Experimental Periodontitis Model Research Articles

REV-ERBα Inhibits Osteoclastogenesis and Protects against Alveolar Bone Loss.

Circadian rhythm disruption is thought to be associated with periodontitis, and molecular clock genes play critical roles in regulating bone homeostasis. However, the specific contribution of molecular clock genes to alveolar bone resorption caused by periodontitis is poorly understood. In this study, we introduced a novel Periodontitis Circadian Rhythm Score (PeriCRS) model that was established through machine learning using periodontal transcriptomic data from periodontitis clinical cohorts in the Gene Expression Omnibus (GEO) database. This approach revealed the potential regulatory role of circadian rhythm disruption in periodontitis and identified key molecular clock genes associated with alveolar bone destruction. Moreover, we established an experimental periodontitis model with circadian rhythm disturbance via periodontal ligation in mice exposed to a 6-h advanced LD12:12 cycle every 2 d. Our bioinformatics analysis revealed that NR1D1, which encodes REV-ERBα, is a pivotal factor in the impact of circadian rhythm disruption on periodontitis in periodontal tissues. Next, we confirmed the abnormal expression of the molecular clock gene Rev-erbα in inflammatory periodontal tissue in mice and confirmed that circadian rhythm disruption altered REV-ERBα expression. Furthermore, the activation of REV-ERBα with the agonist SR9009 notably decreased RANKL-induced osteoclast differentiation and suppressed the expression of osteoclast-related factors. Subsequent in vivo experiments demonstrated that SR9009 mitigated alveolar bone loss caused by periodontitis. Mechanistically, we found that the IL-22-STAT3 pathway inhibited REV-ERBα expression and modulated RANKL-induced osteoclast differentiation in vitro. Our results elucidate the role of REV-ERBα in osteoclastogenesis and suggest a potential new therapeutic avenue for addressing alveolar bone resorption associated with periodontitis.

Detecting structural and functional mitochondrial abnormalities in periodontal tissues using an experimental periodontitis model

Background: Recent research indicates that mitochondrial dysfunction plays a significant role in the development and progression of oral inflammation, such as periodontitis and pulpitis. Aim: To assess structural and functional mitochondrial abnormalities in periodontal tissues using an experimental periodontitis model in rats. Materials and methods: The study used male Wistar rats aged 4 months, with a body weight of 221.0±7.5 g. Simple randomization was used to divide the animals into two groups (n=10 each). Group 1 (control) consisted of intact animals, while Group 2 consisted of animals with experimentally induced periodontitis. A ligature-induced periodontitis model was used, with a non-absorbable polyfilament thread sutured into the gum near the mandibular incisors. Histological examinations were used to validate the experimental periodontitis model. The following molecular genetic and biochemical parameters were assessed: nuclear DNA and mitochondrial DNA (mtDNA) damage, abundance and heteroplasmy of mtDNA, mitochondrial gene expression, and levels of hydrogen peroxide (H2O2), malondialdehyde, and reduced glutathione. Results: The resulting experimental periodontitis model revealed histological changes in periodontal tissues, indicating periodontitis in the animals. On day 14 after ligation, histological findings showed that Group 2 had more significant mtDNA damage and heteroplasmy in periodontal tissues than Group 1 (control). Moreover, Group 2 showed a decrease in the expression of mtDNA genes involved in adenosine triphosphate synthesis. This group also had lower glutathione levels and higher H2O2 and malondialdehyde levels than the control group. Conclusion: The experimental periodontitis model in rats revealed structural and functional mitochondrial abnormalities in periodontal tissues. New approaches to assessing mitochondrial function in periodontitis may facilitate the diagnosis and treatment of the disease and its complications.

Moisei Schwartz: A military surgeon, educator, and researcher

The personnel of the Department of Dentistry (after 1942, the Department of Maxillofacial Surgery and Dentistry) of the Military Medical Academy named after S.M. Kirov made major contributions to the advancement of dentistry and maxillofacial surgery in Russia. This essay focuses on the professional life of Associate Professor Moisei Schwartz, Major of the Medical Service, whose research significantly contributed to experimental dentistry and periodontology. The essay uses the data presented in Russian literature on dentistry and maxillofacial surgery, as well as the recollections of dentists who knew Moisei Schwartz personally: V.P. Zabelin, Colonel of the Medical Service, and Associate Professor V.V. Fiolkovsky, Colonel of the Medical Service. Associate Professor Moisei Schwartz, Major of the Medical Service, performed several studies of the functional properties of jaw vessels, primarily mandibular vessels. He was the first to use the Kravkov–Pisemsky technique in dentistry to assess the effect of biogenic amines on isolated jaw vessels. His comparative studies of ear and jaw vessels in dogs using this technique revealed that jaw vessels are hypersensitive to acetylcholine, histamine, and adrenaline. In 1936, Moisei Schwartz developed an experimental periodontitis model using chronic irritation (ligation) of the inferior alveolar nerve in dogs. Moisei Schwartz fought in the Soviet-Finnish War of 1939–1940, as well as the World War II (1941–1945). Moisei Schwartz was a prominent scientist and experienced doctor with good manual skills, including surgical care in maxillofacial wounds. Moreover, he was a passionate researcher and educator. His contributions to experimental dentistry and periodontology are impossible to exaggerate.

