rgpA DNA Vaccine Induces Antibody Response and Prevents Alveolar Bone Loss in Experimental Peri‐Implantitis

Background: Peri‐implantitis is one of many reasons for dental implant failure. This study is designed to prevent experimental peri‐implantitis by arginine‐specific gingipain A (rgpA) DNA vaccine. Methods: The bilateral mandibular second and third premolars from 15 male beagle dogs were extracted, and 60 implants were immediately implanted. Three months after implantation, the animals were randomly divided into groups A, B, and C and immunized with plasmid vector–rgpA, heat‐killed Porphyromonas gingivalis, and plasmid vector, respectively. Cotton ligatures infiltrated with P. gingivalis were placed in the submarginal position around the neck of the implants to induce peri‐implantitis. Clinical measurements, including probing depth (PD) and bleeding on probing, were recorded every 2 weeks postoperatively, and P. gingivalis–specific immunoglobulin G (IgG) in serum and secretory IgA (sIgA) in saliva were quantitatively analyzed by enzyme‐linked immunosorbent assay at the same time. Animals were sacrificed after 6 weeks, 50‐μm undecalcified histologic sections were prepared using methylene blue dye, and bone loss around implants was measured. Results: Higher levels of IgG in serum and sIgA in saliva could be measured in groups A and B but not in group C after immunization. There were statistical differences (P <0.05) between, before, and after immunization, but no difference was found between groups A and B (P >0.05). Both peri‐implant PD and bone loss in group A were significantly less than in groups B and C. Conclusions: IgG and sIgA could be generated by immunization with rgpA DNA vaccine, which could significantly slow down bone loss in the experimental peri‐implantitis canine model.     

Unaltered expression of the major protease genes in a non‐virulent recA‐defective mutant of Porphyromonas gingivalis W83

Porphyromonas gingivalis FLL32, a recA mutant, was isolated during construction of a recA defective mutant of P. gingivalis W83 by allelic exchange mutagenesis. In contrast to W83 and FLL33, the typical recA^? mutant previously reported, FLL32 was non‐pigmented, lacked β‐hemolytic activity on blood agar and produced significantly less proteolytic activity. The proteolytic activity in FLL32 was mostly soluble. Expression of the rgpA, rgpB and kgp protease genes was unaltered in FLL32 when compared to FLL33 and the wild‐type strain. FLL32 exhibited reduced virulence in a murine model and partially protected the animals immunized with that strain against a subsequent lethal challenge by the wild‐type strain. These results indicate that the reduced level of proteolytic activity in FLL32 may be due to a defect in the processing of the proteases. Further, immunization with a non‐virulent recA defective mutant of P. gingivalis can partially protect against a lethal wild‐type challenge. The results from this study suggest that the recA locus may be involved in expression and regulation of proteolytic activity in P. gingivalis.     

Porphyromonas gingivalis gingipains cause G1 arrest in osteoblastic/stromal cells

Introduction: The program for mammalian cell growth and division consists of four successive phases; G₁, S, G₂, and M. Porphyromonas gingivalis may manipulate the host cell cycle to benefit bacterial virulence expression, which likely causes the cell and tissue tropism observed in chronic periodontal infections. We examined P. gingivalis for its effects on cell‐cycle modulation in mouse ST2 osteoblastic/stromal cells. Methods: Synchronized ST2 cells were infected with P. gingivalis ATCC33277 (wild‐type, WT), gingipain‐mutants [KDP136 (ΔrgpAΔrgpBΔkgp), KDP129 (ΔrgpAΔrgpB), and KDP133 (Δkgp)], and a fimbria‐deficient mutant (KDP150) for 24 h, then the cell cycle was evaluated using flow cytometry. Cell‐cycle‐related molecule expression was examined with a microarray, as well as with quantitative real‐time polymerase chain reaction and Western blotting assays. Results: Both the WT and KDP150 strains significantly inhibited cellular proliferation and arrested the cell cycle in the G₀/G₁ phase, while the expression levels of the cell‐cycle regulatory molecules cyclin D and cyclin E were also decreased. In contrast, KDP136 did not show any effects. G₁ arrest was also clearly induced by KDP129 and KDP133, with KDP129 being more effective. Conclusion: The present findings suggest that P. gingivalis gingipains reduce cyclin expression and cause early G₁ arrest, leading to the inhibition of cellular proliferation.     

