Lipocalin 2, synthesized using a cell-free protein synthesis system and encapsulated into liposomes, inhibits the adhesion of Porphyromonas gingivalis to human oral epithelial cells.


Effects of Candidalysin Derived from Candida albicans on the Expression of Pro-Inflammatory Mediators in Human Gingival Fibroblasts

International Journal of Molecular Sciences


Biological Roles of Fibroblasts in Periodontal Diseases.

Periodontal diseases include periodontitis and gingival overgrowth. Periodontitis is a bacterial infectious disease, and its pathological cascade is regulated by many inflammatory cytokines secreted by immune or tissue cells, such as interleukin-6. In contrast, gingival overgrowth develops as a side effect of specific drugs, such as immunosuppressants, anticonvulsants, and calcium channel blockers. Human gingival fibroblasts (HGFs) are the most abundant cells in gingival connective tissue, and human periodontal ligament fibroblasts (HPLFs) are located between the teeth and alveolar bone. HGFs and HPLFs are both crucial for the remodeling and homeostasis of periodontal tissue, and their roles in the pathogenesis of periodontal diseases have been examined for 25 years. Various responses by HGFs or HPLFs contribute to the progression of periodontal diseases. This review summarizes the biological effects of HGFs and HPLFs on the pathogenesis of periodontal diseases.


Dental students' awareness after clinical training between dental treatment and systemic health: A questionnaire-based survey


Gan-Lu-Yin (Kanroin), traditional Chinese herbal extracts, reduces osteoclast differentiation in vitro and prevents alveolar bone resorption in rat experimental periodontitis

Gan-Lu-Yin (GLY), a traditional Chinese herbal medicine, shows therapeutic effects on periodontitis, but that mechanism is not well known. This study aims to clarify the precise mechanism by investigating the inhibitory effects of GLY extracts on osteoclastogenesis in vitro and on bone resorption in periodontitis in vivo. RAW264.7 cells are cultured with soluble receptor activator of nuclear factor-kappa B (sRANKL) and GLY extracts (0.01-1.0 mg/mL), and stained for tartrate-resistant acid phosphatase (TRAP) to evaluate osteoclast differentiation. Experimental periodontitis is induced by placing a nylon ligature around the second maxillary molar in rats, and rats are administered GLY extracts (60 mg/kg) daily for 20 days. Their maxillae are collected on day 4 and 20, and the levels of alveolar bone resorption and osteoclast differentiation are estimated using micro-computed tomography (CT) and histological analysis, respectively. In RAW264.7 cells, GLY extracts significantly inhibit sRANKL-induced osteoclast differentiation at a concentration of more than 0.05 mg/mL. In experimental periodontitis, administering GLY extracts significantly decreases the number of TRAP-positive osteoclasts in the alveolar bone on day 4, and significantly inhibits the ligature-induced bone resorption on day 20. These results show that GLY extracts suppress bone resorption by inhibiting osteoclast differentiation in experimental periodontitis, suggesting that GLY extracts are potentially useful for oral care in periodontitis.


Inhibitory Effects of Sudachitin on the Production of Inflammation-related Molecules in Human Gingival Fibroblasts Cultured under High Glucose Conditions

高グルコース条件下における歯肉線維芽細胞のカルプロテクチン誘導性炎症関連因子の産生におけるスダチチンの抑制効果

The Japanese Journal of Conservative Dentistry


Disequilibrium of Plasma Protease/Anti-Protease Due to Severe Periodontal Disease Contributes to Human Subarachnoid Hemorrhage


Advanced glycation end-products increase lipocalin 2 expression in human oral epithelial cells.


Evaluation of educational effects of periodontology by lectures and phantom training in dental students

講義および実習試験の評価による歯周病学教育のあり方の考察

 Purpose: In the undergraduate educational curriculum of dentistry, there are two main teaching/learning methods for dental students before clinical training: traditional lectures and phantom training. The purpose of this study was to explore effective educational systems for periodontology by examining the test results of both lectures and phantom training.

 Methods: Both a written test and a practical test of periodontology were administered to students of dentistry. A total of 43 dental students of Tokushima University Dental School were recruited. Written tests were performed before and after the lectures on periodontology. A practical test of scaling and root planing was performed after phantom training. Associations between the results of the written test and practical test were analyzed statistically. P values of less than 0.05 were considered statistically significant.

 Results: The results of the written tests after lectures were higher than those before lectures. A significant positive correlation was found between the results of the written test and practical test.

 Conclusion: In the students' education on periodontology, the results of the practical test were correlated statistically to the results of the written test. These findings may contribute to establishing effective educational systems for periodontology in dentistry.

 目的 : 一般的に, 臨床実習前の歯学部生に対する歯周病学の教育は, 座学講義と基礎 (模型) 実習を主体とする. 本研究の目的は, 歯周病学の講義および実習の評価試験の結果の関連性を調べ, 将来の歯学部生に対する歯周病学教育のあり方について考察することである.

 方法 : 2018年度後期から2019年度前期に徳島大学歯学部に在籍し, 歯周病学を履修する歯学部3∼4年次生43名を対象とした. 歯周病学の講義前後に当該分野に関する筆記試験 (5肢2択形式) を行い, さらに歯周病学の基礎 (模型) 実習の終了時に, 顎模型を用いてスケーリング・ルートプレーニングの実習試験を行った. 講義による学生の理解度・習得度の評価は, カイ二乗検定を用いて判定した. 筆記試験および実習試験の結果の関連性は, スピアマンの順位相関検定を用いて判定した. また, 筆記試験の点数を25点満点で9点以下, 10∼12点, 13点以上の3群に分類し, 各群間の実習試験点数の差異の有無について, ANOVA-Tukey HSD解析を用いて判定した. なお, p値が0.05未満を有意差ありと判定した.

 結果 : 歯周病学の講義後の筆記試験の点数は, 出題内容によって差はあるものの, 講義前と比較しておおむね有意に上昇した. 筆記試験の結果と実技試験の結果は, 有意に相関した (r=0.34, p=0.025). また筆記試験の点数が9点以下の群における実習試験の点数は, 筆記試験の点数が10∼12点, 13点以上の群と比較して有意に低かった (p<0.05). 一方, 筆記試験の点数が10∼12点と13点以上の群間における実習試験の点数に有意差は認められなかった (p=0.76).

 結論 : 臨床実習前の歯周病学の学生教育において, 筆記試験の点数と基礎 (模型) 実習試験の点数は有意に相関した. このことは, 将来の効果的な歯周病学の教育システム構築の一助となる可能性を示唆する.


S100A9 Increases IL-6 and RANKL Expressions through MAPKs and STAT3 Signaling Pathways in Osteocyte-Like Cells.


Relationship of oral conditions to the incidence of infective endocarditis in periodontitis patients with valvular heart disease: a cross-sectional study.

The patients with IE had fewer remaining teeth, more advanced bone resorption compared with those of patients without IE. These findings suggest a possible association between the occurrence of IE and periodontal infection.

No significant differences were observed between patients with or without VHD in oral conditions. A significant increase in the percentage of alveolar bone loss in VHD patients with IE was observed compared with that of patients without IE. The ratio of both Porphyromonas gingivalis (Pg) IgG titer > 1.68 and Pg fimA type II genotype in patients with IE was significantly higher than in patients without IE. There was a significant correlation between the occurrence of IE and clinical oral findings (number of remaining teeth: OR, 0.17; rate of alveolar bone loss > 40%: OR, 11.8).


) is associated with increased atherosclerosis, diabetes mellitus, and other systemic diseases through blood stream. On the other hand, carotenoids belong among phytochemicals that are responsible for different colors of the foods. It is important to examine whether carotenoids are effective to the inhibition of periodontal infection/inflammation cascades. This review summarizes the advanced state of knowledge about suppression of periodontal infection by several carotenoids. A series of findings suggest that carotenoids intake may provide novel strategy for periodontitis treatment, although further study will be needed.


6-Shogaol inhibits advanced glycation end-products-induced IL-6 and ICAM-1 expression by regulating oxidative responses in human gingival fibroblasts

Advanced glycation end-products (AGEs) cause diabetes mellitus (DM) complications and accumulate more highly in periodontal tissues of patients with periodontitis and DM. AGEs aggravate periodontitis with DM by increasing the expression of inflammation-related factors in periodontal tissues. 6-Shogaol, a major compound in ginger, has anti-inflammatory and anti-oxidative activities. However, the influence of shogaol on DM-associated periodontitis is not well known. In this study, the effects of 6-shogaol on AGEs-induced oxidative and anti-oxidative responses, and IL-6 and ICAM-1 expression in human gingival fibroblasts (HGFs) were investigated. When HGFs were cultured with 6-shogaol and AGEs, the activities of reactive oxygen species (ROS) and antioxidant enzymes (heme oxygenase-1 [HO-1] and NAD(P)H quinone dehydrogenase 1 [NQO1]), and IL-6 and ICAM-1 expressions were investigated. RAGE expression and phosphorylation of MAPKs and NF-κB were examined by western blotting. 6-Shogaol significantly inhibited AGEs-induced ROS activity, and increased HO-1 and NQO1 levels compared with the AGEs-treated cells. The AGEs-stimulated expression levels of receptor of AGE (RAGE), IL-6 and ICAM-1 and the phosphorylation of p38, ERK and p65 were attenuated by 6-shogaol. These results suggested that 6-shogaol inhibits AGEs-induced inflammatory responses by regulating oxidative and anti-oxidative activities and may have protective effects on periodontitis with DM.


Advanced glycation end-product 2 and Porphyromonas gingivalis lipopolysaccharide increase sclerostin expression in mouse osteocyte-like cells

Sclerostin is a secreted glycoprotein that is mainly expressed in osteocytes, exerts negative effects on bone formation, and is present at elevated levels in diabetes mellitus (DM). Periodontitis is an infectious disease caused by periodontopathic bacteria, a complication of DM, and sometimes associated with severe inflammation and alveolar bone resorption. Advanced glycation end-products (AGEs) are a major pathogen in DM complications and adversely influence periodontitis in DM patients. In the present study, the effects of AGE2 and Porphyromonas gingivalis lipopolysaccharide (P-LPS) on the expression of sclerostin in mouse osteocyte-like cells (MLO-Y4-A2 cells) and its function in osteoblast differentiation were investigated. AGE2 and P-LPS up-regulated the expressions of receptor of AGE (RAGE) and Toll-like receptor 2 (TLR2), respectively, and significantly up-regulated that of sclerostin and interleukin 6 (IL-6) in osteocytes. Sclerostin, RAGE and TLR2 levels were synergistically increased by AGE2 and P-LPS. The siRNAs of RAGE and TLR2 significantly inhibited AGE2- and P-LPS-induced sclerostin expression. AGE2 up-regulated sclerostin expression in osteocyte-like cells via the RAGE, ERK and JNK, and NF-κB signal pathways. On the other hand, P-LPS elevated sclerostin levels via the TLR2, JNK and p38, and NF-κB signal pathways. When osteocytes pre-treated with AGE2 and P-LPS and osteoblastic cells (MC3T3-E1) were co-cultured in the medium with a sclerostin-neutralizing antibody, AGE2- and P-LPS-induced decreases in alkaline phosphatase activity and Runx2 expression in osteoblastic cells were significantly inhibited by the sclerostin-neutralizing antibody. These results suggest that AGE2 and P-LPS influence bone metabolism and inflammation through the regulation of sclerostin expression, and may aggravate periodontitis with DM.


