Intermittent parathyroid hormone enhances the healing of medication-related osteonecrosis of the jaw lesions in rice rats.

Medication-related osteonecrosis of the jaw (MRONJ) is a potentially severe adverse event in patients treated with antiresorptives. Management of MRONJ is challenging, and no non-antibiotic, established medical treatment exists. Intermittent parathyroid hormone (iPTH) has been used off-label to treat MRONJ with favorable results. However, its medical efficacy has rarely been substantiated in clinical or preclinical experiments. Using a validated rice rat, infection-based model of MRONJ, we evaluated the effects of iPTH on established MRONJ. We hypothesize that iPTH contributes to MRONJ resolution by enhancing alveolar bone turnover and healing oral soft tissues. Eighty-four rice rats began a standard rodent chow diet at age 4 weeks to induce localized periodontitis. Rats were simultaneously randomized to receive saline (vehicle, VEH) or zoledronic acid (ZOL, 80 µg/kg IV) every 4 weeks. Oral exams were conducted bi-weekly to assign a gross quadrant grade (GQG, 0-4) to evaluate any lesion at the lingual aspect of the interdental space between maxillary molar (M2) and M3. 14 of 20 VEH-treated rice rats (70%) developed maxillary localized periodontitis with GQG 2-3 after 30 ± 10 weeks of saline. Additionally, 40 of 64 ZOL-treated rice rats with periodontitis developed MRONJ-like lesions after 30 ± 10 weeks of ZOL treatment. Rice rats with localized periodontitis or MRONJ-like lesions were treated with saline or iPTH (40 µg/kg) subcutaneously (SC) 3 times/week For 6 weeks until euthanasia. We found that iPTH -treated ZOL rats had a lower prevalence of MRONJ (p < 0.001), with lower severity extent of oral lesions (p = 0.003) and percentage of empty osteocyte lacunae (p < 0.001). ZOL rats treated with iPTH displayed a higher osteoblast surface (p < 0.001), more osteoblasts (p < 0.001), higher osteoclast surface (p < 0.001) and more osteoclasts (p = 0.002) at alveolar bone surfaces than ZOL/VEH rats. Greater gingival epithelial thickness and epithelial cell proliferation rate was found in the oral mucosa and gingiva of ZOL/PTH rats than in ZOL/VEH rats (p < 0.001). Our data suggest that iPTH is an efficacious non-operative medicinal therapy that accelerates oral healing and enhances the resolution of MRONJ lesions in ZOL-treated rice rats.

Gingival Fibroma: An Emerging Distinct Gingival Lesion with Well-Defined Histopathology.

Gingival growths, barring a few are mostly reactive and seldom exhibit significant true neoplastic potential. The common etiology is local irritation from dental plaque/calculus, trauma as well as medication-related overgrowth. Such lesions are easily distinguishable and categorized into diagnoses such as pyogenic granuloma, peripheral ossifying fibroma, etc. We present a previously undescribed, but commonly encountered, reactive gingival growth with unique histologic features and suggest the diagnostic term "gingival fibroma." An IRB approved retrospective review of the University of Florida Oral Pathology Biopsy Service encompassing years 2010-2019, was performed to select cases. Demographics, clinical data, and microscopic diagnoses were recorded and analyzed. Four board-certified oral and maxillofacial pathologists agreed upon and established the diagnostic criteria. These are: a prominent fibromyxoid stroma, variable cellularity, a whorled or storiform pattern of arrangement of the cellular elements, lack of significant inflammation or vascularity, and complete absence of calcification, and/or odontogenic islands. A total of 60 cases met all criteria and were included in the study. Age range in years was 14-87 with the mean at 45.11 years. A striking female predilection (90%) was noted. Approximately 62% of cases were reported on the maxillary gingiva, followed by 38.3% in the mandibular gingiva. Majority, 66.7% were in the anterior incisor region followed by 11.7% in the canine/first premolar areas. All lesions were submitted as excisional biopsy, and 4 cases recurred within 2-3 years of excision. In all cases, lesional tissue appeared to extend to the surgical base of the specimen. We present 60 cases of a histologically unique entity occurring exclusively on the gingiva and introduce the diagnostic term "Gingival Fibroma" for these lesions. Further studies with adequate clinical follow-up may help understand the exact clinical behavior of these lesions.

