A reduced level of the long non-coding RNA SNHG8 activates the NF-kappaB pathway by releasing functional HIF-1alpha in a hypoxic inflammatory microenvironment

Background A series of biochemical responses, including hypoxia and aseptic inflammation, occur in periodontal ligament cells (PDLCs) during periodontal tissue remodeling of orthodontic tooth movement (OTM). However, the role of long non-coding RNA (lncRNA) in these responses is still largely unknown. We investigated the role of the lncRNA SNHG8 in hypoxic and inflammatory responses during OTM, and explored the underlying mechanisms. Methods The expression pattern of SNHG8, and hypoxic and inflammatory responses under compressive force, were analyzed by qRT-PCR, immunohistochemistry, and western blotting, in vivo and in vitro. The effect of overexpression or knockdown of SNHG8 on the nuclear factor-kappaB (NF-κB) pathway was evaluated. RNA sequencing was performed for mechanistic analysis. The interaction between SNHG8 and hypoxia-inducible factor (HIF)-1α was studied using catRAPID, RNA immunoprecipitation, and RNA pulldown assays. The effect of the SNHG8–HIF-1α interaction on the NF-κB pathway was determined by western blotting. Results The NF-κB pathway was activated, and HIF-1α release was stabilized, in PDLCs under compressive force as well as in OTM model rats. The SNHG8 level markedly decreased both in vivo and in vitro. Overexpression of SNHG8 decreased the expression levels of inflammatory cytokines, the phosphorylation of p65, and the degradation of IκBα in PDLCs, whereas knockdown of SNHG8 reversed these effects. Mechanically, RNA sequencing showed that differentially expressed genes were enriched in cellular response to hypoxia after SNHG8 overexpression. SNHG8 binds to HIF-1α, thus preventing HIF-1 from activating downstream genes, including those related to the NF-κB pathway. Conclusion SNHG8 binds to HIF-1α. During OTM, the expression of SNHG8 dramatically decreased, releasing free functional HIF-1α and activating the downstream NF-κB pathway. These data suggest a novel lncRNA-regulated mechanism during periodontal tissue remodeling in OTM.

WNT-5a and SOST Levels in Gingival Crevicular Fluid Depend on the Inflammatory and Osteoclastogenic Activities of Periodontal Tissues

Background and Objectives: Wnt signaling leads to stimulation of osteoblasts and it reduces osteoclastogenesis and bone resorption via the regulation of the osteprotegrin and receptor activator of nuclear factor kappa-Β ligan (RANKL). Wnt signaling pathways are regulated by their physiological antagonists such as sclerostin (SOST) as well as WNT-5a. The aim of this study was to determine the total amount of Sclerostin and WNT-5a in the gingival crevicular fluid (GCF) in sites with a continuum from a healthy to diseased periodontium. Materials and Methods: In this cross-sectional study, a total of 20 patients with generalized periodontitis, 10 subjects with gingivitis as well as 14 individuals with a healthy periodontium were recruited upon clinical and radiographic periodontal examination. In patients diagnosed with periodontitis, GCF samples were collected from periodontitis, gingivitis and healthy sites, while gingivitis patients provided samples from gingivitis and healthy sites. In healthy patients, only healthy sites were sampled. Protein total amount of SOST and WNT-5a were quantified by sandwich enzyme-linked immunosorbent assay (ELISA). Results: A total of 108 GCF samples were collected from a total of 44 individuals. When all periodontitis (n = 51), gingivitis (n = 12) and healthy (n = 45) sites were analyzed regardless of the patient diagnosis, periodontitis sites demonstrated significantly elevated WNT-5a total amounts (p = 0.03) when compared to gingivitis sites. Gingivitis sites demonstrated a trend of more total SOST (p = 0.09) when compared to periodontitis and healthy sites. Within each patient diagnostic category, sites showed similar SOST and WNT-5a total amounts (p > 0.05). Conclusions: WNT-5a levels in GCF depend on the stage of periodontitis sites. SOST trended higher in the GCF of gingivitis sites but similar in chronic periodontitis and healthy sites. WNT-5a and SOST play a crucial role in periodontal tissue remodeling and depend on the inflammatory and osteoclastogenic activities.

