Diss. (sammanfattning) Linköping : Univ., 2003. / Härtill 4 uppsatser.

The forces applied during orthodontic treatment will cause effects on the teeth and adjacent tissues, depending on the intensity and time of application. Changes caused in the pulp tissues are not fully clarified in the literature by the scarcity of studies, the variety of methodologies, conflicting results and difficulty in conducting research that simulate clinical situations. Given this fact, the present study aimed to evaluate the human dental pulp after application of orthodontic intrusion force through histological examination. The selected sample consisted of 34 first human premolars, with orthodontic indication of extraction, from 17 young individuals (12-19 years old, both genders). On each patient, was applied intrusion force of 60g, randomly in only one of the dental elements, forming Group-Experimental (GE). The counterpart did not receive force, constituting the Control Group (CG), characterizing a split-mouth study. After 21 days, the premolars were extracted, stored in 10% formaldehyde, submitted to histotechnical preparation and stained with hematoxylineosin. The analysis of the histological slides was carried out in optical microscopy by a single experienced calibrated examiner. The paired Fisher exact test (p ≤ 0.05), showed significant increase of fibrous tissue in the GE. The nonparametric Wilcoxon test (p ≤ 0.05), paired, showed significant increase in the number of pulpal nodules in the elements of GE and showed no difference in the number of blood vessels between the groups. In 8 elements of the GE was observed the presence of congested large caliber vessels. The orthodontic intrusion force in these conditions, caused vascular changes in the pulp tissue, increase of fibrosis and of the number of pulpal calcifications in the experimental elements. / As forças aplicadas durante o tratamento ortodôntico ocasionam efeitos nos dentes e tecidos adjacentes, conforme a intensidade e tempo de aplicação. As alterações causadas nos tecidos pulpares não estão totalmente esclarecidas na literatura pela escassez de estudos, variedade de metodologias empregadas, resultados conflitantes e dificuldade de realização de pesquisas que simulem situações clínicas. Diante desse fato, o presente estudo teve como objetivo avaliar a polpa dental humana, após a aplicação de força ortodôntica de intrusão, por meio de exame histológico. A amostra selecionada foi de 34 primeiros pré-molares humanos, com indicação ortodôntica de exodontia, de 17 indivíduos jovens (12 a 19 anos, ambos os sexos). Em cada paciente, foi aplicada força de intrusão de 60g aleatoriamente em apenas um dos elementos dentais, constituindo o Grupo- Experimental (GE). O homólogo não recebeu força constituindo o Grupo-Controle (GC), caracterizando um estudo de boca dividida. Passados 21 dias, os pré-molares foram extraídos, armazenados em formol a 10%, submetidos ao preparo histotécnico e corados com hematoxilina-eosina. A análise das lâminas histológicas foi feita em microscópio óptico, por um único examinador experiente calibrado. O teste exato de Fischer (p≤ 0,05), pareado, mostrou significativo aumento da presença de tecido fibroso no GE. O teste não-paramétrico de Wilcoxon (p≤ 0,05), pareado, mostrou significativo aumento no número de nódulos pulpares nos elementos do GE e não mostrou diferença no número de vasos sanguíneos entre os grupos. Em 8 elementos do GE foi observada a presença de vasos calibrosos e congestos. A força ortodôntica de intrusão, nessas condições, causou alterações vasculares no tecido pulpar, aumentou a presença de áreas de fibrose e o número de calcificações pulpares nos elementos experimentais.