Hyperglycemia Exacerbates Periodontal Destruction via Systemic Suppression of Regulatory T Cell Number and Function.

Diabetes is a significant risk factor that exacerbates the pathological progression of periodontal disease. In recent years, attention has focused on the effect of regulatory T cells (Tregs), which play a central role in immune tolerance, on inflammatory processes in periodontal tissue, suggesting a link with diabetes-associated periodontitis. In this study, we examined the dynamics of Tregs in periodontal tissue of mice with streptozotocin (STZ)-induced hyperglycemia. Eleven-week-old male C57BL/6J mice were divided into four treatment groups: Untreated (C group), ligature placed around the maxillary second molars with silk sutures (PD group), intraperitoneal administration of STZ (HG group), and ligature placed after STZ administration (PHG group). Establishment of hyperglycemia was assessed 14 days after STZ administration, and ligation was performed 7 days later. After another 7 days of ligation, the mice were euthanized. The right side of the maxilla was observed histopathologically, whereas the palatal gingiva on the left side of the maxilla was analyzed genetically, and the microstructure of the alveolar bone was also assessed. In addition, lymphocytes from peripheral blood, spleen, and periodontal tissue were analyzed using flow cytometry. In bone structure analyses, alveolar bone height, bone volume/tissue volume (BV/TV), and bone mineral density (BMD) were lower in the PHG group than the PD group. In the gingival tissue, expression of the Foxp3 gene was up-regulated in the PHG group compared with the C group, and IL-17a was up-regulated in the PHG group compared with the PD group. Flow cytometry analyses showed that the number of Tregs (CD4+CD25+Foxp3+ cells) in the blood and gingival tissue was significantly higher in the PD and PHG groups than the C group. The number of CD4+CD25-Foxp3+ cells, which are reportedly functionally attenuated as Tregs, was increased in blood of the PHG group. Immunofluorescence staining of periodontal tissue showed that the number of CD25+Foxp3+ cells was significantly increased only in the PD group, whereas a trend toward an increased number of CD25-Foxp3+ cells was observed in the PHG group. The present study showed that STZ-induced hyperglycemia numerically and functionally attenuates Tregs in a mouse model of experimental periodontitis. Furthermore, impaired immune tolerance capacity appears to be involved in exacerbating inflammation and bone destruction in periodontal tissue.

Comparison of Micro-CT and Morphometric Outcomes in a Modified Experimental Rat Model of Periodontitis.

To assess the correlation between micro-computed tomography (micro-CT) and linear morphometric measurements in terms of mandibular bone levels in a modified experimental periodontitis model in rodents to study the mechanisms of association between periodontal destruction and neuroinflammation. The proposed invivo experimental periodontitis model involves the administration of oral rinses with Porphyromonas gingivalis and Fusobacterium nucleatum, four times per week during 4, 8 or 12 weeks, in 24 male Wistar Hannover rats (180 g, 5 weeks old). After euthanasia, hemi-mandibles were collected. One hemi-mandible was analysed using morphometry, while the other was assessed with micro-CT. Linear measurements were taken at the buccal aspect and furcation level for both techniques, and volumetric measurements were also obtained with micro-CT. Passing-Bablok regression analysis was used to compare the results of both techniques, with morphometric measurements serving as the reference. Moreover, Lin's Concordance correlation coefficient was calculated to assess the level of agreement. Periodontal clinical variables with neuroinflammatory parameters from the frontal cortex were used to evaluate the association between the resulting condition and neuroinflammation. Twenty-one out of the initial 24 rats were analysed. The micro-CT linear measurements demonstrated high concordance values with the linear morphometric measurements at the buccal surfaces of the roots in molars (r = 0.714) but not at the furcation area (r = 0.052). At 12 weeks, there was a significant impact on neuroinflammation with significant decreases in iNOS levels and p-mTOR levels at 4 and 8 weeks. The proposed invivo experimental periodontitis model demonstrated a high degree of correlation between morphometric and micro-CT measurements in buccal areas but not at the furcation level. Concomitantly, there was a significant temporary modulation of the neuroinflammatory response.

