SPOCK1 induces adipose tissue maturation: New insights into the function of SPOCK1 in metabolism.

SPOCK1 is a calcium-binding matricellular proteoglycan that has been extensively studied in several cancer cells. Previously, we generated a mouse line overexpressing SPOCK1 (Spock1-Tg mouse) and showed that SPOCK1 might play an important role in drug-induced gingival overgrowth, indicating that it possesses physiological functions in non-cancer diseases as well. Although SPOCK1 was reported to be secreted from human adipocytes, its role in adipocyte physiology has not been addressed yet. In this study, SPOCK1 protein expression was confirmed in pancreas, adipose tissues, spleen, and liver of normal diet (ND)-fed mice. Interestingly, SPOCK1 was up-regulated in the pancreas and adipose tissues of the high-fat diet (HFD)-fed mice. Spock1-Tg mice fed with ND showed increased maturation in epididymal and inguinal adipose tissues. In addition, Spock1 overexpression strongly decreased expression of UCP-1 in adipose tissues, suggesting that SPOCK1 might regulate thermogenic function through suppression of UCP-1 expression. Finally, exogenous SPOCK1 treatment directly accelerated the differentiation of 3T3-L1 adipocytes, accompanied by the up-regulation of adipocyte differentiation-related gene expression. In conclusion, we demonstrated for the first time that SPOCK1 induced adipocyte differentiation via the up-regulation of adipogenesis-related genes.

Luteolin Is a Potential Immunomodulating Natural Compound against Pulpal Inflammation.

Aim: The present study evaluated the therapeutic effects of luteolin in alleviating pulpitis of dental pulp- (DP-) derived microvesicles (MVs) via the inhibition of protein kinase R- (PKR-) mediated inflammation. Methodology. Proteomic analysis of immortalized human dental pulp (DP-1) cell-derived MVs was performed to identify PKR-associated molecules. The effect of luteolin on PKR phosphorylation in DP-1 cells and the expression of tumor necrosis factor-α (TNF-α) in THP-1 macrophage-like cells were validated. The effect of luteolin on cell proliferation was compared with that of chemical PKR inhibitors (C16 and 2-AP) and the unique commercially available sedative guaiacol-parachlorophenol. In the dog experimental pulpitis model, the pulps were treated with (1) saline, (2) guaiacol-parachlorophenol, and (3) luteolin. Sixteen teeth from four dogs were extracted, and the pulp tissues were analyzed using hematoxylin and eosin staining. Immunohistochemical staining was performed to analyze the expression of phosphorylated PKR (pPKR), myeloperoxidase (MPO), and CD68. Experimental endodontic-periodontal complex lesions were established in mouse molar through a silk ligature and simultaneous MV injection. MVs were prepared from DP-1 cells with or without pretreatment with 2-AP or luteolin. A three-dimensional microcomputed tomography analysis was performed on day 7 (n = 6). Periodontal bone resorption volumes were calculated for each group (nonligated-ligated), and the ratio of bone volume to tissue volume was measured. Results: Proteomic analysis identified an endogenous PKR activator, and a protein activator of interferon-induced PKR, also known as PACT, was included in MVs. Luteolin inhibited the expressions of pPKR in DP-1 cells and TNF-α in THP-1 cells with the lowest suppression of cell proliferation. In the dog model of experimental pulpitis, luteolin treatment suppressed the expression of pPKR-, MPO-, and CD68-positive cells in pulp tissues, whereas guaiacol-parachlorophenol treatment caused coagulative necrosis and disruption. In a mouse model of endodontic-periodontal complex lesions, luteolin treatment significantly decreased MV-induced alveolar bone resorption. Conclusion: Luteolin is an effective and safe compound that inhibits PKR activation in DP-derived MVs, enabling pulp preservation.

Effect of Periodontal Treatment on Reducing Chronic Inflammation in Systemically Healthy Patients With Periodontal Disease.

