Background

Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection (Singer et al., 2016). It is a major public health problem worldwide and is the leading cause of multiple organ dysfunction syndrome (MODS), which is associated with high in-hospital mortality (Torio and Moore, 2016; Kellum et al., 2019). When sepsis was first defined in 1991, the focus was on the systemic inflammatory response (ACCP/SCCMCC et al., 1992). Since then, studies of pro- and anti-inflammatory molecules have advanced and have begun to reveal that the host adaptive responses go beyond immunological phenomena (Singer et al., 2016). In the early or acute phase, sepsis is characterized by a hyperinflammatory host response accompanied by pro- and anti-inflammatory processes that lead to cell death by apoptosis. In the late or immunosuppressive phase, it is characterized by depletion of immune cells (Hotchkiss et al., 2013). In the early phase, many physiological systems are compromised, hemodynamic and cardiopulmonary alterations being the most clinical significant symptoms (Rello et al., 2017). Despite the use of several modern treatment strategies, including fluid and antimicrobial therapy, the mortality rates associated with sepsis remain high. Therefore, experimental animal models have been developed apace with the evolution of the knowledge of sepsis and advances in its treatment. The establishment of an animal model that replicates the features of sepsis in humans is essential for understanding the pathogenic mechanisms and for developing new treatment strategies. Various experimental rodent models have been described, including intraperitoneal or intravenous infusion of purified endotoxin (lipopolysaccharide), intravascular infusion of certain bacterial species, the abscess model, and the cecal ligation and puncture (CLP) model (Baker et al., 1983; Parker et al., 2001; Fink, 2014).

The CLP model is a polymicrobial model of bacterial peritonitis that is considered the gold-standard method of replicating features of sepsis in humans, which is easily reproducible, and can be adapted to mimic specific clinical events. It has been widely used as a research tool for more than 30 years (Wichterman et al., 1980). The use of this model produces a hyperdynamic circulatory state, mitochondrial dysfunction, and endothelial alterations, as well as having acute renal, pulmonary, and cardiac effects. It also stimulates receptor molecules present in immune cells, such as Toll-like receptors (TLRs), and cytokine production, as observed in pre-clinical studies (Rodrigues et al., 2012; Moreira et al., 2014; Cóndor et al., 2016; Capcha et al., 2020). The CLP model can be modified to produce varying degrees of severity, depending on the cecum ligation site and the number of punctures made (Singleton et al., 2003).

In our version of the CLP model, antibiotics and crystalloid solution are administered at 6 h after the procedure, in accordance with intensive care unit (ICU) guidelines for the restoration of blood volume and pressure (Singer et al., 2016). Although early recognition of sepsis is particularly important for appropriate patient management, as well as to achieve favorable outcomes, not all patients have timely access to treatment. Therefore, the time to the start of treatment in our model is intended to simulate the delay that occurs when patients have limited access to health care. In this rat model, 5-day mortality is typically 30-60%.

Here, we describe the CLP model in detail in order to disseminate the methodology, as well as to contribute to a better understanding of the mechanisms involved in the development of sepsis.