Thesis (doctoral)--Lund University, 1995. / Added t.p. with thesis statement inserted.

Thesis (doctoral)--Lund University, 1995. / Added t.p. with thesis statement inserted.

Bisphosphonate-Related Osteonecrosis of the Jaw (BRONJ) is a chronic necrosis of the jawbone that occurs in ≈1-5% of patients receiving bisphosphonate medication for conditions such as osteoporosis and certain cancers. Although the pathogenesis of BRONJ is still uncertain, several recent theories have emerged; these include vascular disruption of the jaw tissue; inappropriate osteoclast activation; and direct cytotoxicity of jaw epithelium. However, despite bisphosphonates having well-documented stimulatory effects on immune cells, and BRONJ being associated with oral microbial infections, an immune-mediated pathology for BRONJ has largely been ignored. Bisphosphonates activate human γδ T cells, specifically those that use T cell receptor (TCR) γ-chain variable-region-9, and TCR δ-chain variable-region-2 (Vγ9Vδ2 T cells). These unconventional T cells typically account for ≈ 1-5% of circulating lymphocytes, make protective responses to microbial challenge, and are promising candidates for cancer immunotherapy. In the context of BRONJ, we have hypothesised that bisphosphonate-mediated activation of jaw-associated Vγ9Vδ2 T cells, in the presence of oral microbiota, may drive the disruption of bone turnover that is central to the disease process. On initiation of these studies, it quickly became apparent that traditional characterisation of Vγ9Vδ2 T cells using CD27/CD45RA was demonstrably sub-optimal, while phenotypic analysis of 63 healthy volunteers revealed an unexpected and surprising degree of Vγ9Vδ2 T cell heterogeneity; both of which are likely to confuse assessment of disease scenarios. Thus, this thesis describes the novel phenotypic and functional characterisation of Vγ9Vδ2 T cells using alternative surface markers and methods of analysis. This reveals that different sets of individuals have different Vγ9Vδ2 T cell “profiles” that predict very different functional capabilities and responses to immunotherapeutic interventions. Using this improved definition of Vγ9Vδ2 T cells, this thesis then describes preliminary investigations of Vγ9Vδ2 T cell development in the human neonatal thymus, and assesses the involvement of Vγ9Vδ22 T cells in a small cohort of eight BRONJ patients.

Orientador: Edilson Ervolino / Coorientador: Juliano Milanezi de Almeida / Banca: Elias Naim Kassis / Banca: Rogerio Leone Buchaim / Resumo: O objetivo do presente estudo foi avaliar a terapia fotodinâmica antimicrobiana (aPDT), como monoterapia ou como coadjuvante à raspagem e alisamento radicular (RAR) no tratamento da periodontite experimental (PE) em ratos tratados com dose oncológica de zoledronato. Cento e vinte ratos foram aleatoriamente distribuídos em quatro grupos experimentais, cada um composto por 30 ratos. Durante 8 semanas, foi administrado 100 µg/Kg de zoledronato com intervalo de três dias entre as injeções. No 14º dia do início do protocolo medicamentoso, foi instalada uma ligadura nos primeiros molares inferiores para indução de PE. Após 14 dias a ligadura foi removida e foram realizados os seguintes tratamentos: exclusivamente uma sessão de RAR (grupo PE-RAR), exclusivamente três sessões de aPDT (grupo PE-aPDT) e uma sessão de RAR associada à três sessões de aPDT (grupo PE-RAR-aPDT). As sessões de aPDT foram realizadas aos 0, 2 e 4 dias pós tratamento local. No grupo PE-NTL não