치주염 유발 세균 Aggregatibacter actinomycetemcomitans와 Porphyromonas gingivalis에 의한 committed osteoclast precursor 분화 증가

박옥진 ( Ok-jin Park ),권영각 ( Yeongkag Kwon ),윤철희 ( Cheol-heui Yun ),한승현 ( Seung Hyun Han ) 한국미생물생명공학회(구 한국산업미생물학회) 2016 한국미생물·생명공학회지 Vol.44 No.4

치주질환은 만성염증성 질환으로 치조골소실을 일으켜 성인치아상실을 유발하는 요인 중 하나이다. 그람 음성세균인Aggregatibacter actinomycetemcomitans와 Porphyromonas gingivalis는 치주질환환자의 병소에서 쉽게 동정된다. 지질 다당체(Lipopolysaccharide; LPS)는 그람 음성세균의 핵심 독력인자로 알려져 있다. 이러한 세균과 LPS는 파골세포에 의한 골소실을 조절하는 요인 중 하나이다. 그러므로 본 연구에서는 동물모델을 활용하여 A. actinomycetemcomitans와 P. gingivalis의 의한 골소실 여부를 확인하고, 기전규명을 위하여 A. actinomycetemcomitans, P. gingivalis, A. actinomycetemcomitans와 P. gingivalis에서 분리한 LPS에 의한 파골세포분화 영향을 연구하였다. 열사멸한 A. actinomycetemcomitans (HKAa)와 열사멸한 P. gingivalis (HKPg)가 복강으로 투여된 쥐의 대퇴골은 대조군에 비해 감소된 골량을 보여주었다. 이러한 골소실의 증가가 파골세포분화 때문인지 확인하기 위해 파골세포분화를 연구한 결과, bone marrow-derived macrophage (BMM)의 RANKL-매개 파골세포분화를 감소시켰으나, committed osteoclast precursor의 파골세포분화를 유도함을 확인하였다. 세균에 의한 파골세포분화 결과와 동일하게 A. actinomycetemcomitans와 P. gingivalis에서 분리한 LPS 역시 RANKL-매개 파골세 포분화는 감소시키고, committed osteoclast precursor의 파골세포분화를 유도하였다. 결과적으로 치주원인균인 A. actinomycetemcomitans와 P. gingivalis는 committed osteoclast precursor의 파골세포분화를 증가시키는데, 이 세균들의 LPS가 핵심 역할을 수행하는 것으로 판단되며 이를 통해 골 흡수를 유발함을 알 수 있었다. Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis are gram-negative bacteria frequently found in lesions from patients with periodontitis manifesting alveolar bone loss. Lipopolysaccharides are a major virulence factor of gram-negative bacteria. Bone resorption is known to be regulated by bacteria and their virulence factors. In the present study, we investigated the effects of A. actinomycetemcomitans and P. gingivalis on bone resorption. Heat-killed A. actinomycetemcomitans (HKAa) and heatkilled P. gingivalis (HKPg) induced bone loss in the femurs of mice after intraperitoneal administration. HKAa and HKPg augmented the differentiation of committed osteoclast precursors into osteoclasts, while they inhibited the differentiation of bone marrow-derived macrophages into osteoclasts. Concordant with the effects of the heat-killed whole cells, LPS purified from A. actinomycetemcomitans and P. gingivalis also augmented osteoclast differentiation from committed osteoclast precursors but attenuated it from bone marrow-derived macrophages. Taken together, these results suggest that the whole cells and lipopolysaccharides of A. actinomycetemcomitans and P. gingivalis induce the differentiation of committed osteoclast precursors into osteoclasts, potentially contributing to bone resorption in vivo.

