Les métastases osseuses sont une complication majeure des cancers du sein. Les bisphosphonates (BPs) bloquent la progression des lyses osseuses. Des effets anti-tumoraux ont été mis en évidence mais à des doses importantes incompatibles avec un usage clinique. La forte affinité des BPs pour le minéral osseux limite leur biodisponibilité et amoindrit donc leur potentiel antitumoral in vivo. Mon travail de thèse s'est inscrit dans le cadre de la potentialisation des effets anti-tumoraux des BPs. Les BPs ont été combinés avec des ultrasons de faible intensité (LIUS) dont le rôle est d'induire une stimulation mécanique et une augmentation modérée de la température du milieu insonifié sans effet de cavitation. Les LIUS favorisent la pénétration des BPs dans des cellules tumorales en augmentant le phénomène d'endocytose. In vivo, un traitement répété aux LIUS associé à une dose clinique de BPs entraine une diminution des lyses osseuses ainsi que de la masse tumorale. L'accumulation d'un marqueur, retrouvé dans les moelles des souris traitées aux LIUS, suggère que la pénétration des BPs a été favorisée sous l'effet des LIUS. L'effet de l'association BPs avec les LIUS a ensuite été évalué sur un modèle de tumeur sous-cutanée mammaire. Un ralentissement de la croissance tumorale lors des premiers jours de traitements a été mis en évidence. Une étude menée avec de la doxorubicine et des BPs a permis d'élargir le champ d'application des LIUS pour le traitement du cancer du sein. En conclusion, ces travaux montrent que les LIUS apparaissent comme une solution d'intérêt pour augmenter la pénétration de drogues dans des tumeurs osseuses et mammaires et augmenter leur potentiel antitumoral / Bone metastases are common complications of advanced breast cancer. They increase morbidity of patients and alter their quality of life. Bisphosphonates (BPs) stop the progression of osteolysis. However, BPs do not affect the tumor burden located inside the bone marrow cavity. Antitumoral effects have been shown but with high doses incompatibles with a clinical use. BPs bind also avidly to bone mineral which limits their bioavailability and reduce their antitumoral potential in vivo. This work is incorporated within the framework of the enhancement of antitumoral effects of BPs. BPs were combined with low intensity ultrasound (LIUS), which are known to induce a mechanical stimulation and a slight increase of temperature without involving cavitationnal effect. Initially, LIUS were found to increase the penetration of BPs inside several mammary tumor cell lines without affecting their viability by increasing endocytosis. In vivo, a daily repeated treatment of LIUS associated with a single and clinical dose of BPs lead to a decrease in osteolysis as well as tumor burden. The accumulation of unprenylated Rap1A form was found in bone marrow of mice suggesting that LIUS promote BPs penetration inside cells of the bone cavity. The effect of BPs and LIUS was evaluated in a subcutaneous mammary tumor xenograft. Tumor growth was slowed during the first days of LIUS treatment. A study was performed using doxorubicin and BPs leading to a better penetration of both compounds when LIUS were used. This last result allows increasing the field of application of LIUS for breast cancer treatment. In conclusion, this work showed that LIUS are an interesting method to enhance the penetration of drugs inside bone and mammary tumors leading to an increase of their antitumoral activity