大黄素-8-O-β-D-吡喃葡萄糖苷对牙龈卟啉单胞菌唾液酸酶活性及其毒力基因表达影响研究

目的    探讨大黄素-8-O-β-D-吡喃葡萄糖苷对牙龈卟啉单胞菌（P. gingivalis）唾液酸酶活性及其毒力基因表达的影响。方法    使用不同质量浓度的大黄素-8-O-β-D-吡喃葡萄糖苷（0.2、0.5、2、5、10 mg/mL）处理P. gingivalis W83（实验组）,用未加药物的P. gingivalis W83作对照（对照组）,采用荧光法检测大黄素-8-O-β-D-吡喃葡萄糖苷对P. gingivalis唾液酸酶活性的作用。5 mg/mL大黄素-8-O-β-D-吡喃葡萄糖苷作用于P. gingivalis W83,Real-time PCR法检测毒力基因fimA、fimR、fimS、kgp、rgpA和rgpB的表达情况。结果    大黄素-8-O-β-D-吡喃葡萄糖苷对P. gingivalis唾液酸酶活性产生了抑制作用,当其质量浓度为0.2、0.5、2、5、10 mg/mL时,对唾液酸酶活性的抑制率分别为11.4%、32.23%、40.21%、73.54%、84.31%。与对照组比较,实验组（5 mg/mL大黄素-8-O-β-D-吡喃葡萄糖苷处理）的fimA、fimR、fimS、kgp、rgpA和rgpB基因表达均下降,差异均有统计学意义（均P < 0.05）。结论    大黄素-8-O-β-D-吡喃葡萄糖苷可有效抑制P. gingivalis唾液酸酶活性,其抑制作用会降低细菌毒力基因表达,有望成为预防及治疗牙周炎的新型药物。     

Antibody responses to Porphyromonas gingivalis infection in a murine abscess model--involvement of gingipains and responses to re-infection

BACKGROUND: Porphyromonas gingivalis is one of the most important periodontopathogens. It produces cysteine proteinases named gingipains. We previously examined the effect of gingipains on abscess formation in a murine model. The rgpA rgpB double and kgp mutants induced smaller abscesses than the wild type. Moreover, the rgpA rgpB kgp triple (gingipain-null) mutant hardly showed lesion formation at all under the experimental conditions used, indicating that genes encoding gingipains are important for P. gingivalis virulence. OBJECTIVES: Here, we further report the humoral immune responses induced by P. gingivalis strains. METHODS: After the lesions were apparently cured, sera were collected from the mice and immunoglobulin G (IgG) responses against the whole cell antigens of wild-type P. gingivalis were measured. RESULTS: Wild-type strain was found to induce a strong antibody reaction. On the other hand, the rgpA rgpB kgp triple and kgp mutants induced significantly lower antibody responses compared to the wild type. Western blotting analysis confirmed the differences in antibody production. Next, these mice were re-infected with wild-type strain. Mice that were first infected with wild-type strain showed significantly smaller lesion formation than control mice that were first infected with medium only. On the other hand, mice that were first infected with mutant strains devoid of gingipain activities did not show resistance to re-infection and immunoglobulins directed against gingipains may be protective. CONCLUSIONS: These results suggest that gingipains play an important role in abscess formation in mice, and humoral immune responses seem to be partly responsible for the resistance to re-infection by P. gingivalis.     

Unaltered expression of the major protease genes in a non-virulent recA-defective mutant of Porphyromonas gingivalis W83

Porphyromonas gingivalis FLL32, a recA mutant, was isolated during construction of a recA defective mutant of P. gingivalis W83 by allelic exchange mutagenesis. In contrast to W83 and FLL33, the typical recA- mutant previously reported, FLL32 was non-pigmented, lacked beta-hemolytic activity on blood agar and produced significantly less proteolytic activity. The proteolytic activity in FLL32 was mostly soluble. Expression of the rgpA, rgpB and kgp protease genes was unaltered in FLL32 when compared to FLL33 and the wild-type strain. FLL32 exhibited reduced virulence in a murine model and partially protected the animals immunized with that strain against a subsequent lethal challenge by the wild-type strain. These results indicate that the reduced level of proteolytic activity in FLL32 may be due to a defect in the processing of the proteases. Further, immunization with a non-virulent recA defective mutant of P. gingivalis can partially protect against a lethal wild-type challenge. The results from this study suggest that the recA locus may be involved in expression and regulation of proteolytic activity in P. gingivalis.     