Development and Optimization of a Rapid Colorimetric Membrane Immunoassay for Porphyromonas gingivalis

( Jiyon Lee ),( Myoung-kwon Choi ),( Jinju Kim ),( Sechul Chun ),( Hong-gyum Kim ),( Hosung Lee ),( Jinsoo Kim ),( Dongwook Lee ),( Seung-hyun Han ),( Do-young Yoon ) 한국미생물생명공학회(구 한국산업미생물학회) 2021 Journal of microbiology and biotechnology Vol.31 No.5

Porphyromonas gingivalis (P. gingivalis) is a major bacterial pathogen that causes periodontitis, a chronic inflammatory disease of tissues around the teeth. Periodontitis is known to be related to other diseases, such as oral cancer, Alzheimer’s disease, and rheumatism. Thus, a precise and sensitive test to detect P. gingivalis is necessary for the early diagnosis of periodontitis. The objective of this study was to optimize a rapid visual detection system for P. gingivalis. First, we performed a visual membrane immunoassay using 3,3′,5,5′-tetramethylbenzidine (TMB; blue) and coating and detection antibodies that could bind to the host laboratory strain, ATCC 33277. Antibodies against the P. gingivalis surface adhesion molecules RgpB (arginine proteinase) and Kgp (lysine proteinase) were determined to be the most specific coating and detection antibodies, respectively. Using these two selected antibodies, the streptavidin-horseradish peroxidase (HRP) reaction was performed using a nitrocellulose membrane and visualized with a detection range of 10³-10⁵ bacterial cells/ml following incubation for 15 min. These selected conditions were applied to test other oral bacteria, and the results showed that P. gingivalis could be detected without cross-reactivity to other bacteria, including Streptococcus mutans and Escherichia fergusonii. Furthermore, three clinical strains of P. gingivalis, KCOM 2880, KCOM 2803, and KCOM 3190, were also recognized using this optimized enzyme immunoassay (EIA) system. To conclude, we established optimized conditions for P. gingivalis detection with specificity, accuracy, and sensitivity. These results could be utilized to manufacture economical and rapid detection kits for P. gingivalis.

Fusobacterium nucleatum modulates serum binding to Porphyromonas gingivalis biofilm

최점일,김성조,김수진,Choi, Jeom-Il,Kim, Sung-Jo,Kim, Soo-Jin The Korean Academy of Periodontoloy 2001 Journal of Periodontal & Implant Science Vol.31 No.4

P. gingivalis를 단독면역하거나 또는 Fusobacterium nucleatum 선면역 후 P. gingivalis 항혈청을 각각 얻어냈다. 두 종류의 항혈청이 P. gingivalis biofilm을 침투해 들어가는 능력을 confocal laser scanning microscope를 이용하여 비교 감증하였다. 항혈청의 P. gingivalis에 대한 avidity index도 측정하였다. 결과적으로 F. nucleatum의 선면역은 P. gingivalis 특이 항혈청에 대해 세균성 biofilm의 침투능력을 저하시키고, 동일한 세균에 대한 avidity도 감소시켰다. Anti-P. gingivalis immune sera were obtained from mice immunized with either P. gingivalis alone, or F. nucleaturm followed by P. gingivalis. Two groups of immune sera were examined for binding capacity to P. gingivalis biofilm by confocal laser scanning microscope, Antibody avidity index was also determined for each immune sera. The results indicated that prior immunization of mice with F. nucleaturm impaired P. gingivalis-specific immune sera in binding capacity to biofilm and antibody avidity to P. gingivalis. Elevated antibody responses in patients with destructive periodontal disease has often been related to suboptimal level of protective antibody $(opsonophagocytosis)^{1-3)}$ while post-immune sera obtained with experimental animals using a single periodontal pathogen demonstrated satisfactory levels of protective function against the homologous bacterial $challenge^{4,5)}$.The reason is unclear why elevated IgG responses in periodontal patients to periodontal pathogens do not necessarily reflect their protective function. Such an immune deviation might be derived from the fact that destructive periodontal disease is cumulative result of immunopathologic processes responding to an array of different colonizing microorganisms sequentially infecting in the subgingival environmental niche. Fusobacterium nucleaturm is one of the key pathogens in gingivitis, in the transitional phase of conversion of gingivitis into destructive periodontitk, and in adult $periodontitis^{6-8)}$. It also plays a central role in coaggregation with other important microbial species in subgingival $area^{6,9,10)}$ as well as in $biofilm^{11)}$, especially with Porphyromonas gingjvalis in synergism of virulence in human periodontal disease or in animal $models^{12-14)}$. This organism has also been reported to have immune modulating activity for secondary immune response to Actinobacillus $actinomycetemcomitans^{15)}$. It is presumed that sequential colonization and intermicrobial coaggregation between intermediate and late colonizers could potentially modulate the immune responses and development of specific T cell phenotypes in periodontal lesions. We have recently demonstrated the skewed polarization of P. gingivalis-specific helper T cell clones in mice immunized with F. nucleaturm followed by P. $gingivalis.^{16)}$. Consequently F. nucleaturm may initially prime the immune cells and modify their responses to the successive organism, P. gingivalis. This could explain why one frequently observes non-protective serum antibodies to P. gingivalis in periodontal patients in contrast with those obtained from animals that were immunized with $P.gingivalis\;alone^{17)}$. The present study was performed to investigate the immune modulating effect of F. nucleatum on serum binding to experimental biofilms and the avidity of anti-P. gingivalis antibody.