Master of Science / Department of Chemistry / Santosh Aryal / Nanomedicine-based therapeutics have exhibited clear benefits when compared to unmodified drugs, which include improved pharmacokinetics, drug retention, targeting efficiency, and minimizes toxicity. Every year thousands of bone cancer cases are diagnosed in the United States. Moreover, development of bone metastasis occurs in over 80% to 90% of various cancers that metastasize and signals the entry of the disease into an incurable phase. Cancer in bones can cause pain, fractures, hypercalcemia, and compression of the spinal cord, due to deposits that can erode into the bone using bone-absorbing cells. Bisphosphonates are drugs that reduce the activity of bone-absorbing cells and targets overexpressed calcium. They are characterized pharmacologically to inhibit bone resorption, skeletal distribution, and renal elimination. In addition, they can target bone microenvironment and bind strongly with calcium. The goal of this thesis is to engineer targeted nanomedicine drug with the ability to spatiotemporally control therapeutics delivery to the bone. Herein we synthesized biopolymers with functional end group moieties as alendronate (a molecular member of bisphosphate), which can target overexpressed calcium ions at the vicinity of the bone lesion where bone resorption takes place. In order to achieve our goal, a ring opening polymerization of cyclic L-lactide initiated by ALE in the presence of catalytic amount of stannous octoate was conducted in an inert environment. Thus, formed polymers are characterized for their chemistry and physicochemical properties using various analytical tools. These polymers were characterized by nuclear magnetic resonance (¹H-NMR) and Fourier Transfer Infrared Spectrometer (FT-IR), which shows monomer conversion and the presence of amide and phosphate moiety. Thereafter we engineered bone-homing polymeric nanoparticles of 80nm diameter by nanoprecipitation for controlled delivery of Dox, a first line anticancer drug used in clinics. The in-vitro results show that the nanoparticles have the ability to accumulate and internalized into the bone cancer cells, deliver drugs efficiently, and are least toxic. Therefore, innovative and efficient bisphosphonate functionalized Poly-l-lactide polymers were synthesized to target bone microenvironment.

Osteopenia and/or Osteoporosis (OOS) is becoming an increasingly prevalent chronic disease among Beta Thalassemia Major (BTM) patients, especially now that life expectancy in these patients has considerably improved through regular blood transfusions and iron chelation therapy. With several, complex genetic and acquired factors involved in its pathogenesis, coupled with the heterogeneity in the clinical response of BTM patients to different pharmacological agents, OOS has proven to be particularly difficult to treat. The great majority of treatment options currently available are not curative, but instead are aimed towards managing the symptoms and progression of the disease in patients. General preventative measures, such as iron chelation therapy and hormonal replacement therapy (HRT), are instrumental aspects of the treatment plan; however, the incredible complexity of OOS necessitates an individualized, multidisciplinary approach to management, with a principal therapy that is safe and effective in patients, and that is accompanied by these other supportive measures. This review, through a comprehensive analysis of current literature, includes data from randomized, placebo-controlled trials, double blind and observational clinical studies, and suggests optimal therapeutic interventions for first-line management of OOS. It also addresses treatment options for BTM patients in whom resistance to the recommended first-line therapy develops, or who display secondary endocrine conditions contributing to OOS. In addition to providing a current synopsis of OOS management and the potential of emerging treatment options, this analysis highlights some of the limitations of traditional therapies. In this way, the paper effectively illustrates the current status of TM-induced OOS; it describes what is or isn’t working, as well as underscores the diagnostic and therapeutic challenges continually faced by patients, researchers and clinicians.