High Glucose-Mediated Cytokine Regulation in Gingival Fibroblasts and THP-1 Macrophage: a Possible Mechanism of Severe Periodontitis with Diabetes.

Diabetic conditions such as HG induce IL-1 and sIL-6R production from macrophages in inflammatory periodontal tissues and may exacerbate the periodontitis synergistically via MMP-1 production from HGFs.


Association between oral frailty and geriatric conditions


Advanced glycation end-products increase IL-6 and ICAM-1 expression via RAGE, MAPK and NF-kB pathways in human gingival fibroblasts

Diabetes mellitus (DM) is a risk factor for periodontal diseases and may exacerbate the progression of the pathogenesis of periodontitis. Advanced glycation end-products (AGEs) cause DM complications relative to levels of glycemic control and larger amounts accumulate in the periodontal tissues of patients with periodontitis and DM. In the present study, we investigated the effects of AGEs on the expression of inflammation-related factors in human gingival fibroblasts (HGFs) to elucidate the impact of AGEs on DM-associated periodontitis. HGFs were cultured with or without AGEs. Cell viability was examined, and RNA and protein fractions were isolated from AGE-treated cells. The expression of interleukin (IL)-6, intercellular adhesion molecule-1 (ICAM-1), and the receptor for AGE (RAGE) was investigated using reverse transcription-polymerase chain reaction, quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay, and reactive oxygen species activity was measured using a kit with 2',7'-dichlorofluorescin diacetate. Human monocytic cells (THP-1) labeled with a fluorescent reagent were co-cultured with HGFs treated with AGEs and IL-6 siRNA, and the adhesive activity of THP-1 cells to HGFs was assessed. The expression of IL-6 and ICAM-1 was examined when HGFs were pretreated with recombinant human IL-6, the siRNAs of RAGE and IL-6, and inhibitors of MAPK and NF-κB, and then cultured with and without AGEs. The phosphorylation of MAPK and NF-κB was assessed using western blotting. AGEs increased the mRNA and protein expressions of RAGE, IL-6, ICAM-1 and reactive oxygen species activity in HGFs, and promoted the adhesion of THP-1 cells to HGFs, but had no effect on cell viability until 72 hours. Recombinant human IL-6 increased ICAM-1 expression in HGFs, while the siRNAs of RAGE and IL-6 inhibited AGE-induced IL6 and ICAM1 mRNA expression, and IL-6 siRNA depressed AGE-induced THP-1 cell adhesion. AGEs increased the phosphorylation of p38 and ERK MAPKs, p65 NF-κB and IκBα, while inhibitors of p38, ERK MAPKs and NF-κB significantly decreased AGE-induced IL-6 and ICAM-1 expression. AGEs increase IL-6 and ICAM-1 expression via the RAGE, MAPK and NF-κB pathways in HGFs and may exacerbate the progression of the pathogenesis of periodontal diseases.


Clinical effects of low body mass index on geriatric status in elderly patients.

Low BMI might be a useful parameter to evaluate the geriatric status, and the viewpoint would contribute to decide the care plan for the good end-of-life of elderly.

). Significant associations of low BMI to several geriatric factors such as loss of posterior occlusion, cognitive impairment were observed in both male and female. FIM scores in above cut-off point group were significantly higher than in below cut-off point group in female (FIM gain, P = 0.0005; FIM efficiency, P = 0.0025, Mann-Whitney U test). On the other hand, there were no significant differences between low and above BMI cut-off point in FIM scores of male patients.


Relationship of aspiration pneumonia to cognitive impairment and oral condition: a cross-sectional study.

Early and simple evaluation of the oral condition and cognitive function can predict the risk of aspiration pneumonia.


b-carotene suppresses Porphyromonas gingivalis lipopolysaccharide-mediated cytokine production in THP-1 monocytes cultured with high glucose condition.

Periodontitis is associated with development of diabetes mellitus. Although lipopolysaccharide (LPS) of Porphyromonas gingivalis (Pg), a major pathogen of periodontitis, may lead the progression of diabetes complications, the precise mechanisms are unclear. We, therefore, investigated the effects of β-carotene on production of Pg LPS-induced inflammatory cytokines in human monocytes cultured high glucose (HG) condition. THP-1 cells were cultured under 5.5 mM or 25 mM glucose conditions, and cells were stimulated with Pg LPS. To investigate the productivity of TNF-α, IL-6, and MCP-1, cell supernatants were collected for ELISA. To examine the effects of NF-kB signals on cytokine production, Bay11-7082 was used. HG enhanced Pg LPS-induced production of TNF-α, IL-6, and MCP-1 via NF-kB signals in THP-1. β-carotene suppressed the enhancement of the Pg LPS-induced cytokine production in THP-1 via NF-κB inactivation. Our results suggest that β-carotene might be a potential anti-inflammatory nutrient for circulating Pg LPS-mediated cytokine production in diabetic patients with periodontitis.


Swallowing impairment is a significant factor for predicting life prognosis of elderly at the end of life.

For judgement of life prognosis, the condition of being frail such as impaired swallowing function might be a useful factor, and the viewpoint would contribute to decide the treatment plan for the good end-of-life care of elderly.

A total of 320 elderly patients was enrolled (male/female 151/169; averaged age: male 84.7 ± 5.9 year, female 86.8 ± 6.3 year) and retrospective analyses were performed. The elderly patients were classified as either: (1) with or without past illness of aspiration pneumonia; (2) with or without incidence of cerebrovascular disorder; (3) impaired or normal cognitive function; (4) impaired or normal swallowing function, and performed Kaplan-Meier survival analysis. Swallowing function was examined using video endoscopic (VE) evaluation method. The Kaplan-Meier analysis of the number of days from implementation of VE test (day 0) to death was evaluated with the log-rank Mantel-Cox test. The maximum follow-up time recorded was 180 days.

Aging Clinical and Experimental Research


Local administration of calcitonin inhibits alveolar bone loss in an experimental periodontitis in rats.

Calcitonin (CTN), a calcium regulatory hormone, promotes calcium diuresis from the kidney and suppresses bone resorption. The objective of this study was to evaluate whether the topical and intermittent application of CTN inhibits alveolar bone resorption using ligature-induced experimental periodontitis in rats. Experimental periodontitis was induced by placing a nylon ligature around maxillary molars of 8-week-old male Wistar rats for 20 days. Thirty-two rats were divided into four groups: basal sham control group, periodontitis group, periodontitis plus 0.2 U CTN (low dose), and periodontitis plus 1.0 U CTN (high dose) group. To investigate the effects of CTN on alveolar bone resorption, CTN was topically injected into the palatal gingivae every 2 days after ligature removal (day 0). Micro-computed tomography (CT) analysis was performed for linear parameter assessment of alveolar bone on day 5 and day 14. Periodontal tissues were examined histo-pathologically to assess the differences among the study groups. Micro-CT images showed that alveolar bone resorption was induced statistically around the molar of ligatured rats on day 5 and day 14. The amount of bone resorption was more severe on day 14 than that on day 5. On day 5, only high-dose CTN treatment significantly suppressed bone resorption. In addition, both doses of CTN significantly suppressed bone resorption on day 14. Histological examination clarified that there were fewer TRAP-positive cells in the CTN treatment groups than in the periodontitis group on day 5. Local administration of CTN decreased alveolar bone resorption by regulating osteoclast activation in rats with periodontitis.


Advanced glycation end-products and Porphyromonas gingivalis lipopolysaccharide increase calprotectin expression in human gingival epithelial cells.

Accumulation of advanced glycation end-products (AGEs) in periodontal tissues of patients with diabetes mellitus aggravates periodontitis, but the mechanisms are unknown. Calprotectin, a heterocomplex of S100A8 and S100A9 proteins, is a constitutive cytoplasmic component of healthy gingival epithelial cells. This study aimed at investigating the effects of AGE and Porphyromonas gingivalis lipopolysaccharide (PgLPS) on calprotectin expression in the human gingival epithelial cell line OBA-9. AGE and PgLPS increased the expression of S100A8 and S100A9 mRNAs, and AGE + PgLPS co-stimulation amplified their expression in OBA-9 cells. A higher concentration of calprotectin in cell lysates was also induced by stimulation with AGE and/or PgLPS. S100A8 was mainly translocated from the nucleus to the cytoplasm by AGE stimulation, while cytoplasmic localization of S100A9 was not altered following stimulation with AGE and/or PgLPS. Calprotectin was found in the cytoplasm of BSA-treated cells, but cytoplasmic and nuclear localization was observed following stimulation with AGE and/or PgLPS. AGE-induced S100A8, and S100A9 mRNA expression was partially suppressed by RAGE-specific siRNA. In contrast, PgLPS-induced S100A8 and S100A9 mRNA expression was strongly suppressed by TLR2-specific siRNA. Furthermore, the inhibition of p38, JNK MAPK, and NF-κB attenuated AGE- and PgLPS-induced S100A8 and S100A9 mRNA expression. Taken together, these results demonstrate that AGE acts in synergy with PgLPS to stimulate RAGE and TLR2 expression and activate p38, JNK MAPK, and NF-κB signaling pathways, resulting in increased activation of calprotectin (S100A8/S100A9) in human gingival epithelial cells. Our results suggest that calprotectin may be involved in the pathogenesis of diabetic periodontitis. This article is protected by copyright. All rights reserved.


Useful Immunochromatographic Assay of Calprotectin in Gingival Crevicular Fluid for Diagnosis of Diseased Sites in Patients with Periodontal Diseases.

Determination of GCF calprotectin using a novel IC chip system is useful for diagnosis of periodontal diseases.


Association between Oral Conditions and Returning Home after Discharge in Elderly Patients


Clinical Significance of GCF sIL-6R and Calprotectin to Evaluate the Periodontal Inflammation.

Both GCF sIL-6R and calprotectin levels are significant bio-markers to evaluate the periodontal inflammation.


Calprotectin Induces IL-6 and MCP-1 Production via Toll-Like Receptor 4 Signaling in Human Gingival Fibroblasts.