Porphyromonas gingivalis가 일부 구강미생물의 형광 발현에 미치는 영향

김세연 ( Se-yeon Kim ),우동협 ( Dong-hyeob Woo ),이민아 ( Min-ah Lee ),김지수 ( Ji-soo Kim ),이정하 ( Jung-ha Lee ),정승화 ( Seung-hwa Jeong ) 대한예방치과·구강보건학회 2017 大韓口腔保健學會誌 Vol.41 No.1

Objectives: Dental plaque is composed of 700 bacterial species. It is known that some oral microorganisms produce porphyrin, and thus, they emit red fluorescence when illuminated with blue light at a specific wavelength of <410 nm. Porphyromonas gingivalis belongs to the genus Porphyromonas , which is characterized by the production of porphyrin. The aim of this study was to evaluate red fluorescence emission of some oral microorganisms interacting with P. gingivalis. Methods: Five bacterial strains (P. gingivalis, Streptococcus mutans, Lactobacillus casei, Actinomyces naeslundii, and Fusobacterium nucleatum) were used for this study. Tryptic soy agar medium supplemented with hemin, vitamin K3, and sheep blood was used as a growth medium. The fluorescence emission of bacterial colonies was evaluated under 405 nm-wavelength blue light using a Quantitative Light-induced Fluorescence Digital (QLF-D) camera system. Each bacterium was cultured alone and cocultured in close proximity with P. gingivalis. The red/green (R/G) ratio of fluorescence image was calculated and the differences of R/G ratio according to each growth condition were compared using the Mann-Whitney test (P<0.05). Results: Single cultured S. mutans , L. casei and A. naeslundii colonies emitted red fluorescence (R/ G ratio=2.15±0.06, 4.31±0.17, 5.52±1.29, respectively). Fusobacterium nucleatum colonies emitted green fluorescence (R/G ratio=1.36±0.06). The R/G ratios of A. naeslundii and F. nucleatum were increased when P. gingivalis was co-cultured with each bacterium (P<0.05). In contrast, the R/G ratios of S. mutans and L. casei were decreased when P. gingivalis was co-cultured with each bacterium (P=0.002, 0.003). Conclusions: This study confirmed that P. gingivalis could affect the red fluorescence of other oral bacteria under 405 nm-wavelength blue light. Our findings concluded that P. gingivalis has an important role for red fluorescence emission of dental biofilm.

