• Journal of dental hygiene science
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• v.23 no.4
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• pp.296-301
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• 2023

Background: Periodontal disease is a major cause of tooth loss in adults and is a representative oral disease commonly suffered by most people around the world. Mainly the proliferation of Gram-negative bacteria and secreted virulence factors cause an inflammatory response and destroy periodontal tissue. Gossypetin, isolated from Hibiscus sabdariffa L, is known to have various pharmacological effects, including antibacterial and anticancer activities. We aimed to confirm the anti-inflammatory effect of gossypetin through interleukin-6 (IL-6) regulation in human gingival fibroblasts (HGFs) stimulated with lipopolysaccharide (LPS) of Porphyromonas gingivalis, a major cause of adult periodontitis. Methods: CCK-8 assay was performed to confirm the concentration-dependent cytotoxicity of gossypetin against HGFs. The secretion level and mRNA expression of IL-6, an inflammation-related cytokine, and the effect of gossypetin on these in HGFs stimulated with P. gingivalis LPS were confirmed by ELISA and qRT-PCR analysis, respectively. Results: Up to a concentration of 100 µM gossypetin with or without P. gingivalis LPS, the survival rate for HGFs was maintained at over 95% and showed no toxicity. ELISA and qRT-PCR analysis results showed that P. gingivalis LPS increased IL-6 secretion and mRNA levels in HGFs compared to the control group. However, this increase in IL-6 was significantly down-regulated by gossypetin treatment in a dose-dependent manner. In particular, 80 µM gossypetin inhibited IL-6 production to the level of the control group. Conclusion: These results indicated that gossypetin attenuated IL-6 production in HGFs stimulated by P. gingivalis LPS, which may ultimately suppress the inflammatory response in periodontal tissue. Therefore, gossypetin may have potential as a natural ingredient for the prevention and treatment of periodontal disease.

• Restorative Dentistry and Endodontics
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• v.22 no.1
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• pp.76-92
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• 1997

Porpilyromonas endodontalis is specifically involved in endodontic infections. The bacterium can be isolated almost exclusively only from infected rool canals. P. gingivalis also has been implicated in endodontic infection. Pathogemcity of P. gingival is is attributed to a variety of virulence factors, especially proteases, produced by the bacterium. Importance of P. endodontalis in endodontic infection has been revealed. However, the pathogenic property of P. endodontalis has not been extensively studied. The present study was undertaken to characterize the proteolytic activity of P. endodontalis and compare the activity with that of P. gingivalis which has the most potent and diverse proteases among oral bacteria. For this purpose, culture supematants(SUP) and cell extracts(CE) were obtained from these two bacteria and were subjected to zymography using 15% polyacrylamide gel copolymerized with gelatin, type I, IV collagens or albumin. Hydrolysis of the collagens was further investigated by the cleavage assay using native type I and IV collagens in solution-phase. The results were as follows: 1. P. endodontalis apparently has a proteolytic activity that is comparable with that of P. gingivalis. 2. SUP and CE obtained from P. endodontalis and P. gingival is showed the strongest activity for gelatin, followed by type I and IV collagens, and albumin. 3. In the zymography, no noticeable difference in proteolytic activity for gelatin and albumin between the SUP and CE was observed, but in the cleavage assay using native collagens, the SUP showed a stronger collagenolytic activity than the CE. 4. The gelatinolytic activity of both the SUP and CE from these two bacteria was diminished in the presence of $CaCl_2$ or reducing agents such as ${\beta}$-mercaptoethanol and dithiothreitol(DTT). 5. Type I(calf skin and human placenta) collagenolytic activity of P. endodontalis and P. gingivalis was reduced by DTT but not affected by $CaCl_2$. The inhibitory effect of DTT, however, was reduced to some extent by $CaCl_2$. 6. Type IV collagenolytic activity of these two bacteria was not affected by $CaCl_2$ but increased to some extent in association with the reducing agents. 7. Hydrolysis of albumin by P. endodontalis and P. gingivalis was demonstrated only in the presence of the reducing agents. The overall results indicate that with respect to proteolytic activity, P. endodontalis appears to be as potent as P. gingivalis, or maybe more, and its proteolytic characteristic is similar to that of P. gingivalis. This suggests that P. endodontalis has so potent proteolytic activity that can participate by itself in endodontic infections and apical periodontitis, causing tissue destruction.