Made available in DSpace on 2014-06-11T19:30:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2005-12-16Bitstream added on 2014-06-13T19:01:06Z : No. of bitstreams: 1 fernandes_rg_dr_sjc.pdf: 1447209 bytes, checksum: f3ae23b178b1c3e233c4acb1b08af663 (MD5) / Esta pesquisa avaliou o efeito do uso local do alendronato sódico, da hidroxiapatita e da associação alendronato mais hidroxiapatita em diferentes concentrações molares, no processo de reparação de defeitos ósseos em fêmures de ratos. Foi confeccionado no fêmur de 168 ratos (84 machos e 84 fêmeas) um defeito ósseo medindo 2,5mm de diâmetro. Estes animais foram divididos em grupos: controle, amido, alendronato um mol, alendronato dois moles, hidroxiapatita um mol, hidroxiapatita dois moles e alendronato mais hidroxiapatita, de acordo com o material de preenchimento utilizado. Nos animais do grupo controle o defeito ficou preenchido apenas pelo coágulo proveniente do defeito. Os animais foram sacrificados aos sete e 21 dias, quando o fêmur era removido, fixado e descalcificado, para a confecção de lâminas histológicas. Foi realizada análise histológica e histomorfométrica, e os dados obtidos foram submetidos à análise estatística ANOVA. Aos sete dias, observava-se trabéculas ósseas imaturas, contendo grandes osteócitos. Notava-se neoformação óssea em todos os grupos, exceto nos animais machos onde o alendronato se fazia presente. Nos grupos que receberam a hidroxiapatita, visualizava-se as imagens negativas dos grânulos da hidroxiapatita. Aos 21 dias, as trabéculas praticamente fechavam o defeito da maioria dos espécimes estudados. Estatisticamente, houve diferenças entre machos e fêmeas, entre os períodos de observação e com relação ao uso do alendronato. Concluiu-se que a aplicação local do alendronato sódico interferiu negativamente na reparação óssea, que a hidroxiapatita e o alendronato mais a hidroxiapatita não interferiram na reparação e que a reparação óssea foi maior nas fêmeas independentemente do período estudado. / This research evaluated the effect of the local use of sodium alendronate, of hydroxyapatite and the association alendronate more hydroxyapatite in different molars concentrations, in the repair of bone defects in femurs of mices. We made in the femur of 168 mices (84 males and 84 females) a bone defect measuring 2,5mm of diameter. We divided these animals in groups: control, starch, alendronate one mol, alendronate two mols, hydroxyapatite one mol, hydroxyapatite two mols and alendronate more hydroxyapatite, in accordance with the material of fulfilment used. In the animals of the control group, the defect was just filled by the clot originating from defect. The animals were sacrificed at seven and 21 days, when the femur was removed, fixed and decalcified, to making histologics laminae. Histological and histomorphometric analyses were performed and the results obtained were submitted to statistical analysis ANOVA. At seven days, it was observed immature bone trabeculae with larges osteocyties. It was noticed bone formation in all groups, exceptin the male animals where the alendronate was present. In the group that received hydroxyapatite, it was visualized negatives images of the hydroxyapatite's granules. At 21 days, the trabeculae practically closed the defect of most studied specimens. Statistically, there were differences between males and females, between the observation periods and with relationship of the alendronate use. It was concluded that the local application of sodium alendronate interfered negatively in bone repair, that the hydroxyapatite and alendronate more hydroxyapatite didn't interfere in the repair and that the bone repair was larger in the females independently of studied period.

Made available in DSpace on 2014-06-11T19:30:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-09-29Bitstream added on 2014-06-13T21:01:51Z : No. of bitstreams: 1 canettieri_acv_dr_sjc_prot.pdf: 1776887 bytes, checksum: 80a7e878725233d8cf3256bd1985da11 (MD5) / Este trabalho avaliou a ação local do alendronato sódico, da hidroxiapatita e da associação alendronato com hidroxiapatita na reparação de defeitos ósseos em fêmures de ratas ovariectomizadas. Noventa e oito animais foram divididos em sete grupos: controle (C), amido (Am), alendronato 1mol (A1), alendronato 2moles (A2), hidroxiapatita 1 mol (HA1), hidroxiapatita 2moles (HA2) e associação alendronato e hidroxiapatita (A+HA). As ratas pesando, aproximadamente, 250g foram ovariectomizadas e, após trinta dias, os defeitos ósseos, medindo 2,5mm de diâmetro, foram confeccionados nos fêmures esquerdos. Os defeitos foram preenchidos com alendronato sódico, hidroxiapatita e/ou com ambos, sendo que o grupo C não recebeu material de preenchimento e o grupo Am apenas o amido. Os animais foram sacrificados sete e 21 dias após a cirurgia. Foram realizadas análise histológica e histomorfométrica da área do defeito ósseo e os resultados submetidos à análise