Cavalcanti, BN. Resposta de fibroblastos de polpa humanos frente a substâncias liberadas por capeadores pulpares diretos [Tese de Doutorado]. São Paulo: Faculdade de Odontologia da USP; 2003. RESUMO O objetivo do presente estudo foi o de avaliar os efeitos citotóxicos de substâncias liberadas durante a aplicação de materiais utilizados em capeamento pulpar direto, sobre fibroblastos de polpa dentária humana. Utilizou-se para o experimento meios condicionados pelas substâncias a serem testadas, divididas nos grupos a seguir: grupo I: controle (meio de cultivo sem condicionamento); grupo II: cimento de hidróxido de cálcio; grupo III: adesivo dentinário; grupo IV: ácido ortofosfórico a 37%. O condicionamento foi realizado, colocando-se meio de cultivo fresco sobre os materiais de modo que a presa (grupo II), polimerização (grupo III) ou o contato direto (grupo IV) liberassem substâncias para esse meio de cultivo. Esse meio era colocado sobre as células durante todo o experimento, excetuando-se o grupo IV, onde o contato foi feito por um período de 15 segundos, conforme recomendações clínicas. Posteriormente foram realizadas contagens em hemocitômetro pelo método de exclusão por azul de Trypan, que cora somente as células mortas. As contagens foram realizadas em períodos de 0, 6, 12 e 24 horas para o experimento de viabilidade celular (curto prazo), onde se avaliou o percentual de células vivas sobre o total de células, e em períodos de 1, 3, 5 e 7 dias para o experimento de sobrevivência celular, no qual se avaliou o número absoluto de células vivas. Observou-se que as substâncias liberadas pelo adesivo dentinário são citotóxicas em qualquer período, diminuindo consideravelmente a viabilidade celular e afetando suas curvas de crescimento. Aquelas liberadas pelo ácido ortofosfórico a 37% provocam diminuição da viabilidade somente nos primeiros momentos do contato com as células, enquanto as substâncias liberadas durante a presa do hidróxido de cálcio não são citotóxicas em nenhum momento. / The purpose of the present study was to evaluate the cytotoxic effects of substances leached during the use of direct pulp capping materials, on human pulp fibroblasts. There were used cell culture mediums conditioned by the test materials, as follows: group I: control (fresh medium without conditioning); group II: calcium hydroxide cement; group III: bonding system; group IV: 37% orthophosphoric acid. The medium conditioning was made, pouring the fresh conditioning medium on the materials, in order that its setting (group II), polymerization (group III) or the direct contact (group IV) would be able to leach substances to this culture medium. These conditioned mediums were put on the cells for the entire experiment, excepting the group IV, in which the mediums were put in contact with the cells for 15 seconds, following clinical recommendations. Cell counting was performed in hemocytometer, using the Trypan blue exclusion method, which mark only the dead cells. These counting was made at experimental times of 0, 6, 12 and 24 hours for the cell viability assay (short term), where it is evaluated the percentage of live cells on the total number of cells, and at experimental times of 1, 3, 5 and 7 days for the survival assay, in which is evaluated the absolute number of live cells. It was observed that the substances leached by the bonding system are cytotoxic at all experimental times, decreasing significantly the cell viability and affecting its growing rate. Those leached by the 37% orthophosphoric acid decreased the cell viability only at the first contact with the cells, and the substances leached during the setting of the calcium hydroxide cement are not cytotoxic.