The effect of a new 3-hydroxypyridine derivative LHT-2-20 on free radical oxidation in experimental periodontitis

Aim. To study the effect of a new complex compound LHT-2-20 (2-ethyl-6-methyl-3-hydroxypyridine-2-(3benzoyl phenyl)-propanoate) on free radical oxidation in experimental periodontitis.Materials and methods. The experimental study was performed on 195 white mongrel mice weighing 19–23 g and 137 white mongrel rats weighing 180–220 g. The effect of a new complex compound LHT-2-20 (2-ethyl-6methyl-3-hydroxypyridine-2-(3-benzoyl phenyl)-propanoate) on the intensity of free radical oxidation and the local state of periodontal tissues during a course of intragastric administration was studied on the experimental model of periodontitis. Statistical processing of the results was carried out using the SPSS Statistics 20.0 software package with the analysis of variance (ANOVA) and the parametric Tukey’s test.Results. The LHT-2-20 compound reduced elevated levels of primary and secondary lipoperoxidation products (conjugated dienes, malondialdehyde in plasma and in erythrocytes during spontaneous and iron-induced oxidation) already at the early stages of the experiment, bringing the studied parameters closer to the reference values by the end of the course of treatment. The use of the compound LHT-2-20 contributed to an increase in the activity of the main antioxidant enzymes (catalase and superoxide dismutase), normalizing them to baseline values by the end of the experiment. With the correction of free radical processes, the use of LHT-2-20 limited the local inflammatory response in periodontal tissues, which was confirmed by a decrease in gingival edema and hyperemia, bleeding, depth of periodontal pockets, and tooth mobility.Conclusion. The results of this study confirm the anti-inflammatory potential of the compound and the multiplicity of its effects due to the impact on the mechanisms of oxidative stress. The expediency of further study of the drug is justified by the prospect of creating a new drug and its subsequent wide clinical application as part of the complex therapy of periodontal inflammation.

Soluble epoxide hydrolase inhibition impairs triggering receptor expressed on myeloid cells-1 in periodontal tissue.

Periodontitis is a prevalent inflammatory disorder affecting the oral cavity, driven by dysbiotic oral biofilm and host immune response interactions. While the major clinical focus of periodontitis treatment is currently controlling oral biofilm, understanding the immune response is crucial to prevent disease progression. Soluble epoxide hydrolase (sEH) inhibition has shown promise in preventing alveolar bone resorption. Triggering receptors expressed on myeloid cells (TREMs) play pivotal roles in regulating inflammation and bone homeostasis, and dysregulation of TREM signaling is implicated in periodontitis. Here, we investigated the impact of sEH inhibition on TREM 1 and 2 expression, associated with inflammatory cytokines, and histologically assessed the inflammatory infiltrate in periodontal tissue. The experimental periodontitis model was induced by placing a ligature around the upper second molar. For 14 days, animals were treated daily with a sEH inhibitor (TPPU) or vehicle. The alveolar bone loss was examined using a methylene blue stain. Gingival tissues were used to measure the mRNA expression of TREM-1, TREM-2, IKKβ, NF-κB, IL-1β, IL-6, IL-8, and TNF-α by RT-qPCR. Another set of experiments was performed to determine the histological inflammatory scores. In a ligature-induced periodontitis model, sEH inhibition prevented alveolar bone loss and reduced TREM1 expression, albeit with a slight elevation compared to the disease-free group. In contrast, TREM2 expression remained elevated, suggesting sustained immunomodulation favoring resolution. The inhibition of sEH reduced the expression of NF-κB, IL-1β, and TNF-α, while no differences were found in the expression of IL-6, IL-8, and IKKβ. In histological analysis, sEH inhibition reduced the inflammatory leukocyte infiltrate in periodontal tissues close to the ligature. These findings underscore the potential of sEH inhibition to modulate periodontal inflammation by regulating TREM-1 alongside decreased IL-1β and TNF-α expression, highlighting a promising therapeutic approach for periodontitis management.