estatística. Histologicamente, as principais diferenças ocorreram após 21 dias. Os grupos C, Am, HA1 e HA2 apresentaram fechamento linear do defeito ósseo em todos espécimes e a maioria dos animais dos grupos A1, A2 e A+HA não exibiu neoformação óssea na região central do defeito, permanecendo este preenchido por tecido conjuntivo fibroso. No período de sete dias não houve diferença estatística significante entre todos os grupos experimentais em relação a neoformação óssea e, após 21 dias, o grupo HA2 apresentou a maior quantidade de osso neoformado. Estatisticamente, não houve diferença entre os grupos A1, A2 e A+HA nos dois períodos de estudo. Concluiu-se que o alendronato sódico, isolado ou associado com a hidroxiapatita, prejudicou a reparação óssea neste modelo experimental e a hidroxiapatita utilizada mostrou-se biocompatível e osteocondutora, com os melhores resultados observados no grupo HA2. / This work evaluated the action of sodium alendronate, of hydroxyapatite and the association alendronate with hydroxyapatite in the repair of bone defects in ovariectomized rats femurs. Ninety eight animals were divided into seven groups: control (C), starch (Am), alendronate 1mol (A1), alendronate 2moles (A2), hydroxyapatite 1 mol (HA1), hydroxyapatite 2 moles (HA2) and the association alendronate and hydroxyapatite (A+HA). The rats weighing, approximately, 250g were ovariectomized and, after thirty days, bone defects, measuring 2,5mm, were created in the lefts femurs. The bone defects were filled with alendronate, hydroxyapatite or with both, but the group C not received none material, and the group Am, only starch. The animals were sacrified at seven and 21 days after surgery. Histological and histomorphometric analyses were performed and the results obtained were submitted to statistical analysis. Histologically, the principal differences occurred after 21 days, with the groups C, Am, HA1 and HA2 showing a linear closure of bone defect in every specimen. The most of animals of the groups A1, A2 and A+HA did not show central bone neoformation in bone defects, and there was fibrous connective tissue in this region. After seven days, there was not significance statistical difference among all experimental groups in relation to bone neoformation and, after 21 days, the group HA2 showed the most quantity of new bone formation. Statistically, there were no differences among the groups A1, A2 and A+HA in both studied period. It was concluded that the sodium alendronate, alone or combinated with hydroxyapatite, harmed the bone repair in this experimental model and the hydroxyapatite was biocompatible and osteoconductive, with the best results in group HA2.

Many things might happen in the body when a titanium implant is inserted into bone. Examples are activation of the immune system and imbalance between bone formation and bone resorption, which might lead to damaged bone around the implant and at worse, loosening of the implant. Bisphosphonates, BP’s, is a class of drugs that is able to decrease the osteoclast (bone resorption cell) activity and thereby strengthen the bone. FibMat2.0 is a fibrinogen matrix and consists of a thin protein layer which can be applied on an implant and act as a local drug delivery system. The work in this thesis was divided into two parts where aim of the first part was to study FibMat2.0 with integrated BP’s, and their effect in the presence of blood. The aim for the second part was to determine whether it was possible to incorporate antithrombotic drugs into the fibrinogen matrix. No detection method for the amount of drugs incorporated into the fibrinogen matrix was used but the fact that the drugs gave effect was verifying that it is possible to integrate other drugs than BP’s into FibMat2.0. Methods that have been used in the experiments in presence of blood are imaging of coagulation, fluorescence microscopy and cone-and-plate. For the first part, the results showed that surfaces incubated with fibrinogen and fibrinogen with integrated BP’s act alike in regard to coagulation and platelet adhesion. Compared to titanium, which is known to be a biocompatible material, the surfaces with fibrinogen and fibrinogen with BP’s behave similar in regard to platelet adhesion. When it comes to coagulation, the surfaces coated with fibrinogen with or without an addition of BP’s have shown a longer coagulation time compared to the clean titanium surface. For the second part, some conclusions have been drawn according to the results. Heparin and hirudin have shown anticoagulant effects when integrated in the matrix. The platelet inhibitor cangrelor seemed to have better effect when added in blood and incubated compared to incubation with the platelet inhibitor on the surface before incubation in blood. Finally, when combining heparin and cangrelor, very clear differences in regard to formation of fibrin network could be seen. It seems promising to be able to load different kind of drugs in FibMat2.0.