Calprotectin, a heterodimer of S100A8 and S100A9 molecules, is associated with inflammatory diseases such as inflammatory bowel disease. We have reported that calprotectin levels in gingival crevicular fluids of periodontitis patients are significantly higher than in healthy subjects. However, the functions of calprotectin in pathophysiology of periodontitis are still unknown. The aim of this study is to investigate the effects of calprotectin on the productivity of inflammatory cytokines in human gingival fibroblasts (HGFs). The HGFs cell line CRL-2014® (ATCC) were cultured, and total RNAs were collected to examine the expression of TLR2/4 and RAGE mRNA using RT-PCR. After the cells were treated with S100A8, S100A9, and calprotectin, supernatants were collected and the levels of IL-6 and MCP-1 were measured using ELISA methods. To examine the intracellular signals involved in calprotectin-induced cytokine production, several chemical inhibitors were used. Furthermore, after the siRNA-mediated TLR4 down-regulated cells were treated with S100A8, S100A9, and calprotectin, the levels of IL-6 and MCP-1 were also measured. HGFs showed greater expression of TLR4 mRNA, but not TLR2 and RAGE mRNA compared with human oral epithelial cells. Calprotectin increased significantly the production of MCP-1 and IL-6 in HGFs, and the cytokine productions were significantly suppressed in the cells treated with MAPKs, NF-B, and TLR4 inhibitors. Furthermore, calprotectin-mediated MCP-1 and IL-6 production were significantly suppressed in TLR4 down-regulated cells. Taken together, calprotectin induces IL-6 and MCP-1 production in HGFs via TLR4 signaling that involves MAPK and NF-B, resulting in the progression of periodontitis. J. Cell. Physiol. 232: 1862-1871, 2017. © 2016 Wiley Periodicals, Inc.


Association between Periodontal Condition and Kidney Dysfunction in Japanese Adults: A Cross-Sectional Study

Clinical and Experimental Dental Research


Effects of new over-the-counter periodontal ointment-containing applicator with single-tuft brush on cytokine levels in gingival crevicular fluid during supportive periodontal therapy phase: a randomized double-blind clinical trial.

This OTC medication is biochemically effective for steady chronic periodontitis in the supportive periodontal therapy phase.

The levels of IL-1β, IL-6, IL-8 and TNF-α remained significantly lower in the test group compared to the placebo group. In the placebo group, when the probing pocket depth at baseline was 4 mm, IL-1β increased, particularly in the second molar tooth, and the greatest increase was seen when PPD at baseline was 5-6 mm. In the test group, IL-1β decreased markedly in cases with furcation involvement and low bleeding on probing at baseline. In both groups, IL-1β, IL-6 and TNF-α were closely correlated with each other.


Inhibitory Effects of Cryptotanshinone on Inflammation-related Molecules in Human Gingival Fibroblasts

培養歯肉線維芽細胞におけるクリプトタンシノンによる炎症関連分子の産生抑制効果

Purpose: Cryptotanshinone (CPT) isolated from the root of an Asian medicinal plant, Salvia miltiorrhiza bunge, acts as a potent anti-tumor or anti-inflammatory agent in vitro. However, the effects of CPT on the progression of periodontitis are still unknown. This study investigated the effects of CPT on the production of inflammation-related molecules in human gingival fibroblasts. Methods: Human gingival fibroblasts (HGFs), CRL-2014TM (ATCC), were used. Cytotoxicity of the cells by CPT was examined. Dead cells stained by trypan blue were counted and analyzed. Next, the levels of IL-6 and VEGF in IL-1β-treated cells with or without CPT were measured using ELISA kits. The productivity of cathepsin L in IL-1β-treated cells with or without CPT was examined using Western blotting. Results: The ratio of viable cell numbers was significantly decreased by 100 μmol/l CPT. Next, the increase of IL-6 and cathepsin L induced by IL-1β was significantly suppressed by the pretreatment of 10 μmol/l CPT in HGFs. On the other hand, productivity of VEGF induced by IL-1β was not changed by the pretreatment of 10 μmol/l CPT. Conclusion: CPT suppresses IL-1β-induced IL-6 and cathepsin L production in HGFs, and might regulate the progression of periodontitis.

目的 : 漢方薬 (生薬) タンジンの主成分であるクリプトタンシノンは, 抗がん作用や抗炎症作用をもつことが知られている. しかしながら, クリプトタンシノンが歯周組織の炎症に対してどのような影響を及ぼすのかは不明である. 本研究の目的は, 歯肉線維芽細胞を標的として, その炎症関連分子の産生に及ぼすクリプトタンシノンの影響を調べることである. 方法 : 細胞は歯肉線維芽細胞CRL-2014TM (ATCC) を用いた. クリプトタンシノンの濃度依存的な細胞障害作用の有無は, トリパンブルー染色法を用いて各濃度間における生細胞率の差を比較検討して評価した. 歯肉線維芽細胞のサイトカイン産生は, 市販のELISAキットを用いて, インターロイキン (IL) -1βで細胞を24時間刺激した後のIL-6およびVEGF産生量を測定して検討した. 一方, タンパク質分解酵素カテプシンLの産生は, IL-1βで細胞を24時間刺激した後に回収した全細胞タンパク質を用いて, ウエスタンブロット法によって検討した. なお, クリプトタンシノンはIL-1β添加の1時間前に細胞培養系に添加した. 各群における差異は, ANOVA Tukey-KramerのHSD検定によって有意差を検討した. 結果 : クリプトタンシノンは, 10μmol/lまでは歯肉線維芽細胞の生細胞率に影響は与えないが, 100μmol/lでは有意に生細胞率を低下させた. また, 歯肉線維芽細胞におけるIL-1β誘導性のIL-6産生およびカテプシンL産生は, 10μmol/lクリプトタンシノンの添加によって著明に減少した. 一方, IL-1β誘導性のVEGF産生は, クリプトタンシノンの添加によっても変化しなかった. 結論 : クリプトタンシノンは, 歯肉線維芽細胞におけるIL-1β誘導性IL-6およびカテプシンLの産生を抑制することで, 歯周組織の炎症を抑制する可能性が示唆された.

The Japanese Journal of Conservative Dentistry


YKL-40 level in gingival crevicular fluid from patients with periodontitis and type 2 diabetes

YKL-40 is a chitin-binding glycoprotein, the level of which increases in inflammatory diseases, diabetes mellitus (DM), cardiovascular diseases, and tumors. Gingival crevicular fluid (GCF) contains many proteins and markers of periodontitis. The purpose of this study was to investigate YKL-40 level in GCF from patients with periodontitis and DM and the association between YKL-40 level and chronic periodontitis (CP) or DM. The subjects were 121 patients with DM, CP, DM and periodontitis (DM-P), and healthy subjects (H). GCF was collected using paper strips after the sites for GCF collection were clinically evaluated for probing depth (PD), gingival index (GI), and bleeding on probing (BOP). YKL-40 in GCF was identified by Western blotting, and its level was determined by ELISA. YKL-40 was contained in GCF samples from H, DM, CP, and DM-P sites, and its levels (amount and concentration) in CP and DM-P were significantly higher than those in H and DM. GCF YKL-40 level significantly correlated with PD and GI, and its level in BOP-positive sites was significantly higher than that in BOP-negative ones. GCF YKL-40 level was elevated in periodontitis, but not DM. YKL-40 in GCF may be an inflammatory marker for periodontitis.


肥満糖尿病ラット歯周炎の歯肉組織におけるオステオポンチンの局在

肥満糖尿病ラット歯周炎の歯肉組織におけるオステオポンチンの局在

Journal of the Japanese Association of Periodontology


Predictors of improved functional outcome in elderly inpatients after rehabilitation: a retrospective study.

Age and disorder of oral function may be significant predictors of improved functional capacity after rehabilitation for elderly inpatients.


Gingival overgrowth caused by vitamin C deficiency associated with metabolic syndrome and severe periodontal infection: a case report.

It has been suggested that vitamin C deficiency/scurvy is associated with gingival inflammatory changes; however, the disorder is very infrequently encountered in the modern era. Here, we report a case of extensive gingival overgrowth caused by vitamin C deficiency associated with metabolic syndrome and severe periodontal infection.


ADL，認知機能の低下及び低栄養を併発した老年症候群患者は摂食嚥下機能が低下する

全国自治体病院協議会雑誌

全国自治体病院協議会雑誌


Combined therapeutic effects of adenoviral vector-mediated GLIPR1 gene therapy and radiotherapy in prostate and bladder cancer models.

Combining AdGlipr1 (AdGLIPR1) with radiotherapy may achieve additive or synergistic tumor control in selected prostate and bladder tumors, and additional therapeutic effects may result with repeated treatment cycles.

Urologic Oncology: Seminars and Original Investigations


Secreted caveolin-1 enhances periodontal inflammation by targeting gingival fibroblasts.

Caveolin-1 (Cav-1) is a membrane protein. Recently, it has been reported that secreted Cav-1 induces angiogenesis in inflammatory microenvironment. However, it is unclear that Cav-1 regulates gingival inflammation. Therefore, we investigated the Cav-1 function to periodontal cells. Expression of Cav-1 in human periodontitis tissues was examined pathologically. Secretion of Cav-1 from human gingival fibroblasts (HGFs) or human periodontal ligament cells (HPLFs) treated with IL-1 and TNF- was examined using Western blotting. Likewise, intracellular signals induced by Cav-1 were examined. Finally, we examined whether the secreted Cav-1 induces production of inflammatory mediators in HGFs using ELISA or qRT-PCR. Pathologically, high expression of Cav-1 was observed in human periodontitis tissues. Cav-1 secretion increased in both cultured HGFs and HPLFs treated with IL-1 and TNF-. Cav-1 induced phosphorylation of JNK and ERK, but not Stat3 in HGFs. Furthermore, Cav-1 increased proMMP-1 and VEGF secretion in HGFs, and the VEGF secretion was statistically suppressed by JNK inhibitor SP600125, but not ERK inhibitor PD98059. ProMMP-1 secretion was suppressed statistically by both SP600125 and PD98059. In addition, Cav-1 increased significantly MMP-1, -10 and -14 mRNA expressions, whereas no increase of TIMPs mRNA was observed in HGFs treated with Cav-1. These data suggest that secreted Cav-1 derived from periodontal fibroblastic cells enhances inflammation-related several proteases and VEGF secretion in HGFs via MAPKs pathway, resulting in progression of periodontitis through induction of tissue degradation or angiogenesis.


Enhancement of gingival inflammation induced by synergism of IL-1 and IL-6.

