Preventive effect of probiotics on periodontal ligament in a rat model of anorexia nervosa

Abstract

The rising prevalence of anorexia nervosa (AN), especially among adolescents, and the limited understanding of its effects on the periodontium often hinder decision making in dentistry and periodontology. As an adjunct to periodontal therapy, probiotic administration has shown promising effects on oral health by decreasing pathogen counts and altering the immune response. This study thus investigates changes in morphology and remodelling of alveolar bone and periodontal ligament (PDL) due to an AN-like condition, along with potential protective effects of probiotics, using a rat model. Three-week-old female Wistar rats were divided into three groups: a control group with ad libitum food and two groups undergoing an activity-based anorexia (ABA) model. One ABA group received oral multi-strain probiotics during starvation. After five weeks, all rats were sacrificed for ex-vivo micro-CT scans of the maxilla and mandible to assess alveolar bone and PDL morphology, as well as histological evaluations for PDL fibre vitality and structural organization. The data were statistically evaluated by one-way ANOVA with Tukey's post-hoc test or by Kruskal-Wallis with Dunn's test for parametric or nonparametric data, respectively. The results showed no structural changes in alveolar bone caused by either ABA or probiotic treatment; however, the ABA group exhibited a significant reduction in PDL thickness, which could not be reversed by probiotic treatment. Despite this, histological analysis indicated improved connectivity and density of PDL fibres in the probiotic group compared with the ABA-only group. No differences were found between the mandible and maxilla. In conclusion, while probiotics did not prevent PDL thinning, they enhanced its composition/vitality compared to the ABA condition alone.

Keywords: Activity-based anorexia model; Anorexia nervosa; Dentistry; Periodontal and alveolar bone remodeling; Periodontal ligament; Periodontology; Probiotics; Tomography.

Fig. 1

Graphical representation of the animal experiment. After 8 days of habituation with ad libitum access to food, the ABA and ABA-P groups underwent the acute and later chronic ABA phase with controlled feeding and daily activity. Both groups received daily oral interventions with water (ABA) or probiotics (ABA-P).

Fig. 2

Evaluation of the alveolar bone morphology in the M1 and M2 sockets of the maxilla and mandible: (A) morphological parameters were estimated from the cylindrical volume of interest (VOI-1) of M1 and M2. Statistical evaluation of the bone parameters revealed relatively stable alveolar bone morphology under ABA condition independent of probiotic therapy: (B) BV/TV—bone volume/total volume, (C) CONN—trabecular connectivity, (D) Tb.N.—trabecular number, (E) Tb.Th.—trabecular thickness and (F) Tb.Sep.—trabecular separation, for all studied groups: C (control), activity-based anorexia model (ABA) and ABA + P (ABA + probiotics). M1—1st molar, M2—2nd molar, M3—3rd molar, D—dentin, AB—alveolar bone, PDL—periodontal ligament; *p < 0.05.

Fig. 3

Analysis of PDL thickness and volume of M1 tooth in maxilla and mandible. A strong reduction of PDL thickness (A) and PDL volume (B) was found in the ABA group in both maxilla and mandible. This reduction could not be restored by probiotic administration. (C) 3D rendering of the VOI-2 with a color mapped PDL thickness shows the thinning of the PDL (predominantly blue/green areas) in the ABA and ABA + P groups compared to the control (prevalently red). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Fig. 4

The PDL space around the 1st molar (M1) was evaluated histologically using HE staining and three magnifications. The PDL area was clearly reduced in both activity-based anorexia (ABA) (B, E, H) and ABA + P (ABA + probiotics) (C, F, I) groups compared to the control (A, D, G). Histograms with the highest magnifications (scale 50 μm) revealed that the ABA group showed a strong loss of PDL fibres compared to the control group. In the ABA + P group, this structure was visibly repaired by the concomitant probiotic treatment. AB—alveolar bone; PDL—periodontal ligament; D—dentin.

Fig. 5

Collagen staining of the 1st molar space revealed less dense PDL fibres in the activity-based anorexia model (ABA) group while its structure remained unchanged under probiotic supplementation (ABA + P). (A–C) Overview of transverse sections of the periodontium in control, ABA and ABA + P groups, respectively. (D) Quantitative estimation of collagen intensity confirmed the preventive effect of probiotics against collagen degradation under ABA conditions. (F, I) At higher magnification, a strong loss of collagen fibres in the ABA group compared to the control (E, H) can be seen. In the ABA + P group (G, J) this structure was visibly repaired by the additional probiotic treatment. AB—alveolar bone; PDL—periodontal ligament; D—dentin. ****p < 0.0001.