Introdução: Doador de rim em vida é uma importante fonte de órgão para os pacientes portadores de doença renal crônica (DRC). Os doadores experimentam uma redução abrupta da taxa de filtração glomerular (TFG) e adaptações ao metabolismo mineral demandam estudos nesta população. Nós avaliamos prospectivamente esta adaptação em doadores de rim em vida. Métodos: Entre janeiro de 2010 a agosto de 2011, no hospital das Clínicas de São Paulo e na Universidade de Miami, realizamos a avaliação prospectiva do metabolismo mineral e da função renal por 1 ano em 74 doadores de rim em vida. Medimos a taxa de filtração glomerular (TFG), fósforo (Pi), cálcio (Ca), paratohormônio (PTH), fibroblast Growth Factor 23 (FGF23) e a fração de excreção do fósforo (FePO4) no pré-operatório e nos dias 1, 2, 14, 180 e 360 do pós-operatório. Resultados: Observamos uma redução, aproximadamente, de 40% da TFG nos dois primeiros dias após a cirurgia. No décimo quarto dia após a nefrectomia, observamos o início da recuperação da TFG, chegando ao máximo da recuperação com 1 ano, quando se atingiu 68,6% da função renal se comparado com o dia anterior a doação (75,3 ml/min/1,73m2, p < 0,001). O cálcio sérico apresentou seu nadir no dia 1 (7,99 mg/dL; p < 0,01) e o Pi sérico atingiu seu nadir no dia 2 (2,61 mg/dL; p < 0,01). Já no dia 14, os valores de Ca e Pi retornaram aos valores basais tendo o fósforo evoluído novamente com valores inferiores ao basal no último dia de seguimento (3,36mg/dL; p < 0,001). FGF23 e PTH apresentaram elevação no D1 (111,0144,6 percentil 25-75: 16-63 RU/ml 64,9 30,3pg/mL; p < 0,01). Os valores de FGF23 se mantiveram elevados até o final do estudo enquanto que o PTH retornou aos valores de base no segundo dia e, a partir de então, manteve sem diferença do valores basais até o último dia de estudo. FePO4 elevou de 11,45,2% para 15,28,1% entre o pré-operatório e D365 (p < 0,01). Conclusão: A nefrectomia para doação de rim em 74 pacientes saudáveis elevou os valores de FGF23 durante todo o estudo juntamente com a FePO4. O fósforo, cálcio e PTH séricos apresentaram queda nos seus valores na primeira semana após a nefrectomia, e, com duas semanas após a cirurgia, retornaram aos valores basais mantendo-se estáveis até o final do estudo / Introduction: Living kidney donors (LKDs) experience an abrupt decline in glomerular filtration rate (GFR). Mineral metabolism adaptations in early CKD are still debated and not well studied in LKDs. We prospectively studied acute and long term mineral metabolism adaptation of LKDs. Materials and Methods: We measured renal function and mineral metabolites longitudinally for 1 year (days (D) 1, 2, 14, 180, & 365 post-operatively) in 74 healthy individuals who underwent kidney live donation. Results: eGFR (MDRD) decreased to 59% of its baseline on day 2 and started to increase at day 3, to its maximum at day 360 (75.3±15.6 ml/min/1.73m2, p < 0.01) wile FGF23 increased from 60.6 (25th-75th percentile 19-81 RU/mL) at baseline to 111.0±144.6 (p < 0.01) on day 1 and keep higher than baseline throwout the study. PTH rose maximally on day 1 (64.9 ± 30.3pg/ml) and returned to its base line on D2 and did not change after that. Total serum Calcium levels decreased from 9,40±0,48 mg/dL to a nadir of 7.99±0,51 mg/dL on day 1 (p < 0.001). Serum Phosphate levels reached their nadir on day 2 (2.61±0,52 mg/dL; p < 0.01). At D14 total calcium and phosphate levels had returned to baseline, but phosphate levels returned down on D360 (3.36±0,52 mg/dL; p < 0.001). Phosphate excretion fraction (FePO4) increased from base line (11.4±5.2%) up to 15.2±8.1% until D360 (p < 0.001). Conclusions: Abrupt reduction in eGFR induces physiological increases in FGF23 and PTH, and decreases in serum Ca and Pi in the first week. The changes in FGF23 and Pi urinary fractional excretion of Pi remain modestly yet significantly different from baseline throughout the first year after nephrectomy. Wile Ca, PTH and Pi serum levels are not significantly different from the baseline