Bmi‐1 alleviates alveolar bone resorption through the regulation of autophagy

AbstractBackgroundB‐cell‑specific Moloney MLV insertion site‐1(Bmi‐1)is a crucial osteopenic target molecule. The aim of this study is to explore the effects of Bmi‐1 on alveolar bone resorption and the underlying mechanisms in vitro and vivo.MethodsA Bmi‐1‐knockout (Bmi‐1−/−) mouse model was used to investigate the effect of Bmi‐1 on alveolar bone metabolism, with micro‐computed tomography imaging, histology, and immunohistochemistry staining. Furthermore, we utilized a ligature‐induced experimental periodontitis model to examine the impact of Bmi‐1‐knockdown (Bmi‐1±) on inflammatory alveolar bone resorption. Finally, we stimulated human periodontal ligament stem cells (hPDLSCs) with lipopolysaccharide (LPS) to explore the potential mechanism of Bmi‐1 overexpression in the process of osteogenesis.ResultsCompared with wild‐type mice, Bmi‐1−/− mice demonstrated more alveolar bone resorption by inhibiting osteogenesis, which was characterized by decreases in Runt‐related transcription factor 2 and type 1 collagen formation. In addition, Bmi‐1−/− mice had lower levels of autophagy markers such as Parkin and LC3, but higher levels of inflammation‐related factors such as interleukin (IL)‐6 and IL‐1β in periodontal tissues. In addition, Bmi‐1‐knockdown aggravated ligature‐induced alveolar bone loss. Under in vitro inflammatory conditions, Bmi‐1 overexpression stimulated osteoblast differentiation and inhibited the production of inflammatory factors, as well as the autophagy and apoptosis in hPDLSCs stimulated with LPS. When 3‐methyladenine (3‐MA), an autophagy inhibitor, was added, the osteogenic effect of Bmi‐1 was further enhanced.ConclusionsBmi‐1 alleviates alveolar bone resorption by regulating autophagy, indicating that it could be a potential target for periodontitis prevention and treatment.Plain Language SummaryPeriodontitis is a chronic inflammatory disease, which leads to progressive destruction of periodontal tissues, manifested as periodontal pocket formation, loss of periodontal attachment and alveolar bone resorption. Currently, there is a lack of effective treatments to regenerate damaged periodontal tissues. Therefore, it is of great clinical significance to explore new mechanisms of periodontitis and effective intervention targets. B‐cell‑specific Moloney MLV insertion site‐1 (Bmi‐1) is involved in the regulation of the cell cycle, DNA damage repair, autophagy, bone metabolism, tumor, and other physiopathological processes. Autophagy, as an important mechanism of intracellular self‐regulation, plays an indispensable role in the destruction and repair of periodontal tissues. The aim of this study was to investigate the role of Bmi‐1 on periodontal tissues and its intrinsic mechanism. The results revealed that Bmi‐1 regulates autophagy to protect periodontal tissues, suggesting that it may be a potential target for the prevention and treatment of periodontitis.

The impact of omega-3 fatty acids in a combined experimental model of periodontitis and metabolic syndrome

Relevance. Periodontitis and metabolic syndrome are interrelated conditions that often aggravate each other through shared pathogenic mechanisms, highlighting the need for integrated immunomodulatory therapeutic approaches. Despite this, existing literature presents inconsistent data regarding the interplay between periodontitis and metabolic syndrome, as well as limited insights into the potential benefits of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) as an adjuvant therapy. Specifically, the role of ω-3 PUFAs in modulating the inflammatory response and mitigating bone resorption remains inadequately explored. Therefore, this study aims to evaluate the inflammatory response and osteoresorption in periodontal tissues under the combined conditions of periodontitis and metabolic syndrome, while also assessing the therapeutic effects of ω-3 PUFAs.Materials and methods. This experimental study was conducted using 30 male Wistar rats, which were randomly assigned to one of five groups: a control group, a periodontitis group, a combined periodontitis and metabolic syndrome group, a periodontitis group treated with omega-3 fatty acids, and a combined periodontitis and metabolic syndrome group treated with omega-3 fatty acids. The study employed several research methodologies. Biochemical analyses were performed to confirm the presence of metabolic syndrome. Histopathological and morphometric assessments were conducted to evaluate the inflammatory response, the extent of osteoresorption, and the number of osteoclasts in the periodontal tissues. Additionally, molecular genetic analysis was used to measure the relative mRNA expression levels of key inflammatory and osteoclastogenic markers, including TNFα, IL-1β, RANK, and OPG.Results. The data demonstrated that metabolic syndrome in laboratory animals significantly exacerbates both the pro-inflammatory response and the extent of lacunar osteoclastic bone resorption. Administration of ω-3 PUFAs at a dose of 40 mg/kg over 30 days led to a statistically significant reduction in inflammatory infiltration within the periodontal tissues (3.17 ± 0.21 in the control group vs. 1.83 ± 0.21 in the treated group, р = 0.001) and a decrease in the number of osteoclasts (3,75 ± 0,45 in the control group vs. 1,75 ± 0,35 in the treated group, р = 0.003). Additionally, there was a notable reduction in the expression levels of TNFα, IL-1β, and RANK mRNA. Similar but less pronounced effects were observed in the group with combined metabolic syndrome following ω-3 PUFA administration.Conclusion. These findings enhance the current understanding of the interplay between periodontitis and metabolic syndrome, highlighting the potential of ω-3 PUFAs as a preventive or therapeutic intervention. ω-3 PUFAs may be effective in mitigating inflammatory responses and bone resorption, both in cases of isolated periodontitis and when accompanied by metabolic syndrome.