Η ακτινοθεραπεία αποτελεί σημαντική θεραπευτική προσέγγιση για ασθενείς με διάφορους τύπους καρκίνου συμπεριλαμβανομένων των καρκίνων μαστού και πνεύμονα. Τόσο στο μεταστατικό καρκίνο του μαστού, όσο και στο μεταστατικό καρκίνο του πνεύμονα και καρκίνο σταδίου IIIB, η ακτινοθεραπεία έχει παρηγορητικό ρόλο βελτιώνοντας την ποιότητα ζωής των ασθενών. Και τα δύο αυτά είδη καρκίνου μπορούν να δώσουν οστικές μεταστάσεις, για την αντιμετώπιση των οποίων, τα τελευταία χρόνια, χορηγούνται στους ασθενείς διφωσφονικά. Προ-κλινικές μελέτες έχουν δείξει ότι τα διφωσφονικά έχουν αντιαγγειογενετική και αντινεοπλασματική δράση. Σκοπός της παρούσας εργασίας είναι η μελέτη της δράσης του συνδυασμού της ακτινοβόλησης και των διφωσφονικών σε καρκινικά κύτταρα μαστού και πνεύμονα, λόγω του ότι τόσο η ακτινοθεραπεία, όσο και τα διφωσφονικά έχουν επίδραση στους οστεοκλάστες, πράγμα που ενισχύει το γεγονός ότι οι δύο αυτές θεραπείες μπορούν να έχουν συνεργική δράση. Για τα in vitro πειράματα χρησιμοποιήθηκαν οι ανθρώπινες καρκινικές σειρές μαστού MCF-7 και MDA-MB-231 και οι ανθρώπινες καρκινικές σειρές μη μικροκυτταρικού καρκίνου του πνεύμονα H23 και H358. Το ζολενδρονικό οξύ χορηγήθηκε στα κύτταρα σε διάφορες συγκεντρώσεις (0,1, 1, 10, 20 και 100 μΜ). Στα πειράματα συνδυασμού ακτινοβόλησης και ζολενδρονικού οξέος, το φάρμακο προστέθηκε σε συγκέντρωση 10 μΜ, 24 ώρες πριν την ακτινοβόληση. Τα κύτταρα ακτινοβολήθηκαν σε θερμοκρασία δωματίου με διάφορες δόσεις ακτινών Χ: 0, 0,5, 1, 2 και 5 Gy (σε γραμμικό επιταχυντή, 6 MV). Ο πολλαπλασιασμός των κυττάρων προσδιορίστηκε 2, 4 και 6 ημέρες μετά την ακτινοβόληση με τη μέθοδο του μεθυλ-τετραζολίου (MTT). Στη συνέχεια, με τη μέθοδο της κυτταρομετρίας ροής (FACS), μελετήθηκε το ποσοστό της επαγωγής της απόπτωσης και της νέκρωσης των καρκινικών κυττάρων μετά την εφαρμογή σε αυτά του συνδυασμού της ακτινοβόλησης και του ζολενδρονικού οξέος. Η προσθήκη του ζολενδρονικού οξέος πριν την ακτινοβόληση ευαισθητοποιεί τα κύτταρα στις ακτίνες Χ. Πιο συγκεκριμένα, στις καρκινικές σειρές μαστού, η ακτινοβόληση προκαλεί μείωση του αριθμού των κυττάρων, δράση που ενισχύεται όταν οι ακτίνες Χ συνδυάζονται με ζολενδρονικό οξύ. Στο μη μικροκυτταρικό καρκίνο του πνεύμονα, η ανασταλτική δράση των ακτινών Χ επίσης ενισχύεται παρουσία του ζολενδρονικού οξέος. Σε όλες τις περιπτώσεις, η ανασταλτική δράση του ζολενδρονικού οξέος, των ακτινών Χ και του συνδυασμού τους δεν οφείλεται σε επαγωγή της απόπτωσης ή της νέκρωσης των κυττάρων. Συμπερασματικά, η συνδυαστική εφαρμογή ακτινών Χ και ζολενδρονικού οξέος ευαισθητοποιεί τα καρκινικά κύτταρα μαστού και μη μικροκυτταρικού καρκίνου του πνεύμονα μειώνοντας τον αριθμό τους, είτε σε μικρότερους χρόνους είτε χρησιμοποιώντας μικρότερες δόσεις ακτινών Χ, αντίστοιχα, χωρίς να επάγει την απόπτωση ή τη νέκρωση των κυττάρων. / Radiotherapy is an important treatment for patients suffering from various types of cancer, including breast and lung cancer. Especially in metastatic breast and lung cancer, radiotherapy is a palliative treatment, which improves the quality of life of patients. Bone metastases are common in both breast and