<i>Porphyromonas gingivalis</i> traffics into endoplasmic reticulum-rich-autophagosomes for successful survival in human gingival epithelial cells

Lee, Kyulim,Roberts, JoAnn S.,Choi, Chul Hee,Atanasova, Kalina R.,Yilmaz, Ö,zlem TaylorFrancis 2018 Virulence Vol.9 No.1

<P><B>ABSTRACT</B></P><P><I>Porphyromonas gingivalis</I>, an opportunistic pathogen usurps gingival epithelial cells (GECs) as primary intracellular niche for its colonization in the oral mucosa. However, the precise characterization of the intracellular trafficking and fate of <I>P. gingivalis</I> in GECs remains incomplete. Therefore, we employed high-resolution three-dimensional-transmission-electron-microscopy to determine the subcellular location of <I>P. gingivalis</I> in human primary GECs upon invasion. Serial sections of infected-GECs and their tomographic reconstruction depicted ER-rich-double-membrane autophagosomal-vacuoles harboring <I>P. gingivalis</I>. Western-blotting and fluorescence confocal microscopy showed that <I>P. gingivalis</I> significantly induces LC3-lipidation in a time-dependent-manner and co-localizes with LC3, ER-lumen-protein Bip, or ER-tracker, which are major components of the phagophore membrane. Furthermore, GECs that were infected with FMN-green-fluorescent transformant-strain (PgFbFP) and selectively permeabilized by digitonin showed rapidly increasing large numbers of double-membrane-vacuolar-<I>P. gingivalis</I> over 24 hours of infection with a low-ratio of cytosolically free-bacteria. Moreover, inhibition of autophagy using 3-methyladenine or ATG5 siRNA significantly reduced the viability of intracellular <I>P. gingivalis</I> in GECs as determined by an antibiotic-protection-assay. Lysosomal marker, LAMP-1, showed a low-degree colocalization with <I>P. gingivalis</I> (∼20%). PgFbFP was used to investigate the fate of vacuolar- versus cytosolic-<I>P. gingivalis</I> by their association with ubiquitin-binding-adaptor-proteins, NDP52 and p62. Only cytosolic-<I>P. gingivalis</I> had a significant association with both markers, which suggests cytosolically-free bacteria are likely destined to the lysosomal-degradation pathway whereas the vacuolar-<I>P. gingivalis</I> survives. Therefore, the results reveal a novel mechanism for <I>P. gingivalis</I> survival in GECs by harnessing host autophagy machinery to establish a successful replicative niche and persistence in the oral mucosa.</P>

Porphyromonas gingivalis accelerates atherosclerosis through oxidation of high-density lipoprotein

Hyun-Joo Kim,Gil Sun Cha,김형준,Eun-Young Kwon,이주연,최점일,주지영 대한치주과학회 2018 Journal of Periodontal & Implant Science Vol.48 No.1

Purpose: The aim of this study was to evaluate the ability of Porphyromonas gingivalis (P. gingivalis) to induce oxidation of high-density lipoprotein (HDL) and to determine whether the oxidized HDL induced by P. gingivalis exhibited altered antiatherogenic function or became proatherogenic. Methods: P. gingivalis and THP-1 monocytes were cultured, and the extent of HDL oxidation induced by P. gingivalis was evaluated by a thiobarbituric acid-reactive substances (TBARS) assay. To evaluate the altered antiatherogenic and proatherogenic properties of P. gingivalis-treated HDL, lipid oxidation was quantified by the TBARS assay, and tumor necrosis factor alpha (TNF-α) levels and the gelatinolytic activity of matrix metalloproteinase (MMP)-9 were also measured. After incubating macrophages with HDL and P. gingivalis, Oil Red O staining was performed to examine foam cells. Results: P. gingivalis induced HDL oxidation. The HDL treated by P. gingivalis did not reduce lipid oxidation and may have enhanced the formation of MMP-9 and TNF-α. P. gingivalis-treated macrophages exhibited more lipid aggregates than untreated macrophages. Conclusions: P. gingivalis induced HDL oxidation, impairing the atheroprotective function of HDL and making it proatherogenic by eliciting a proinflammatory response through its interaction with monocytes/macrophages.