foi realizado nenhum tratamento local. Após 7, 14 e 28 dias pós tratamento periodontal local foram realizadas as eutanásias. Foi realizada análise microtomografica (micro-CT), análise histopatológica, histométrica da porcentagem tecido ósseo (PTO) e porcentagem de tecido ósseo não vital (PTONV) e imunoistoquímica para fator de crescimento transformador beta (TGFβ) nos tecidos periodontais. Os dados obtidos foram submetidos à análise quantitativa e estatística com nível de significancia de 5%. Não houve diferença estatisti... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of the present study was to evaluate the antimicrobial photodynamic therapy (aPDT) as a monotherapy or adjunct to scaling and root planning (SRP) for the treatment of experimental periodontitis (EP) in rats treated with oncological dosage of zoledr onate. A hundred and twenty rats were randomly divided in four experimental groups, each one of them composed by 30 rats. During a period of 8 weeks, 100 μg/Kg of zoledronate was administered every three days. After 14 days of the beginning of the drug pro tocol, a ligature was installed in the lower first molars for EP induction. After 14 days, ligature was removed and the following treatments were performed: a single session of SRP, exclusively (group EP - SRP), three sessions of aPDT, exclusively (group EP - aPDT), and one session of SRP associated to the three sessions of aPDT (group EP - SRP - aPDT). aPDT sessions were performed at days 0, 2, and 4 after the local treatment. In the fourth group, no local treatment was made (EP - NLT). After 7, 114, and 28 days aft er the local periodontal treatment, the animals were euthanized. Microcomputed tomography (micro - CT) analysis, histopathologic analysis, percentage of bone tissue (PBT), percentage of non - vital bone (PNVB), and immunohistochemistry for transforming growth factor beta ( TGFβ ) in the periodontal tissues were performed. Obtained data underwent quantitative analysis and statistics considering 5% level of significance. No statistical difference was detected i... (Complete abstract click electronic access below) / Mestre

Submitted by DANIELA PEREIRA DE SÁ (daniela.sa@hotmail.com) on 2018-04-25T20:08:16Z No. of bitstreams: 1 DISSERTAÇAO MESTRADO DANIELA SÁ PDF....pdf: 2506715 bytes, checksum: bf68fa58e403e9ad16de3c31ff7d3b9d (MD5) / Approved for entry into archive by Ana Paula Rimoli de Oliveira null (anapaula@foa.unesp.br) on 2018-04-25T20:29:09Z (GMT) No. of bitstreams: 1 sa_dp_me_araca_int.pdf: 2373246 bytes, checksum: 97cbbe6e1db525a114babf3166227ff5 (MD5) / Made available in DSpace on 2018-04-25T20:29:09Z (GMT). No. of bitstreams: 1 sa_dp_me_araca_int.pdf: 2373246 bytes, checksum: 97cbbe6e1db525a114babf3166227ff5 (MD5) Previous issue date: 2018-04-16 / O objetivo do presente estudo foi avaliar a terapia fotodinâmica antimicrobiana (aPDT), como monoterapia ou como coadjuvante à raspagem e alisamento radicular (RAR) no tratamento da periodontite experimental (PE) em ratos tratados com dose oncológica de zoledronato. Cento e vinte ratos foram aleatoriamente distribuídos em quatro grupos experimentais, cada um composto por 30 ratos. Durante 8 semanas, foi administrado 100 µg/Kg de zoledronato com intervalo de três dias entre as injeções. No 14º dia do início do protocolo medicamentoso, foi instalada uma ligadura nos primeiros molares inferiores para indução de PE. Após 14 dias a ligadura foi removida e foram realizados os seguintes tratamentos: exclusivamente uma sessão