A terapia com laser em baixa intensidade, conhecida como Low-intensity laser therapy (LILT), tem sido alvo de pesquisas que buscam esclarecer os mecanismos pelos quais o laser age na cicatrização de feridas. Este processo é dependente da transmissão de sinais entre epitélio e conjuntivo, os quais influenciam a proliferação e a migração celular, onde participam fatores de crescimento. O objetivo desta pesquisa foi verificar os efeitos da radiação com lasers vermelho e infravermelho em baixa intensidade na produção de fatores de crescimento por fibroblastos in vitro. Foi obtida uma cultura primária de fibroblastos gengivais humanos (linhagem FGH). As células foram cultivadas em placas de 96 poços (5 x 103 células /poço), em estado de quiescência (meio de cultura suplementado com 1% de soro fetal bovino) e irradiadas com lasers de diodo (AlGaAs - 660nm, e AlGaInP - 780nm). O laser em modo contínuo, foi aplicado em contato e na forma pontual. A potência utilizada foi de 40mW numa área de 0,042cm2 e com densidades de energia de 2, 3, 4, 5 e 6J/cm² com respectivos tempos de aplicação de 2, 3, 4, 5 e 6s. As culturas foram irradiadas duas vezes com 6h de intervalo. Os grupos controle positivo e negativo foram cultivados com 10% e 1% de soro fetal bovino, respectivamente e não receberam irradiação. A produção do fator de crescimento de queratinócitos (KGF) e do fator de crescimento de fibroblastos básico (bFGF) foi analisada e quantificada por ensaio imunoenzimático (método ELISA) no meio condicionado pelas células. Os dados foram comparados estatisticamente pela análise de variância e teste de Kruskal-Wallis complementados pelos testes de Tukey e Dunn, respectivamente (p<0,05). Culturas de todos os grupos produziram KGF e bFGF. Controles positivos produziram maior quantidade de KGF e bFGF que controles negativos. A produção de KGF foi similar em todos os grupos experimentais. A produção de bFGF foi significativamente maior nos grupos tratados com o laser infravermelho nas densidades de energia de 3J/cm2 e 5J/cm2 quando comparados com os controles. Baseados nas condições deste estudo, concluiu-se que a LILT não influencia a produção de KGF, porém, influencia positivamente a produção de bFGF de maneira dependente do comprimento de onda e da densidade de energia empregados. O infravermelho, nas densidades de energia de 3J/cm2 e 5J/cm2, é capaz de aumentar a produção deste fator de crescimento. Portanto, a utilização da LILT nestes parâmetros poderá constituir papel relevante na cicatrização de feridas. / Low-intensity laser therapy (LILT) has been the subject of researches to understand the mechanisms by which the laser acts stimulating wound healing. This process is dependent of a signaling transfer between epithelium and connective tissue and it influences cellular proliferation and migration. Growth factors are directly involved in these mechanisms. The aim of this study was to analyze the effects of red and infrared low-level laser on production of fibroblast growth factors by human gingival fibroblasts in vitro. The cells were obtained by primary culture (FGH cell line). They were grown in 96 wells plates (5x10³ cells / well) in a quiescence environment (1% fetal bovine serum) and then they were irradiated with a diode laser (GaAlAs - 660nm, and InGaAlP - 780nm). The continuous laser was applied in contact and in a punctual mode. The power was 40mW, the area of irradiation was 0,042cm2, the energy densities were 2, 3, 4, 5 and 6J/cm² with 2, 3, 4, 5 and 6s of time application, respectively. The cultures were irradiated twice with 6 h-interval. The positive and negative control groups were grown in 10% and 1% of bovine fetal serum, respectively and didn?t receive any irradiation. The production of keratinocyte growth factor (KGF) and basic fibroblast growth factor (bFGF) was analyzed and quantified by immunoenzymatic test (ELISA) in the conditioned medium. The data were compared statistically by analysis of variance and Kruskal-Wallis, complemented by Tukey?s and Dunn?s tests, respectively (p<0,05). Positive controls produced more KGF and bFGF than negative controls. The KGF production was similar in all experimental groups. The bFGF production was significantly higher in groups treated with infrared laser with energy densities of 3J/cm² and 5J/cm² when compared to controls. Based on these study conditions, authors concluded that LILT doesn?t influence KGF production although it positively influences bFGF production in a dependent manner from wavelength and energy density. Infrared laser with energy densities of 3J/cm² and 5J/cm² is capable of improving the production of this growth factor. Therefore the LILT in these parameters can constitute a relevant role on wound healing.

