Porphyromonas gingivalis induces exacerbated periodontal disease during pregnancy

Highlights

• A ligature-induced periodontal disease model was used to investigate the aggravated disease during pregnancy.

• P. gingivalis infection induced higher gingival inflammation and alveolar bone loss in pregnant mice.

• P. gingivalis infection down-regulated anti-inflammatory cytokine expression in pregnant mice.

• There was no significant difference in oral bacterial number between the pregnant mice and non-pregnant mice.

Abstract

Although pregnant women are prone to gingival inflammation, its mechanism remains unclear. Animal models are ideal for investigating immunological mechanisms in the periodontal disease. A murine model for ligature-induced periodontal disease has been modified and utilized to determine the susceptibility to periodontal inflammation and tissue damage in pregnant mice. Expression of different inflammatory mediators in the gingivae was determined by quantitative real-time PCR (qPCR). Inflammatory bone loss was determined by measuring the distance from the cementoenamel junction to the alveolar bone crest (CEJ-ABC). Oral bacterial number was determined by the CFU (Colony Forming Units) count from anaerobic culture of oral swabs. In our experiments, ligation itself did not cause higher gingival inflammation and bone loss in pregnant mice than non-pregnant mice, while ligation combined with P. gingivalis infection led to increased gingival inflammation and periodontal bone loss, accompanied by lower gingival expression of anti-inflammatory cytokines in pregnant mice. Our results indicated that P. gingivalis infection was important in inducing more severe periodontal diseases during pregnancy, which might be attributed to the down-regulated anti-inflammatory mechanisms, but not be associated with higher oral bacterial burden.

Introduction

Pregnancy is a unique stage accompanied by various immunological changes, during which the women are more susceptible to some infectious diseases [1]. Pregnant women are prone to developing gingival inflammation [[2], [3], [4], [5], [6], [7], [8], [9], [10]]. Some women may even develop more severe clinical attachment loss when they are pregnant, especially the ones with pre-existing periodontal disease [[11], [12], [13], [14]]. Inflammation and oral pathogenic bacteria infiltration into circulation are detrimental to pregnant women's health and negatively affect their quality of life. For example, it has been suggested that periodontal disease is associated with low birth weight and preterm birth [[15], [16], [17], [18], [19], [20]]. Therefore, it is critical to understand the pathogenesis of exacerbated gingival inflammation and/or periodontal disease during pregnancy.

Elevated gingival inflammation can be simply attributed to increased vascularity and vascular flow, or result from the changes in immune responses to oral pathogens during pregnancy. While many reports showed that the periodontal inflammation during pregnancy is more restricted in gingival tissues and wanes after delivery [4,5,10,21], other reports demonstrated irreversible periodontal damage during pregnancy [[11], [12], [13], [14]]. The periodontal disease during pregnancy shares many aspects with the disease in non-pregnancy environment. Periodontal pathogens can induce host immune responses and cause inflammation and collateral tissue damage. While reports showed that the gingiva without pre-existing inflammation remains healthy during pregnancy, implying that periodontal pathogens possess special capabilities to promote inflammation during pregnancy [11,22,23], other clinical studies exhibited differently and found that even the pregnant women with low dental plaque accumulation developed gingival inflammation [9,24]. One explanation is that oral pathogens are still present in the women with low dental plaque, and cause inflammation during the pregnancy in a pathogenic environment.

Porphyromonas gingivalis (P. gingivalis) is one of the most widely investigated oral pathogens and can induce periodontitis in animal models [25]. P. gingivalis, as well as other periodontal pathogens such as Prevotella intermedia and Treponema denticola, are present in the pregnant women with gingival inflammation [2,6,[26], [27], [28]]. P. gingivalis has been shown to be able to regulate the host immunity and oral microbiota dysbiosis. The role of periodontal pathogens and the mechanisms of more severe gingival inflammation and periodontal tissue damage during pregnancy are still unclear, partially due to the lack of an efficient animal model. In order to understand the role of P. gingivalis infection and periodontal inflammation during pregnancy, a mouse model is optimal since P. gingivalis is not a natural inhabitant in mouse oral cavity [[29], [30], [31]]. In a well-established ligature-induced murine periodontal disease model, a silk ligature is placed around molar teeth to induce massive local accumulation of bacteria and the disease [[32], [33], [34]]. In this model, the disease can be initiated at a specific time with enhanced bacteria-mediated inflammation in a short period [35]. In current manuscript, we modified and utilized this murine model and induced periodontal inflammation and tissue damage during pregnancy (Fig. 1). In order to test the role of P. gingivalis in the process, we performed experiments with or without P. gingivalis infection (Fig. 1). In the mice without P. gingivalis infection, there are indigenous “normal” oral bacteria present in the oral cavity of the mice. Ligation with the indigenous bacteria is able to induce gingival inflammation and bone loss [35]. Comparing the mice infected with P. gingivalis infection and the ones without P. gingivalis infection, we illustrated the role of P. gingivalis in promoting the disease.

We found that P. gingivalis infection is important to induce more severe gingival inflammation and inflammatory bone loss in pregnant mice. Moreover, we found that P. gingivalis infection induced lower gingival anti-inflammatory cytokine expression in pregnant mice than non-pregnant mice, implying that the mechanism of the disease might be attributed to impaired anti-inflammatory regulation.