necessary advancing knowledge regarding septic phenomena observed clinically today.