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.

Rupture of the anterior cruciate ligament (ACL) of the knee leads to chronic joint instability and reduced range of motion while the long term results are marred by a high prevalence of degenerative joint disease especially osteoarthritis. Bundles of collagen threads have been widely investigated for the repair of torn ACL, but are limited by insufficient tissue ingrowth to repopulate and completely regenerate these grafts. We have developed a novel in vitro method of characterizing fiber-based thread matrices by probing their ability to promote tissue ingrowth from a wound margin as a measure of their ability to promote repopulation and regeneration. This method is useful in the optimization of thread scaffolds, and is sensitive enough to distinguish between subtle variations in biopolymer chemistry and organization. Furthermore, this method was used to characterize the effects of crosslinking on the cell outgrowth and correlated the findings with the mechanical properties of collagen threads. The results suggest that crosslinking is required to achieve sufficient mechanical properties for high stress applications such as ACL replacement, but regardless of technique, crosslinking attenuated the cell outgrowth properties of the threads. To improve the regenerative capacity of these scaffolds, novel fibrin microthread matrices were developed with a similar morphology to collagen threads and sufficient mechanical strength to be incorporated in composite thread scaffold systems. These fibrin microthreads were loaded with FGF-2, a potent mitogen and chemotactic agent that works synergistically with fibrin in regulating cell signaling and gene expression. Increases in fibroblast migration and proliferation in FGF-2-loaded fibrin threads were successfully demonstrated with the concomitant promotion of oriented, aligned, spindle-like fibroblast morphology. These results suggest that fibrin-FGF-2 microthreads have distinct advantages as a biomaterial for the rapid regeneration of injured tissues such as the ACL.

Fibroblast growth factor 23 (FGF-23) has recently been shown to be involved in phosphate regulation and bone mineralization. This study evaluated the effect of FGF-23 on three human cell lines (Caco-2, HK-2, SaOS-2) representing three different sites of phosphate regulation (small intestine, kidney proximal tubules, and bone, respectively). FGF-23 induced gene expression was studied using Clontech human Atlas glass microarrays containing various assortments of genes and by a custom designed oligo microarray containing specific genes selected for their biological relevance to FGF-23's potential function. FGF-23 induced differential gene expression in all three cell types, suggesting that FGF-23 may be capable of acting on these three primary sites of phosphate regulation. Human small intestine-like endothelial cell line, Caco-2, showed upregulation of several genes including parathyroid hormone receptors 1 and 2. FGF-23 inhibited the expression of water channel transporters aquaporin 5 and 6 in human osteoblast-like SaOS-2 cells while upregulating aquaporin expression in HK-2 cells. Somatostatin receptors 1-4 were identified to be upregulated in the human kidney, HK-2 cell line. Mucin 2, a gene that is linked to abnormal cellular growth, was consistently induced by FGF-23 in all three cell lines. Families of aquaporins, somatostatins, parathyroid hormones, and other identified differentially expressed genes are involved in different signaling pathways that are associated with phosphate and calcium regulation. Selected candidates were analyzed further by real-time RT-PCR. These data support FGF-23 induced regulation of aquaporin 5 mRNA in HK-2 cells and 1-alpha-hydroxylase mRNA in Caco cells. FGF-23 induced changes in mRNA analysis of four additional genes was less than two-fold in triplicate analysis of selected samples. Taken together, these results suggest that each cell type may have responded to FGF-23, but additional validation of the array data set will be required to identify those genes specifically regulated by FGF-23. Further refinement of this data set will undoubtedly uncover additional functions of FGF-23 and may provide valuable insight into designing therapeutic approaches for phosphate specific disorders.