Preventing or controlling periodontitis reduces the occurrence of osteonecrosis of the jaw (ONJ) in rice rats (Oryzomys palustris).

INTRODUCTION: Osteonecrosis of the jaw (ONJ) is an adverse event that requires association of both systemic risk factors, such as powerful anti-resorptives (pARs; e.g. zoledronic acid [ZOL]), and local oral risk factors (e.g. tooth extraction, periodontitis). Whereas optimal oral health prior to initiate pARs is recognized as critically important for minimizing ONJ risk, the efficacy of preventive/maintenance measures in patients who are taking pARs is understudied. Rice rats fed a standard diet (STD), rich in insoluble fiber, develop localized periodontitis. STD-rats with localized periodontitis treated with ZOL for 18-24 wk develop ONJ. Hence, we hypothesized that controlling/preventing localized periodontitis in the ZOL-treated rats, reduces ONJ occurrence. METHODS: We used two approaches to attempt reducing periodontitis prevalence: 1) periodontal cleaning (PC); and 2) replacing the STD-diet with a nutritionally-equivalent diet high in soluble fiber (SF). 75 four-week-old male rats were weight-randomized into five groups (n = 15) in a 24-week experiment. Three groups ate the STD-diet and two the high SF-diet. STD-diet groups received intravenous (IV) vehicle (VEH) q4wks (STD + VEH), 80 µg/kg ZOL q4wks IV (STD + ZOL), or ZOL plus PC q2wks (STD + ZOL + PC). The SF-diet groups received VEH (SF + VEH) or ZOL (SF + ZOL). Jaws were processed for histopathology and evaluated for ONJ prevalence and tissue-level periodontitis. RESULTS: 1) 40% of STD + VEH rats developed maxillary localized periodontitis with no ONJ; 2) 50% of STD + ZOL rats developed ONJ; 3) 7% of STD + ZOL + PC rats developed ONJ (p < 0.01 vs. STD + ZOL); and 4) one SF + ZOL rat developed localized periodontitis, and no SF + VEH or SF + ZOL rats developed ONJ (p < 0.001 vs. STD + ZOL). CONCLUSIONS: 1) Periodontal cleaning in ZOL-treated rats decreases localized periodontitis severity and reduces ONJ prevalence; and 2) feeding a SF-diet to ZOL-treated rats reduces both incidence of localized periodontitis and ONJ. Our data indicates strong oral microbial community shifts according to oral health condition and trends in the shifts associated with diet.

Odontogenic Myxoma: A 23-Year Retrospective Series of 38 Cases.

Odontogenic myxoma (OM) is an uncommon benign odontogenic tumor arising in the jaw. Though it has slight histologic overlap with other entities, definitive diagnosis is imperative considering the tumor's aggressive nature, high recurrence rate, and necessity of radical surgical intervention in large-sized lesions. With IRB approval, a retrospective search of the University of Florida College of Dentistry Oral Pathology Biopsy Service archives from 1994 to 2017 for diagnosis of OM of the jaw was performed. Biopsy reports and original slides for each case were assessed and reviewed along with any accompanying radiographs to confirm the diagnosis. Immunohistochemical staining was utilized to exclude entities with histologic overlaps, such as intraosseous myxoid neurofibroma. A total of 38 cases were included. The patients' age ranged from 6 to 84 years, with a mean age of 37.47 years. Females comprised two-thirds of the cases (n = 25) versus males (n = 13). The mandible was the most affected at 60.5% (n = 23), followed by maxilla 39.4% (n = 15). Posterior jaw involvement was higher than anterior in both the mandible (n = 20 versus n = 3) and the maxilla (n = 11 versus n = 4). Most lesions presented clinically as expansile masses with variable radiographic appearance. The clinical impression from submitting providers included "gelatinous masses", abscesses, odontogenic lesions, fibro-osseous lesions, soft tissue or bone neoplasms, and reactive gingival lesions. A consensus of odontogenic myxoma as a diagnosis was rendered for 30 cases (79%), while in 8 cases (21%) that exhibited a more fibrous stroma was identified as fibromyxoma. OM may exhibit a varied demographic and clinical profile with a wide spectrum of histologic presentations. Pathologists should be sentient of this variability in order to arrive at an accurate diagnosis and correctly manage these patients.

An Analysis of Clinical and Histopathologic Features of Fibrous Dysplasia of the Jaws: A Series of 40 Cases and Review of Literature.