Proliferation of smooth muscle cells stimulated by Porphyromonas gingivalis is inhibited by apple polyphenol

Background: Porphyromonas gingivalis (Pg) is thought to be involved in the progression of occlusive arterial lesions, whereas vascular smooth muscle cell (SMC) proliferation is considered to be involved in occlusive arterial disease. We previously showed that bacteremia caused by Pg infection induced proliferation of mouse aortic SMCs. Furthermore, human SMCs stimulated with human plasma incubated with Pg showed a marked transformation from the contractile to proliferative phenotype. In the present study, we examine the involvement of Pg gingipains and fimbriae in induction of the SMC transformation and proliferation, and effective inhibitors. Methods: Pg strains including gingipain‐ and fimbria‐null mutants were incubated in human plasma, after which the bacteria were removed and the supernatants were added to cultured SMCs. To evaluate the effects of inhibitors, Pg organisms were incubated in plasma in the presence of apple polyphenol (AP), epigallocatechin gallate, KYT‐1 (Arg‐gingipain inhibitor), and KYT‐36 (Lys‐gingipain inhibitor). Results: Plasma supernatants from wild‐type and fimbria‐mutant cultures markedly stimulated cellular proliferation, whereas those containing gingipain‐null mutants showed negligible effects. SMC proliferation was also induced by plasma treated with trypsin. Furthermore, plasma supernatants cultured in the presence of KYT‐1/KYT‐36 and AP showed significant inhibitory effects on SMC proliferation, whereas cultures with epigallocatechin gallate did not. Conclusion: Our results suggest that Pg gingipains are involved in the induction of SMC transformation and proliferation, whereas this was inhibited by AP.     

Microbial Characteristics of Peri‐Implantitis: A Case‐Control Study

Background: The aim of this case‐control study is to compare oral microbiologic characteristics of patients with healthy peri‐implant conditions and patients with peri‐implantitis and to explore the influence of various patient‐ and implant‐related factors on microbiologic characteristics. Methods: Peri‐implant submucosal microbial samples were collected from 85 patients with peri‐implantitis (cases) and from 69 patients with only implants with healthy peri‐implant conditions (controls). Samples were analyzed using culturing techniques. Multivariable logistic regression was used to explore the association of disease status and various patient‐ and implant‐related factors (sex, patient age, smoking, number of remaining teeth, percentage of teeth with bone loss, implant function time, implant surface, and presence of plaque) with microbiologic characteristics. Results: Peri‐implant disease status was significantly associated with the submucosal presence of Porphyromonas gingivalis (Pg), Prevotella intermedia (Pi), Tannerella forsythia (Tf), and Fusobacterium nucleatum (Fn). The association with disease status was most obvious for Pi (odds ratio [OR]: 15.1; 95% confidence interval [CI]: 5.1 to 45.3) and Tf (OR: 13.3; 95% CI: 5.4 to 32.5). The prevalence of Aggregatibacter actinomycetemcomitans and Staphylococcus species was very low. Conclusions: The periodontal pathogens Pg, Pi, Tf, and Fn are associated with peri‐implantitis. A. actinomycetemcomitans and Staphylococcus species do not seem to play an important role in peri‐implantitis.     

大鼠实验性牙周炎中牙龈卟啉单胞菌rgpB和kgp变化的研究

目的:通过建立大鼠实验性牙周炎模型,检测不同时段大鼠上颌第一磨牙龈下人工定植的牙龈卟啉单胞菌、牙龈素rgpB和kgp相对含量的改变,动态观察牙周炎发展不同阶段牙龈卟啉单胞菌致病基因的变化。方法:选择6周的成年雄性大鼠13只,应用钢丝结扎法和细菌种植法建立大鼠实验性牙周炎模型。分别在4周和8周取上颌第一磨牙龈下菌斑,提取细菌DNA,应用PCR方法进行牙龈卟啉单胞菌、牙龈素基因rgpB和kgp特异引物扩增,应用SPSS13.0统计软件,分析大鼠牙周炎不同时段rgpB和kgp的相对变化。结果:大鼠实验性牙周炎模型,龈下菌斑中牙龈卟啉单胞菌的相对含量实验组明显高于对照组,8周组高于4周组;实验组牙龈卟啉单胞菌牙龈素rgpB和kgp明显高于对照组,4周组和8周组rgpB和kgp相对含量无显著性差异。结论:rgpB基因和kgp基因与牙周炎的致病性有关而与牙周炎的严重程度可能无直接相关。     