Um dos fatores que regula o córtex adrenal é o hormônio adrenocorticotrópico, ACTH, no entanto, o fator de crescimento de fibroblastos do tipo 2, FGF2, e os peptídeos N-terminais da pro-opiomelanocortina, N-POMC, também podem estar envolvidos. As vias de sinalização das proteínas quinases: ERK, JNK e p38, juntamente com outras vias como PKA, PKC e PI3K/Akt são importantes para a definição trófica das células. Nós analisamos a importância destas vias de sinalização e sua influência na viabilidade, proliferação e morte celular, induzidas pelo ACTH, FGF2 e N-POMC, utilizando inibidores farmacológicos e moleculares em culturas primárias de células adrenocorticais, células glomerulosa e fasciculadas/reticulares. Nossos resultados mostram que as vias mediadoras envolvidas na resposta proliferativa do FGF2 e da N-POMC são, respectivamente, as vias ERK/JNK e ERK/JNK/Akt. Por outro lado, a resposta pró-apoptótica promovida pelo ACTH é mediada pela via p38, provavelmente associada à ausência de ativação das vias relacionadas com a sobrevivência, como as vias ERK e JNK. / One of the factors that regulate adrenal cortex is the adrenocorticotrophic hormone, ACTH, however, the fibroblast growth factor type 2, FGF2) and pro-opiomelanocortin N-terminal peptides, N-POMC, might also be involved. The mitogen-activated protein kinase pathways: ERK, JNK and p38, together with other signaling pathways such as PKA, PKC and PI3K/Akt are important for cells trophic definition. We analyze the importance of these pathways and their influence in viability, proliferation and cell death stimulated by ACTH, FGF2 and N-POMC, using pharmacological and molecular inhibitors in primary culture of adrenocortical cells, glomerulosa and fasciculata/reticularis cells. Our results show that the mediating signaling pathways involved in FGF2 and N-POMC proliferative effects are, respectively, ERK/JNK and ERK/JNK/Akt. On the other hand, the pro-apoptotic response promoted by ACTH is through p38 signaling, probably associated with the absence of activation of other pathways involved with cell survival, like ERK and JNK.

Le cheval est souvent victime de plaies traumatiques, dont la guérison est fréquemment problématique, et ce, principalement quand la plaie survient sur le membre. Il est courant de voir chez le cheval le développement d’un tissu de granulation exubérant ou « bouton de chair », qui mène à une cicatrisation excessive due à la surproduction de tissu fibreux. Ce tissu cicatriciel, non épithélialisé, est caractérisé par une occlusion au niveau de la microcirculation due à l’hypertrophie des cellules endothéliales, qui laisse supposer la présence d’hypoxie tissulaire. Une hypoxie relative a effectivement été mesurée par spectroscopie dans le proche infrarouge au niveau des plaies appendiculaires prédisposées au développement de tissu de granulation exubérant, par rapport aux plaies corporelles. De plus, une étude thermographique a révélé un patron spatial similaire de la perfusion. Au niveau moléculaire, la littérature rapporte que le facteur de transcription «hypoxia inducible factor» (HIF) est à l’origine de plusieurs changements dans les niveaux d’expression de divers gènes régulés par l’hypoxie. L’objectif du présent projet de recherche était de définir la contribution de l’hypoxie à la guérison cutanée chez le cheval. Le premier volet (in vivo) du projet visait à mesurer l’expression protéique temporelle du HIF1A dans des échantillons tissulaires en provenance de plaies cutanées guérissant normalement et d’autres développant une cicatrisation excessive, selon divers sites anatomiques (tronc, membre). Les résultats obtenus suggèrent que la mesure de HIF1A, dans les échantillons pluricellulaires de cette étude, reflète l’épithélialisation de la plaie plutôt que les niveaux