Local Induction of B Cell Interleukin-10 Competency Alleviates Inflammation and Bone Loss in Ligature-Induced Experimental Periodontitis in Mice

Abstract

Interleukin-10 (IL-10)-producing B cells (B10 cells) play a critical role in the immune system balance by negatively regulating inflammatory responses. This study was conducted to determine the effect of local B10 cell induction on periodontal inflammation and bone loss in ligature-induced experimental periodontitis in vivo. Purified spleen B cells from C57BL/6J mice (8 to 10 weeks old) were cultured with CD40 ligand (CD40L) and the Toll-like receptor 9 (TLR9) agonist cytidinephosphate-guanosine oligodeoxynucleotide (CpG) to determine effective IL-10 induction in vitro. Silk ligatures (size 7-0) were tied around the mouse maxillary second molars on day 0, followed by the injection of CD40L and CpG into the palatal gingiva on days 3, 6, and 9. All the mice were sacrificed, and samples were collected on day 14. CD40L and CpG significantly increased the level of IL-10 production by B cells in vitro, although the frequencies of CD1d hi CD5 ϩ and IL-10-producing (IL-10 ϩ ) CD45 ϩ cells were decreased. IL-10 was predominantly produced by the CD1d hi CD5 ϩ subpopulation of B cells. In vivo, both IL-10 mRNA expression and the number of IL-10 ϩ CD45 ϩ cells were significantly increased after gingival injection of CD40L and CpG. Periodontal bone loss was significantly decreased and the gingival expression of IL-1␤, tumor necrosis factor alpha, and RANKL was significantly reduced. The number of multinucleated tartrate-resistant acid phosphatase-positive cells along the alveolar bone surface was significantly decreased after gingival injection of CD40L and CpG. This study indicates for the first time that the local induction of B10 cell activity could inhibit periodontal inflammation and bone loss. 1-4). One of the most widely studied Breg subsets is termed interleukin-10 (IL-10)-producing B cells (B10 cells) (5) due to their pivotal capability to produce the anti-inflammatory cytokine IL-10. Currently, due to the lack of unique phenotypic markers, these IL-10-competent B cells are functionally identified by their production of IL-10 (6). Spleen B10 cells are predominantly enriched within the CD1d hi CD5 ϩ subpopulation, where among them more than 50% are competent to express 7). Although the frequency is low (1 to 3% of splenic B cells in mice and less than 1% of circulating B cells in humans), there is emerging evidence demonstrating that a deficiency of B10 cells exacerbates the

miR-146a regulates inflammatory cytokine production in Porphyromonas gingivalis lipopolysaccharide-stimulated B cells by targeting IRAK1 but not TRAF6

“…Studies have demonstrated that IL-10 is a potent anti-inflammatory cytokine [25, 26]. IL-10 can maintain bone mass through inhibiting osteoclastic bone resorption and enhancing osteoblastic bone formation.…”

“…Also, IL-10 was involved in local B cell response and detected in inflamed gingival tissues [19]. IL-10 could be secreted by T cells, B cells and other periodontal cells [10, 26, 28]. Studies demonstrated that miR-146a inhibition did not change IL-10 production in human gingival fibroblasts [10].…”

Osteoimmunology: Inflammatory osteolysis and regeneration of the alveolar bone

“…There is also evidence for a beneficial role of B cells in periodontal disease. In mouse models, IL10‐producing B cells in the gingiva decreased inflammation and alveolar bone resorption (Yu et al., ). These IL10‐producing cells increased in rheumatoid arthritis patients with disease remission but switched to RANKL‐producing B10 cells under pathological conditions (Hu et al., ).…”

OC‐STAMP promotes osteoclast fusion for pathogenic bone resorption in periodontitis via up‐regulation of permissive fusogen CD9

“…Total RNA was isolated from cells using RNA-Bee isolation reagent (AMS Biotechnology, Cambridge, MA, USA), following the manufacturer's instructions, and subjected to reverse transcription with the Verso cDNA Synthesis Kit (Thermo Fisher Scientific) in the presence of random primers and oligo-dT. Gene expression was quantified using the LightCycler 480 SYBR Green Mouse model of ligature-induced periodontitis OC-STAMP-KO mice (The Jackson Laboratory) or wild-type (WT) mice (6-8 wk old, C57BL/J male, n = 5-6) were placed with a 5-0 silk ligature (Ethicon, Guaynabo, Puerto Rico, USA) on the upper left second molar, whereas the upper right second molar without ligature was used as a control, following the protocol that was developed by our group (12,(24)(25)(26)(27). Because we used mice raised from different breeding cages, especially those of OC-STAMP-KO strains, to avoid the effect of different oral flora, the male mice used for the experiment were cohoused for a week so that all mice harbored similar oral flora, following the protocol reported by other groups (28,29).…”

“…Ligatures were removed, and the maxillae were defleshed using dermestid beetles (30). Alveolar bone resorption was measured as previously described (12,(24)(25)(26)(27)31). Briefly, the distance from the cementoenamel junction (CEJ) to the alveolar crest (AC) on the buccal side of each root was measured with a dissection microscope.…”