Internleukin-1 (IL-1) and IL-6 are the most potent proinflammatory cytokines being involved in inflammatory diseases such as periodontitis. The objective of this study was to examine the synergistic effects of IL-1 and IL-6 on gingival inflammation by targeting cultured human gingival fibroblasts (HGFs). HGFs were treated with IL-1 or IL-6/soluble IL-6R (sIL-6R), and total RNA and total cell lysate were collected to examine expression of gp130 known as a signal transducer of IL-6 using qRT-PCR and Western blotting. IL-1-mediated IL-6 productivity in HGFs was examined using ELISA method. Likewise, after HGFs and THP-1 macrophages were treated with IL-1, TNF- and IL-6, sIL-6R productivity was examined. Next, HGFs were treated with IL-6/ sIL-6R after pretreatment of IL-1, and the intracellular signals were examined using Western blotting. Finally, various mRNA/protein expressions in HGFs treated with IL-6/sIL-6R after pretreatment of IL-1 were examined using qRT-PCR and ELISA method. IL-1 increased significantly both gp130 and IL-6 expression in HGFs. IL-6 increased significantly sIL-6R production in THP-1 macrophages but not HGFs. Co-stimulation with IL-1 and IL-6/sIL-6R induced dramatically the phosphorylation of Stat3, ERK and JNK in HGFs. Interestingly, expression of various inflammation- related molecules such as MMP-1, MCP-1, IL-1ra, bFGF and VEGF were enhanced by co-stimulation with IL-1 and IL-6/sIL-6R in HGFs. Gingival inflammation is regulated by HGFs affected by both IL-1 and IL-6/sIL-6R synergistically through induction of gp130 expression, resulting in progression of periodontitis.


Assessment of the plasma/serum IgG test to screen for periodontitis.

Chronic periodontitis is a silent infectious disease prevalent worldwide and affects lifestyle-related diseases. Therefore, efficient screening of patients is essential for general health. This study was performed to evaluate prospectively the diagnostic utility of a blood IgG antibody titer test against periodontal pathogens. Oral examination was performed, and IgG titers against periodontal pathogens were measured by ELISA in 1,387 individuals. The cut-off value of the IgG titer was determined in receiver operating characteristic curve analysis, and changes in periodontal clinical parameters and IgG titers by periodontal treatment were evaluated. The relationships between IgG titers and severity of periodontitis were analyzed. The best cut-off value of IgG titer against Porphyromonas gingivalis for screening periodontitis was 1.682. Both clinical parameters and IgG titers decreased significantly under periodontal treatment. IgG titers of periodontitis patients were significantly higher than those of healthy controls, especially in those with sites of probing pocket depth over 4 mm. Multiplied cut-off values were useful to select patients with severe periodontitis. A blood IgG antibody titer test for Porphyromonas gingivalis is useful to screen hitherto chronic periodontitis patients.


Relationship between periodontitis-related antibody and frequent exacerbations in chronic obstructive pulmonary disease.

Normal-IgG titer for periodontitis-related antibody can be an independent predictor of frequent exacerbations. Measuring periodontitis-related antibody titers might be useful to identify patients with susceptibility to frequent exacerbations so that an aggressive prevention strategy can be designed.

The numbers of exacerbations and the rate of individuals with frequent exacerbations (at least two per year) were significantly lower in patients with higher IgG titer than those with normal IgG titer (0.8 vs. 1.2 per year, p= 0.045 and 14.3 vs. 38.6%, p= 0.009, respectively). Multivariate logistic regression analysis showed that being normal-IgG titer for periodontitis-related antibody significantly increased the risk of frequent exacerbations (relative risk, 5.27, 95% confidence interval, 1.30-25.7; p = 0.019) after adjusting for other possible confounders, such as a history of exacerbations in the past year, disease severity, COPD medication and smoking status.


Specific in situ visualization of plasma cells producing antibodies against Porphyromonas gingivalis in gingival radicular cyst: application of the enzyme-labeled antigen method.

The enzyme-labeled antigen method was applied to visualize plasma cells producing antibodies to Porphyromonas gingivalis, flora of the human oral cavity. Antibodies to P. gingivalis have reportedly been detected in sera of patients with periodontitis. Biotinylated bacterial antigens, Ag53, and four gingipain domains (Arg-pro, Arg-hgp, Lys-pro, and Lys-hgp) were prepared by the cell-free protein synthesis system using the wheat germ extract. In paraformaldehyde-fixed frozen sections of rat lymph nodes experimentally immunized with Ag53-positive and Ag53-negative P. gingivalis, plasma cells were labeled with biotinylated Arg-hgp and Lys-hgp. Antibodies to Ag53 were detected only in the nodes immunized with Ag53-positive bacteria. In two of eight lesions of gingival radicular cyst with inflammatory infiltration, CD138-positive plasma cells in frozen sections were signalized for Arg-hgp and Lys-hgp. An absorption study using unlabeled antigens confirmed the specificity of staining. The AlphaScreen method identified the same-type antibodies in tissue extracts but not in sera. Antibodies to Ag53, Arg-pro, and Lys-pro were undetectable. In two cases, serum antibodies to Arg-hgp and Lys-hgp were AlphaScreen positive, whereas plasma cells were scarcely observed within the lesions. These findings indicate the validity of the enzyme-labeled antigen method. This is the very first application of this novel histochemical technique to human clinical samples.

The Journal of Histochemistry and Cytochemistry


Immune responses to porphyromonas gingivalis infection suppress systemic inflammatory response in experimental murine model.

Periodontitis is a localized infectious disease caused by periodontopathic bacteria such as Porphyromonas gingivalis (P. gingivalis), and the severity correlates to significance of immune responses. Recently, it has been reported that periodontitis is associated with the development of systemic disease such as diabetes and atherosclerosis because of increasing invasion of oral pathogens to the circulation. However, the association between local and systemic infectious responses is still unclear. In the present study, we examined the differences of biological responses in animals with or without bacterial infection. After Balb/c mice were infected subcutaneously with live P. gingivalis W83, serum, skin and liver were collected according to experimental protocol. The skin and liver tissues were observed pathologically by haematoxylin-eosin staining, and serum IL-6 levels were measured using ELISA method. Throughout the experimental period, conditions of the mice were observed continuously. As expected, severe infiltration of leukocytes were observed at inflamed skin corresponding to the number of bacterial challenges. Although no inflammatory appearance of skin was observed, serum IL-6 levels were increased dramatically (P <0.01, Student's t-test) and liver tissues were injured in the mice without bacterial challenge. Interestingly, although severe inflammatory appearance of the skin was observed, serum IL-6 levels were not increased and no inflammatory responses were observed in the liver of the 3-times bacterially challenged group. Importantly, immunoglobulin G against P. gingivalis W83 was detected in the blood of mice with 3-times bacterial challenge corresponding to improvement of weight loss and survival. In conclusion, although multiple infections develop severe localized inflammation, the immune system should be sufficient to protect the systemic inflammatory responses.

Journal of Biological Regulators and Homeostatic Agents


臨床研究 医科歯科連携医療における歯周病原細菌に対する血清IgG抗体価検査の導入

Journal of Japanese Society for Evidence and the Dental Professional


Prognosis of periodontitis recurrence after intensive periodontal treatment using examination of serum IgG antibody titer against periodontal bacteria.

Chronic periodontitis is associated with systemic diseases such as atherosclerosis. In this study, we evaluated the efficacy of serum IgG antibody titer to periodontal bacteria for prognosis of periodontitis recurrence during supportive periodontal therapy (SPT) phase. The 139 patients during SPT phase were selected and divided to two groups as follows: "Stable" and "Recurrence" group at SPT phase for case-control study: "High IgG titer" and "Normal IgG titer" group before transition to SPT phase for cohort study. We examined whether clinical findings or serum IgG antibody titers to periodontal bacteria are risk factors for the development of periodontitis recurrence. Case-control study showed that there were significant differences between the stable and recurrence groups in age and number of teeth. The serum IgG antibody titer to Eikenella corrodens FDC1073, Porphyromonas gingivalis SU63, and Campylobacter rectus ATCC33238 was significantly higher in the recurrence group. Next, we found, that the recurrence ratio in the high IgG titer group to Gram-negative obligate anaerobe, Prevotella intermedia, Treponema denticola, and C. rectus was significantly higher than that of the normal IgG titer group. Taken together, serum IgG antibody titer test is useful in the prognosis of periodontitis recurrence during the SPT phase.


Cathepsin B, D, and L regulation in cyclosporin A-mediated gingival hyperplasia of a patient with sarcoidosis.

Cyclosporin A (CsA) is an immunosuppressant with side effects including gingival hyperplasia. Sarcoidosis is a systemic disease characterized by granulomas. Here, we report on a rare case of sarcoidosis with gingival hyperplasia to clarify whether clinical observation corresponds to in vitro results. Gingival fibroblasts (HGFs) were isolated from healthy gingiva and cultured with CsA. Total RNA was collected and expression of mRNAs examined using semi-quantitative RT-PCR analysis. Cathepsin B, D, and L expression in overgrown gingiva of the patient was examined by immunohistochemistry. Cathepsin D, L, and vascular endothelial growth factor (VEGF)165 mRNA were markedly suppressed in CsA-treated HGFs, whereas cathepsin B, matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA were not reduced. Next, the decrease of cathepsin B and L expression in enlarged gingiva was observed, whereas an increase of cathepsin D expression was observed. Clinically, the enlarged gingival lesions were fully resolved by performing oral infection control. Cathepsins regulation might be an important factor in the development of CsA-mediated gingival hyperplasia.


Clinical and microbiological effect of newly developed OTC periodontal ointment-containing applicator with brush on the SPT phase

サポーティブペリオドンタルセラピー期における塗布用ブラシ一体型一般用歯周病薬の臨床的および細菌学的な効果

It is important for maintaining improved periodontal tissue health during supportive periodontal therapy (SPT) to restrain infection caused by periodontopathic bacteria by the patients' own self control in addition to periodic professional intervention. We developed an over-the-counter (OTC) medication, named MC1, consisting of a one-tuft brush containing the compound cetylpyridinium chloride (CPC) as a bactericidal agent, dipotassium glycyrrhizate (GK2) as an anti-inflammatory drug, and allantoin (ALT) as a promoter of cell proliferation and wound healing, for delivery to hardly-brushed regions. In this study, we evaluated the effectiveness of MC1 clinically and microbiologically in maintaining improved periodontal tissue status during SPT.
This study was designed as a double-blind clinical trial targeting chronic periodontitis patients during the SPT phase. Sixty-one subjects (test group: 27 subjects, placebo group: 28 subjects, dropout: 6 subjects) participated in this study, and applied MC1 to 2 molar periodontal pockets 4mm twice a day for 12 weeks. The clinical indicators used were the probing pocket depth (PPD), bleeding on probing (BOP), plaque control record (PCR), gingival index (GI), and weight of the gingival crevicular fluid (GCF). The microbiological indicators were the number of total bacteria, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola. These were evaluated at the baseline, and at the 6- and 12-week time-points.
All of the clinical indicators other than PCR were significantly improved in both groups in a time-dependent manner, and earlier improvement tended to be noted in the test group with GI and GCF volume than in the placebo group. On the other hand, the numbers of all of the examined bacteria at sites with a PPD 5mm were significantly improved in the test group as compared with those in the placebo group.
It is suggested that MC1 is effective in maintaining the state of the periodontal tissue clinically and microbiologically in chronic periodontitis patients during the SPT phase.
Nihon Shishubyo Gakkai Kaishi(J Jpn Soc Periodontol)52(3) : 225-238, 2010.