• BMB Reports
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• v.54 no.6
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• pp.323-328
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• 2021

Periodontal diseases have been reported to have a multidirectional association with metabolic disorders. We sought to investigate the correlation between periodontitis and diabetes or fatty liver disease using HFD-fed obese mice inoculated with P. gingivalis. Body weight, alveolar bone loss, serological biochemistry, and glucose level were determined to evaluate the pathophysiology of periodontitis and diabetes. For the evaluation of fatty liver disease, hepatic nonalcoholic steatohepatitis (NASH) was assessed by scoring steatosis, inflammation, hepatocyte ballooning and the crucial signaling pathways involved in liver metabolism were analyzed. The C-reactive protein (CRP) level and NASH score in P. gingivalis-infected obese mice were significantly elevated. Particularly, the extensive lobular inflammation was observed in the liver of obese mice infected with P. gingivalis. Moreover, the expression of metabolic regulatory factors, including peroxisome proliferator-activated receptor γ (Pparγ) and the fatty acid transporter Cd36, was up-regulated in the liver of P. gingivalis-infected obese mice. However, inoculation of P. gingivalis had no significant influence on glucose homeostasis, insulin resistance, and hepatic mTOR/AMPK signaling. In conclusion, our results indicate that P. gingivalis can induce the progression of fatty liver disease in HFD-fed mice through the upregulation of CD36-PPARγ axis.

• Biomedical Science Letters
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• v.29 no.2
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• pp.88-94
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• 2023

In oriental medicine, the fruit of Actinidia polygama has long been used to alleviate the symptoms of gout, arthritis, and inflammation. In this study, it was to designed to analyze the antibacterial activity of A. polygama ethanol extract (APEE) against Streptococcus mutans, one of the major strains for dental caries, and Porphyromonas gingivalis, one of the critical strains for periodontal disease. The antibacterial activity of APEE was analyzed by disk diffusion, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) assays. In addition, it was also analyzed the inhibitory effect of APEE on bacterial growth and biofilm formation against both oral pathogens. APEE exhibited its antibacterial effect through the inhibited bacterial diffusion as well as low concentration of MIC and MBC. In addition, APEE significantly inhibited not only bacterial growth but also biofilm formation in a dose-dependent manner. Consequently, APEE showed potent antibacterial activity against both S. mutans and P. gingivalis, which indicates that APEE might be used as a potential antibacterial material for the improvement of oral healthcare.

• Journal of Periodontal and Implant Science
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• v.42 no.6
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• pp.185-195
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• 2012

Purpose: (-)-epigallocatechin-3-gallate (EGCG) has been reported to exert anti-inflammatory and antibacterial effects in periodontitis. However, its exact mechanism of action has yet to be determined. The present in vitro study evaluated the anti-in-flammatory effects of EGCG on human periodontal ligament fibroblasts (hPDLFs) and human periodontal ligament stem cells (hPDLSCs) affected by bacterial lipopolysaccharide (LPS) extracted from Porphyromonas gingivalis. Methods: hPDLFs and hPDLSCs were extracted from healthy young adults and were treated with EGCG and/or P. gingivalis LPS. After 1, 3, 5, and 7 days from treatment, cytotoxic and proliferative effects were evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and bromodeoxyuridine assay, respectively. And then, the gene expressions of hPDLFs and hPDLSCs were observed for interleukin (IL)-$1{\beta}$, IL-6, tumor necrosis factor (TNF)-${\alpha}$, osteoprotegerin (OPG), receptor activator of nuclear factor kappa-B ligand (RANKL), and RANKL/OPG using real-time polymerase chain reaction (PCR) at 0, 6, 24, and 48 hours after treatment. The experiments were performed with the following groups for hPDLFs and hPDLSCs; 1) No treat, 2) EGCG alone, 3) P. gingivalis LPS alone, 4) EGCG+P. gingivalis LPS. Results: The 20 ${\mu}M$ of EGCG and 20 ${\mu}g/mL$ of P. gingivalis LPS had the lowest cytotoxic effects, so those concentrations were used for further experiments. The proliferations of hPDLFs and hPDLSCs increased in all groups, though the 'EGCG alone' showed less increase. In real-time PCR, the hPDLFs and hPDLSCs of 'EGCG alone' showed similar gene expressions to those cells of 'no treat'. The gene expressions of 'P. gingivalis LPS alone' in both hPDLFs and hPDLSCs were highly increased at 6 hours for IL-$1{\beta}$, IL-6, TNF-${\alpha}$, RANKL, and RANKL/OPG, except the RANKL/OPG in hPDLSCs. However, those increased gene expressions were down-regulated in 'EGCG+P. gingivalis LPS' by the additional treatment of EGCG. Conclusions: Our results demonstrate that EGCG could exert an anti-inflammatory effect in hPDLFs and hPDLSCs against a major pathogen of periodontitis, P. gingivalis LPS.