Fibrous dysplasia (FD) is a rare condition commonly involving the jaws. While FD has a typical clinical and histological presentation, considerable variation exists. Moreover, overlap of features with other disorders is possible. This study serves to characterize the features of a large case series of FD of the jaws. With IRB approval, the University of Florida Oral Pathology Biopsy Service archive was retrospectively searched from 1994 to 2015 for cases of FD. Epidemiological data, location, duration, clinical and radiographic appearance, clinical impression and exact microscopic diagnosis were recorded. The average age was 37.3 years (range 7-87 years) with majority of cases in females (67.5%). The most common ethnicity was Caucasian. Maxillary location was predominant (59%), followed by mandible (38%) and multiple locations (3%). Expansion was reported in 78% of cases. Radiographically, most cases exhibited ground glass opacity, however some presented with a mottled or mixed radiopaque/radiolucent appearance. Histologically, a wide variation in terms of stromal cellularity, presence of osteoblastic rimming, and presence of calcified material mimicking cemento-osseous dysplasia was observed. Clinicians and pathologists should be cognizant of the significant variability in clinical, histopathologic, and radiographic presentation of FD, which may pose a diagnostic challenge.

Anti-vascular endothelial growth factor antibody monotherapy causes destructive advanced periodontitis in rice rats (Oryzomys palustris).

OBJECTIVE: Angiogenesis inhibitors (AgI) are commonly used in combination chemotherapy protocols to treat cancer, and have been linked to osteonecrosis of the jaw (ONJ). However, it is unknown if AgI therapy alone is sufficient to induce ONJ. We have previously established an ONJ model in rice rats with localized periodontitis that receive zoledronic acid (ZOL). The purpose of this study was to use this model to determine the role of anti-vascular endothelial growth factor A (anti-VEGF) antibody treatment of rice rats with localized maxillary periodontitis. We hypothesized that rice rats with localized maxillary periodontitis given anti-VEGF monotherapy will develop oral lesions that resemble ONJ, defined by exposed, necrotic alveolar bone. METHODS: At age 4 weeks, 45 male rice rats were randomized into three groups (n = 15): 1) VEH (saline), 2) ZOL (80 µg/kg body weight, intravenously once monthly), and 3) anti-VEGF (5 mg B20-4.1.1/kg body weight, subcutaneously twice weekly). After 24 weeks, rats were euthanized, jaws were excised and a high-resolution photograph of each quadrant was taken to assign a severity grade based on gross appearance. Jaws were then fixed, scanned by MicroCT, decalcified and sectioned for histopathologic and immunohistochemical analyses. RESULTS: 40-80% of the rats in the three groups developed gross oral lesions. 50% of ZOL rats developed ONJ. In contrast, 80% of the anti-VEGF rats developed destructive advanced periodontitis that was characterized by extreme alveolar bone loss and fibrosis. Anti-VEGF rats never developed exposed, necrotic bone. Furthermore, only anti-VEGF rats developed mild to severe mandibular periodontitis. Compared to VEH rats, more T-cells were found in periodontal lesions of anti-VEGF rats and more cells of the monocyte lineage were found in ONJ lesions of ZOL rats. CONCLUSIONS: Anti-VEGF monotherapy administered to a validated rodent model of ONJ caused a destructive advanced form of periodontitis that differed significantly from ONJ.

Lichenoid Characteristics in Premalignant Verrucous Lesions and Verrucous Carcinoma of the Oral Cavity.

Verrucous hyperkeratosis (VH), verrucous carcinoma (VC) and the relentless, truly pre-malignant variant proliferative verrucous leukoplakia often exhibit lichenoid histologic features that may create a diagnostic dilemma for pathologists. This study aims to evaluate and categorize the frequency and the histopathologic patterns of lichenoid features seen in these lesions. Following IRB approval, cases of VH and VC from 1994 to 2014 were retrieved from the archives of UF Oral Pathology Biopsy Service. A panel of 4 board-certified oral and maxillofacial pathologists reviewed and scored the presence or absence of 5 lichenoid features: band-like infiltrate (BLI), saw tooth rete ridges (STRR), interface stomatitis (IS), civatte bodies (CB), and basement membrane degeneration (BMD). Cases not fulfilling the stringent selection criteria were excluded. A total of 70 cases of VH and 56 cases of VC were included. Approximately 25% of both VH and VC cases exhibited 3 or more lichenoid features. By Chi square testing, BLI (p = 0.000), IS (p = 0.005), and CB (p = 0.026) were significantly more common in VC than VH. Gingival lesions had significantly less frequent BLI (p = 0.004) and IS (p = 0.024) versus other sites. However, STRR was significantly more common in VH than VC (p = 0.000) in the gingiva. (p = 0.002). Statistical analysis revealed that the only significant valid association was the increased presence of band-like infiltrate in VC over VH (p = 0.001). Lichenoid features are common in both VH and VC and may represent a nonspecific inflammatory response to the dysplasia or malignancy rather than concomitant lichenoid disease. This could lead to significant under diagnoses of these premalignant or potentially malignant lesions by pathologists.