As alterações pulpares e periapicais decorrentes de sobrecarga oclusal são problemas que acometem considerável parcela da população. Geralmente causadas por contato prematuro de restaurações com material em excesso, provocam um trauma de baixa intensidade, porém de alta frequência. Objetivo: Avaliar as alterações pulpares e periapicais, induzidas por esta sobrecarga oclusal associada ou não à doença periodontal induzida em dentes de ratas. Método: Foram utilizadas vinte ratas (Wistar) com noventa dias de idade e peso aproximado de 250g divididas em quatro grupos (n=5): GC (controle), GT (trauma), GL (ligadura) e GLT (ligadura e trauma), com sobrecarga oclusal e periodontite induzida restritas aos primeiros molares inferiores, que receberam a ligadura. As superfícies oclusais dos segundos e terceiros molares superiores receberam desgastes, sem que houvesse exposição pulpar e os animais foram observados por um período de quatro semanas. Depois de anestesiados, estes foram decapitados com uma guilhotina e, após a eutanásia, as hemi-mandíbulas foram enviadas para a análise histológica do tecido pulpar e periapical, com coloração por hematoxicilina-eosina, para avaliação da presença de nódulos pulpares, lacunas de reabsorção e células inflamatórias, além de submetidos à análise histoquímica pela Fosfatase Ácida Tartarato Resistente (TRAP) para avaliação da presença de células clásticas e pré-clásticas. Resultados: Histologicamente, os dentes submetidos à ligadura apresentaram presença moderada de fibras e vasos, enquanto os submetidos ao trauma e ao conjunto trauma-ligadura apresentaram grande aumento de vasos e fibras além de infiltrado neutrofílico e, por vezes, nódulos. Histoquimicamente, notou-se certa normalidade no GC, uma irregularidade na região cervical externa no GL e maior presença de lacunas dentinárias e células multinucleadas no GT e GLT. Conclusão: A sobrecarga oclusal provoca alterações pulpares e periapicais assim como a doença periodontal induzida. Mas a associação da doença periodontal e da sobrecarga é mais lesiva aos tecidos pulpares e radiculares. / Pulp and periapical changes resulting from occlusal overload are problems that affect a considerable portion of the population. Generally caused by premature contact of restorations with material in excess, cause a trauma of low intensity, however high frequency. Objective: To evaluate the changes of pulpal and periapical induced by this overload occlusal associated or not to the induced periodontal disease in teeth of rats. Method: Used twenty rats (Wistar) with ninety days old and weighing approximately 250g divided into four groups (n = 5): GC (control), GT (overload), GL (ligature) and GLT (ligature and overload) with occlusal overload and induced periodontitis restricted to lower first molars, which received the ligature. The occlusal surfaces of upper second and third molars were worn away, with no pulp exposure and the animals were observed for a period of four weeks. Once anesthetized, they were beheaded with a guillotine and, after euthanasia, the hemi-mandibles were sent for histological analysis of pulp and periapical with hematoxylin-eosin staining for evaluation of nodules pulp, resorption gaps and inflammatory cells, and analyzed by immunohistochemistry Tartrate Resistant Acid Phosphatase (TRAP) for assessing the presence of clastic cells and pre-clastic. Results: Histologically, the teeth underwent to ligature showed the presence of fibers and vessels, while the other submitted to overload and overload-ligature group showed a large increase in vessels and fibers as well as neutrophil infiltration and sometimes nodules. Histochemically, it was noted some degree of normality in the GC, an irregularity in the external cervical region in the GL and greater presence of dentinal gaps and multinucleated cells in the GT and GLT. Conclusion: The occlusal overload causes changes pulp and periapical as well as induced periodontal disease. But the association between periodontal disease and the overload occusal is more damage to pulp and root tissues.

Indiana University-Purdue University Indianapolis (IUPUI) / The effects of hydroxyapatite in a DAP-loaded MC carrier on dental pulp stem cells Introduction: Regenerative endodontic procedures (REP) require disinfection techniques to eliminate bacteria from the infected immature root canal system and promote new growth of the pulp-dentin complex. Double antibiotic paste (DAP), a mixture of ciprofloxacin and metronidazole, has shown efficacy in doing so while minimizing cytotoxicity on dental pulp stem cells (DPSC). Stem cells, scaffolding, and growth factors are necessary in the maturation, proliferation, and differentiation of mesenchymal stem cells into the root canal system. Nano-hydroxyapatite (n-HA) has a history of biocompatibility and, in addition, has shown promising effects as a tissue bioengineering material. Objective: The aim of this in vitro study was to investigate the proliferation and mineralization of DPSC in the presence of 1% DAP and methylcellulose (MC) with varying concentrations of nano-hydroxyapatite. Materials and Methods: DPSC were plated in 24-well plates containing culture media. The next day, semi-permeable 0.1 mm Transwell chambers were inserted into the wells to separate the reservoirs for