Effects of periodontal pathogen-induced intestinal dysbiosis on transplant immunity in an allogenic skin graft model

Abstract

Periodontal disease can induce dysbiosis, a compositional and functional alteration in the microbiota. Dysbiosis induced by periodontal disease is known to cause systemic inflammation and may affect transplant immunity. Here, we examined the effects of periodontal disease-related intestinal dysbiosis on transplant immunity using a mouse model of allogenic skin graft in which the mice were orally administered the periodontal pathogen Porphyromonas gingivalis (Pg). For 6 weeks, the Pg group orally received Pg while the control group orally received phosphate-buffered saline solution. After that, both groups received allogenic skin grafts. 16 s rRNA analysis of feces revealed that oral administration of Pg significantly increased three short chain fatty acids (SCFAs) producing genera. SCFA (acetate and propionate) levels were significantly higher in the Pg group (p = 0.040 and p = 0.005). The ratio of regulatory T cells, which are positively correlated with SCFAs, to total CD4+ T cells in the peripheral blood and spleen was significantly greater (p = 0.002 and p < 0.001) in the Pg group by flowcytometry. Finally, oral administration of Pg significantly prolonged skin graft survival (p < 0.001) and reduced pathological inflammation in transplanted skin grafts. In conclusion, periodontal pathogen-induced intestinal dysbiosis may affect transplant immunity through increased levels of SCFAs and regulatory T cells. (198 words).

Figure 1

Study design. All mice used in this experiment were given antibiotics in their drinking water for one week. After that, mice in the Porphyromonas gingivalis group received Pg orally twice per week for 6 weeks, while mice in the control group received PBS orally twice per week for 6 weeks. (a) Both groups were sacrificed before transplantation. (b) Both groups received allogenic skin grafts and were monitored until skin graft rejection. (c) Both groups received allogenic skin grafts. On day 8 after skin grafting, the mice in both groups were sacrificed.

Figure 2

Analysis of the microbiota in both groups. (a) Unweighted UniFrac distances between the groups (red: Pg; green: control). (b) Taxonomic bar plots at the order level for both groups. (c–h) There was a significant change in the relative abundance of six genera between groups (four genera increased, and two genera decreased). (c) Alloprevotella, (d) Lachnospiracae_NK4A136_group, (e) Oscillibacter, (f) Parasutterella, (g) Enterorhabdus, (h) Lachnospiracae_FCS020_group. Each group: n = 5. *p < 0.05, ** p < 0.005.

Figure 3

SCFAs in both groups. Short-chain fatty acid (SCFA) levels in the feces of mice from the Pg group and the control group, as determined by gas chromatography (each group: n = 5). (a) Acetate, (b) propionate, (c) butyrate, (d) valerate, (e) total SCFAs (total concentrations of acetate, propionate, butyrate, valerate, and caproate). *p < 0.05, **p < 0.005.

Figure 4

Flow cytometry analysis of cells from both groups. The proportion of Tregs out of CD4+ T cells in the peripheral blood (a) and spleen (c) in the Pg group and the control group before transplantation, as determined by flow cytometry (each group: n = 6). Statistical analysis was performed to identify differences between the groups. (b) Peripheral blood, (d) spleen. *p < 0.05, **p < 0.005.

Figure 5

Skin graft survival and pathological changes in skin grafts in both groups. (a) Skin graft survival between the groups (each group: n = 7). On day 8 after skin grafting, the degree of inflammation of the skin graft was evaluated pathologically (hematoxylin/eosin staining). (b) × 2 magnification. The bar scale is 500 µm, × 10 magnification The bar scale is 100 µm. (c) Percentage of inflammatory cell infiltration in both groups (each group: n = 3). *p < 0.05, **p < 0.005.