Induced Periodontitis in Rats With Three Ligature Types: An Exploratory Study.

The placement of ligatures in the cervical area of rat molars is considered as a predictable model to induce periodontitis. The present explorative study aimed to compare the efficacy of metal wires (MWs), without or with sandblasting, versus silk ligatures (SLs) in inducing periodontal bone loss in rats. Twenty-four Wistar rats were randomly divided into three groups of eight rats that received three different types of ligatures (MW, sandblasted wire [SMW], and SL) around their first right mandibular molar, while the contralateral tooth was left without the ligature and served as a control. Bone loss was assessed by measuring the distance from the cementoenamel junction (CEJ) to the bone crest at the distal aspect of the first molar on central mesiodistal sections generated from micro-CT scans taken 24 and 35 days after ligature placement. In the SL group, only in two rats the ligatures were retained until the end of the 24-day period; in all other animals, the ligatures were lost at some time point. In the SMW, the ligatures were retained only for the 24-day period. In the MW group, no ligatures were lost. Irrespective of the group or experimental period, the difference in the crestal bone level between ligated and control teeth was in most cases z < 0.20 mm, that is, in 19 out of 25 pairs of teeth. In a few cases, the bone crest was more apically located at the control teeth compared to the ligated ones (four cases each, during both 24- and 35-day experimental periods). Bone loss was minimal during the experimental period, with no significant differences between the test and control teeth, or among the three types of ligatures. MWs, not even roughened, do not seem to be a better alternative to SLs for inducing bone loss in the experimental periodontitis model in the rat. This assumption, however, has to be confirmed in a larger, well-powered study.

Neural epidermal growth factor-like 1 protein (Nell-1) alleviates periodontal tissue destruction in periodontitis by regulating the ratio of M2/M1 macrophage phenotypes

BackgroundPeriodontitis is a common oral disease with high prevalence worldwide. Neural epidermal growth factor-like 1 protein (Nell-1) has recently been reported to have anti-inflammation effects and may be a drug candidate for osteoarthritis. However, its immunotherapeutic effects in periodontitis remain unknown. Therefore, this study aimed to investigate the effects of Nell-1 on periodontitis in terms of macrophage polarization and analyze its possible underlying mechanism. MethodsA rat ligation-induced experimental periodontitis model was established and locally injected with Nell-1 (n = 6/group). Periodontal tissue destruction and macrophage polarization in vivo were analyzed using micro-CT, histology analysis, and western blot. Enzyme-linked immunosorbent assay was used to evaluate serum inflammatory cytokines. Then, the RAW 264.7 macrophage cells were treated with lipopolysaccharide (LPS), Nell-1, and the c-Jun N-terminal kinases (JNK) inhibitor (SP600125). RT-PCR, western blot, and flow cytometry were performed to further analyze the effect of Nell-1 on macrophage polarization and the underlying mechanism in vitro. ResultsLocal treatment with Nell-1 significantly alleviated the destruction of alveolar bone and fibers in periodontitis, and upregulated the ratio of M2/M1 macrophages in periodontal tissues (P < 0.05). In vitro, Nell-1 at the concentrations of 200 and 500 ng/mL could significantly inhibit the expression of M1-related inflammatory factors in LPS-stimulated macrophages, and increase the expression of M2-related markers, regulating the macrophage phenotype switch into M2 (P < 0.05). The mRNA of JNK and relative protein level of phospho-JNK/JNK were also upregulated by Nell-1 (P < 0.05). Additionally, the JNK inhibitor (SP600125) could reverse the effect of Nell-1 on macrophage polarization (P < 0.05). ConclusionsNell-1 could modulate the ratio of M2/M1 macrophages possibly through the JNK/MAPK signaling pathway, subsequently attenuating the inflammation and destruction of periodontal tissues caused by periodontitis.

Prunin Laurate Derived from Natural Substances Shows Antibacterial Activity against the Periodontal Pathogen Porphyromonas gingivalis.