de RAR (grupo PE-RAR), exclusivamente três sessões de aPDT (grupo PE-aPDT) e uma sessão de RAR associada à três sessões de aPDT (grupo PE-RAR-aPDT). As sessões de aPDT foram realizadas aos 0, 2 e 4 dias pós tratamento local. No grupo PE-NTL não foi realizado nenhum tratamento local. Após 7, 14 e 28 dias pós tratamento periodontal local foram realizadas as eutanásias. Foi realizada análise microtomografica (micro-CT), análise histopatológica, histométrica da porcentagem tecido ósseo (PTO) e porcentagem de tecido ósseo não vital (PTONV) e imunoistoquímica para fator de crescimento transformador beta (TGFβ) nos tecidos periodontais. Os dados obtidos foram submetidos à análise quantitativa e estatística com nível de significancia de 5%. Não houve diferença estatisticamente significante nos seguintes parâmetros microtomográficos: perda óssea alveolar, volume ósseo na furca, número e espessura de trabéculas ósseas. O processo de reparação tecidual pós tratamento foi mais favorável em PE-aPDT e PE-RAR-aPDT. Não houve diferença estatisticamente significante na PTO, todavia, a PTONV foi menor tanto em PE-aPDT quanto em PE-RAR-aPDT. A imunomarcação para TGFβ foi maior em PE-RAR, PE-aPDT e PE-RAR-aPDT aos 7 dias pós tratamento. Conclui-se que o emprego da aPDT, quando empregada como monoterapia ou como terapia coadjuvante à RAR, se mostrou mais efetiva e segura para tratamento da PE em ratos. / The aim of the present study was to evaluate the antimicrobial photodynamic therapy (aPDT) as a monotherapy or adjunct to scaling and root planning (SRP) for the treatment of experimental periodontitis (EP) in rats treated with oncological dosage of zoledr onate. A hundred and twenty rats were randomly divided in four experimental groups, each one of them composed by 30 rats. During a period of 8 weeks, 100 μg/Kg of zoledronate was administered every three days. After 14 days of the beginning of the drug pro tocol, a ligature was installed in the lower first molars for EP induction. After 14 days, ligature was removed and the following treatments were performed: a single session of SRP, exclusively (group EP - SRP), three sessions of aPDT, exclusively (group EP - aPDT), and one session of SRP associated to the three sessions of aPDT (group EP - SRP - aPDT). aPDT sessions were performed at days 0, 2, and 4 after the local treatment. In the fourth group, no local treatment was made (EP - NLT). After 7, 114, and 28 days aft er the local periodontal treatment, the animals were euthanized. Microcomputed tomography (micro - CT) analysis, histopathologic analysis, percentage of bone tissue (PBT), percentage of non - vital bone (PNVB), and immunohistochemistry for transforming growth factor beta ( TGFβ ) in the periodontal tissues were performed. Obtained data underwent quantitative analysis and statistics considering 5% level of significance. No statistical difference was detected in the following microtomographic parameters: alveolar b one loss, bone volume in the furcation, number and thickness of the trabecullaes. Tissue healing after the treatment was more favorable in EP - aPDT and EP - RAR - aPDT groups. No significant differences were detected in PTO; however, PNVB was lower both in EP - a PDT and EP - SRP. Immunolabeling for TGFβ was higher in EP - SRP, EP - aPDT and EP - SRP - aPDT at day 7 post - treatment. It is concluded that when aPDT is used as monotherapy or as adjunct therapy associated with SRP, it is more effective and safe for the treatment of EP in rats.