Expression of Wnt10b, β-catenin, Rankl and Runx2 in the Periodontal Tissues During Orthodontic Tooth Movement in Rats

Objectiveto investigate the expression of Wnt10b, Rankl, β-catenin and Runx2 mRNA in periodontal tissues during orthodontic tooth movement (OTM) in rats; Materials and methods36 healthy male SD rats were selected and divided into six groups randomly according to the time of the force applied (0 hour, 12 hours, 24 hours, 5 days, 7 days, 14 days), with 6 rats in each group. 0hour group served as the control group. Except for control group, an orthodontic force of 50g was applied to the upper-left first molar with a nickel-titanium tension spring. The expression of Wnt10b, Rankl, β-catenin and Runx2 during OTM were detected by Real-time PCR (RT-PCR). ResultsCompared with the control group, Wnt10b mRNA expression in the pressure side was inhibited at the initial stage, and then increased, reached peak at day 5. A strong expression of Rankl mRNA in the pressure side can be seen from 12 hours to 14 days. The expression level was consistently higher than control group, and it peaked at day 7. The initial expression of β-catenin and Runx2 mRNA in the periodontal tissues of the experimental groups were small in the tension side, but still higher than the control group. The expression increased gradually with the extension of time, and reached the peak at day 7 after force treatment. ConclusionWnt10b, β-catenin, Rankl and Runx2 are related to periodontal tissues remodeling during OTM in rats; Wnt10b/β-catenin signaling pathways participate in periodontal tissue remodeling during OTM.

Long Non-coding RNA FER1L4 Mediates the Autophagy of Periodontal Ligament Stem Cells Under Orthodontic Compressive Force via AKT/FOXO3 Pathway

Orthodontic tooth movement is achieved by periodontal tissue remodeling triggered by mechanical force. It is essential to investigate the reaction of periodontal ligament stem cells (PDLSCs) for improving orthodontic therapeutic approaches. Autophagy is an endogenous defense mechanism to prevent mechanical damage of environmental change. Long non-coding RNAs (lncRNAs) are key regulators in gene regulation, but their roles are still largely uncharacterized in the reaction of PDLSCs during orthodontic tooth movement. In this study, we showed that autophagy was significantly induced in PDLSCs under compressive force, as revealed by the markers of autophagy, microtubule-associated protein light chain 3 (LC3) II/I and Beclin1, and the formation of autophagosomes. After the application of compressive force, lncRNA FER1L4 was strongly upregulated. Overexpression of FER1L4 increased the formation of autophagosome and autolysosomes in PDLSCs, while knockdown of FER1L4 reversed the autophagic activity induced by mechanical force. In mechanism, FER1L4 inhibited the phosphorylation of protein kinase B (AKT) and subsequently increased the nuclear translocation of forkhead box O3 (FOXO3) and thus mediated autophagic cascades under compressive strain. In mouse model, the expression of Lc3 as well as Fer1l4 was increased in the pressure side of periodontal ligament during tooth movement. These findings suggest a novel mechanism of autophagy regulation by lncRNA during periodontal tissue remodeling of orthodontic treatment.

Biomechanical Interfaces of Corticotomies on Periodontal Tissue Remodeling during Orthodontic Tooth Movement

Corticotomy is an effective approach in accelerating orthodontic tooth movement (OTM) in clinical treatment. Corticotomy causes regional acceleratory phenomenon (RAP) in the alveolar bone of surgical sites. However, the molecular mechanism of RAP after corticotomy remains unclear. Herein, we established a mouse model to study the biomechanical interfaces of corticotomy-assisted OTM and to investigate the histological responses and underlying cellular mechanism. A total of 144 male C57BL/6 mice were randomly assigned into four groups: corticotomy alone (Corti), sham operation (Sham), corticotomy with tooth movement (Corti + TM), and sham operation with tooth movement (Sham + TM). Nickel–titanium orthodontic springs were applied to trigger tooth movement. Mice were sacrificed on Post-Surgery Day (PSD) 3, 7, 14, 21, and 28 for radiographic, histological, immunohistochemical, and molecular biological analyses. The results reveal that corticotomy significantly promoted alveolar bone turnover and periodontal tissue remodeling. During orthodontic tooth movement, corticotomy significantly promoted osteogenic and proliferative activity, accelerated tooth movement, and eliminated root resorption by upregulating Wnt signal pathway.

Effects of LL-37 on Gingival Fibroblasts: A Role in Periodontal Tissue Remodeling?

Mounting evidence suggests that the host defence peptide, LL-37, plays a role in both inflammation and in wound healing; however, the role of this peptide in the remodeling and maintenance of oral tissues is not yet fully understood. Fibroblasts are the most abundant cell type within the periodontal tissues, and gingival fibroblasts play an important role in maintaining and repairing the gingival tissues which are constantly exposed to external insults. In this study we examined the direct effects of LL-37 treatment on gingival fibroblasts and found that LL-37 significantly increased secretion of both interleukin 8 (IL-8) and IL-6 from these cells. LL-37 tended to decrease matrix metalloproteinase (MMP) activity in gingival fibroblasts, but this decrease did not reach statistical significance. LL-37 significantly increased tissue inhibitor of metalloproteinase-1 (TIMP-1) production by gingival fibroblasts, but had no significant effect on TIMP-2 levels. LL-37 was also shown to significantly increase production of basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and keratinocyte growth factor (KGF) in gingival fibroblasts. Taken together, these results suggest an important role for the host defence peptide, LL-37, in modulating the fibroblast response to remodeling in periodontal tissues.