Mixed infection of Porphyromonas gingivalis and Bacteroides forsythus in a murine abscess model: involvement of gingipains in a synergistic effect

Several microorganisms including Porphyromonas gingivalis and Bacteroides forsythus have been implicated to be etiologically important agents of periodontal disease. In this study, we determined the ability of combinations of periodontopathogenic microorganisms to cause tissue destruction in a murine abscess model. Although all bacterial combinations used in this study produced larger abscesses than did monoinfection of each bacterium, the combination of P. gingivalis and B.forsythus showed a synergistic effect on abscess formation. Since these two bacteria have been frequently found together in lesions of periodontitis, these results suggest the significance of their co-infection in the progression of periodontitis. P. gingivalis produces extracellular and cell-associated cysteine proteinases (gingipains) which appear to be involved in its virulence. The rgpA rgpB double and kgp mutants induced significantly smaller abscesses than the wild type. Moreover, the rgpA rgpB kgp triple (gingipain-null) mutant hardly showed lesion formation at all with the experimental conditions used in this study, indicating that these genes encoding gingipains are important for virulence of P. gingivalis. Mixed infection of these P. gingivalis mutants with B. forsythus showed an additive effect on abscess formation, indicating that the gingipains of P. gingivalis may play an important role in the pathological synergism between P. gingivalis and B. forsythus.     

A Novel Rat Model of Polymicrobial Peri‐Implantitis: A Preliminary Study

Background: Peri‐implantitis is a complex polymicrobial biofilm‐induced inflammatory osteolytic gingival infection that results in orofacial implant failures. To the best knowledge of the authors, there are no preclinical in vivo studies in implant dentistry that have investigated the inflammatory response to known microbial biofilms observed in humans. The aim of this study is to develop a novel peri‐implant rat model using an established model of polymicrobial periodontitis. Methods: Wistar rats were used for the study of experimental peri‐implantitis. One month after extraction of maxillary first molars, a titanium mini‐implant was inserted. Two months after implant healing, implants were uncovered, and abutment fixing was done using cyanoacrylate to prevent abutment loosening. Rats were separated into two groups (group A: polymicrobial‐infected and group B: sham‐infected). One week after healing of abutments, rats were infected with Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia for 12 weeks. Bacterial colonization, bone resorption, and implant inflammation were evaluated by polymerase chain reaction (PCR), microcomputed tomography, and histology, respectively. Results: Three rats with four implants in the infection group and two rats with three implants in the sham‐infection group were analyzed. PCR analysis revealed presence of bacterial genomic DNA, and infection elicited significant immunoglobulin (Ig)G and IgM antibody responses, indicating bacterial colonization/infection around implants. Infection induced an enhanced mean distance from implant platform to the first bone‐to‐implant contact, extensive peri‐implantitis with advanced bone resorption, and extensive inflammation with granulation tissue and polymorphonuclear leukocytes. Conclusions: To the best knowledge of the authors, this is the first study to develop a novel rat model of polymicrobial peri‐implantitis. With modifications to improve implant retention it could offer significant advantages for studies of initiation and progression of peri‐implantitis.     

Sequence variations in rgpA and rgpB of Porphyromonas gingivalis in periodontitis

Objective:  The aim of the present study was to determine sequence variations in the active centre of the Arg-X-specific protease encoding genes rgpA and rgpB of clinical Porphyromonas gingivalis isolates and to analyse their prevalence in periodontitis patients before and 3 months after mechanical periodontal therapy.
Background:  Genetic diversity at nucleotides 281, 283, 286 and 331 has been shown to result in amino acid substitutions in the catalytic domain of RgpA and RgpB that affect the substrate specificity and thus may influence the efficacy of Arg-X-protease specific inhibitors.
Methods:  Sequence analysis of rgpA and rgpB genes in clinical P. gingivalis strains isolated from subgingival plaque samples of 82 periodontitis patients before and 3 months after mechanical supra- and subgingival debridement was performed.
Results:  No specific variation within the rgpA sequence was observed. However, the rgpB sequence in the region of the active centre showed five different rgpB genotypes, which were named NYPN, NSSN, NSSK, NYPK and DYPN according to the derived amino acid substitution. Porphyromonas gingivalis genotype NYPN was detected in 27 patients (32.9%) before and in 8 patients (9.8%) after therapy, NSSN in 26 (31.7%) and 10 (12.2%), NSSK in 22 (26.8%) and 2 (2.4%), NYPK in 5 (6.2%) and 1 (1.2%), and DYPN in 1 patient (1.2%) and 0 patients (0%), respectively. Only one patient (1.2%) harboured two P. gingivalis rgpB genotypes (NSSK/NYPN) before treatment; these were no longer detected after therapy.
Conclusion:  The results indicate that five rgpB genotypes are maintained in natural populations of P. gingivalis. These data may be of importance with regard to the development of specific rgpB inhibitors.     