lung cancer and in recent years, bisphosphonates have become a standard treatment for metastatic bone disease. Pre-clinical research has proved that bisphosphonates have antiangiogenic and antitumor effects. The purpose of this particular research is to investigate the effect of the combination of X-rays and bisphosphonates in human breast cancer and in human non-small cell lung cancer cells, due to the fact that both radiotherapy and bisphosphonates have common action on osteoclasts, which implies a potential synergistic activity. The cell lines used in the in vitro experiments were MCF-7 and MDA-MB-231 human breast cancer cells and H23 and H358 human non-small cell lung cancers cells. Cells were treated with various doses of zoledronic acid (0.1, 1, 10, 20 and 100 nM). In case of the combination of radiotherapy and zoledronic acid, the drug dose was 10 μM and it was added 24 hours prior to radiotherapy. The cells were irradiated at room temperature and the doses of irradiation were 0, 0.5, 1, 2 and 5 Gy (linear accelerator, 6MV). The cell number was determined by the MTT method, 2, 4 and 6 days after the irradiation. Apoptosis and necrosis were estimated by using flow cytometry. Addition of zoledronic acid to the cells before irradiation rendered cells more sensitive to irradiation. In breast cancer cell lines, irradiation decreased the number of cells, an effect that was enhanced when the cells were irradiated in the presence of zolendronic acid. In non-small cell cancer cell lines, the inhibitory activity of irradiation was also enhanced in the presence of zoledronic acid. In all cases, the inhibitory effect of zolendronic acid, X-rays and their combination was not due to induction of cell apoptosis or necrosis. In summary, the combination of radiotherapy and zoledronic acid renders tumor cells more sensitive to irradiation by reducing their number in less time or in smaller irradiation doses, without inducing cell apoptosis or necrosis.

Recombinant human bone morphogenetic proteins (rhBMPs) are increasingly used for reconstructing bony defects, fracture non-unions, and augmenting existing bone volumes. For the numerous patients taking bisphosphonates, the impact of prior bisphosphonate treatment on rhBMP bone induction is not well understood. Objective: to evaluate the effect of the prior treatment with zoledronate on rhBMP induced bone formation in mice. Methods: 42 mice were pre-treated with 0, 2, and 20 µg of zoledronate/mouse. The osteoinductive activity of a bioimplant, containing rhBMP-2, was assessed using the mouse muscle pouch assay, and analyzed with micro CT and histology. Results: micro CT demonstrated that BMP bioimplants placed in mice pretreated with 20 µg of zoledronate, formed bony ossicles of greater volume but reduced bone density compared to controls. Histologically, the heterotopic ossicles from the 20 µg group consisted of more immature bone than those from the other groups. Conclusion: bone induced by rhBMP-2 in mice pre-treated with a high concentration of zoledronate was immature as evidenced by radiographic and histologic appearance.