サポーティブペリオドンタルセラピー(SPT)期における歯周組織の安定には, 定期的な専門的管理に加え, 患者自身の日々のプラークコントロールが重要である．今回我々は, 患者によるプラークコントロールを強化する目的で, 殺菌作用のある塩化セチルピリジニウム, 抗炎症効果のあるグリチルリチン酸二カリウムおよび組織創傷治癒効果をもつアラントインの3種の薬効成分を配合した薬剤をタフトブラシと一体化した塗布用ブラシ一体型歯周病薬(MC1)を開発した．本研究では, SPT期の歯周組織の維持・改善に対するMC1の有効性を臨床的および細菌学的指標を用いて評価した．
本研究は, SPT期の慢性歯周炎患者を対象とした二重盲検法にて実施した．被験者として61名が参加し(試験群 : 27名, プラセボ群 : 28名, 脱落 : 6名), 4 mm以上の歯周ポケットを残す臼歯2歯の辺縁歯肉に, 1日2回12週間, MC1を塗布した．MC1の効果は, ベースライン時, 6および12週間目の3時点について, 臨床指標(PPD, BOP, PCR, GIおよびGCF量), および細菌学的指標(GCF中の総菌数, Porphyromonas gingivalis, Tannerella forsythia およびTreponema denticola)を算出し, 統計学的に群間比較して検討した．
PCRを除く全ての臨床指標は2群ともに経時的に有意に改善した．特に試験群では, GIおよびGCF量が早期に改善する傾向にあった．一方, 5mm以上の歯周ポケットを有する部位において, 試験群では調べた3菌種すべての菌数が, プラセボ群よりも有意に減少した．
以上のことから, MC1はSPT期の慢性歯周炎患者の病状安定において, 臨床的および細菌学的に有効であることが示唆された．
日本歯周病学会会誌(日歯周誌)52(3) : 225-238, 2010

Journal of the Japanese Association of Periodontology


Antigenic group II chaperonin in Methanobrevibacter oralis may cross-react with human chaperonin CCT.

Methanobrevibacter oralis is an archaeal species frequently isolated from sites of severe periodontitis. However, its pathogenic roles remain unclear. Here, we aimed to isolate group II chaperonin from M. oralis and examine its antigenicity. The genes encoding two chaperonin subunits (Cpn-1 and Cpn-2) were cloned from M. oralis using polymerase chain reaction and genome walking procedures. Recombinant proteins Cpn-1 and Cpn-2 were generated, and the reactivities of sera from patients with periodontitis were examined by Western immunoblotting. The open reading frames of Cpn-1 and Cpn-2 genes consisted of 1641 and 1614 base pairs, respectively. Putative ATP-binding domains conserved among the chaperonin family were observed in both genes. The deduced amino acid sequences of the two genes showed 28.8-40.0% identity to each of the subunits of human CCT (CCT1-8). Thirty and 29 of 36 patients' sera reacted with the recombinant Cpn-1 and recombinant Cpn-2, respectively. Western immunoblotting using antiserum against human CCT subunits indicated that anti-CCT3 and anti-CCT8 antibodies recognized recombinant Cpn-1. In addition, anti-CCT1, CCT3, CCT6, and CCT8 antibodies recognized an antigen of approximately 60 kDa in M. oralis. The results suggested that the chaperonin subunits of M. oralis were antigenic molecules that were recognized by periodontitis patients and that may cross-react with human chaperonin CCT.


Rapid detection of mecA and spa by the loop-mediated isothermal amplification (LAMP) method.

The LAMP offers an alternative detection assay for mecA and spa with a great advantage of the rapidity.


臨床研究 周術期患者に対する口腔管理システムの樹立と評価

臨床研究 周術期患者に対する口腔管理システムの樹立と評価


Highly expressed genes in a rough-colony-forming phenotype of Aggregatibacter actinomycetemcomitans: implication of a mip-like gene for the invasion of host tissue.

Aggregatibacter actinomycetemcomitans, a potent pathogen of periodontitis, typically grows as a rough and adherent colony on primary isolated cultures. The colony transforms into a smooth phenotype during repeated subculture. In this study, we aimed to identify highly expressed genes in the rough-colony-forming phenotype for isolation of host-induced genes. Using a cDNA-subtractive hybridization technique, three genes, homologous to a macrophage infectivity potentiator gene (mip), peroxiredoxin gene (prx) and outer membrane protein gene (ompA), were identified. The expression levels of these genes in the rough-colony-forming phenotype were 4-10-fold higher as compared with the smooth-colony-forming phenotype. Attention was focused on the mip-like gene, and a recombinant protein and a deficient mutant were constructed. The recombinant protein reacted with sera from patients with periodontitis, suggesting the production of the Mip-like protein in periodontal lesions. Viable quantitative invasion assay demonstrated that the viable cell counts of the wild-type strain that invaded HeLa cells were more than fourfold as compared with the mip-deficient mutant. The expression of the mip-like gene, prx-like gene and ompA-like gene may be enhanced in the host, and the mip-like gene may play an important role in the infection of A. actinomycetemcomitans, especially in its invasion of the epithelium.

FEMS Immunology and Medical Microbiology


Functional analysis of secreted caveolin-1 in mouse models of prostate cancer progression.

Previously, we reported that caveolin-1 (cav-1) is overexpressed in metastatic prostate cancer and that virulent prostate cancer cells secrete biologically active cav-1. We also showed that cav-1 expression leads to prosurvival activities through maintenance of activated Akt and that cav-1 is taken up by other cav-1-negative tumor cells and/or endothelial cells, leading to stimulation of angiogenic activities through PI-3-K-Akt-eNOS signaling. To analyze the functional consequences of cav-1 overexpression on the development and progression of prostate cancer in vivo, we generated PBcav-1 transgenic mice. Adult male PBcav-1 mice showed significantly increased prostatic wet weight and higher incidence of epithelial hyperplasia compared with nontransgenic littermates. Increased immunostaining for cav-1, proliferative cell nuclear antigen, P-Akt, and reduced nuclear p27(Kip1) staining occurred in PBcav-1 hyperplastic prostatic lesions. PBcav-1 mice showed increased resistance to castration-induced prostatic regression and elevated serum cav-1 levels compared with nontransgenic littermates. Intraprostatic injection of androgen-sensitive, cav-1-secreting RM-9 mouse prostate cancer cells resulted in tumors that were larger in PBcav-1 mice than in nontransgenic littermates (P = 0.04). Tail vein inoculation of RM-9 cells produced significantly more experimental lung metastases in PBcav-1 males than in nontransgenic male littermates (P = 0.001), and in cav-1(+/+) mice than in cav-1(-/-) mice (P = 0.041). Combination treatment with surgical castration and systemic cav-1 antibody dramatically reduced the number of experimental metastases. These experimental data suggest a causal association of secreted cav-1 and prostate cancer growth and progression.


Porphyromonas gingivalis fimbriae-dependent interleukin-6 autocrine regulation by increase of gp130 in endothelial cells.

Our findings indicate that P. gingivalis fimbriae are important factors in the autocrine regulation of IL-6, by increasing gp130 in endothelial cells.


歯周病スクリーニング検査としての歯周病原細菌に対する指尖血漿IgG抗体価の有用性

Journal of Japanese Society for Evidence and the Dental Professional


Therapeutic effects of gelatin matrix-embedded IL-12 gene-modified macrophages in a mouse model of residual prostate cancer.

We evaluated the potential use of intraoperative gelatin matrix hemostatic sealant (GMHS; FloSeal; Baxter Healthcare) embedded with macrophages (Mphi) transduced with murine interleukin (IL)-12 recombinant adenoviral vector (G/Mphi/AdmIL-12) for prevention of recurrence of prostate cancer following radical prostatectomy. Application of G/Mphi/AdmIL-12 resulted in significant suppression of tumor growth and spontaneous lung metastases, a statistically significant survival advantage of the G/Mphi/AdmIL-12-treated animals, more efficient trafficking of Mphi to lymph nodes draining from the prostate and generation of systemic natural killer cell activity and tumor-specific cytolytic T lymphocyte responses compared to the controls in a preclinical mouse model of residual prostate cancer. Our data recommend this treatment as a novel adjuvant for prevention of local recurrence of prostate cancer following radical prostatectomy.


IL-6/sIL-6R enhances cathepsin B and L production via caveolin-1-mediated JNK-AP-1 pathway in human gingival fibroblasts.

Interleukin (IL)-6 has an important role in inflammatory diseases. Lysosomal enzymes cathepsins are widely expressed as cysteine proteases regulating inflammatory process. Caveolin-1 (Cav-1) is a scaffolding/regulatory membrane protein that interacts with signaling molecules. In this study, we investigated the role of Cav-1 on (1) the productivity, and (2) the enzymatic activity of cathepsin B and L in human gingival fibroblasts (HGFs) treated with IL-6 in the presence of soluble form of IL-6 receptor (sIL-6R). At first, we established the siRNA-mediated Cav-1 down-regulating in vitro systems by transient transfection of Cav-1 siRNA. The siRNA-mediated Cav-1 down-regulated cells were treated with IL-6/sIL-6R for indicated times. Then, cell lysates were collected, and examined the IL-6-induced signaling pathway, cathepsin B and L production, and measurement of cathepsins activity. To investigate the cathepsin L activity, cathepsin-(B + L) activity was measured after pretreatment with CA-074Me, a specific inhibitor for cathepsin B. We found that IL-6/sIL-6R enhanced significantly both production and activity of cathepsin B and L in HGFs. Interestingly, IL-6-mediated phosphorylation of both p44/42 MAPK and JNK was dramatically suppressed in Cav-1 down-regulated HGFs treated with IL-6/sIL-6R. In addition, both production and activity of cathepsin B and L were also significantly suppressed. Importantly, we demonstrated that JNK inhibition, but not p44/42 MAPK inhibition, significantly diminished IL-6/sIL-6R-induced cathepsin B and L production. Taken together, we concluded that IL-6/sIL-6R enhances cathepsin B and L production via IL-6/sIL-6R-mediated Cav-1-JNK-AP-1 pathway in HGFs. Our findings indicate that Cav-1 might be a therapeutic target for IL-6-mediated tissue degradation in periodontitis.