BACKGROUND: We determined the effects and an accurate marker of periodontal treatment on serum interleukin (IL)-6 and high-sensitivity C-reactive protein (HsCRP) levels in systemically healthy individuals with periodontal disease. METHODS: This multicenter study included systemically healthy individuals with periodontal disease who received initial periodontal treatment and had no periodontal treatment history. Periodontal parameters, including periodontal inflamed surface area, masticatory efficiency, and periodontal disease classification; serum IL-6 and HsCRP levels; and serum immunoglobulin (Ig)G titers against periodontal pathogens were evaluated at baseline and after treatment. Subjects were classified as low or high responders (group) based on periodontal inflamed surface area changes. RESULTS: There were 153 participants. Only periodontal inflamed surface area changes were markedly different between low and high responders. Periodontal treatment (time point) decreased both serum IL-6 and HsCRP levels. The interaction between group and time point was remarkable only for serum IL-6 levels. Changes in serum immunoglobulin (Ig)G titers against periodontal pathogens were not associated with IL-6 changes in high responders. We analyzed the indirect effect of serum anti-Porphyromonas gingivalis type 2 IgG titer changes using mediation analysis and found no significance. However, the direct effect of group (low or high responder) on IL-6 changes was considerable. CONCLUSIONS: Periodontal treatment effectively decreased serum IL-6 levels, independent of periodontal pathogen infection, in systemically healthy individuals with periodontal disease.

Epithelial-to-mesenchymal transition, inflammation, subsequent collagen production, and reduced proteinase expression cooperatively contribute to cyclosporin-A-induced gingival overgrowth development.

Drug-induced gingival overgrowth (DIGO), induced by certain immunosuppressive drugs, antihypertensive agents, and antiepileptic drugs, may contribute to the formation of deeper periodontal pockets and intractableness in periodontitis. To date, multiple factors such as enhanced matrix production, inflammation, and reduced matrix degradation might be involved in the pathogenesis of DIGO. We have previously reported that SPOCK-1, a heparan sulfate proteoglycan, could affect gingival thickening by promoting epithelial-to-mesenchymal transition (EMT) in gingival keratinocytes. However, few studies have investigated whether a combination of these factors enhances the DIGO phenotype in animal models. Therefore, we investigated whether SPOCK-1, periodontal inflammation, and cyclosporin-A (CsA) could cooperatively promote gingival overgrowth. We first confirmed that Spock-1 overexpressing (Spock1-Tg) mice showed significantly thicker gingiva and greater alveolar bone loss than WT mice in response to ligature-induced experimental periodontitis. DIGO was induced by the combination of CsA administration and experimental periodontitis was significantly enhanced in Spock1-Tg mice compared to that in WT mice. Ligature-induced alveolar bone loss in CsA-treated Spock1-Tg mice was also significantly greater than that in CsA-treated WT mice, while being accompanied by an increase in Rankl and Col1a1 levels and a reduction in matrix metalloprotease expression. Lastly, SPOCK-1 promoted RANKL-induced osteoclast differentiation in both human peripheral blood mononuclear cells and murine macrophages, while peritoneal macrophages from Spock1-Tg mice showed less TNFα and IL-1ß secretion than WT mice in response to Escherichia coli lipopolysaccharide. These results suggest that EMT, periodontal inflammation, and subsequent enhanced collagen production and reduced proteinase production contribute to CsA-induced DIGO pathogenesis.

A Case Report of Improved Palmoplantar Pustulosis following Periodontal Treatment and Possible Association with Diminished Systemic Subclinical Inflammation.

Palmoplantar pustulosis (PPP) is a recurrent pustular dermatosis located on the palms and soles. Focal infection may exacerbate the symptoms of PPP, but the etiology is not fully clear. A 56-year-old woman with PPP was diagnosed with severe chronic periodontitis. Initial treatment for periodontitis combined with topical application of antibiotics and surgical treatment was performed. In this case, attention was paid to the relevance of systemic inflammation caused by periodontitis with the clinical symptoms of PPP. With periodontal treatment, the symptoms of PPP and periodontitis, high-sensitivity C-reactive protein (hs-CRP) level, and periodontal inflamed surface area (PISA) improved. This case highlights the importance of comprehensive dental examinations, including those for oral infections, such as periodontitis and other unrecognized sources of infection, and dental treatment in the overall management of PPP.

The Link Between Periodontal Inflammation and Obesity.