Recombinant human bone morphogenetic proteins (rhBMPs) are increasingly used for reconstructing bony defects, fracture non-unions, and augmenting existing bone volumes. For the numerous patients taking bisphosphonates, the impact of prior bisphosphonate treatment on rhBMP bone induction is not well understood. Objective: to evaluate the effect of the prior treatment with zoledronate on rhBMP induced bone formation in mice. Methods: 42 mice were pre-treated with 0, 2, and 20 µg of zoledronate/mouse. The osteoinductive activity of a bioimplant, containing rhBMP-2, was assessed using the mouse muscle pouch assay, and analyzed with micro CT and histology. Results: micro CT demonstrated that BMP bioimplants placed in mice pretreated with 20 µg of zoledronate, formed bony ossicles of greater volume but reduced bone density compared to controls. Histologically, the heterotopic ossicles from the 20 µg group consisted of more immature bone than those from the other groups. Conclusion: bone induced by rhBMP-2 in mice pre-treated with a high concentration of zoledronate was immature as evidenced by radiographic and histologic appearance.

Submitted by CRISTIAN STATKIEVICZ (c.statkievicz@gmail.com) on 2016-10-15T03:12:53Z No. of bitstreams: 1 DISSERTAÇÃO CRISTIAN VERSÃO FINAL - REPOSITÓRIO.pdf: 7053318 bytes, checksum: 7e3743d0506776749bc5bf1702e95116 (MD5) / Approved for entry into archive by Juliano Benedito Ferreira (julianoferreira@reitoria.unesp.br) on 2016-10-20T18:34:04Z (GMT) No. of bitstreams: 1 statkievicz_c_me_araca.pdf: 7053318 bytes, checksum: 7e3743d0506776749bc5bf1702e95116 (MD5) / Made available in DSpace on 2016-10-20T18:34:04Z (GMT). No. of bitstreams: 1 statkievicz_c_me_araca.pdf: 7053318 bytes, checksum: 7e3743d0506776749bc5bf1702e95116 (MD5) Previous issue date: 2016-08-30 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / OBJETIVOS: Os objetivos do presente estudo foram avaliar o efeito da terapia com laser em baixa intensidade (LLLT) e da terapia fotodinâmica antimicrobiana (aPDT) no processo de reparo alveolar de ratas com os principais fatores de risco para a osteonecrose dos maxilares associada à terapia medicamentosa (ONM-M). MATERIAIS E MÉTODOS: Ratas senis (n=29) foram distribuídas nos grupos: SAL, ZOL, ZOL/LLLT e ZOL/aPDT. Durante sete semanas, a cada dois dias, administrou-se pela via intraperitoneal 0,45ml de NaCl 0,9% (SAL) ou 0,45ml desta mesma solução acrescida de 100μg/Kg de zoledronato (ZOL, ZOL/LLLT e ZOL/aPDT). Decorridas três semanas, realizou-se a exodontia do primeiro molar inferior esquerdo. Nos grupos ZOL/LLLT e ZOL/aPDT as ratas foram submetidas respectivamente a LLLT ou aPDT aos 0, 2 e 4 dias pós exodontia. A eutanásia foi realizada 28 dias após a exodontia. No sítio de extração dental foram realizadas análises: histopatológica do processo de reparação tecidual, histométrica da área de tecido ósseo neoformado (ATO) e imunoistoquímica direcionada para os seguintes biomarcadores: PCNA, BAX, C3C, TNFα, IL-1β, IL-6, HIF-1α, VEGF, CD31, BMP2/4, RUNX-2, OCN, OPG, RANKL e TRAP. RESULTADOS: Os grupos tratados com zoledronato apresentaram maior imunomarcação para OPG e menor imunomarcação para RANKL e TRAP. Em ZOL observou-se áreas de osteonecrose, comprometimento da reparação tecidual, menor ATO, menor imunomarcação para