Streptococcus mutans와 Porphyromonas gingivalis에 대한 상백피(Morus alba root bark) 에탄올 추출물의 항균 효과

박충무(Chung Mu Park),윤현서(Hyun-Seo Yoon) 한국차학회 2023 한국차학회지 Vol.29 No.4

본 연구는 구강질환을 유발하는 대표적인 두 균주인 Streptococcus mutans와 Porphyromonas gingivalis에대한 상백피 에탄올 추출물(MAEE)의 항균 활성을 분석하였다. Griess reaction과 WST-1 assay를 통해lipopolysaccharide (LPS)로 자극한 생쥐대식세포인 RAW 264.7 세포에서 MAEE의 nitric oxide (NO) 억제 및 세포독성 효과를 분석하였다. 그리고 디스크 확산법, 최소억제농도(minimum inhibition concentration, MIC), 최소살균농도(minimum bactericidal concentration, MBC), 성장억제 효과 분석을 통해 MAEE의 항균활성을 평가하였다. 그 결과 MAEE는 LPS로 자극한 RAW 264.7 세포에서 세포독성없이 농도 의존적으로 NO 생성을 억제하였다. 그리고 MAEE는 강력한 항균 활성을 나타냈는데, 100 mg/mL의 MAEE는 S. mutans와 P. gingivalis에 대해 각각 13.94 mm와 15.65 mm의 clear zone을 형성하는 것으로 보다 MAEE는 P. gingivalis보다 S. mutans에 더강한 항균활성을 보이는 것으로 생각된다. 그리고, S. mutans와 P. gingivalis의 MIC는 각각 0.4 mg/mL와 0.4~0.8 mg/mL였고, 두 병원균의 MBC는 각각 0.4 mg/mL와 0.8 mg/mL로 나타났다. 또한 MAEE 처리는 두 병원균의성장을 유의하게 억제하였으나 농도 의존적이지는 않았다. 특히, 24시간동안 MAEE를 처리한 S. mutans의OD600이 3.2 mg/mL에서 0.146인 반면 대조군의 흡광도는 0.509를 보였다. 그리고 같은 용량의 MAEE 처리된 P.gingivalis를 24시간 동안 배양했을 때 대조군의 흡광도 0.486 대비 0.154로 나타나 강한 성장억제효과를 보였다.결론적으로, MAEE는 두 구강 내 병원균에 대해 강력한 항균 활성을 보였으며, MAEE는 P. gingivalis보다 S.mutans에 대해 더 강력한 항균 활성을 나타내는 것을 확인할 수 있었다. 이러한 결과를 종합해 볼 때 상백피는다양한 형태로 가공하여 음용 시 항당뇨, 고혈압, 콜레스테롤 개선 외에도 본 연구를 통해 치아우식증과 치주질환의 예방 및 치료제로 활용 가능성을 검증하였기에 상백피의 차로 활용성을 높인 것으로 생각된다. This study aimed to analyze the antibacterial activity of the ethanol extract of Morus alba root bark (MAEE) against two critical oral pathogens, Streptococcus mutans and Porphyromonas gingivalis. The Griess reaction and WST-1 assays were used to analyze the nitric oxide (NO) inhibitory and cytotoxic effects of MAEE on lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. The antibacterial activity of MAEE was evaluated using a disk diffusion assay and a growth inhibition assay by determining minimum inhibition concentrations (MICs) and minimum bactericidal concentrations (MBCs). MAEE treatment dose-dependently inhibited NO production and had no cytotoxic effect on LPS-stimulated RAW 264.7 cells. Furthermore, MAEE exhibited potent antibacterial activity and more potently generated a clear zone in the presence of S. mutans than P. gingivalis. The MICs of MAEE for S.mutans and P. gingivalis were 0.4 and 0.4~0.8 mg/mL, and its MBCs were 0.4 and 0.8 mg/mL, respectively. In addition, MAEE significantly inhibited the growths of both pathogens, but no dose-dependency was observed. The OD600 of S. mutans treated with MAEE at 3.2 mg/mL for 24 h was 0.146 compared to a control OD600 of 0.509. Absorbance at 600 nm for P. gingivalis treated with MAEE at 3.2 mg/mL for 24 h was 0.154 compared to a control OD600 of 0.486. In conclusion, MAEE showed potent antibacterial activity against both oral pathogens but affected S.mutans stronger than P. gingivalis. The study indicates that MAEE might help prevent dental caries and periodontal diseases.