Introduction First described in 2002, osteonecrosis of the jaw (ONJ, or avascular necrosis of the jaw) is an uncommon but potentially serious side effect of treatment with bisphosphonates. Although typically identified in patients with multiple myeloma and other malignancies, a few cases have been reported in patients taking bisphosphonates - a potent drug class used in the treatment of osteoclast-mediated bone resorption issues, including postmenopausal osteoporosis, Paget's disease, multiple myeloma, and malignant hypercalcemia. The clinical diagnosis of ONJ can be obscured by jaw pain, abscess, swelling, and fistulas, but exposed bone is a distinctive sign. This reports a case of ONJ secondary to bisphosphonate use in a 65-year-old woman and clinical management complications. Case Presentation A 65-year-old lady with history of age-related osteoporosis and compression fractures on alendronate for 4 years, squamous cell carcinoma of neck status post excision and radiotherapy 11-years prior, Sjogren's syndrome and discoid lupus on hydroxychloroquine, diabetes, hypertension, stroke and multiple dental abscesses presents with persistent neck pain. Initial CT neck with contrast showed diffuse fat stranding. Subsequently, alendronate was discontinued due to jaw necrosis suspicion. Eight months later, repeat CT scan showed new non-mass-like soft tissue thickening in the subcutaneous fat abutting the right anterior mandible with mandibular teeth cavities and periapical lucencies, likely to be periodontal cellulitis versus radiation osteonecrosis. Later, patient complained of a piece of bone penetrating the skin of her chin and presented with continuous drainage from sinus tract in her mandible, which was diagnosed as osteonecrosis attributed to bisphosphonates, previous radiation therapy, and dental abscesses. Patient was started on abaloparatide, an osteo-anabolic medication for osteoporosis and enrolled in hyperbaric oxygen therapy which immensely helped in controlling sinus drainage. Patient is currently awaiting mandibular reconstruction surgery. Discussion ONJ, often associated with pain, swelling, exposed bone, local infection, and pathologic fracture of the jaw, is a rare complication of bisphosphonate therapy. Currently, no prospective data exists to advise the benefits of therapy discontinuation however most clinical practices tend to discontinue at least temporarily. The incidence increases with longer treatment duration, particularly when therapy exceeds four years. Risk factors for developing ONJ while taking bisphosphonates include IV administration, anticancer therapy, dose and duration of exposure, dental extractions/implants, glucocorticoids, smoking, diabetes, and preexisting dental disease. Case reports and series suggest benefit from hyperbaric oxygen therapy in wound healing, pain, and quality of life at three months, however no significant differences exist with outcomes beyond three months. Patients being considered for therapy with a bisphosphonate should be thoroughly evaluated for dental issues, prior to initiating therapy. Conservative management with limited debridement, antibiotic therapy as needed, and topical mouth rinses rather than aggressive surgical resection are recommended. Conservative therapy may result in healing in a significant proportion of patients. Surgical resection of necrotic bone should be reserved for refractory or advanced cases. Providers should remain cautious while prescribing high doses of bisphosphonates in patients with increased risk factors to prevent, timely diagnose and treat this condition. References Edwards BJ, Gounder M, McKoy JM, et al. Pharmacovigilance and reporting oversight in US FDA fast-track process: bisphosphonates and osteonecrosis of the jaw. Lancet Oncol 2008; 9:1166. Khosla S, Burr D, Cauley J, et al. Bisphosphonate-associated osteonecrosis of the jaw: report of a task force of the American Society for Bone and Mineral Research. J Bone Miner Res 2007; 22:1479. Hoff AO, Toth BB, Altundag K, et al. Frequency and risk factors associated with osteonecrosis of the jaw in cancer patients treated with intravenous bisphosphonates. J Bone Miner Res 2008; 23:826.