PURPOSE OF REVIEW: Obesity is a trigger for multiple diseases such as diabetes mellitus, hypertension, and cardiovascular diseases. Epidemiological studies have shown that obesity may be a risk factor for periodontal disease. Recently, there have been reports of presumed mechanisms of the associations between periodontitis and lipid metabolism or thermogenesis. This review aims to discuss the link between periodontal disease and energy regulatory function based on recent findings. RECENT FINDINGS: It has been demonstrated that activation of the C-C motif chemokine ligand/C-C chemokine receptor 7 pathway in adipose tissue induces inflammation and impairment of lipid metabolism and energy regulation in mice. Porphyromonas gingivalis administration has been shown to induce further weight gain and increased adipose tissue in diet-induced obese mice. Additionally, it has been reported that Porphyromonas gingivalis-induced endotoxemia potentially affect obesity by altering endocrine functions in brown adipose tissue in mice. Several cohort studies have shown that obesity is associated with tooth loss 5 years later, and periodontal conditions of obese individuals are significantly worse 2 and 6 months after the treatment compared with those of non-obese individuals. It has also been reported that body mass index is positively associated with the periodontal inflamed surface area index, a measure of periodontal inflammation. These results suggest that not only the enhancement of inflammation due to obesity but also the activation of inflammatory signaling may affect energy regulation. SUMMARY: Loss of adipose tissue homeostasis induces increase and activation of immune cells in adipose tissue, leading to impaired immune function in obesity. Various cytokines and chemokines are secreted from obese adipose tissue and promote inflammatory signaling. Some of these signaling pathways have been suggested to affect energy regulation. The combination of obesity and periodontitis amplifies inflammation to levels that affect the whole body through the adipose tissue. Obesity, in turn, accelerates the exacerbation of periodontitis.

Anti-inflammatory effects of miRNA-146a induced in adipose and periodontal tissues.

MicroRNA (miRNA) plays an important role in diverse cellular biological processes such as inflammatory response, differentiation and proliferation, and carcinogenesis. miR-146a has been suggested as a negative regulator of the inflammatory reaction. Although, it has been reported as expressed in inflamed adipose and periodontal tissues, however, miR-146a's inhibitory effects against inflammatory response in both the tissues, are not well understood. Therefore, in this study, the inhibitory effects of miR-146a on both adipose and periodontal inflammation, was investigated. In vitro study has revealed that miR-146a transfection into either adipocytes or gingival fibroblasts, has resulted in a reduced cytokine gene expression, observed on co-culturing the cells with macrophages in the presence of lipopolysaccharides (LPS), in comparison to the control miRNA transfected. Similarly, miR-146a transfection into macrophages resulted in a reduced expression of TNF-α gene and protein in response to LPS stimulation. In vivo study revealed that a continuous intravenous miR-146a administration into mice via tail vein, protected the mice from developing high-fat diet-induced obesity and the inflammatory cytokine gene expression was down-regulated in both adipose and periodontal tissues. miR-146a appeared to be induced by macrophage-derived inflammatory signals such as TNF-α by negative feed-back mechanism, and it suppressed inflammatory reaction in both adipose and periodontal tissues. Therefore, miR-146a could be suggested as a potential therapeutic molecule and as a common inflammatory regulator for both obesity-induced diabetes and related periodontal diseases.

SPOCK1 is a novel inducer of epithelial to mesenchymal transition in drug-induced gingival overgrowth.

Few studies have investigated the role of extracellular-matrix proteoglycans in the pathogenesis of drug-induced gingival overgrowth (DIGO). SPOCK1 is an extracellular proteoglycan that induces epithelial to mesenchymal transition (EMT) in several cancer cell lines and exhibits protease-inhibitory activity. However, the role of SPOCK1 in non-cancerous diseases such as DIGO has not been well-addressed. We demonstrated that the expression of SPOCK1, TGF-ß1, and MMP-9 in calcium channel blocker-induced gingival overgrowth is higher than that in non-overgrowth tissues. Transgenic mice overexpressing Spock1 developed obvious gingival-overgrowth and fibrosis phenotypes, and positively correlated with EMT-like changes. Furthermore, in vitro data indicated a tri-directional interaction between SPOCK1, TGF-ß1, and MMP-9 that led to gingival overgrowth. Our study shows that SPOCK1 up-regulation in a noncancerous disease and SPOCK1-induced EMT in gingival overgrowth occurs via cooperation and crosstalk between several potential signaling pathways. Therefore, SPOCK1 is a novel therapeutic target for gingival overgrowth and its expression is a potential risk of EMT induction in cancerous lesions.