サポーティブペリオドンタルセラピーによる歯周炎の再発防止と進行抑制の効果に関する統計学的検討


Mice with cav-1 gene disruption have benign stromal lesions and compromised epithelial differentiation.

Caveolin-1 (cav-1) is a major structural protein of caveolae, small invaginations of the plasma membrane that integrate and regulate signaling pathways involved in cell growth and differentiation. We previously generated a genetically engineered mice that are homozygous for a null mutation in exon 2 of cav-1 and documented increased incidence of urolithiasis in young male cav-1(-/-) mice. We attributed this, in part, to improper localization of plasma membrane calcium/calmodulin-dependent calcium ATPase in the distal convoluted tubules of the kidney. To document pathologies related to cav-1 function, we maintained cav-1(-/-) and control cav-1(+/+) mice for an extended time period. We report here that cav-1(-/-) mice demonstrate organ-specific growth-related disorders in stromal cells that normally have high levels of cav-1 expression. In many of these organs, epithelial cell growth/differentiation abnormalities were also observed, yet in most of these sites the epithelial cells normally express low to non-detectable levels of cav-1. We propose that loss of cav-1 function in stromal cells of various organs directly leads to a disorganized stromal compartment that, in turn, indirectly promotes abnormal growth and differentiation of adjacent epithelium.


cAMP-response element binding protein (CREB) regulates cyclosporine-A-mediated down-regulation of cathepsin B and L synthesis.

Cyclosporin A (CsA) is an immunosuppressant with severe side effects including gingival overgrowth. We have previously reported that CsA impairs the activity of the lysosomal enzymes cathepsin B and L in human gingival fibroblasts (HGFs). Here, we have examined the effects of CsA on the DNA-binding activity of the cyclic AMP response element-binding protein (CREB) and cell viability, and the effects of CREB on cathepsin B and L synthesis and activity in HGFs. We have confirmed that CsA down-regulates cathepsin B and L synthesis. Further, CsA has no effect on cell viability and dramatically impairs CREB-DNA binding activity. Importantly, the synthesis of cathepsin B and L is down-regulated, and their activity is also significantly impaired in HGFs transfected with plasmid expressing dominant-negative CREB. These results suggest that CREB is essential for the CsA-mediated down-regulation of cathepsin B and L synthesis in HGFs.


IL-12 gene-modified bone marrow cell therapy suppresses the development of experimental metastatic prostate cancer.

To investigate the immunomodulatory effects of interleukin-12 (IL-12) for treatment of metastatic prostate cancer, we administered adult bone marrow cells (BMC) that were genetically modified by retroviral vector-mediated IL-12 gene transduction in an experimental mouse model of prostate cancer metastasis. This therapy produced significant anti-metastatic effects in bone and lung and prolonged animal survival. Flow cytometric analysis indicated donor BMC could effectively home to bone and lung after treatment. Intensive infiltration of CD4 and CD8T cells in lung metastases and increased systemic natural killer and cytotoxic T lymphocyte activities indicated induction of a significant anti-metastatic immune response after treatment with IL-12 transduced BMC. Our results demonstrate the therapeutic potential of gene-modified BMC gene therapy.


Gene profiles during root canal treatment in experimental rat periapical lesions.

The purpose of this study was to profile gene expression in periapical lesions during root canal treatment (RCT). Periapical lesions were induced experimentally by exposing the pulp in Sprague-Dawley rats. After 3 wk, the animals received root canal filling (RCF) and were sacrificed 1 or 4 wk later. From the periapical tissues, total RNA was extracted and processed for cDNA-microarray analysis. The lesions were histologically and radiographically confirmed to expand 4 wk after pulp exposure (inflammation phase) and to stabilize 4 wk after RCF (healing phase). In approximately 30,000 genes on the microarray, 203 genes were up-regulated to more than 5-fold (e.g., IL-1beta), and 864 genes were down-regulated to less than 20% of baseline level (e.g., caspase 8) in inflammation phase. Compared with inflammation phase, we found that 133 genes were up-regulated (e.g., IL-1alpha) and 50 genes were down-regulated (e.g., defensin alpha5) in healing phase. Corresponding to the gene expression profiles, accumulation of IL-1alpha and IL-1beta was observed in the periapical lesions by immunohistochemistry. These gene profiles might be useful in diagnosing the healing process of periapical lesions.


Cooperative effects of adenoviral vector-mediated interleukin 12 gene therapy with radiotherapy in a preclinical model of metastatic prostate cancer.

We investigated the potential benefits of combining adenoviral vector mediated in situ interleukin-12 (AdmIL-12) gene therapy with radiation therapy (XRT) to enhance therapeutic efficacy. In a metastatic mouse prostate cancer cell line, 178-2 BMA, AdmIL-12+XRT demonstrated enhanced therapeutic activities in vitro as determined by clonogenic survival, apoptosis, and mIL-12 levels. At the molecular level, increased expression of tumor necrosis factor-alpha mRNA was specific for the combined therapy. In a subcutaneous 178-2 BMA in vivo model, the combination of AdmIL-12+XRT produced statistically significant tumor growth suppression compared to control vector Adbetagal, Adbetagal XRT, or AdmIL-12 as monotherapy. In addition, significant prolongation of survival was demonstrated for the combination of AdmIL-12+XRT. The combination of AdmIL-12+XRT significantly suppressed both spontaneous and pre-established lung metastases, and led to a prolonged elevation of serum IL-12 and significantly increased natural killer (NK) activities. Importantly, in vivo depletion of NK cells resulted in significant attenuation of the antimetastatic activities of AdmIL-12 alone or AdmIL-12+XRT. These combined effects suggest that AdIL-12 gene therapy together with radiotherapy may achieve maximal tumor control (both local and systemic) in selected prostate cancer patients via radio-gene therapy induced local cytotoxicity and local and systemic antitumor immunity.


Cloning and characterization of lipopolysaccharide-induced tumor necrosis factor alpha factor promoter.

We have recently identified lipopolysaccharide tumor-induced tumor necrosis factor alpha factor (LITAF) as a novel transcription factor controlling necrosis factor (TNF)-alpha expression in the human monocytic cell line, THP-1. To characterize the human (h) LITAF promoter, we isolated a 1.2-kb DNA fragment and followed this by a screening of human genomic DNA with a hLITAF cDNA probe. A 34-bp sequence domain located from nucleotides -74 to -43 in the hLITAF promoter exhibited the highest basal reporter gene activity; however, the activity was not elevated by lipopolysaccharide (LPS) stimulation. The sequence domain included a consensus sequence for hepatocyte nuclear factor (HNF)-3alpha, regulating the transcription of many kinds of genes. Interestingly, the DNA sequence position between -542 and -538 in the hLITAF promoter contained the CTCCC motif, which has been reported to act as a specific binding site for hLITAF protein. Electrophoretic mobility shift assays demonstrated that LPS induced the binding of THP-1 nuclear factors to a 22 bp probe containing the CTCCC motif. In addition, hLITAF mRNA and nuclear hLITAF protein increased significantly in the THP-1 cells immediately after LPS stimulation. These results suggest that the consensus sequence for HNF-3alpha, or a nuclear binding protein to the CTCCC motif, may play an important role in regulating LPS-dependent LITAF transcription.

FEMS Immunology and Medical Microbiology


Adenoviral vector-mediated RTVP-1 gene-modified tumor cell-based vaccine suppresses the development of experimental prostate cancer.

We previously identified a novel p53 target gene, RTVP-1, that possesses unique cytotoxic and immunostimulatory activities which make it potentially useful for cancer gene therapy. To test the therapeutic potential of RTVP-1 in a gene-modified tumor cell-based vaccine model, we used an adenoviral vector capable of efficient transduction and expression of RTVP-1 (AdRTVP-1), together with a highly metastatic mouse prostate cancer cell line (178-2 BMA). A vaccine was prepared with 178-2 BMA cells transduced with AdRTVP-1 or a control adenoviral vector expressing beta-galactosidase (Adbetagal). After irradiation of the cells, syngeneic 129/Sv mice were vaccinated three times at weekly intervals. After 3 weeks, they were challenged with orthotopic 178-2 BMA cells. After 21 days, fewer than 60% of the RTVP-1-cell-vaccinated mice developed tumors compared to 100% of the control mice. The RTVP-1-cell vaccine significantly reduced primary tumor wet weight compared with control Adbetagal-cell vaccine (P<0.0001 at 7 and 14 days). Experimental metastasis to lung was also significantly reduced (P=0.0377), and survival significantly increased (P=0.0002). In addition, significantly increased NK and CTL activities were demonstrated in the AdRTVP-1-cell-vaccinated mice. These findings indicate that RTVP-1 gene-modified cell-based vaccines may be useful in the prevention of recurrent prostate cancer.


Sustained long-term immune responses after in situ gene therapy combined with radiotherapy and hormonal therapy in prostate cancer patients.

Sustained long-term (up to 8 to 12 months) systemic T-cell responses were noted after combined radio-gene-hormonal therapy for prostate cancer. Prolonged use of hormonal therapy does not suppress this response. These results suggest the potential for sustained activation of cell-mediated immune responses against cancer.

International Journal of Radiation Oncology*Biology*Physics


Long-term cyclosporin A exposure suppresses cathepsin-B and -L activity in gingival fibroblasts.

The decreased ability of protein degradation by not only cathepsin-L but also cathepsin-B is, at least, one of the several factors developing the cyclosporin A-induced gingival overgrowth.


Systemic up-regulation of sTNFR2 and IL-6 in Porphyromonas gingivalis pneumonia in mice.

Aspiration pneumonia is a common cause of death in older people, and the pathophysiology is a chronic respiratory failure with a mild airway inflammation. In this study, we established a mild inflammatory pneumonia model using Porphyromonas gingivalis (Pg) pathogen-infected mice. It elucidated the effects of Pg-infected pneumonia on proinflammatory cytokines tumor necrosis factor (TNF)-alpha, interleukin-6 (IL-6), and IL-1beta production in both lung tissue and serum. We also elucidated production of soluble (s) TNF receptor (R) s, because TNF-alpha is considered to be a dominant inflammatory mediator. Lung TNF-alpha levels significantly increased at 2 h after infection, and rapidly returned to basal level at 24 h. Consistent with increase of TNF-alpha, remarkable increase of sTNFR2 but not sTNFR1 was detected in lung tissue from 2 to 72 h. Interestingly, sTNFR2/sTNFR1 ratio was significantly enhanced at 2 h in serum. In addition, lung IL-1beta and IL-6 levels also significantly increased from 2 to 24 h. Importantly, we found that IL-6 levels in serum reflected its local level. These results may suggest that systemically produced sTNFR2 and IL-6 could be a key role to modulate proinflammatory activities of TNF-alpha in Pg-induced lung inflammation simulated aspiration pneumonia.