• Journal of Life Science
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• v.29 no.12
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• pp.1386-1392
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• 2019

This study investigated the potential of dye plants as natural oral health products. The antibacterial activity of ethanol stem extracts of A. japonica, R. verniciflua Stokes, G. jasminoides, D. morbifera, P. amurense Rupr., and S. japonica against P. gingivalis KCTC 5352, S. mutans KCTC3065, S. downei KCTC3634, S. sanguinis KCTC3284, and S. gordonii KCTC 3286 was confirmed. Among the stem extracts from 6 dye plants grown in Korea, ethanol extract from A. japonica stem (1 mg/disc) showed the highest antibacterial activity against P. gingivalis KCTC5352. The A. japonica stem extracts showed antibacterial activity similar to chlorhexidine, which was used as a positive control. The MIC and MBC of P. gingivalis KCTC5352 were 0.4 mg/ml and 0.6 mg/ml, respectively. The biofilm production rate and cell growth of P. gingivalis KCTC5352 in the cultures treated with 0.2-2.0 mg/ml of A. japonica extract were significantly decreased in a concentration-dependent manner. In addition, the mRNA expression of the superoxide dismutase and fimA associated with fimbriae formation in these cultures was suppressed, also in a concentration-dependent manner. Based on these results, it is concluded that A. japonica stem extracts can be used as an oral health product derived from natural materials, as demonstrated by its antibacterial action against and inhibition of biofilm formation of P. gingivalis KCTC5352.

• International Journal of Oral Biology
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• v.33 no.4
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• pp.163-177
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• 2008

Periodontal disease, a form of chronic inflammatory bacterial infectious disease, is known to be a risk factor for cardiovascular disease (CVD). Porphyromonas gingivalis has been implicated in periodontal disease and widely studied for its role in the pathogenesis of CVD. A previous study demonstrating that periodontopathic P. gingivalis is involved in CVD showed that invasion of endothelial cells by the bacterium is accompanied by an increase in cytokine production, which may result in vascular atherosclerotic changes. The present study was performed in order to further elucidate the role of P. gingivalis in the process of atherosclerosis and CVD. For this purpose, invasion of human aortic smooth muscle cells (HASMC) by P. gingivalis 381 and its isogenic mutants of KDP150 ($fimA^-$), CW120 ($ppk^-$) and KS7 ($relA^-$) was assessed using a metronidazole protection assay. Wild type P. gingivalis invaded HASMCs with an efficiency of 0.12%. In contrast, KDP150 failed to demonstrate any invasive ability. CW120 and KS7 showed relatively higher invasion efficiencies, but results for these variants were still negligible when compared to the wild type invasiveness. These results suggest that fimbriae are required for invasion and that energy metabolism in association with regulatory genes involved in stress and stringent response may also be important for this process. ELISA assays revealed that the invasive P. gingivalis 381 increased production of the proinflammatory cytokine interleukin (IL)-$1{\beta}$ and the chemotactic cytokines (chemokine) IL (interleukin)-8 and monocyte chemotactic (MCP) protein-1 during the 30-90 min incubation periods (P<0.05). Expression of RANTES (regulation upon activation, normal T cell expressed and secreted) and Toll-like receptor (TLR)-4, a pattern recognition receptor (PRR), was increased in HASMCs infected with P. gingivalis 381 by RT-PCR analysis. P. gingivalis infection did not alter interferon-$\gamma$-inducible protein-10 expression in HASMCs. HASMC nonspecific necrosis and apoptotic cell death were measured by lactate dehydrogenase (LDH) and caspase activity assays, respectively. LDH release from HASMCs and HAMC caspase activity were significantly higher after a 90 min incubation with P. gingivalis 381. Taken together, P. gingivalis invasion of HASMCs induces inflammatory cytokine production, apoptotic cell death, and expression of TLR-4, a PRR which may react with the bacterial molecules and induce the expression of the chemokines IL-8, MCP-1 and RANTES. Overall, these results suggest that invasive P. gingivalis may participate in the pathogenesis of atherosclerosis, leading to CVD.