Clear cell changes in salivary gland neoplasms: A 20-year retrospective study.

BACKGROUND: Clear cells are observed histopathologically in both benign and malignant neoplasms but their presence in salivary gland tumors has not been extensively documented. MATERIAL AND METHODS: With IRB approval, the archive of the University of Florida College of Dentistry oral pathology biopsy service was retrospectively searched from 1994-2014 for all benign and malignant salivary tumors. Epidemiological data, tumor location and duration, and type of tumor were recorded. A four reviewer panel examined the original slides. Reviewers scaled each case as 0 (no clear cells present), 1 (few to focal clear cells), 2 (less than 50% clear cells), and 3 (greater than 50% clear cells). RESULTS: A total of 535 cases were included of which 48% of tumors displayed 0 clear cells (257/535), 31.4% (168/535) scored 1, 13.6% (73/535) scored 2, and 7% (37/535) scored 3. Of the 251 (47%) malignant neoplasms, 64% (160/251) demonstrated 0-1 clear cell change, while 36% (91/251) showed a score of 2-3. For the total 284 (53%) benign tumors, 93% (265/535) scored 0-1 and 7% (19/535) scored a 2-3 range. No statistical difference was noted for gender, age, or duration of time present in regards to presence or absence of clear cells. Statistically significant differences in clear cell presence were found between location groups, between benign and malignant diagnosis, and between specific diagnostic groups. CONCLUSIONS: This study demonstrates the frequent presence of increased numbers of clear cells in oral salivary malignancies and highlights salivary gland differential diagnoses when presented with clear cell changes.

Direct immunofluorescence testing results in cases of premalignant and malignant oral lesions.

OBJECTIVES: Oral premalignant and malignant lesions may mimic oral lichen planus (OLP) clinically and microscopically. OLP often shows basement membrane fibrinogen positivity on direct immunofluorescence testing (DIF). This study examined fibrinogen positivity in oral premalignant lesions and squamous cell carcinoma. STUDY DESIGN: The University of Florida Oral Pathology Biopsy Service records were searched for the years 2003 to 2013 for oral premalignant lesions and squamous cell carcinoma with concurrent DIF testing. Demographic, clinical, and DIF or histologic information was collected and analyzed. RESULTS: Sixty-eight fibrinogen positive lesions were identified within a total of 164 cases. Low-grade dysplasia and premalignant verrucous lesions made up the majority of the fibrinogen positive lesions (combined n = 43; 63.2%), and the most common locations in positive cases were the buccal mucosa, tongue, and gingiva. A lichenoid distribution of the inflammatory infiltrate significantly predicted fibrinogen positivity (P < .0005). CONCLUSIONS: Fibrinogen positivity may be seen in premalignant and malignant oral lesions increasing the risk of misdiagnosis.

Desmoplastic fibroma of the mandible: a series of three cases and review of literature.

The desmoplastic fibroma (DF) is a rare, fibroblastic lesion of bone that histologically resembles the desmoid tumor of soft tissue. Although classified as benign, it frequently demonstrates aggressive behavior, often causing tooth mobility, extensive bone destruction, and has a moderate to high recurrence rate. We present three cases of DF in the mandible: the first in a 13 year old female involving the mandibular body in the region of teeth #s 27-#28, the second in a 57 year old female with a lesion apical to tooth #30, and the third in a 20-year-old female involving the left posterior mandible. Clinical, histologic, immunohistochemical (IHC) and radiographic features of this rare neoplasm are discussed. The challenges encountered in establishing an accurate diagnosis due to significant microscopic overlap with other spindle cell lesions are also detailed. Additionally, the findings of IHC stains including vimentin, smooth muscle actin, S-100 protein, ß-catenin, HHF-35 and proliferation marker, Ki-67 on 3 cases are reported. The potential for misdiagnosis is high, especially in early lesions, since immunohistochemistry has been reported in literature to be inconsistent when differentiating DFs from other spindle cell lesions. A comparative review of DF and similar entities in the jaws with current considerations in treatment and prognosis is presented.