Periodontal disease is an inflammatory disease caused by infection with periodontopathogenic bacteria. Oral care is essential to prevent and control periodontal disease, which affects oral and systemic health. However, many oral hygiene products currently on the market were developed as disinfectants, and their intense irritation makes their use difficult for young children and older people. This study investigated the antibacterial effects of prunin laurate (Pru-C12) and its analogs on periodontopathogenic bacteria, Porphyromonas gingivalis (P. gingivalis). Pru-C12 and its analogs inhibited in vitro bacterial growth at more than 10 μM and biofilm formation at 50 µM. Among its analogs, only Pru-C12 showed no cytotoxicity at 100 µM. Three of the most potent inhibitors also inhibited the formation of biofilms. Furthermore, Pru-C12 inhibited alveolar bone resorption in a mouse experimental periodontitis model by P. gingivalis infection. These findings may be helpful in the development of oral hygiene products for the prevention and control of periodontal disease and related disorders.

Capsaicin attenuates Porphyromonas gingivalis-suppressed osteogenesis of periodontal ligament stem cells via regulating mitochondrial function and activating PI3K/AKT/mTOR pathway.

Prevention of periodontal bone resorption triggered by Porphyromonas gingivalis (P. gingivalis) is crucial for dental stability. Capsaicin, known as the pungent ingredient of chili peppers, can activate key signaling molecules involved in osteogenic process. However, the effect of capsaicin on osteogenesis of periodontal ligament stem cells (PDLSCs) under inflammation remains elusive. P. gingivalis culture suspension was added to mimic the inflammatory status after capsaicin pretreatment. The effects of capsaicin on the osteogenesis of PDLSCs, as well as mitochondrial morphology, Ca2+ level, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and osteogenesis-regulated protein expression levels were analyzed. Furthermore, a mouse experimental periodontitis model was established to evaluate the effect of capsaicin on alveolar bone resorption and the expression of osteogenesis-related proteins. Under P. gingivalis stimulation, capsaicin increased osteogenesis of PDLSCs. Not surprisingly, capsaicin rescued the damage to mitochondrial morphology, decreased the concentration of intracellular Ca2+ and ROS, enhanced MMP and activated phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway. The invivo results showed that capsaicin significantly attenuated alveolar bone loss and augmented the expression of bone associated proteins. Capsaicin increases osteogenesis of PDLSCs under inflammation and reduces alveolar bone resorption in mouse experimental periodontitis.

HA2-FimA DNA Vaccine Treats Experimental Periodontitis.

To study the therapeutic effect of hemagglutinin-2 and fimbrial (HA2-FimA) vaccine on experimental periodontitis in rats. The first batch of rats was divided into two groups and immunised with pure water or pVAX1-HA2-FimA at the age of 6, 7, and 9 weeks. After sacrificing the animals, total RNA was extracted from the spleens for RNA high-throughput sequencing (RNA-Seq) analysis. The second batch of rats was divided into four groups (A, B, C, D), and an experimental periodontitis rat model was established by suturing silk thread around the maxillary second molars of rats in groups B, C, and D for 4 weeks. The rats were immunised with pure water, pVAX1-HA2-FimA vaccine, empty pVAX1 vector, and pure water at 10, 11, and 13 weeks of age, respectively. Secretory immunoglobulin A (SIgA) antibodies and cathelicidin antimicrobial peptide (CAMP) levels in saliva were measured by enzyme-linked immunosorbent assay (ELISA). All rats were euthanised at 17 weeks of age, and alveolar bone loss was examined using micro-computed tomography (Micro-CT). Through sequencing analysis, six key genes, including Camp, were identified. Compared with the other three groups, the rats in the periodontitis+pVAX1-HA2-FimA vaccine group showed higher levels of SIgA and CAMP (p < 0.05). Micro-CT results showed significantly less alveolar bone loss in the periodontitis+pVAX1-HA2-FimA vaccine group compared to the periodontitis+pVAX1 group and periodontitis+pure water group (p < 0.05). HA2-FimA DNA vaccine can increase the levels of SIgA and CAMP in the saliva of experimental periodontitis model rats and reduce alveolar bone loss.