Orientador: Edilson Ervolino / Coorientador: Juliano Milanezi de Almeida / Banca: Idelmo Rangel Garcia Junior / Banca: Paula Lazilha Faleiros / Resumo: O objetivo do presente estudo foi avaliar a progressão da periodontite experimental (PE) e a resposta tecidual periodontal frente à raspagem e alisamento radicular (RAR) durante tratamento com dose oncológica de zoledronato. Foram utilizados 100 ratos (6 meses de idade) distribuídos aleatoriamente em 4 grupos experimentais: SAL (n=30), ZOL (n=30), SAL-RAR (n=20) e ZOL-RAR (n=20). O plano de tratamento medicamentoso teve duração de 8 semanas. Os ratos receberam injeções intraperitoneais de 0,45 ml de solução de cloreto de sódio 0,9% (Grupos SAL e SAL-RAR) ou 0,45 ml da mesma solução acrescida de 100 µg/Kg de zoledronato (ZOL e ZOL-RAR) com um intervalo de três dias entre as aplicações. Decorridas duas semanas de tratamento medicamentoso, foi instalada uma ligadura de algodão ao redor do primeiro molar inferior esquerdo. Nos grupos SAL e ZOL essa ligadura permaneceu até o final do experimento. Nos grupos SAL-RAR e ZOL-RAR após outras duas semanas a ligadura foi removida, e foi efetuada a RAR. A eutanásia foi efetuada nos grupos SAL e ZOL aos 14, 21 e 42 dias pós instalação da ligadura e nos grupos SAL-RAR e ZOL-RAR aos 7 e 28 dias pós tratamento local com RAR. Foi executado o processamento histológico das hemi-mandíbulas e os cortes histológicos foram submetidos à coloração pela hematoxilina-eosina (HE). Na região de furca do primeiro molar inferior foram efetuadas: análise histopatológica dos tecidos peridontais e análise histométrica da porcentagem de osso na região de furca ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The present study aims to assess the progression of experimental periodontitis (EP) and the periodontal tissue response to scaling and root planing (SRP) during treatment with oncological dose of zoledronate. A total of 100 rats were used (6 months old) randomically distributed into 4 experimental groups: SAL (n=30), ZOL (n=30), SAL-SRP (n=20) e ZOL-SRP (n=20). Drug treatment plan lasted for 8 weeks. Intraperitoneal injections of 0,45 ml of sodium chloride solution 0,9% (Groups SAL and SAL-SRP) or 0,45 ml of the same solution adding 100 μg/Kg of zoledronate (ZOL and ZOL-SRP) were given to the rats within intervals of three days between injections. A cotton ligature was installed around the first left lower molar after two weeks of medicative treatment. At SAL and ZOL groups such ligature endured until the end of the experiment. At the groups SAL-SRP and ZOL-SRP ligature was removed two more weeks later and SRP was executed. Euthanasia was done at the groups SAL and ZOL after 14, 21 and 42 days from the ligature installation and at the groups SAL-SRP and ZOL-SRP after 7 and 28 days after local treatment with SRP. Hemimandibula was histological processing and executed and histological sections went under Hematoxylin and eosin staining (HE). At the furcal region of the first lower molar were executed: histopathological analysis of periodontal tissues and histometric analysis of bone percentage at the furcal region (BPF) and necrotic bone percentage at the furcal region (NPF). Results went under statistical analysis. ZOL group presented increased local inflammatory response, higther BPF and higher NPF at days 14, 21 and 42 after ligature installation in relation to SAL group. At ZOL-SRP, inflammatory response was increased, there was no BFP alteration, and NPF was higher throughout time in comparison to the group... / Mestre

Os bisfosfonatos têm sido indicados para o tratamento de doenças ósseas líticas. Atualmente, seu emprego terapêutico aumentou e com ele os efeitos adversos, sendo um dos mais importante a indução da osteonecrose dos maxilares, uma complicação de difícil tratamento e solução. Até o presente, não se sabe ao certo qual o mecanismo de desenvolvimento da osteonecrose e nem qual deve ser o melhor tratamento estabelecido perante essa manifestação. Este trabalho teve como objetivo avaliar o efeito do alendronato sódico sobre a viabilidade e proliferação de osteoblastos e fibroblastos em cultura. Foram utilizados osteoblastos-símile linhagem OSTEO 1 e fibroblastos de mucosa bucal humana linhagem FMM1. Após serem submetidos aos testes de citotoxicidade com concentrações do alendronato sódico variando de 10-2M a 10-8M os fibroblastos apresentaram diminuição significante de viabilidade celular apenas na concentração de 10-2M (p<0,01). Os osteoblastos demonstraram viabilidade celular no grupo controle significantemente maior