Induction of Periodontitis via a Combination of Ligature and Lipopolysaccharide Injection in a Rat Model

Periodontitis (PD) is the sixth most prevalent public health condition worldwide, affecting approximately 11% of the total population, being an advanced, irreversible, and destructive form of periodontal disease^1,2. PD is an inflammatory process that affects the gingival and periodontal tissues, which results in gingiva recession, apical migration of the junctional epithelium with pocket development, and the loss of alveolar bone³. Furthermore, PD is associated with several systemic diseases, including cardiovascular disease, obesity, diabetes, and rheumatoid arthritis, for which environmental and host-specific factors play a significant role^4,5.

Hence, PD is a multifactorial disease primarily initiated by the accumulation of microbial plaque – resulting from dysbiosis of microbial communities – and by an exaggerated host immune response to periodontal pathogens, which leads to the breakdown of periodontal tissue^4,6. Among several periodontal bacteria, the gram-negative anaerobic bacterium Porphyromonas gingivalis is one of the key pathogens in PD⁴. P. gingivalis contains a complex lipopolysaccharide (LPS) in its walls, a molecule known to induce polymorphonuclear leukocyte infiltration and vascular dilatation in inflamed periodontal tissues⁷. This results in the production of inflammatory mediators, such as interleukin 1 (IL-1), IL-6, and IL-8, tumor necrosis factor (TNF), or prostaglandins, with a subsequent osteoclast activation and bone resorption, leading to tissue destruction and ultimate tooth loss³.

Among the different advantages of animal models include the capacity to mimic cellular complexities as in humans, or to be more accurate than in vitro studies, which are carried out on plastic surfaces with limited cell types⁸. For modeling PD experimentally in vivo, different animal species have been used, like non-human primates, dogs, pigs, ferrets, rabbits, mice, and rats⁹. However, rats are the most extensively studied animal model for the pathogenesis of PD because they are inexpensive and easy to handle¹⁰. Their dental gingival tissue has similar structural features to human gingival tissue, with a shallow gingival sulcus and junctional epithelium attached to the tooth surface. Furthermore, as in humans, the junctional epithelium facilitates the passage of bacterial, foreign materials, and exudates from inflammatory cells ⁹.

One of the most reported experimental models of PD induction in rats is the placement of ligatures around the teeth, which is technically challenging but reliable¹⁰. The ligature placement facilitates dental plaque and bacterial accumulation, generating a dysbiosis in the gingival sulci, that causes periodontal tissue inflammation and destruction¹¹. Loss of periodontal attachment and resorption of alveolar bone could occur in 7 days in this rat model⁸.

Another animal model for PD consists of the injection of LPS into the gingival tissue. As a result, osteoclastogenesis and bone loss are stimulated. The histopathological features of this model are similar to human-established PD, characterized by higher levels of proinflammatory cytokines, collagen degradation, and alveolar bone resorption^6,8.

Thus, the aim of this study was to describe a simple rat model of experimental PD based on the techniques of P. gingivalis-LPS (Pg-LPS) injections, combined with ligature placement around the first maxillary molars (M1). This is a model with similar characteristics to those observed in human PD disease, which could be used in the study of disease progression mechanisms and future possible treatments.