d’oxygène tissulaire. En effet, le HIF1A semble réguler l’homéostasie et la prolifération des kératinocytes. Le second volet (in vitro), consistait en la mise en culture de fibroblastes dermiques équins provenant du tronc ou du membre, en condition de normoxie ou d’hypoxie (à 1% d’O2 ou à l’aide d’un mimétique, le CoCl2) afin d’en étudier le comportement (capacités de prolifération et de synthèse protéique). Les résultats obtenus soutiennent une contribution de l’hypoxie à la cicatrisation extensive chez le cheval puisque l’hypoxie favorise la prolifération des fibroblastes en plus d’encourager la synthèse de collagène de type 1 et de diminuer la synthèse de la métalloprotéinase de type 2. Les changements observés semblent dépendre de facteurs extrinsèques (environnementaux) car les fibroblastes dermiques se comportent de façon similaire indépendamment de la provenance anatomique. En somme, les deux volets de l’étude ont permis d’élucider une part des mécanismes sous-jacents à la formation du tissu de granulation exubérant lors de guérison cutanée chez le cheval. La poursuite des recherches dans ce domaine mènera à une meilleure compréhension de la pathologie et ainsi, permettra de développer des méthodes de traitement spécifiques à la condition. / The horse is often victim of traumatic wounds for which healing can be problematic, mainly when the wound occurs on the limb. The development of exuberant granulation tissue, also known clinically as “proud flesh”, leads to extensive scarring characterized by overproduction of fibrous tissue and the absence of an epithelial cover. This scar tissue suffers from occlusion of the microcirculation within the residual granulation tissue, due to endothelial cell hypertrophy, suggesting tissue hypoxia. The presence of relative hypoxia in limb wounds of horses was recently confirmed using near infrared spectroscopy. Additionally, thermography showed decreased perfusion in limb wounds. Abundant literature incriminates "hypoxia inducible factor” (HIF) in the regulation of expression of a number of genes in response to hypoxia. The overall objective of this research project was to define the contribution of hypoxia to problematic wound healing in horses. The first phase of the project (in vivo) aimed to measure the temporal expression of HIF1A protein in tissue samples taken from skin wounds healing normally and others developing exuberant granulation tissue and subsequent extensive scarring, according to various anatomical sites (trunk, limb). Results suggest that the expression of HIF1A within the pluricellular tissue samples of this study reflects wound epithelialization rather than tissue oxygen levels. Indeed, HIF1A appears to regulate the homeostasis and proliferation of skin keratinocytes. The second phase of the project (in vitro) consisted in the culture of equine dermal fibroblasts from the body or the limb, under conditions of normoxia or hypoxia (1% O2 or using a mimetic, CoCl2) in an effort to study their behavior (proliferation and protein synthesis). Results corroborate the contribution of hypoxia to over-scarring in the horse since hypoxia promotes both the proliferation of dermal fibroblasts and the synthesis of collagen type 1 while decreasing the synthesis of matrix metalloproteinase 2. Extrinsic factors (environmental) appear to govern the behavior of equine dermal fibroblasts since these cells behave similarly regardless of their anatomic origin (body or limb). In summary, the two phases of the study contributed to the elucidation of a portion of the mechanisms underlying the development of exuberant granulation tissue during wound healing in horses. Further research in this area will provide a better understanding of the pathology and thus aid in the design and development of targeted therapies.