Unraveling ferroptosis in osteogenic lineages: implications for dysregulated bone remodeling during periodontitis progression

Periodontitis is a highly prevalent disease characterized by inflammation and destruction of tooth-supporting tissues that leads to tooth loss in extreme situations. Elucidating the underlying mechanisms of periodontitis pathogenesis and progression will establish the groundwork for developing effective treatment strategies. Recently, evidence concerning the role of ferroptosis in periodontitis progression has emerged. Osteogenic lineage cells are key regulators of bone remodeling. Osteogenic cell death, as observed in experimental periodontitis models, disrupts the balance between bone resorption and bone formation. However, whether the osteogenic lineage undergoes ferroptosis during periodontitis and the corresponding effect on periodontitis progression remain elusive. Here, we investigated cell-specific ferroptosis within the alveolar bone in a murine periodontitis model. Through immunofluorescence double staining and immunohistochemistry, we identified ferroptotic osteocytes and osteoblasts in inflammatory alveolar bone. Next, in vivo administration of erastin or liproxstatin-1 was conducted to either induce or inhibit ferroptosis, respectively. Severe bone resorption and inflammation, accompanied by increased osteoclast formation and impaired osteogenic potential were detected following ferroptosis activation. Subsequently, we carried out in vitro experiments on osteocytes and further verified that ferroptosis enhanced the osteocytic expression of RANKL and IL-6. These findings suggest that ferroptosis occurring within the osteogenic lineage acts as a catalyst in the progression of periodontitis by stimulating osteoclastogenesis through the secretion of inflammatory cytokines and inhibiting osteoblastic function, providing insights into ferroptosis-induced alterations in microenvironment-based intercellular communication. Ferroptosis is a promising target for controlling inflammation and preventing bone resorption in periodontitis.

Transcriptomic features of programmed and inflammatory cell death in gingival tissues.

This study used an experimental periodontitis model to detail the gingival transcriptome related to cell death processes of pyroptosis, necroptosis, ferroptosis, and cuproptosis. Healthy Macaca mulatta primates stratified by age, ≤3 years (young), 7-12 years (adolescent), 12-15 years (adult), and 17-23 years (aged), provided gingival tissue biopsies for microarray analysis focused on 257 genes representative of the four cell death processes and bacterial plaque samples for 16S rRNA gene analysis. Age differences in the profiles of gene expression in healthy tissues were noted for cuproptosis, ferroptosis, necroptosis, and pyroptosis. Major differences were then observed with disease initiation, progression, and resolution also related to the age of the animals. Distinct bacterial families/consortia of species were significantly related to the gene expression differences for the cell death pathways. These results emphasized age-associated differences in the gingival tissue molecular response to changes in the quality and quantity of bacteria accumulating with the disease process reflected in regulated cell death pathways that are both physiological and pathophysiological.

IRE1α regulates macrophage polarization in type 2 diabetic periodontitis through promoting endoplasmic reticulum stress

ObjectivesThe aim of this study was to investigate the effect of 4μ8c, an inhibitor targeting the endoplasmic reticulum stress-associated factor IRE1α, on macrophage polarization in an experimental model of diabetic periodontitis through ex vivo experiments. Materials and methodsLocal alveolar bone parameters were evaluated using Micro-CT following intraperitoneal administration of 4μ8c in mice with experimental diabetic periodontitis. Surface markers indicating macrophage polarization were identified using immunofluorescence. In vitro experiments were performed employing bone marrow-derived macrophages and gingival fibroblasts. Macrophage polarization was determined using flow cytometry. Principal impacted signaling pathways were identified through Western blot analysis. ResultsResults from both in vitro and in vivo experiments demonstrated that 4μ8c mitigated alveolar bone resorption and inflammation in mice with diabetic periodontitis. Furthermore, it modulated macrophage polarization towards the M2 phenotype and augmented M2 macrophage polarization through the MAPK signaling pathway. ConclusionsThese findings suggest that inhibiting IRE1α can modulate macrophage polarization and alleviate ligature-induced diabetic periodontitis via the MAPK signaling pathway. This unveils a novel mechanism, offering a scientific foundation for the treatment of experimental diabetic periodontitis.

SorCS2 is involved in promoting periodontitis-induced depression-like behaviour in mice.

Emerging evidence supports the association between periodontitis and depression, although the mechanisms are unclear. This study investigated the role of SorCS2 in the pathogenesis of periodontitis-induced depression. An experimental periodontitis model was established using SorCS2 knockout mice and their wild-type littermates, and depression-like behaviour was evaluated. The expression of proBDNF signalling, neuronal activity, and glutamate-associated signalling pathways were further measured by western blotting and immunofluorescence. In addition, neuroinflammatory status, astrocytic and microglial markers, and the expression of corticosterone-related factors were measured by immunofluorescence, western blotting, and enzyme-linked immunosorbent assays. SorCS2 deficiency alleviated periodontitis-induced depression-like behaviour in mice. Further results suggested that SorCS2 deficiency downregulated the expression of pro-BDNF and glutamate signalling and restored neuronal activities in mice with periodontitis. Neuroinflammation in the mouse hippocampus was triggered by experimental periodontitis but was not affected by SorCS2 deficiency. The levels of corticosterone and the expression of glucocorticoid receptors were also not altered. Our study, for the first time, reveals the critical role of SorCS2 in the pathogenesis of periodontitis-induced depression. The underlying mechanism involves proBDNF and glutamate signalling in the hippocampus, providing a novel therapeutic target for periodontitis-associated depression.