que todos os demais grupos; o grupo tratado com o alendronato na concentração de 10-4M apresentou viabilidade celular semelhante a de todos os grupos, exceto o grupo de concentração 10-2M e a viabilidade celular dos demais grupos foi semelhante entre si (p<0,01). As concentrações de alendronato sódico superiores a 10-5M impediram a proliferação dos osteoblastos. Foi possível concluir que o alendronato sódico é citotóxico para células osteoblastos-símile e fibroblastos em cultura em função de sua concentração. Os fibroblastos são menos sensíveis a concentrações maiores de alendronato sódico que os osteoblastos. / Bisphosphonates have been therapeutically used for the management of lytic bone diseases. Their use has been increased nowadays and besides that associated adverse effects have been amplified. Jaw osteonecrosis induced by this drug is perhaps the most important complication because of the great morbidity and difficulty to deal with. Until now the physiopathology of osteonecrosis remains unclear and the treatment that should be established is uncertain. This study aimed to evaluate the effect of sodium alendronate a bisphosphonate used for the treatment of osteoporosis on the viability and proliferation of osteoblasts and fibroblasts in culture. Osteoblast-simile from the lineage OSTEO1 and a human oral mucosa fibroblasts from the lineage FMM1 were used. After being subjected to tests with concentrations of sodic alendronate ranging from 10-8M to 10-2M fibroblasts showed a significant decrease in cell viability at the concentration of 10-2M (p < 0.01). Osteoblasts showed that the cell viability in the control group was significantly higher than all other groups, the group treated with alendronate at a concentration of 10-4M had similar cell viability with all groups except the group of 10-2M concentration and the cell viability of other groups was similar between groups (p < 0.01). The concentrations of alendronate greater than 10-5M prevented the proliferation of osteoblasts. It was possible to conclude that alendronate is cytotoxic to osteoblast-símile cells and fibroblasts in culture due to its concentration. The fibroblasts are less sensitive to higher concentrations of alendronate than osteoblasts.

Bisphosphonate-related osteonecrosis of the jaws (BRONJ) occurs subsequent to intravenous and oral bisphosphonate exposure in a small subset of patients. Evidence of concurrent bacterial colonization at sites of bone necrosis, previous reports of neutrophil-related complications in some patients taking bisphosphonates along with perturbed neutrophil function in bisphosphonate-treated mice suggests an innate immune role in the development of bisphosphonate-related osteonecrosis of the jaws. This study investigates neutrophil function in BRONJ patients to determine if neutrophil functional defects may serve as a potential biomarker for BRONJ susceptibility. Patients with BRONJ and patients beginning intravenous pamidronate were studied. Eighteen patients with BRONJ and five patients beginning pamidronate therapy provided oral and blood neutrophil samples. Neutrophils from the population of patients with bisphosphonate-related osteonecrosis of the jaws and from those post-pamidronate treatment showed lower reactive-oxygen species production. These data suggest that a compromise in neutrophil function may be a potential biomarker for BRONJ susceptibility.

Bisphosphonate-related osteonecrosis of the jaws (BRONJ) occurs subsequent to intravenous and oral bisphosphonate exposure in a small subset of patients. Evidence of concurrent bacterial colonization at sites of bone necrosis, previous reports of neutrophil-related complications in some patients taking bisphosphonates along with perturbed neutrophil function in bisphosphonate-treated mice suggests an innate immune role in the development of bisphosphonate-related osteonecrosis of the jaws. This study investigates neutrophil function in BRONJ patients to determine if neutrophil functional defects may serve as a potential biomarker for BRONJ susceptibility. Patients with BRONJ and patients beginning intravenous pamidronate were studied. Eighteen patients with BRONJ and five patients beginning pamidronate therapy provided oral and blood neutrophil samples. Neutrophils from the population of patients with bisphosphonate-related osteonecrosis of the jaws and from those post-pamidronate treatment showed lower reactive-oxygen species production. These data suggest that a compromise in neutrophil function may be a potential biomarker for BRONJ susceptibility.