INTRODUÇÃO: Tumores indutores de osteomalácia (TIOs) são raros, geralmente apresentam origem mesenquimal, têm produção excessiva de fosfatoninas sendo a mais comum o FGF23 (Fibroblast Growth Factor 23) que, em níveis elevados, provoca osteomalácia hipofosfatêmica. A cura dos TIOs envolve a remoção completa do tumor, o que torna essencial sua localização. OBJETIVOS: (1) caracterizar nove pacientes com TIO ao diagnóstico e avaliá-los evolutivamente em longo prazo; (2) avaliar a eficácia da cintilografia com Octreotida (Octreoscan®) e a da cintilografia de corpo inteiro com Mibi (MIBI) na detecção dos TIOs. MÉTODOS: O acompanhamento dos pacientes consistiu na avaliação clínica, na avaliação laboratorial com ênfase no metabolismo ósseo e na realização de exames de imagem para caracterização das deformidades esqueléticas. Para a localização dos TIOs, os pacientes foram submetidos a exames de Octreoscan®, MIBI, ressonância magnética (RM) e tomografia computadorizada (TC). RESULTADOS: O período de observação dos pacientes variou de dois a 25 anos. Ao diagnóstico, todos exibiam fraqueza muscular, dores ósseas e fraturas de fragilidade. Em relação à avaliação laboratorial, apresentavam: hipofosfatemia com taxa de reabsorção tubular de fosfato reduzida, fosfatase alcalina aumentada e níveis elevados de FGF23. O Octreoscan® permitiu a identificação dos TIOs nos nove pacientes e o MIBI possibilitou a localização dos TIOs em seis pacientes, sendo que ambos os exames foram concordantes entre si e com os exames topográficos (RM ou TC). Os achados histopatológicos das lesões dos nove pacientes confirmaram tratar-se de oito tumores mesenquimais fosfatúricos (PMTs) benignos e um PMT maligno. Após a primeira intervenção cirúrgica para a remoção dos TIOs, quatro pacientes encontram-se em remissão da doença e cinco evoluíram com persistência tumoral. Dos cinco, quatro foram reoperados e um aguarda nova cirurgia. Dos que foram reoperados, um paciente se mantém em remissão da doença, um foi a óbito por complicações clínicas, uma teve doença metastática e o último apresentou recidiva tumoral três anos após a segunda cirurgia. Deformidades ósseas graves foram observadas nos pacientes cujo diagnóstico e/ou tratamento clínico foram tardios. O tratamento da osteomalácia foi iniciado com fosfato e perdurou até a ressecção tumoral, tendo sido reintroduzido nos casos de persistência/recidiva tumoral. Quatro pacientes que fizerem uso regular desse medicamento por mais de seis anos evoluíram com hiperparatireoidismo terciário (HPT). CONCLUSÕES: O estudo revelou que tanto o Octreoscan® como o MIBI foram capazes de localizar os TIOs. Por isso, incentivamos a realização do MIBI nos locais onde o Octreoscan® não for disponível. Uma equipe experiente é indispensável para o sucesso cirúrgico visto que os tumores, embora benignos, costumam ser infiltrativos. Recomendamos o seguimento por tempo indeterminado em função do risco de recidiva tumoral. Assim como o FGF23, consideramos o fósforo um excelente marcador de remissão, persistência e recidiva dos TIOs. O diagnóstico e o tratamento precoce são fundamentais para a melhora dos sintomas podendo minimizar as deformidades esqueléticas e as sequelas ósseas. O uso prolongado do fosfato no tratamento da osteomalácia hipofosfatêmica foi associado ao desenvolvimento do HPT / BACKGROUND: Tumor-induced osteomalacia (TIO) is rare. The tumor usually has mesenchymal origin and produces excessive phosphatonins, most commonly FGF23 (Fibroblast Growth Factor 23), which at high levels causes hyphophostatemic osteomalacia. The cure for TIO is achieved through complete removal of the tumor. It is therefore essential identify its location. OBJECTIVES: (1) to characterize nine patients with TIO at diagnosis and to evaluate their follow-up over a long-term period; (2) to evaluate the efficacy of whole-body scintigraphy 111In-octreotide (Octreoscan®) and 99mTc-sestamibi (MIBI) in TIO detection. METHODS: Evaluations consisted of clinical and laboratory testing of bone metabolism and imaging to characterize skeletal deformities. To locate TIO, patients underwent Octreoscan®, MIBI, magnetic resonance (MRI), and computed tomography (TC). RESULTS: Patients were followed-up from two to 25 years. At diagnosis, all patients presented with muscle weakness, bone pain and fragility fractures. Laboratorial evaluation revealed hypophosphatemia with