牙龈卟啉单胞菌精氨酸特异性牙龈素基因疫苗预防比格犬种植体周围炎的实验研究

目的 构建牙龈卟啉单胞菌精氨酸特异性牙龈素基因疫苗pVAX1-rgpA,并对成年犬进行免疫接种,观察该疫苗在防治种植体周围炎发生发展中的作用。方法 构建真核表达质粒pVAX1-rgpA。拔除15只成年犬下颌双侧第二、三前磨牙,随机即刻植入种植体。3个月后,实验犬随机均分成A、B、C组,分别接种重组质粒pVAX1-rgpA、热失活牙龈卟啉单胞菌、空白质粒pVAX1,连续接种3次。接种开始前、接种结束2周后采用酶联免疫吸附试验检测血清IgG和唾液分泌型IgA（sIgA）含量。随机选取一侧采用丝线结扎法构建种植体周围炎,并检测种植体周探诊深度（PD）和探诊出血指数（BOP）。结扎6周后处死所有动物,制作50 μm厚的硬组织切片,亚甲基蓝染色后观察种植体周骨丧失程度。结果 A、B组动物免疫后,IgG、sIgA抗体较未免疫前明显增高（P<0.05）,同时较C组升高（P<0.05）,但A、B组间差异无统计学意义（P>0.05）。丝线结扎第4和6周时,C组结扎侧的PD明显高于A、B组（P<0.05）,A、B组间差异无明显统计学意义（P>0.05）。A组结扎侧骨丧失量明显小于其他两组（P<0.05）。硬组织切片可见,各组种植体周骨丧失区均有大量的炎症细胞和细菌存在,C组结扎侧最严重。结论 精氨酸特异性牙龈素（rgpA）基因疫苗产生的IgG和sIgA,能有效减弱犬种植体周围炎的骨丧失量。     

Periodontal and Systemic Responses in Various Mice Models of Experimental Periodontitis: Respective Roles of Inflammation Duration and Porphyromonas gingivalis Infection

Background: The great variability of periodontal and systemic responses to experimental periodontitis reflects the inherent pathogenic complexity of mice models and could limit the resulting interpretations and their extension to human diseases. This study compared the effect of Porphyromonas gingivalis (Pg) infection and experimental periodontitis duration at local and systemic levels in various models. Methods: Periodontitis was induced in C57BL/6J mice by ligatures previously incubated with Pg (LIGPG group) or not (LIG group) or by oral gavage (GAV) with Pg ATCC 33277. Blood samples were taken, and mice were euthanized at different times. Periodontal tissue destruction, osteoclast number, and inflammation were assessed by histomorphometry, tartrate‐resistant acid phosphatase histoenzymology, and cathepsin B (CATB) and matrix metalloproteinase 9 (MMP9) immunochemistry. Serum levels of interleukin‐6 (IL‐6) and IL‐1β were measured using enzyme‐linked immunosorbent assay bioplex methods. Results: Periodontal tissue destruction and osteoclast numbers were significantly elevated in LIGPG models compared to LIG and GAV models. They increased with time with the exception of osteoclast numbers in the LIG model. CATB and MMP9 expression was related to bone destruction processes and Pg infection. The highest serum levels of IL‐6 and IL‐1β were observed in the LIGPG group. A decrease of IL‐6 and an increase of IL‐1β serum level were observed with time in LIGPG group contrary to LIG group. Conclusions: These data indicate that Pg infection worsened periodontal tissue destruction through specific pathogenic pathways and modified systemic response to periodontal inflammation. Furthermore, the blood cytokine response to ligature models showed their relevance for evaluating the systemic impact of periodontal disease.     