medicaments. Treatment paste composed of methylcellulose containing 1% DAP with either 0.25%, 0.50%, or 1.0% nano-hydroxyapatite was added along with culture media. Methylcellulose alone and calcium hydroxide (Ultracal) were used as control groups. After 3 days, cells were evaluated for cytotoxic effects using an MTS proliferation assay (n = 10, in triplicate). DPSCs were also cultured with these medicaments for 7 days in osteogenic media and evaluated for alkaline phosphatase (ALP) activity and mineralization activity (n = 13, in triplicate). Comparisons between groups for differences in mineralization, BSA, and ALP activity were performed using analysis of variance (ANOVA), with different variances allowed for each group and a random effect included in the model to account for correlation within each of the three trials. A simulation-based model was used to adjust for multiple comparisons. Results: Addition of n-HA treatment groups increased mineralization significantly greater than calcium hydroxide, with MC alone and MC+DAP+0.5% HA providing the greatest effect. Regarding ALP, all HA concentrations performed significantly greater than MC and DAP concentrations. Proliferation demonstrated similar metabolic activity in all experimental groups with few comparisons significant. Conclusion: The challenge in REPs is to maintain survival, and preferably promote the proliferation and development of DPSCs into the pulp-dentin complex with a consistent treatment outcome. The combination of DAP with hydroxyapatite may allow for both disinfection and improved mineralization and cellular differentiation. This contribution has shown significant ability to increase stem cell differentiation into an osteogenic lineage as well as calcium deposition, indicating end goal results of regenerative procedures.

Indiana University-Purdue University Indianapolis (IUPUI) / Introduction: Profound pulpal anesthesia is not always adequate in mandibular teeth after the administration of local anesthesia, especially in the presence of irreversible pulpitis. Failure to achieve anesthesia has been seen in 30–80% of patients in teeth with a diagnosis of irreversible pulpitis. Onpharma™ developed an FDA-approved device that uses sodium bicarbonate to buffer a standard local anesthetic (LA) solution so that its pH may become closer to its pKa. Claims have been made that buffering a local anesthetic increases the anesthetic’s effect. Previous studies on the anesthetic efficacy of Onpharma’s Onset buffering system were inconclusive and may be dependent on the techniques used. Objectives: The aim of this study is to determine whether a buffered local anesthetic can lead to more profound and faster pulpal anesthesia in mandibular molars diagnosed with symptomatic irreversible pulpitis as compared to a standard, unbuffered local anesthetic. Materials and Methods: 40 total subjects completed the study. Screened and eligible subjects with a mandibular molar diagnosed with symptomatic irreversible pulpitis were randomly allocated into 2 groups so 1 group received a total of 3 cartridges of a standard, unbuffered 2% lidocaine with 1:100,000 epinephrine via inferior alveolar nerve block (IANB) followed by supplemental buccal and lingual infiltrations, while the other received the equivalent yet buffered formulation. An electronic pulp tester (EPT) was used to objectively determine baseline pulpal status of the affected tooth, followed by 2-minute interval testing following the administration of all local anesthesia. The onset of pulpal anesthesia was defined by the first of 2 consecutive EPT=80 readings, and the endodontic treatment could begin. Profound pulpal anesthesia was ultimately determined if the patient reported a comfortable pulpotomy as reflected on the Wong-Baker FACES Visual Analog Scale. Null Hypothesis 1: Subjects possessing mandibular molars diagnosed with symptomatic irreversible pulpitis will not achieve pulpal anesthesia more profoundly using buffered 2% lidocaine w/ 1:100,000 epinephrine in comparison to the standard, unbuffered anesthetic formulation. Null hypothesis 2: Subjects possessing mandibular molars diagnosed with symptomatic irreversible pulpitis will not achieve pulpal anesthesia faster using buffered 2% lidocaine w/ 1:100,000 epinephrine in comparison to the standard, unbuffered anesthetic formulation. Results: We observed a local anesthetic success rate of 45% in the buffered group, 70% in the unbuffered group, and ultimately 57.5% between both groups. The findings further indicate that the VAS scores after pulpotomy is significantly different between the 2 groups (p=0.019), with the unbuffered group having a more profound mean VAS score of 1.2 (as opposed to a buffered mean of 3.1). Regarding the time of onset for pulpal anesthesia, there was no statistically significant difference noted between the buffered and unbuffered groups. Conclusion: Based on the findings of this study, the null hypothesis 1 cannot be rejected since unbuffered 2% lidocaine with 1:100,000 epinephrine had a statistically significant increase in profound pulpal anesthesia compared to the buffered equivalent. The null hypothesis 2 cannot be rejected since there was no evidence of a significant difference in the time to pulpal anesthesia between the buffered and unbuffered groups.