구강 질환 유발균인 Streptococcus mutans와 Porphyromonas gingivalis에 대한 모링가(Moringa oleifera) 잎의 항균 효과

박충무,현숙경,윤현서 한국차학회 2023 한국차학회지 Vol.29 No.3

구강질환을 유발하는 두 균주인 Streptococcus mutans와 Porphyromonas gingivalis에 대한 모링가(Moringa oleifera) 잎 에탄올 추출물 (M. oleifera ethanol extract, MOEE)의 항균 효과를 확인하고자 하였다. MOEE의 항균활성은 디스크 확산법, 최소억제농도(minimum inhibition concentration, MIC), 최소살균농도(minimum bactericidal concentration, MBC), 성장억제 효과 분석을 통해서, RAW 264.7 cell에서의 nitric oxide (NO) 생성 억제효과와 세포독성은 Griess reaction과 WST-1 assay를 통해 분석하였다. 실험 결과, lipopolysaccharide (LPS)에 의해유도된 NO 생성은 MOEE의 농도가 높아짐에 따라 유의미하게 억제되었고, 세포독성도 강하게 나타나지 않았다. 그리고 MOEE는 S. mutans와 P. gingivalis에 대해 300 mg/mL에서 19.72 mm, 17.22 mm로 나타나 MOEE는 P. gingivalis를 더 강하게 억제하였다. 또한 S. mutans와 P. gingivalis의 MIC는 0.8 mg/mL과 0.4~0.8 mg/mL였고, MBC는 두 균주 모두에서 0.8 mg/mL로 나타났다. 성장억제 효과는 S. mutans와 P. gingivalis를 0.8 mg/mL의MOEE 농도에서 24시간 배양하였을 때 OD600이 각각 0.219, 0.125로 나타나 MOEE가 P. gingivalis의 성장을 더 강하게 억제하는 것으로 나타났다. 결론적으로, MOEE는 구강질환을 유발하는 두 균주에 대해 유의미한 항균 효과를 나타내었고, 특히 S. mutans와 P. gingivalis의 성장을 더 강하게 억제하였다. 추후 생물막 생성 억제와 다양한 용매 추출물에 대한 활성 검증을 통해 모링가 잎이 치아우식증과 치주질환 예방을 위한 기능성 소재로서 활용될 수있을 기초를 제공할 수 있을 것으로 생각한다. This study examined the antibacterial activity of Moringa oleifera ethanol extract (MOEE) against oral pathogens, Streptococcus mutans and Porphyromonas gingivalis. The antibacterial activity of MOEE was analyzed using a disk diffusion assay, minimum inhibition concentration (MIC), minimum bactericidal concentration (MBC), and growth inhibition assay, and the inhibitory effect of nitric oxide synthesis and cytotoxicity was assessed using the Griess reaction and WST-1 assay in RAW 264.7 cells. NO production by LPS stimulation was inhibited dose-dependently without cytotoxicity. MOEE potently inhibited the disk diffusion of P. gingivalis than S. mutans, which was 19.72 mm and 17.22 mm at 300 mg/mL, respectively. In addition, the MIC of S. mutans and P. gingivalis was 0.8 mg/mL and 0.4~0.8 mg/mL, and the MBC of both pathogens was 0.8 mg/mL for both. MOEE also potently inhibited the growth of P. gingivalis and S. mutans, which was OD600 of 0.8 mg/mL MOEE-treated BHI broth was 0.219 and 0.125, respectively. Consequently, MOEE exhibited potent antibacterial activity, which was stronger against P. gingivalis than S. mutans. MOEE might be used as a functional material for preventing dental caries and periodontal diseases when the inhibitory activity of the biofilm and extracts from various solvents will be conducted in future studies.