Oral bacterial DNA differ in their ability to induce inflammatory responses in human monocytic cell lines

Background: Deoxyribonucleic acids (DNA) of periodontal pathogens, Porphyromonas gingivalis (Pg) and Tannerella forsythia, stimulate cytokine production in human monocytic cells (THP‐1) through Toll‐like receptor 9 (TLR‐9) and nuclear factor‐κB signaling. Fusobacterium nucleatum (Fn) is one of the most frequently isolated bacteria in periodontally diseased tissues and is reported to synergize with Pg, enhancing the pathogenicity. We investigate inflammatory mediator production in THP‐1 cells challenged with Fn and Streptococcus sanguinis (Ss) DNA, a non‐pathogenic oral bacteria, and further assess whether cytokines triggered by whole pathogens or Pg lipopolysaccharide (LPS) are affected by TLR‐9 signaling inhibitors (chloroquine). Methods: THP‐1 cells were stimulated with Pg‐DNA (100 ng/μL), Fn‐DNA (100 ng/μL), Ss‐DNA (100 ng/μL), Pg‐LPS (10 ng/μL), and heat‐killed whole bacteria (multiplicity of infection, 1:100) for 16 hours with or without chloroquine pretreatment (10 μg/mL). Interleukin (IL)‐1β, IL‐6, IL‐8, and tumor necrosis factor‐α levels were determined using enzyme‐linked immunosorbent assay. Statistical analyses included analysis of variance with multiple comparisons using Dunnett or Tukey methods and paired t test. A value of P <0.05 was significant. Results: Inflammatory mediator levels were increased in response to all the stimuli with the exception of Ss‐DNA (P <0.05). Chloroquine pretreatment significantly decreased cytokine production from THP‐1 cells with the exception of IL‐6 production triggered by whole Fn and Ss (P <0.05). Conclusions: Differences exist among oral bacterial DNA in inducing immune responses. By altering the conditions in cytosolic compartments, we can interfere with cellular responses triggered by extracellular receptor activation. Thus, alternative treatment approaches targeted to intracellular receptors might be of benefit in controlling periodontal inflammation.     

The roles of RgpB and Kgp in late onset gingipain activity in the vimA‐defective mutant of Porphyromonas gingivalis W83

Previous studies have shown that VimA, an acetyltransferase, can modulate gingipain biogenesis in Porphyromonas gingivalis. Inactivation of the vimA gene resulted in isogenic mutants that showed a late onset of gingipain activity that only occurred during the stationary growth phase. To further elucidate the role and contribution of the gingipains in this VimA‐dependent process, isogenic mutants defective in the gingipain genes in the vimA‐deficient genetic background were evaluated. In contrast with the wild‐type strain, RgpB and Kgp gingipain activities were absent in exponential phase in the ?rgpA::tetQ‐vimA::ermF mutant. However, these activities increased to 31 and 53%, respectively, of that of the wild‐type during stationary phase. In the ?rgpA::cat‐?kgp::tetQ‐vimA::ermF mutant, the RgpB protein was observed in the extracellular fraction but no activity was present even at the stationary growth phase. There was no gingipain activity observed in the ?rgpB::cat‐?kgp::tetQ‐vimA::ermF mutant whereas Kgp activity in ?rgpA::cat‐?rgpB::tetQ‐vimA::ermF mutant was 24% of the wild‐type at late stationary phase. In contrast to RgpA, the glycosylation profile of the RgpB catalytic domain from both W83 and P. gingivalis FLL92 (vimA::ermF) showed similarity. Taken together, the results suggest multiple gingipain activation pathways in P. gingivalis. Whereas the maturation pathways for RgpA and RgpB are different, the late‐onset gingipain activity in the vimA‐defective mutant was due to activation/maturation of RgpB and Kgp. Moreover, unlike RgpA, which is VimA‐dependent, the maturation/activation pathways for RgpB and Kgp are interdependent in the absence VimA.     

The evaluation of peri‐implant sulcus fluid osteocalcin,osteopontin, and osteonectin levels in peri‐implant diseases

1 Background

Peri‐implant mucositis is an inflammation of the soft tissues surrounding an implant. Peri‐implantitis refers to a process characterized by peri‐implant bone loss along with an inflammation of the soft tissues. Osteocalcin, osteopontin, and osteonectin proteins are related to bone remodeling. The aim of the present study was to investigate peri‐implant sulcus fluid (PISF) osteocalcin, osteopontin, and osteonectin levels in peri‐implant mucositis and peri‐implantitis.