Die Entdeckung adulter dentaler Stammzellen eröffnete ein neues Forschungsfeld im Hinblick auf die Regeneration dentaler Gewebe. Bisher liegen nur wenige Studien vor, in denen das Genexpressionsprofil dentaler Stammzellen im Vergleich zu den Knochenmarkstammzellen analysiert wurde. Diese Untersuchungen wurden vorwiegend an Mischkulturen vorgenommen. Im Gegensatz dazu war es daher das Ziel der vorliegenden Arbeit, das Genexpressionsprofil einer bestimmten Stammzell-Population, nämlich der Stro1-positiven pulpalen mesenchymalen Zahnkeimstammzellen (Stro1+ZK) im Vergleich zu Stro1-positiven mesenchymalen Knochenmarkstammzellen (Stro1+BK), zu untersuchen. Die Genexpression beider Zelltypen wurde anhand von Microarrays ermittelt. Insgesamt gingen 22.454 Gene in die Auswertung ein, wovon bei einem konservativ festgesetzten Schwellenwert einer FDR≤1% 2730 Gene eine hochsignifikant differentielle Expression zeigten. Die Analyse dieser differentiell exprimierten Gene mithilfe der Programme „DAVID“ und „Ingenuity“ ergab, dass in den Stro1+ZK vermehrt Gene heraufreguliert sind, die mit Zellfunktionen wie beispielsweise Proliferationsregulation, der Zell-zu-Zell-Signalleitung und der Organisation des Zytoskeletts verknüpft sind. Die Stro1+BK hingegen exprimieren verstärkt Gene, die mit der Organisation der extrazellulären Knochenmatrix und Zell-Adhäsion assoziiert sind. Des Weiteren findet sich in diesen Zellen eine verstärkte Expression von Genen, die mit der Struktur- und Formgebung des Skeletts in Verbindung stehen. Trotz identischem Stammzellmarker-Typus (Stro1) weisen die untersuchten mesenchymalen Stammzelltypen stark unterschiedliche Hox-Gen-Signaturen auf. Dabei zeigt sich sowohl eine Variation in Anzahl und Art der Hox-Gene als auch in deren Expressionsmuster. Stro1+BK exprimieren verstärkter Hox-Gene der Cluster A bis D (HOXA-D), die Segment- und Positionsinformationen codieren. Hingegen sind in den Stro1+ZK die Hox-Gene BARX1, MSX1, MSX2, DLX1, DLX2, PAX9 und LEF1 hochreguliert, welche eine tragende Rolle in der Zahnentwicklung spielen. So ist z.B. bereits bekannt, dass Mutationen dieser Gene zu Fehlbildungen von Zähnen wie Aplasien oder Hypoplasien führen können. Die vorliegende Arbeit zeigt, dass insbesondere hinsichtlich der Hox-Gene signifikante Unterschiede zwischen den Stro1+ZK und Stro1+BK bestehen. Weiterführende Experimente zur Aufklärung der Funktionsweise von Genen, die in den Stro1+ZK von Bedeutung sein könnten, einschließlich deren Erforschung auf proteinbiochemischer und zellbiologischer Ebene, wären wünschenswert.

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published_or_final_version / Dentistry / Master / Master of Dental Surgery

published_or_final_version / abstract / Dentistry / Master / Master of Dental Surgery