High glucose enhances interleukin-6-induced vascular endothelial growth factor 165 expression via activation of gp130-mediated p44/42 MAPK-CCAAT/enhancer binding protein signaling in gingival fibroblasts.

Diabetic patients are susceptible to severe inflammatory periodontitis manifesting as swollen gingiva with bleeding, but the underlying mechanism is not well understood. Our purpose was to determine the effect of a high glucose (HG) condition on the interleukin-6/soluble interleukin-6 receptor (IL-6/sIL-6R)-induced activation of signaling and vascular endothelial growth factor (VEGF) expression in human gingival fibroblasts (HGFs). In this study, HGFs were cultured for at least two passages under a normal glucose (NG; 5.5 mM) condition or high glucose (25 mM) condition. Importantly, the HG condition significantly induced expression of gp130 mRNA in HGFs compared with levels in control cells. Consistent with the expression of its mRNA, the HG condition also increased the expression of gp130 protein, and phosphorylation of the tyrosine residue by gp130 was enhanced significantly by IL-6/sIL-6R stimulation. Furthermore, the HG condition enhanced the IL-6/sIL-6R-induced phosphorylation of p44/42 MAPK and led to subsequent activation of CCAAT/enhancer binding protein in nuclei. In contrast, there was no significant difference in phosphorylation of JNK between the HG and NG condition. Interestingly, HGFs increased IL-6/sIL-6R-induced VEGF165 mRNA expression and VEGF165 secretion under the HG condition compared with levels under the NG condition. In contrast, the induction of VEGF165 secretion was partially inhibited by PD98059 (selective p44/42 MAPK inhibitor) under the HG condition. In addition, the VEGF165 secretion was completely inhibited by the combination of PD98059 and SP600125 (JNK inhibitor). Our findings suggest that the HG condition indirectly increases VEGF expression via activation of gp130-mediated p44/42 MAPK-CCAAT/enhancer binding protein signaling in HGFs. Thus, elevated VEGF secretion in HGFs under the HG condition may play a role in the development of the severe periodontitis observed in diabetic patients.


C-jun N-terminal kinase (JNK) inhibitor, SP600125, blocks interleukin (IL)-6-induced vascular endothelial growth factor (VEGF) production: cyclosporine A partially mimics this inhibitory effect.

Angiogenesis is a common complication of organ-transplant rejection. One of the primary responsible molecules for enhanced angiogenesis is vascular endothelial growth factor (VEGF). Activated protein (AP)-1 is considered to play a key role in the transcription of VEGF. c-jun N-terminal kinase (JNK), one of the MAP kinase family members, plays a critical role in AP-1 activation. Thus, we tested the effect of a novel JNK inhibitor, SP600125, on VEGF production in fibroblasts. SP600125 significantly suppressed interleukin (IL)-6-induced production of VEGF in cultured fibroblasts. Cyclosporine A (CsA), a known in vitro anti-angiogenic reagent, partially mimicked this suppression. In fact, CsA suppressed IL-6-induced phosphorylation of JNK. The results indicate that although both SP600125 and CsA are anti-angiogenic by inhibiting VEGF production by way of a JNK-dependent pathway, the inhibitory effect was much stronger with the novel inhibitor of JNK than with CsA.


Cathepsin-L, a key molecule in the pathogenesis of drug-induced and I-cell disease-mediated gingival overgrowth: a study with cathepsin-L-deficient mice.

Drug-induced gingival overgrowth, the chronic side effect of calcium antagonists, is frequently seen due to the increase in patients with hypertension, although the etiology of the disease is largely unknown. I-cell disease, which accompanies gingival overgrowth, is characterized by a deficiency in UDP-N-acetyl-glucosamine and is classified as one of the lysosomal storage diseases. Here, we hypothesized that a common mechanism may underlie the etiology of gingival overgrowth seen in patients treated with calcium antagonist and in patients with I-cell disease. A calcium antagonist, nifedipine, specifically suppressed cathepsin-L activity and mRNA expression, but not that of cathepsin-B in cultured gingival fibroblasts. The activity of cathepsin-L was suppressed up to 50% at 24 hours after treatment of the cells with the reagent. The selective suppression of cathepsin-L activity appeared not to be dependent on Ca(2+), since treatment of the cells with thapsigargin suppressed both cathepsin-B and -L activity. Mice deficient in the cathepsin-L gene manifested enlarged gingivae. Histological observation of the gingivae demonstrated typical features of acanthosis, a phenotype very similar to that of experimentally induced gingival overgrowth. Since cathepsin-L deficiency was reported to be associated with thickening of the skin, impaired cathepsin-L activity may play a key role in the establishment of skin and gingival abnormalities seen in I-cell disease. In addition, reduced cathepsin-L activity may play an important role in inducing drug-induced gingival overgrowth.


Impairment of gingival fibroblast adherence by IL-6/sIL-6R.

Interleukin-6 (IL-6) binds to human gingival fibroblasts (HGF) in the presence of a soluble form of IL-6 receptor (sIL-6R). We investigated the effects of IL-6 on the functions of HGF in the presence of sIL-6R. HGF changed their morphology from spindle-shaped to round, and detached from the culture dish by stimulation with IL-6/sIL-6R. In this condition, a signal transducer gp130 and a transcription factor Stat3 were phosphorylated, resulting in activation of transcription factors Stat3 and C/EBPbeta. Cytoskeletal beta-actin and adhesion molecule integrin-alpha5, a subunit of alpha5beta1 integrin (VLA-5), were found to possess potential binding domains for these transcription factors in their promoters. Accumulation of beta-actin and integrin-alpha5 mRNA decreased, contrary to the expectation of the induction of gene transcription. Furthermore, the decrease in their mRNAs was associated with reduced expression of both actin and VLA-5 proteins. These results suggest that the expression of VLA-5 and actin was down-regulated in HGF through an IL-6 signaling pathway, resulting in impairment of HGF adherence.


Serum phenytoin concentration and IgG antibody titre to periodontal bacteria in patients with phenytoin-induced gingival overgrowth.

Epileptic patients taking phenytoin with gingival-overgrowth and those without gingival-overgrowth were compared for daily drug dose, plasma total phenytoin concentration, plasma free-phenytoin concentration and serum IgG antibody titre against 13 periodontal bacteria. Significantly higher daily drug dose was noted in patients with gingival overgrowth (P < 0.05) when compared with those without overgrowth. In addition, both total and free forms of plasma phenytoin concentration were significantly higher in sera of patients with gingival growth than of those without overgrowth (P < 0.01). Strong positive correlation was found between daily drug dose and serum phenytoin concentration in patients with gingival overgrowth, while weak correlation was found in patients without gingival overgrowth, suggesting a difference in drug metabolism in these two groups. However, no differences were found in serum IgG antibody titres to 13 periodontal bacteria examined between two groups. These results suggest that metabolic ability of phenytoin is one of the factors for developing gingival overgrowth, and that periodontal infection may not be a primary causative factor for gingival overgrowth but act as an additive factor which increase tissue mass for this unwanted side effect.

Journal of the International Academy of Periodontology


Tumor necrosis factor-alpha (TNF-alpha)-induced and interleukin-1 beta (IL-1 beta)-induced shedding of TNF receptors from gingival fibroblasts.

Tumor necrosis factor-alpha (TNF-alpha) exerts its functions by binding two different receptors (TNFR55 and TNFR75). Both TNFR55 and TNFR75 exist in cell-associated and soluble forms. Soluble TNF receptors (sTNFR), sTNFR55 and sTNFR75, are proteolytically shed upon inflammatory stimuli and then modulate various TNF-alpha bioactivities. As human gingival fibroblasts (HGF) can be potential targets for TNF-alpha in inflamed gingiva, we hypothesized that HGF partially modulate the cellular responses to TNF-alpha by regulating their own TNFR. In this study, the kinetics of expression of cell-associated and soluble forms of both receptors from cultured HGF in response to proinflammatory cytokines TNF-alpha and interleukin-1 beta (IL-1 beta) were investigated in vitro. Both TNF-alpha and IL-1 beta upregulated the gene expression of TNFR75 and did not affect that of TNFR55. TNF-alpha and IL-1 beta decreased binding of [(125)I]TNF-alpha to HGF. Moreover, TNF-alpha and IL-1 beta upregulated the release of sTNFR75 from HGF but not that of sTNFR55. These results suggest that HGF under inflammatory conditions may contribute to the inactivation of circulating TNF-alpha through the preferential induction and shedding of TNFR75.

Journal of Interferon and Cytokine Research


Induction of intracellular interleukin-1 beta signals via type II interleukin-1 receptor in human gingival fibroblasts.

The type II interleukin-1 receptor (IL-1RII) has been thought to be incapable of transducing signals to cells because of its short intracellular domain, while type I IL-1 receptor (IL-1RI) does transduce signals. Since over-expression of IL-1RII has been demonstrated to inhibit cytokine production in the fibroblastic cell line, it has been proposed to use IL-1RII to prevent IL-1-induced inflammation in connective tissue. In this study, trace amounts of IL-1RII mRNA expression were detected in human gingival fibroblasts (HGFs), which are affected by cytokines in inflammatory periodontal disease. Cloning of the cDNA encoding IL-1RII expressed in HGFs revealed 3 amino acid substitutions in the extracellular domain, when compared with the 408 residues predicted from human B-cells. Over-expression of IL-1RII on HGFs by gene transfer down-regulated the expression of IL-1 beta mRNA and IL-6 mRNA in response to IL-1 beta stimulation, while the expression of IL-8 mRNA was not affected. In the IL-1RII-transfected HGFs, phosphorylation of 25- and 74-kDa proteins was up-regulated upon IL-1 beta stimulation in the transfected HGFs. The phosphorylation of these proteins was suppressed by the addition of a neutralizing antibody against IL-1RII. These results suggest that the IL-1RII may regulate HGFs expression of cytokine mRNA upon IL-1 beta stimulation, possibly by altering the IL-1RI-dependent signals.


High glucose suppresses cathepsin activity in periodontal-ligament-derived fibroblastic cells.