2 Methods

Fifty‐two implants with peri‐implantitis, 46 implants with peri‐implant mucositis, and 47 control implants were included in the study. Clinical parameters including probing depth, modified sulcus bleeding index and modified plaque index were recorded. PISF osteocalcin, osteopontin, and osteonectin levels were analyzed by ELISA kits.

3 Results

There were no significant differences in PISF osteocalcin, osteopontin, and osteonectin total amounts between healthy controls, peri‐implant mucositis and peri‐implantitis groups (P > 0.05). Probing depths were not correlated with PISF osteocalcin, osteopontin, and osteonectin levels in the study groups (P > 0.05).

4 Conclusions

Soft tissue inflammation around dental implants does not cause a change in osteocalcin, osteopontin, and osteonectin levels in PISF. Also, peri‐implantitis does not seem to give rise to an increase in PISF levels of osteocalcin, osteopontin, and osteonectin.     

Effect of Porphyromonas gingivalis Lipopolysaccharide on Bone Marrow Mesenchymal Stem Cell Osteogenesis on a Titanium Nanosurface

Background: Titanium (Ti) dental implants have been widely used for prosthetic reconstruction of dentition. Unfortunately, peri‐implantitis can result in failure of dental implant osseointegration. Lipopolysaccharide (LPS) acts as a chronic inflammatory stimulus and maintains peri‐implant inflammation, worsening the prognosis for implant osseointegration. The purpose of this study is to determine the effects of 10 M NaOH‐modified Ti surface with nanonetwork structure on the proliferation and osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMMSCs) in the context of Porphyromonas gingivalis LPS exposure. Methods: Titanium disks treated with 10 M NaOH solution and control were incubated with BMMSCs and exposed to P. gingivalis LPS (0, 0.1, or 1 μg/mL). The effects of the modified nanonetwork structure on osteogenic differentiation of rat BMMSCs were evaluated in the context of different concentrations of P. gingivalis LPS exposure. Results: Rat BMMSCs on the 10 M NaOH‐modified Ti surface with nanonetwork structure had higher levels of osteogenesis‐related gene expression and significantly greater cell proliferation, alkaline phosphatase activity, and extracellular matrix deposition and mineralization than cells on the untreated Ti surfaces, in all the groups with different doses of P. gingivalis LPS exposure. Conclusion: The 10 M NaOH‐modified Ti surface with nanonetwork structure has better endotoxin tolerance under P. gingivalis LPS exposure than the non‐modified surface.     

Erbium,Chromium:Yttrium‐Scandium‐Gallium‐Garnet Laser Effectively Ablates Single‐Species Biofilms on Titanium Disks Without Detectable Surface Damage

Background: Increasing evidence implicates biofilms, consisting of species such as Porphyromonas gingivalis (Pg), in the etiology of peri‐implantitis. Multiple approaches to ablate biofilms on failing implants have been proposed and include use of lasers, most recently the erbium, chromium:yttrium‐scandium‐gallium‐garnet (Er,Cr:YSGG) laser. The purpose of this study is to establish an in vitro single‐species biofilm model on implant surfaces and determine power settings of the Er,Cr:YSGG laser that remove biofilm without causing physical damage to disks. Methods: Single‐species biofilms consisting of Pg strain 381 were grown on titanium disks, including: 1) sandblasted, large‐grit, acid‐etched (SLA); 2) calcium phosphate nano‐coated (CaP); 3) anodized; or 4) machined surfaces. Power settings from 0 to 1.5 W using an Er,Cr:YSGG laser equipped with radial firing tip were used. Biofilm formation/removal was quantitated using confocal and scanning electron microscopy. Surface changes in temperature, microroughness, and water contact angle were analyzed. Results: Results show confluent Pg biofilm coating all disk surfaces. The laser removed biofilms from all surfaces, with CaP and SLA surfaces requiring power setting of 1.0 to 1.5 W for ablation of bacteria coating the disks. Within this power range, and with water spray, there were no changes in surface temperature, surface roughness, or contact angle on any surfaces tested. Conclusion: The Er,Cr:YSGG laser with radial firing tip and water spray was able to effectively ablate ≥95% of biofilm on all types of tested titanium surfaces, using clinically relevant power settings, without causing measurable physical changes to surfaces.