• Journal of Periodontal and Implant Science
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• v.48 no.2
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• pp.70-83
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• 2018

Purpose: The aim of this study was to evaluate the capacity of single and combined applications of the bark of the stems and roots of Magnolia officinalis Rehd. et Wils. (Magnoliae Cortex) and Zea mays L. (maize) to modulate inflammation in RAW 264.7 cells stimulated with Porphyromonas gingivalis. Methods: RAW 264.7 cells were stimulated with P. gingivalis, and Magnoliae Cortex and/or maize was added. Cytotoxicity and the capacity to modulate inflammation were determined with a methylthiazol tetrazolium (MTT) assay, nitrite production, enzyme-linked immunosorbent assay (ELISA), and western blotting. Results: Treatment with Magnoliae Cortex and/or maize inhibited nuclear transcription factor ${\kappa}B$ ($NF-{\kappa}B$) pathway activation and nuclear p44/42 mitogen-activated protein kinase (MAPK) and inducible nitric oxide synthase (iNOS) protein expression in P. gingivalis-stimulated RAW 264.7 cells. Moreover, the treatments suppressed cytokines (prostaglandin $E_2$ [$PGE_2$], interleukin $[IL]-1{\beta}$, and IL-6) and nitrite production. Conclusions: Both Magnoliae Cortex and maize exerted an anti-inflammatory effect on P. gingivalis-stimulated RAW 264.7 cells, and this effect was more pronounced when the extracts were combined. These findings show that these extracts may be beneficial for slowing the progression of periodontal disease.

• Journal of Periodontal and Implant Science
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• v.30 no.1
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• pp.105-111
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• 2000

본 연구는 Balb/c mice를 이용하여 Porphyromonas gingivalis 381(Pg)로 면역하기 전에 Fusobacterium nucleatum ATCC 10953(Fn)로 면역한 Group 1(N=10)과 Pg 로만 단독 면역을 시행한 Group 2(N=10)로부터 채취한 혈청의 Pg에 대한 식균능력을 비교하는 데 그 목적이 있다. 면역 후 혈청항체는 Pg에 대해 현저히 상승하였으나, 두 그룹간의 평균 항체역가는 통계적으로 차이가 없었다. 식균능력을 비교한 결과, Pg로만 단독 면역한 경우 식균능력이 Fn으로 먼저 면역한 Group 1의 경우보다 현저히 높았으며, 혈청항체역가와 식균지수와는 긴밀한 상관관계를 보였다. 결론적으로 치주세균의 사전 감염은 후속적인 세균감염에 대한 숙주 면역기능(식균능력)에 교란을 가져 올 수 있다.

• Journal of Periodontal and Implant Science
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• v.48 no.1
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• pp.12-21
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• 2018

Purpose: The goal of this study was to develop and validate a standardized in vitro pathogenic biofilm attached onto saliva-coated surfaces. Methods: Fusobacterium nucleatum (F. nucleatum) and Porphyromonas gingivalis (P. gingivalis) strains were grown under anaerobic conditions as single species and in dual-species cultures. Initially, the bacterial biomass was evaluated at 24 and 48 hours to determine the optimal timing for the adhesion phase onto saliva-coated polystyrene surfaces. Thereafter, biofilm development was assessed over time by crystal violet staining and scanning electron microscopy. Results: The data showed no significant difference in the overall biomass after 48 hours for P. gingivalis in single- and dual-species conditions. After adhesion, P. gingivalis in single- and dual-species biofilms accumulated a substantially higher biomass after 7 days of incubation than after 3 days, but no significant difference was found between 5 and 7 days. Although the biomass of the F. nucleatum biofilm was higher at 3 days, no difference was found at 3, 5, or 7 days of incubation. Conclusions: Polystyrene substrates from well plates work as a standard surface and provide reproducible results for in vitro biofilm models. Our biofilm model could serve as a reference point for studies investigating biofilms on different surfaces.