reduced tubular reabsorption of phosphate, increased alkaline phosphatase, and high levels of FGF23. TIO was identified in nine patients through Octreoscan® and in six patients through MIBI. Results of both types of scintigraphies matched one another as well with topographic examination (MR or CT). Histopathological findings of the lesions in the nine patients confirmed the existence of eight benign phosphaturic mesenchymal tumors (PMTs) and one malign PMT. After the first surgery for tumor resection, four patients were in remission, whereas five revealed tumoral persistence. Four of the latter five were re-operated, and one is still waiting for another surgery. Of those four patients, one became in remission, one died of clinical complications, one disclosed metastatic disease, and the last one had tumoral recurrence three years after the second surgery. Severe bone deformations were observed in patients whose diagnosis and/or clinical treatment were delayed. Osteomalacia treatment was initiated with oral phosphate, which continued until tumor resection. In case of tumor persistence or recurrence, oral phosphate was reintroduced. Four patients treated with this medication regularly for six years or more developed tertiary hyperparathyroidism (HPT). CONCLUSIONS: The present study revealed that Octreoscan® and MIBI were able to locating TIO. Therefore, we suggest that MIBI should be encouraged in places where Octreoscan® is not available. An expert team of surgeons is essential to the success of TIO\'s treatment, because of their infiltrative, albeit benign nature. Long-term follow-up is important due to the risk of tumor recurrence. Along with FGF23, phosphorous was considered an excellent hallmarker of TIO remission, persistence and recurrence. Early diagnosis and treatment are essential to improve symptoms and minimize skeletal deformities and skeletal disabilities. Long-term treatment of osteomalacia with oral phosphate was associated with the development of HPT

A terapia com laser em baixa intensidade, conhecida como Low-intensity laser therapy (LILT), tem sido alvo de pesquisas que buscam esclarecer os mecanismos pelos quais o laser age na cicatrização de feridas. Este processo é dependente da transmissão de sinais entre epitélio e conjuntivo, os quais influenciam a proliferação e a migração celular, onde participam fatores de crescimento. O objetivo desta pesquisa foi verificar os efeitos da radiação com lasers vermelho e infravermelho em baixa intensidade na produção de fatores de crescimento por fibroblastos in vitro. Foi obtida uma cultura primária de fibroblastos gengivais humanos (linhagem FGH). As células foram cultivadas em placas de 96 poços (5 x 103 células /poço), em estado de quiescência (meio de cultura suplementado com 1% de soro fetal bovino) e irradiadas com lasers de diodo (AlGaAs - 660nm, e AlGaInP - 780nm). O laser em modo contínuo, foi aplicado em contato e na forma pontual. A potência utilizada foi de 40mW numa área de 0,042cm2 e com densidades de energia de 2, 3, 4, 5 e 6J/cm² com respectivos tempos de aplicação de 2, 3, 4, 5 e 6s. As culturas foram irradiadas duas vezes com 6h de intervalo. Os grupos controle positivo e negativo foram cultivados com 10% e 1% de soro fetal bovino, respectivamente e não receberam irradiação. A produção do fator de crescimento de queratinócitos (KGF) e do fator de crescimento de fibroblastos básico (bFGF) foi analisada e quantificada por ensaio imunoenzimático (método ELISA) no meio condicionado pelas células. Os dados foram comparados estatisticamente pela análise de variância e teste de Kruskal-Wallis complementados pelos testes de Tukey e Dunn, respectivamente (p<0,05). Culturas de todos os grupos produziram KGF e bFGF. Controles positivos produziram maior quantidade de KGF e bFGF que controles negativos. A produção de KGF foi similar em todos os grupos experimentais. A produção de bFGF foi significativamente maior nos grupos tratados com o laser infravermelho nas densidades de energia de 3J/cm2 e 5J/cm2 quando comparados com os controles. Baseados nas condições deste estudo, concluiu-se que a LILT não influencia a produção de KGF, porém, influencia positivamente a produção de bFGF de maneira dependente do comprimento de onda e da densidade de energia empregados. O infravermelho, nas densidades de energia de 3J/cm2 e 5J/cm2, é capaz de aumentar a produção deste fator