Porphyromonas gingivalis의 독성, 대사산물 및 유전자이종성과의 관련성

김강주,정종평,Kim, Kang-Ju,Chung, Chong-Pyoung 대한치주과학회 1993 Journal of Periodontal & Implant Science Vol.23 No.1

P. gingivalis has been implicated as a strong pathogen in periodontal disease and known to have three serotypes of P. gingivalis. The purpose of this study is to investigate on the relationship between virulence, metabolic acids and genetic heterogeneity of P. gingivalis. P. gingivalis W50 standard strain and five strains of P. gingivalis serotype b Korean isolates were used in this study. For in vitro virulence test, lyophilized whole cell P. gingivalis were suspended, and sonicated with ultrasonic dismembranometer. Sonicated samples were applied to cultured cells derived from periodontal ligament, and cell activity was assayed with growth and survival assay. The metabolic acids were also extracted, and determined by High Performance Liquid Chromatography. Pst I-digested bacterial genomic DNA was electrophoresed, and densitometric analysis was performed to study the genetic heterogeneity. All of the P. gingivalis serotype b produced butyric acid. In cell activity study, butyric acid inhibited the cell activity irrespective of its concentration. Densitometric analysis showed restriction fragment length polymorphism. These results suggested that there existed heterogeneity of the metabolic acids and the virulence of P. gingivalis and such heterogeneity might be related to genetic heterogeneity.

Induction of NADPH oxidases and antioxidant proteins by Porphyromonas gingivalis in KB cells

김민정,정현주,박병주,박해령,이태훈,Kim, Min-Jeong,Chung, Hyun-Ju,Park, Byung-Ju,Park, Hae-Ryoung,Lee, Tae-Hun The Korean Academy of Periodontoloy 2006 Journal of Periodontal & Implant Science Vol.36 No.4

Porphyromonas gingivalis는 치주질환을 야기하는 독성세균으로서, 구강상피세포에 p. gingivalis가 감염되었을 때, 세포형태에 변화를 초래함으로 인해 방어기작이 작동하게 된다. 치주질환과 관련되어 생성된 활성 산소종의 소거에 관여하는 항산화성분은 p. gingivalis 이 감염된 구강상피세포에서 그 분포와 발현수준이 달라지리라 예상된다. 따라서 이번 연구에서는 구강상피세포(KB 세포)에 p. gingivalis가 감염되었을 때 야기되는 활성산소종과 이를 소거하는 역할을 하는 항산화단백들의 역할들을 규명하고자 하였다. 활성산소종 형성을 조절하는 NADPH oxidase 중 NOX4와 Rac1 전사체는 구강상피세포에서 p. gingivalis세균에 의해 증가하였으며 $gp91^{phox}$, Rac2, $p47^{phox}$와 $p67^{phox}$는 세균에 의한 변화가 관찰되지 않았다. 반면에 $p40^{phox}$ 전사체는 감소하는 경향을 보였다. NOX1 전사체는 p. gingivalis 처리 30분 후 감소하였다가 60분 후에는 다시 증가하는 양상을 보였다. 같은 시간에 NOX 활성화 단백인 NOXA1은 감소하고, NOX 구성단백질인 NOXO1은 증가하는 경향을 보였다. p. gingivalis가 감염된 구강상피세포를 방어하는 항산화단백 발현수준을 평가한 결과, SOD1, 2, 3 모두 p. gingivalis 처리시간에 따라 증가하는 양상을 보였다. GPx 발현 양상도 SOD와 유사하게 나타났다. $H_2O_2$를 소거하는 Prx는 감염된 KB 세포에서 Prx4와 Prx5가 4-6배 증가하는 것을 알 수 있었다. 반면 endocytosis 과정 중 $H_2O_2$ 생산은 변화되지 않았다. 이번 연구의 결과, p. gingivalis의 감염은 KB 세포의 NOX4와 Rac1의 NADPH oxidase 발현을 증가시켰으며, NOX1은 NOXA1과 NOXO1의 조절에 의해 영향을 받음을 알 수 있었다. 또한 항산화기작으로는 SOD, GPx, Prx가 증가하였는데, 이것은 Prx4와 Prx5가 중요한 역할을 할 것을 시사하였다.