The accumulation of extracellular matrices and integrins by high glucose has been reported in relation to diabetic complications. We previously reported that PDL cells expressed a higher amount of VLA-5 when cultured in high-glucose (4500 mg/L) medium than those cultured in low-glucose (1100 mg/L) medium. In this study, we aimed to address (1) whether this effect was mediated by the transcriptional level of the gene or the degradative level of the protein, and (2) whether this effect was mediated by TGF-beta. The results indicated that the level of mRNA encoding alpha5 integrin did not change in PDL cells regardless of the concentration of glucose. Alternatively, high glucose suppressed cathepsin B+L activity. Additionally, the level of mRNA encoding TGF-beta was not affected by high glucose, nor did an anti-TGF-beta neutralizing antibody have an effect on the expression of beta5 gene or cathepsin activity. Therefore, the effects of high glucose appeared to be mediated by impaired protein degradation, but not by autocrine TGF-beta.


Mast cells are not involved in the development of cyclosporin A-induced gingival hyperplasia: a study with mast cell-deficient mice

A previous study suggested that mast cells (MC) are involved in the development of cyclosporin A-induced gingival hyperplasia, since an increased number of MC were observed in the tissue sections of enlarged gingiva. To determine the role of MC in gingival hyperplasia, an MC-deficient mouse model was used in the current study. MC-deficient mice (WBB6F1xW/Wv) and their littermates (+/+) were fed sucrose-containing diets supplemented with or without varying concentrations (300, 400, 500, 600 mg) of cyclosporin A/kg of diet. After 30 days, the mice were sacrificed and the degree of gingival hyperplasia was evaluated by the appearance of the gingiva. Tissue MC were stained with toluidine blue to confirm the presence or absence of MC in the enlarged gingiva. Both W/Wv and +/+ mice, when fed with 600 mg cyclosporin A/kg diet for 30 days, exhibited a similar degree of gingival hyperplasia, while other test mice or control mice did not. Toluidine blue staining of the tissue sections confirmed the presence of MC in the enlarged gingiva of the +/+ mice, but not the W/Wv mice. These results indicate that mast cells are not necessary in the development of cyclosporin A-induced gingival hyperplasia, and that the increased number of MC observed in the enlarged gingiva may be a secondary effect of gingival hyperplasia. We also conclude that a study of mice lacking certain molecules or cells would be quite useful in determining the molecules or cell types responsible for the pathogenesis of drug-induced gingival hyperplasia.


Phenytoin and cyclosporin A suppress the expression of MMP-1, TIMP-1, and cathepsin L, but not cathepsin B in cultured gingival fibroblasts.

The results indicate that, besides suggested effects of these drugs on gingival fibroblasts and/or on accumulated cells in the gingival tissues, extracellular matrix-degrading ability, particularly that by cathepsin L, is also suppressed by cyclosporin A and phenytoin in gingival fibroblasts, and that lysosomal enzyme plays an important role in the pathogenesis of drug-induced gingival hyperplasia.


Role of soluble interleukin-6 receptor in inflamed gingiva for binding of interleukin-6 to gingival fibroblasts.

Interleukin-6 (IL-6), frequently detected in periodontitis, is known to mediate important signals in the inflammatory cytokine network. Gingival fibroblasts (GF) secrete cytokines upon stimulation with inflammatory mediators. However, it is not clear if GF respond to IL-6. We examined the IL-6 receptor gene expression in GF. Furthermore, we tested whether GF are target cells for IL-6 by examination of binding of IL-6. GF were found to contain trace amounts of mRNA for IL-6 receptor (IL-6R), but had high levels of mRNA for 130-kDa glycoprotein (gp130), which is a signal transducer for IL-6/IL-6R complex. Based on this observation, we hypothesized that IL-6 could bind GF if exogenous soluble forms of IL-6R (sIL-6R) existed in the gingiva or culture condition. Thus, we investigated the existence of sIL-6R in gingiva using enzyme-linked immunosorbent assay and whether sIL-6R influenced the binding of IL-6 to GF in vitro. In inflamed gingiva, sIL-6R was detected and its concentration ranged from 150 to 700 pg/microgram protein. The sIL-6R enhanced the binding of IL-6 to GF in a dose-dependent manner. This enhancement was inhibited by an antibody against gp130, suggesting that the IL-6/sIL-6R complex bound to the fibroblasts via gp130. These data demonstrated that gingival fibroblasts can be target cells for IL-6 in the presence of appropriate amounts of sIL-6R. This situation may exist during inflammation in periodontal tissue.


線維芽細胞におけるIL-6の刺激伝達

線維芽細胞におけるIL-6の刺激伝達


Diagnosis of Periodontal Diseases using Biomarkers in Gingival Crevicular Fluid

歯肉溝滲出液中のバイオマーカーを用いた歯周病診断

Periodontal diseases cause an inflammation and degradation of periodontal tissues and missing of teeth. The incidence rate of periodontal diseases is high in middle-aged and elderly people.A reasonable diagnosis of periodontal diseases is very important to keep teeth, however, conventional examinations of periodontal diseases is not necessarily exact and objective. Gingival crevicular fluid (GCF) is an exudate secreted from periodontal tissues and contains many components including proteolytic enzymes, inflammatory cytokines, blood-associated proteins, cellular and bacterial fragments. Because some proteins in GCF are related to inflammation, tissue degradation and bone metabolism, those proteins have been studying as a diagnostic marker of periodontal diseases. GCF is noninvasively collected using a sterile paper strip and biomarkers are determined using enzyme-linked immunosorbent assay (ELISA) and enzyme activity assay. We identified calprotectin, an inflammationrelated protein, in GCF and calprotectin level in GCF from periodontitis sites was significantly higher than that of healthy control. Calprotectin level in GCF was positively correlated to gingival index and other biomarkers and decreased by periodontal treatments. Resistin is an adipocytokine and its level increases in some inflammatory diseases. Resistin level in GCF from periodontitis sites was high compared to the level of healthy control samples. Procollagen type I C-terminal peptide (PICP) is a biomarker for bone metabolism and its level was high in GCF collected from periodontitis sites. These results suggested that calprotectin, resistin and PICP are useful biomarkers for periodontal diseases. On the other hand, we showed that glycated albumin (GA), a marker of diabetes mellitus (DM), was contained in GCF and GA level in GCF from DM patients was significantly higher than that of non-DM individuals. Components in GCF may be biomarkers of systemic diseases as well as periodontal diseases and their determination will be useful diagnostic examination of some diseases. Recently, we have been studying the determining system of GCF calprotectin, including microchip ELISA, surface plasmon resonance assay and immuno-chromatography assay. When GCF biomarkers are determined using the determining systems, we will simply, exactly and objectively diagnose periodontal diseases at our dental offices.


ある侵襲性歯周炎患者の26年間における歯周病治療の経過 -血清IgG抗体価による歯周病原細菌感染度もモニタリング-


Improvement of pustulosis palmaris et plantaris by periodontal infection control in a patient with chronic periodontitis.

Pustulosis palmaris et plantaris (PPP) is a chronic, inflammatory, autoimmune-type disease characterized by sterile pustules of skin. The skin inflammation is influenced by several factors such as drugs, sunlight, metabolic and psychogenic factors as well as metal allergy. Here, we report a rare case that intensive periodontal treatment might have contributed to the improvement of skin inflammation. Skin inflammation regressed 1 month after intensive periodontal treatment. Both CD4/CD8 ratio and % of B cells in the blood sample were slightly decreased corresponding to the improvement of periodontal inflammation. Infection control of periodontal lesions might be one of attractive therapeutic targets in management of PPP.


A Case of Root of Apexification in Extruded-Immature Permanent Incisor

根未完成脱落歯に対するアペキシフィケーションを目的とした歯髄処置の経過報告

Calcium hydroxide is a suitable reagent for apexification procedures in immature permanent teeth. We report on a case of root apexification in a patient (8-yr, male) with a traumatic extrusion of the maxillary right central incisor. After temporary fixation of the traumatically extruded maxillary right central incisor, tooth necrosis and gingival swelling was observed , thus it was diagnosed to require an apexification procedure. The apexification procedure was successfully performed using calcium hydroxide following endodontic treatment. None of the calcium hydroxide -treated tooth showed any clinical or radiographic pathology. Additional follow-up time (2-3 years) is required to determine the final outcome of root canal therapy applied in cases of traumatized maxillary permanent incisor teeth with incomplete root formation.

水酸化カルシウム製剤は,根未完成歯のアペキシフィケーションに用いる薬剤である.我々は,8歳の男児が転倒時に顔面を殴打し,右側上顎中切歯が完全脱臼した症例を経験した.脱臼歯は早急に整復固定したものの失活歯となり,同時に歯肉腫脹も認めたため,根管治療(アペキシフィケーション)が必要と診断された.根管内の失活歯髄を除去した後,市販の水酸化カルシウム製剤を適応した後,症状は速やかに改善した.今後約2~3年の経過観察の後,最終根管充填処置を行う必要があると考えられる.

Dental Journal of Iwate Medical University


A Case Report of Severe Chronic Periodontitis Evaluated by Serum IgG Antibody Titer Test against Periodontal Pathogens

重度歯周病患者における歯周病原性細菌に対する血清IgG抗体価の変化:症例報告

Chronic periodontitis is a polymicrobial infectious disease which may result in loss of teeth by inflammation-mediated bone resorption. More than 500 individual cultivable species of microbes have been identified in the human mouth, and the microbiological examinations for periodontitis have been available to dental clinicians since the late 1980s. Infection with periodontal pathogens leads to humoral immunological responses and elevates the levels of serum IgG antibody to the antigens. Since serum IgG antibodies levels correspond to the amount of periodontal bacteria, the effects of treatments focused on elimination of bacteria could be evaluated by decrease of serum IgG titer to the pathogens. In the present case report, we propose the clinical usefulness of serum IgG antibody levels as a marker for periodontal treatment effects.

慢性辺縁性歯周炎は複数種の口腔細菌の感染によって発症し,炎症性骨吸収によって結果的に歯を喪失する感染症である.口腔内には500種類以上の細菌種が存在すると言われ,1980年代以降,多くの歯科医師によって歯周病の細菌学的な検討が行われてきた.とくに歯周病原性細菌の感染によって,その歯周病原細菌に対する血清中のIgG抗体価の上昇をきたすことが知られている.血清IgG抗体価のレベルは歯周病原細菌の量と比例するので,歯周病治療によって細菌量が減少すると,次第にIgG抗体価のレベルも減少すると考えられている.我々は重度歯周病患者の治療に相応して,複数種の歯周病原性細菌に対する血清IgG抗体価が減少した事例を経験したので,本症例報告において,血清IgG抗体価検査が歯周病治療の有効なマーカーになり得ることを提唱する.


A Case Report of Rheumatoid Arthritis Patient with Severe Periodontitis Whose Inflammatory Marker Improved by Periodontal Treatment

歯周治療により炎症マーカーが改善したと考察された重度歯周炎を伴うリウマチ患者の症例報告

The Japanese Journal of Conservative Dentistry


Cut-off values of Functional Independence Measure scores for discharge destination.