de crescimento. Portanto, a utilização da LILT nestes parâmetros poderá constituir papel relevante na cicatrização de feridas. / Low-intensity laser therapy (LILT) has been the subject of researches to understand the mechanisms by which the laser acts stimulating wound healing. This process is dependent of a signaling transfer between epithelium and connective tissue and it influences cellular proliferation and migration. Growth factors are directly involved in these mechanisms. The aim of this study was to analyze the effects of red and infrared low-level laser on production of fibroblast growth factors by human gingival fibroblasts in vitro. The cells were obtained by primary culture (FGH cell line). They were grown in 96 wells plates (5x10³ cells / well) in a quiescence environment (1% fetal bovine serum) and then they were irradiated with a diode laser (GaAlAs - 660nm, and InGaAlP - 780nm). The continuous laser was applied in contact and in a punctual mode. The power was 40mW, the area of irradiation was 0,042cm2, the energy densities were 2, 3, 4, 5 and 6J/cm² with 2, 3, 4, 5 and 6s of time application, respectively. The cultures were irradiated twice with 6 h-interval. The positive and negative control groups were grown in 10% and 1% of bovine fetal serum, respectively and didn?t receive any irradiation. The production of keratinocyte growth factor (KGF) and basic fibroblast growth factor (bFGF) was analyzed and quantified by immunoenzymatic test (ELISA) in the conditioned medium. The data were compared statistically by analysis of variance and Kruskal-Wallis, complemented by Tukey?s and Dunn?s tests, respectively (p<0,05). Positive controls produced more KGF and bFGF than negative controls. The KGF production was similar in all experimental groups. The bFGF production was significantly higher in groups treated with infrared laser with energy densities of 3J/cm² and 5J/cm² when compared to controls. Based on these study conditions, authors concluded that LILT doesn?t influence KGF production although it positively influences bFGF production in a dependent manner from wavelength and energy density. Infrared laser with energy densities of 3J/cm² and 5J/cm² is capable of improving the production of this growth factor. Therefore the LILT in these parameters can constitute a relevant role on wound healing.

C-ficocianina (C-PC) é um pigmento solúvel em água e está presente em Arthrospira platensis. Este pigmento é constituído por duas subunidades, &#945 e &#946, com massas moleculares de 16 e 17 kDa, respectivamente. Recentemente foi demonstrado que a subunidade &#946 da C-PC de Anabaena tem atividade antitumoral. Os principais objetivos deste estudo foram clonar e expressar a subunidade &#946 de C-PC de A. platensis em amostras de Escherichia coli e verificar a capacidade da proteína recombinante e da proteína C-PC em induzir ou não apoptose em células tumorais (HEp-2) e não tumorais (fibroblastos). As células foram tratadas com 100 µg das proteínas recombinante e C-PC por 6 h. Após tratamento, as células foram coradas com azul de toluidina (Metódo: Concentração Crítica de Eletrólitos - CCE). As células em apoptose foram detectadas pela coloração e por alterações morfológicas. Os resultados obtidos demonstram que ambas as proteínas, &#946-PC recombinante e C-PC, são capazes de induzir apotose em células HEp-2 e não induzi-la em células fibroblásticas. / C-phycocyanin (C-PC) is a water soluble pigment and is present in Arthrospira platensis. It is consisted of two subunits, &#945 and &#946, with molecular masses of 16 and 17 kDa, respectively. Recently it was shown that the &#946 subunit of C-PC Anabaena has antitumor activity. The main objectives of this study were cloning and expression the &#946 subunit of C-PC (&#946-PC) of A. platensis in Escherichia coli and check the ability of the C-PC and recombinant proteins in apoptotic induction in cancer (HEp-2) and non-cancer (fibroblasts) cells. The fibroblasts and HEp-2 cells were treated with 100 µg of recombinant and C-PC, proteins respectively for 6 h. After treatment, the cells were stained with toluidine blue (Critical Electrolyte Concentration Method - CEC). Apoptosis cells were detected by staining and morphological changes. The results show that the recombinant &#946-PC and C-PC proteins are able to induce apotosis in HEp-2 cells and dont induce it in fibroblasts cells.