Intraoral basal cell carcinoma, a rare neoplasm: report of three new cases with literature review.

Intraoral basal cell carcinoma (IOBCC) is an extremely rare entity that bears close microscopic resemblance to and is often confused with the peripheral ameloblastoma (PA). Basal cell carcinomas are thought to arise from pluripotential basal cells present within surface epithelium and adnexal structures, so theoretically they can arise within the oral cavity. Many of the early cases reported as IOBCC actually represent PA. Most of the well documented cases arise from the gingiva. The histologic features of basal cell carcinoma that help separate it from a PA include: tumor arising from surface epithelium, scattered mitotic figures and apoptotic cells, presence of mucoid ground substance and tumor infiltrating widely throughout the connective tissue and often exhibiting a prominent retraction artifact. Clinically IOBCC resemble carcinomas, compared to the benign and innocuous appearance of the PA and typically presents as surface ulcerations varying from rodent ulcer to an ulcerated erythroplakia appearance. This contrasts with the classic "bump on the gum" appearance of PAs with usually intact surface and appearing as small discrete, sessile, exophytic lesions. Importantly, the proliferative basaloid epithelium demonstrates positive immunoreactivity for the anti-epithelial antibody, Ber-EP4, a cell surface glycoprotein. The IOBCC has the potential for local recurrence and aggressive behavior and should be treated with wide surgical excision and close clinical follow up. We present 3 rare cases of IOBCC and discuss the salient histologic, immunohistochemical and clinical features.

Bis-enoxacin inhibits bone resorption and orthodontic tooth movement.

UNLABELLED: Enoxacin inhibits binding between the B-subunit of vacuolar H(+)-ATPase (V-ATPase) and microfilaments, and also between osteoclast formation and bone resorption in vitro. We hypothesized that a bisphosphonate derivative of enoxacin, bis-enoxacin (BE), which was previously studied as a bone-directed antibiotic, might have similar activities. BE shared a number of characteristics with enoxacin: It blocked binding between the recombinant B-subunit and microfilaments and inhibited osteoclastogenesis in cell culture with IC50s of about 10 µM in each case. BE did not alter the relative expression levels of various osteoclast-specific proteins. Even though tartrate-resistant acid phosphatase 5b was expressed, proteolytic activation of the latent pro-enzyme was inhibited. However, unlike enoxacin, BE stimulated caspase-3 activity. BE bound to bone slices and inhibited bone resorption by osteoclasts on BE-coated bone slices in cell culture. BE reduced the amount of orthodontic tooth movement achieved in rats after 28 days. Analysis of these data suggests that BE is a novel anti-resorptive molecule that is active both in vitro and in vivo and may have clinical uses. ABBREVIATIONS: BE, bis-enoxacin; V-ATPase, vacuolar H(+)-ATPase; TRAP, tartrate-resistant acid phosphatase; αMEM D10, minimal essential media, alpha modification with 10% fetal bovine serum; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; RANKL, receptor activator of nuclear factor kappa B-ligand; NFATc1, nuclear factor of activated T-cells; ADAM, a disintegrin and metalloprotease domain; OTM, orthodontic tooth movement.

Characterization of bilateral trigeminal constriction injury using an operant facial pain assay.

In order to better understand and treat neuropathic pain, scientific study must use methods that can assess pain processing at the cortical level where pain is truly perceived. Operant behavior paradigms can accomplish this. We used an operant task to evaluate changes following chronic constriction injury to the trigeminal nerves. We also relate these behavioral changes to immunohistochemistry of transient receptor potential channels vanilloid 1 and melastatin 8 (TRPV1 and TRPM8) in the trigeminal ganglia. Following nerve injury, successful performance of the operant task was reduced and aversive behaviors were observed with 10 and 37 °C stimulation, indicating cold allodynia and mechanical allodynia respectively. In contrast, while aversive behaviors were observed with 48 °C stimulation, successful performance of the operant task was not substantially hindered following injury. These behavioral changes were accompanied by an increase in TRPV1 positive cells and an increased intensity of TRPM8 staining at 2 weeks post-injury, when cold allodynia is maximal. These findings suggest that the incorporation of operant behavioral assessment in the study of pain may provide insight into the relationship among peripheral changes, motivational drive, and pain. Understanding this relationship will allow us to better treat and prevent chronic neuropathic pain.