PCNA, HIF-1α, VEGF, CD31, BMP2/4 e OCN, e maior imunomarcação para BAX, C3C, TNFα, IL-1β, IL-6 e RUNX-2 em relação ao SAL. ZOL/LLLT apresentou melhora em alguns parâmetros em relação ao ZOL (ATO, PCNA, TNFα, HIF-1α, VEGF, CD31 e RUNX-2), no entanto, poucos foram os parâmetros que se igualaram ao SAL (PCNA, HIF-1α, VEGF, CD31 e RUNX-2). ZOL/aPDT não apresentou áreas de osteonecrose, o processo de reparação tecidual não diferiu significativamente de SAL, assim como, os seguintes parâmetros ATO, PCNA, BAX, C3C, IL-1β, HIF-1α, VEGF, CD31, RUNX-2 e OCN. CONCLUSÕES: O zoledronato compromete o processo de reparação tecidual do sítio de extração dental em ratas com os principais fatores de risco para a ONM-M. LLLT e aPDT são capazes de melhorar eventos relacionados com o processo de reparo alveolar. A aPDT se mostrou a terapia preventiva mais efetiva para evitar a ONM-M. RELEVÂNCIA CLÍNICA: A aPDT pode se constituir em uma alternativa de terapia preventiva para evitar o desencadeamento de ONM-M pós exodontia. / OBJECTIVES: The aim of this study was to evaluate the effects of Low Level Laser Therapy (LLLT) and antimicrobial photodynamic therapy (aPDT) in the alveolar bone repair in rats with the high risk factors for Medication-Related Osteonecrosis of the Jaw (MRONJ). MATERIALS AND METHODS: Senile rats (n = 29) were distributed into four groups: SAL, ZOL, ZOL/LLLT and ZOL/aPDT. For seven weeks, every two days, 0,45ml 0.9% NaCl (SAL) or 0,45ml of this same solution plus 100μg/kg zoledronate (ZOL, ZOL/LLLT and ZOL/aPDT) were administered intraperitoneally. After three weeks, the first lower left molar was extracted. In ZOL/LLLT and ZOL/aPDT groups, the rats were submitted to LLLT or aPDT therapy at 0, 2 and 4 days after tooth extraction. Euthanasia was performed 28 days after tooth extraction. In dental extraction site was performed the histopathology analysis of the tissue repair, histometric analysis of newly formed bone area (NFB) and immunohistochemistry related to the following biomarkers: PCNA, BAX, C3C, TNFα, IL-1β, IL-6, HIF-1α, VEGF, CD31, BMP2/4, RUNX-2, OCN, OPG, RANKL and TRAP. RESULTS: The groups treated with zoledronate showed higher immunolabeling for OPG and lower immunolabeling for RANKL and TRAP. In ZOL group was observed areas of osteonecrosis, impairment of wound healing, lower ATO, lower immunolabeling for PCNA, HIF-1α, VEGF, CD31, BMP2/4 and OCN, and higher immunolabeling for BAX, C3C, TNFα, IL-1β, IL-6 and RUNX-2 when compared to the SAL group. ZOL/LLLT showed improvement in some parameters regarding to ZOL (ATO, PCNA, TNFα, HIF-1α, VEGF, CD31 and RUNX-2), however, few parameters were similar to the SAL (PCNA, HIF-1α, VEGF, CD31 and RUNX-2). ZOL/aPDT did not present areas of osteonecrosis and the tissue repair did not differ significantly from the SAL groups, as well as the following parameters: PCNA, BAX, C3C, IL-1β, HIF-1α, VEGF, CD31, RUNX-2 and OCN. CONCLUSIONS: The zoledronate compromises the tissue repair of dental extraction site in rats with the high risk factors for MRONJ. LLLT and aPDT were able to improved the events related to wound's healing. The aPDT showed to be the most effective preventive therapy to MRONJ. CLINICAL RELEVANCE: The aPDT can be in a preventive therapy to prevent the triggering of MRONJ after dental extraction. / FAPESP: 2014/02199-1