Impact of circadian clock protein Bmal1 on experimentally-induced periodontitis-associated renal injury.

To investigate the mechanism of circadian clock protein Bmal1 (Bmal1) on renal injury with chronic periodontitis, we established an experimental rat periodontitis model. Twelve male Wistar rats were randomly divided into control and periodontitis groups (n=6, each group). The first maxillary molars on both sides of the upper jaw of rats with periodontitis were ligated by using orthodontic ligature wires, whereas the control group received no intervention measures. After 8 weeks, clinical periodontal parameters, including probing depth, bleeding index, and tooth mobility, were evaluated in both groups. Micro-CT scanning and three-dimensional image reconstruction were performed on the maxillary bones of the rats for the assessment of alveolar bone resorption. Histopatholo-gical observations of periodontal and renal tissues were conducted using hematoxylin-eosin (HE) and periodic acid-Schiff (PAS) staining. Renal function indicators, such as creatinine, albumin, and blood urea nitrogen levels, and oxidative stress markers, including superoxide dismutase, glutathione, and malondialdehyde levels, were measured using biochemical assay kits. MitoSOX red staining was used to detect reactive oxygen species (ROS) content in the kidneys. The gene and protein expression levels of Bmal1, nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) in rat renal tissues were assessed using real-time quantitative polymerase chain reaction (RT-qPCR) and immunohistochemical staining. Micro-CT and HE staining results showed significant bone resorption and attachment loss in the maxillary first molar region of the periodontitis group. Histological examination through HE and PAS staining revealed substantial histopathological damage to the renal tissues of the rats in the periodontitis group. The findings of the assessment of renal function and oxidative stress markers indicated that the periodontitis group exhibited abnormal levels of oxidative stress, whereas the renal function levels showed abnormalities without statistical significance. MitoSOX Red staining results showed that the content of ROS in the renal tissue of the periodontitis group was significantly higher than that of the control group, and RT-qPCR and immunohistochemistry results showed that the expression levels of Bmal1, Nrf2, and HO-1 in the renal tissues of the rats in the periodontitis group showed a decreasing trend. Circadian clock protein Bmal1 plays an important role in the oxidative damage process involved in the renal of rats with periodontitis.

Characterization and application of in situ curcumin/ZNP hydrogels for periodontitis treatment

BackgroundPeriodontitis is a chronic inflammatory disease that occurs in tooth-supporting tissues. Controlling inflammation and alleviating periodontal tissue destruction are key factors in periodontal therapy. This study aimed to develop an in situ curcumin/zinc oxide (Cur/ZNP) hydrogel and investigate its characteristics and effectiveness in the treatment of periodontitis.MethodsAntibacterial activity and cytotoxicity assays were performed in vitro. To evaluate the effect of the in situ Cur/ZNP hydrogel on periodontitis in vivo, an experimental periodontitis model was established in Sprague‒Dawley rats via silk ligature and inoculation of the maxillary first molar with Porphyromonas gingivalis. After one month of in situ treatment with the hydrogel, we examined the transcriptional responses of the gingiva to the Cur/ZNP hydrogel treatment and detected the alveolar bone level as well as the expression of osteocalcin (OCN) and osteoprotegerin (OPG) in the periodontal tissues of the rats.ResultsCur/ZNPs had synergistic inhibitory effects on P. gingivalis and good biocompatibility. RNA sequencing of the gingiva showed that immune effector process-related genes were significantly induced by experimental periodontitis. Carcinoembryonic antigen-related cell adhesion molecule 1 (Ceacam1), which is involved in the negative regulation of bone resorption, was differentially regulated by the Cur/ZNP hydrogel but not by the Cur hydrogel or ZNP hydrogel. The Cur/ZNP hydrogel also had a stronger protective effect on alveolar bone resorption than both the Cur hydrogel and the ZNP hydrogel.ConclusionThe Cur/ZNP hydrogel effectively inhibited periodontal pathogenic bacteria and alleviated alveolar bone destruction while exhibiting favorable biocompatibility.