Bizarre parosteal osteochondromatous proliferation (Nora's Lesion) of the mandible. a rare bony lesion.

Bizarre parosteal osteochondromatous proliferation (BPOP) also eponymically called "Nora's lesion", is a rare benign reactive bone lesion first reported in 1983. BPOP occurs classically on the bones of the hands and feet and long bones. This lesion can easily be confused, both clinically and microscopically, with other benign and malignant lesions of bone, including osteochondroma, parosteal osteosarcoma, myositis ossificans and reactive periostitis. BPOP has been reported to have a high rate of recurrence. Only 3 cases of BPOP of the head and neck have been reported in the literature, of which one involved the maxilla. We present a rare case of BPOP involving the mandible in a 10 year old African American male. Microscopically, a fibro-cartilaginous cap giving rise to a proliferation of variably mineralized osteophytic finger-like projections of bone was seen. Multiple trabeculae of "blue bone" were noted as well as numerous atypical appearing chondrocytes. The lesion recurred within 4 months following the initial excision but has not recurred to date after the second local excision. To the best of our knowledge, this is the first report of BPOP arising in the mandible. In addition, we discuss the clinical and microscopic features, differential diagnosis, and prognosis of this rare entity. We present a case of BPOP of the mandible and believe this is the first report of such a case in the mandible.

Association of oxidative stress biomarkers and antioxidant enzymatic activity in male infertility of north-East India.

INTRODUCTION: Oxidative stress is a common pathology seen in approximately half of all infertile men. In a normal situation, the seminal plasma contains antioxidant mechanisms which are likely to quench these reactive oxygen species. However, during infertility complications these antioxidant mechanisms may downplay and create a situation which is called oxidative stress. OBJECTIVE: The aim of the present study was to assess the levels of lipid peroxide (LPO), protein peroxide (PPO) and activities of antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase (GPX) in blood and semen samples of an infertile male population from North-East India. METHOD: We measured LPO, PPO, SOD and GPX in a total of 50 infertile individuals. For the study 20 fertile donors served as the control group. RESULT: Patients with male factor infertility had significantly higher LPO and PPO levels (60.84 ± 3.55 and 72.84 ± 3.66; P < 0.001) compared with controls (40.20 ± 4.33 and 59.93 ± 5.24) in blood. In semen also, the same trend was found with significantly higher LPO and PPO levels (200.27 ± 6.25 and 149.80 ± 11.47; P < 0.001) compared with controls (116.51 ± 5.49 and 59.10 ± 4.62). The SOD and GPX enzymes in blood (3.40 ± 1.06 and 0.16 ± 0.01; P < 0.001) and in semen (2.42 ± 1.32 and 0.24 ± 0.015; P < 0.001) showed a significantly lower activity when compared with their respective controls (4.85 ± 0.78; 0.36 ± 0.05 and 4.24 ± 0.89; 0.65 ± 0.03). The SOD and GPX activity when compared with the LPO and PPO values, showed a positive correlation. CONCLUSION: We conclude that oxidative stress is associated with male factor infertility. This assessment may help in the treatment of this male infertility by suitable antioxidants.

Polymicrobial periodontal pathogen transcriptomes in calvarial bone and soft tissue.

Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia are consistently associated with adult periodontitis. This study sought to document the host transcriptome to a P. gingivalis, T. denticola, and T.forsythia challenge as a polymicrobial infection using a murine calvarial model of acute inflammation and bone resorption. Mice were infected with P. gingivalis, T. denticola, and T. forsythia over the calvaria, after which the soft tissues and calvarial bones were excised. A Murine GeneChip(®) array analysis of transcript profiles showed that 6997 genes were differentially expressed in calvarial bones (P < 0.05) and 1544 genes were differentially transcribed in the inflamed tissues after the polymicrobial infection. Of these genes, 4476 and 1035 genes in the infected bone and tissues were differentially expressed by upregulation. Biological pathways significantly impacted by the polymicrobial infection in calvarial bone included leukocyte transendothelial migration (LTM), cell adhesion molecules, adherens junction, major histocompatibility complex antigen, extracellular matrix-receptor interaction, and antigen processing and presentation resulting in inflammatory/cytokine/chemokine transcripts stimulation in bone and soft tissue. Intense inflammation and increased activated osteoclasts were observed in calvarias compared with sham-infected controls. Quantitative real-time RT-PCR analysis confirmed that the mRNA level of selected genes corresponded with the microarray expression. The polymicrobial infection regulated several LTM and extracellular membrane pathway genes in a manner distinct from mono-infection with P. gingivalis, T. denticola, or T. forsythia. To our knowledge, this is the first definition of the polymicrobially induced transcriptome in calvarial bone and soft tissue in response to periodontal pathogens.