Dental pulp cell-derived powerful inducer of TNF-α comprises PKR containing stress granule rich microvesicles.

It is well known that dental pulp tissue can evoke some of the most severe acute inflammation observed in the human body. We found that dental pulp cells secrete a factor that induces tumor necrosis factor-α production from macrophages, and designated this factor, dental pulp cell-derived powerful inducer of TNF-α (DPIT). DPIT was induced in dental pulp cells and transported to recipient cells via microvesicles. Treatment of dental pulp cells with a PKR inhibitor markedly suppressed DPIT activity, and weak interferon signals were constitutively activated inside the cells. In recipient macrophages, stimulation with DPIT-containing supernatants from pulp cells resulted in activation of both nuclear factor-κB and MAP kinases like JNK and p38. Proteomics analyses revealed that many stress granule-related proteins were present in supernatants from dental pulp cells as well as microvesicle marker proteins like GAPDH, ß-actin, HSPA8, HSPB1, HSPE1, and HSPD1. Furthermore, giant molecule AHNAK and PKR were detected in microvesicles derived from dental pulp cells, and gene silencing of AHNAK in dental pulp cells led to reduced DPIT activity. Thus, it appeared that the core protein of DPIT was PKR, and that PKR was maintained in an active state in stress granule aggregates with AHNAK and transported via microvesicles. The activity of DPIT for TNF-α induction was far superior to that of gram-negative bacterial endotoxin. Therefore, we, report for the first time, that active PKR is transported via microvesicles as stress granule aggregates and induces powerful inflammatory signals in macrophages.

Periodontal disease and hypertriglyceridemia in Japanese subjects: potential association with enhanced lipolysis.

Although periodontal disease may be associated with increased risk for atherosclerosis, the mechanism by which the disease causes atherosclerosis is still unknown. The candidates contributing to atherosclerosis in periodontal disease include low-grade inflammation such as C-reactive protein (CRP) and insulin resistance. A previous study demonstrated that periodontal therapy leads to an improvement in CRP as well as insulin resistance, indicating the relationship between periodontal disease and low-grade inflammation or insulin resistance. On the other hand, we previously demonstrated that serum triglyceride (TG) per se is independently associated with CRP or insulin resistance in Japanese populations with a body mass index (BMI) of 21.5 to 27.0 (midrange BMI). To the best of our knowledge, however, the relationship between periodontal disease and serum TG is not fully clarified. The first aim of the present study is to investigate whether periodontal disease is associated with serum TG in Japanese subjects with midrange BMI. If so, another aim of the study is to determine which mechanism is responsible for the association between periodontal disease and serum TG in these subjects. We have performed a periodontal examination in the Ogaki metabolic syndrome medical examination. One hundred sixty-two participants from 40 to 74 years old (56 men and 106 women; mean age, 66.43 ± 6.25 years) were enrolled in the study. Besides medical examination, oral panoramic radiograph was taken for all participants. Average bone score was also calculated. Periodontal bone destruction increased according to the age of the participants (r = 0.227, P < .004, Spearman correlation coefficient). Periodontal bone destruction was also associated with serum TG levels (r = 0.299, P = .000). This association was more evident in subjects with midrange BMI (r = 0.332, P < .001). In subjects with midrange BMI, TG was not correlated with BMI or waste circumstances. Furthermore, TG was not associated with age itself in the midrange BMI group. We then investigated the lipolytic activity of endotoxin in cocultures of adipocytes and macrophages. Low-dose lipopolysaccharide dose-dependently increased lipolytic activity in cocultures, and this activity was neutralized by anti-tumor necrosis factor α neutralizing antibodies. These results suggest that periodontal infection, especially bacterial endotoxinemia, is associated with enhanced lipolysis and subsequent up-regulation of circulating TG in Japanese with midrange BMI.