Tannerella forsythia infection-induced calvarial bone and soft tissue transcriptional profiles.

Tannerella forsythia is associated with subgingival biofilms in adult periodontitis, although the molecular mechanisms contributing to chronic inflammation and loss of periodontal bone remain unclear. We examined changes in the host transcriptional profiles during a T. forsythia infection using a murine calvarial model of inflammation and bone resorption. Tannerella forsythia was injected into the subcutaneous soft tissue over calvariae of BALB/c mice for 3 days, after which the soft tissues and calvarial bones were excised. RNA was isolated and Murine GeneChip (Affymetrix, Santa Clara, CA) array analysis of transcript profiles showed that 3226 genes were differentially expressed in the infected soft tissues (P < 0.05) and 2586 genes were differentially transcribed in calvarial bones after infection. Quantitative real-time reverse transcription-polymerase chain reaction analysis of transcription levels of selected genes corresponded well with the microarray results. Biological pathways significantly impacted by T. forsythia infection in calvarial bone and soft tissue included leukocyte transendothelial migration, cell adhesion molecules (immune system), extracellular matrix-receptor interaction, adherens junction, and antigen processing and presentation. Histologic examination revealed intense inflammation and increased osteoclasts in calvariae compared with controls. In conclusion, localized T. forsythia infection differentially induces transcription of a broad array of host genes, and the profiles differ between inflamed soft tissues and calvarial bone.

Virulence of major periodontal pathogens and lack of humoral immune protection in a rat model of periodontal disease.

OBJECTIVE: This study was designed to test the hypothesis that periodontal pathogens Tannerella forsythia and Porphyromonas gingivalis are synergistic in terms of virulence potential using a model of mixed-microbial infection in rats. MATERIALS AND METHODS: Three groups of rats were infected orally with either T. forsythia or P. gingivalis in mono-bacterial infections or as mixed-microbial infections for 12 weeks and a sham-infected group were used as a control. This study examined bacterial infection, inflammation, immunity, and alveolar bone loss changes with disease progression. RESULTS: Tannerella forsythia and P. gingivalis genomic DNA was detected in microbial samples from infected rats by PCR indicating their colonization in the rat oral cavity. Primary infection induced significantly high IgG, IgG2b, IgG1, and IgG2a antibody levels indicating activation of mixed Th1 and Th2 immune responses. Rats infected with the mixed-microbial consortium exhibited significantly increased palatal horizontal and interproximal alveolar bone loss. Histological examinations indicated significant hyperplasia of the gingival epithelium with moderate inflammatory infiltration and apical migration of junctional epithelium. The results observed differ compared to uninfected controls. CONCLUSION: Our results indicated that T. forsythia and P. gingivalis exhibit virulence, but not virulence synergy, resulting in the immuno-inflammatory responses and lack of humoral immune protection during periodontitis in rats.

Molecular characterization of Treponema denticola infection-induced bone and soft tissue transcriptional profiles.

Treponema denticola is associated with subgingival biofilms in adult periodontitis and with acute necrotizing ulcerative gingivitis. However, the molecular mechanisms by which T. denticola impacts periodontal inflammation and alveolar bone resorption remain unclear. Here, we examined changes in the host transcriptional profiles during a T. denticola infection using a murine calvarial model of inflammation and bone resorption. T. denticola was injected into the subcutaneous soft tissue over the calvaria of BALB/c mice for 3 days, after which the soft tissues and the calvarial bones were excised. RNA was isolated and analysed for transcript profiling using Murine GeneChip arrays. Following T. denticola infection, 2905 and 1234 genes in the infected calvarial bones and soft tissues, respectively, were differentially expressed (P