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Objetivo: Avaliar a quantificação de armadilhas extracelulares de neutrófilos, TLR2 e TLR4 em crianças com doença falciforme, após estímulo com PMA. Metodologia: Este projeto foi aprovado pelo Comitê de Ética em Pesquisa do Hospital de Clínicas da Universidade Federal do Triângulo Mineiro (CEP – HC UFTM). O grupo de estudo foi composto por 25 crianças com idades de 1 a 13 anos, diagnosticadas com doença falciforme, atendidas no Ambulatório de Hematologia e Hemoterapia do Hemocentro Regional de Uberaba/MG. Foi coletado sangue periférico, por venopunção, em tubos contendo heparina. O isolamento de neutrófilos foi realizado por meio de centrifugação em gradiente de concentração por Ficoll-Paque™. Os neutrófilos isolados foram mantidos por 4 horas em incubadora úmida a 37°C em meio RPMI-1640 contendo 10% de soro fetal bovino, com e sem estímulo por PMA (phorbol myristate acetate). Posteriormente, as amostras foram marcadas com Sytox Green e CD66a para quantificação da netose; CD66a e marcador TLR2 para quantificar o receptor TLR2 e CD66a e marcador TLR4 para quantificar o receptor TLR4. As quantificações da netose e dos receptores foram realizadas por citometria de fluxo em citômetro FACSCanto II. A análise estatística foi realizada utilizando o programa GraphPad Prism e a significância estatística foi definida como p < 0,05. Resultados: O estudo foi composto por 13 (52%) crianças do sexo masculino e 12 (48%) feminino, com mediana de idade de 7 anos (1 – 13). Em relação aos genótipos ,: 11 (44%) crianças apresentavam genótipo HbSS, 10 (40%) genótipo HbSC e 4 (16%) outros genótipos. Houve diferença estatística (p = 0,0001) na avaliação da netose, sendo observada maior expressão no grupo de amostras tratadas com PMA em relação ao grupo de amostras não tratadas. Houve diferença estatística (p = 0,0015) na quantificação do receptor TLR2, sendo observada maior quantidade desse receptor no grupo de amostras tratadas com PMA em relação ao grupo de amostras não tratadas. Houve diferença estatística (p = 0,002) na quantificação do receptor TLR4, sendo observada maior quantidade de receptor TLR4 no grupo de amostras tratadas em relação ao grupo de amostras não tratadas. Discussão: A doença falciforme é um distúrbio hematológico caracterizado por um processo inflamatório crônico. Tem sido demonstrado que a adesão de células falciformes e neutrófilos ao endotélio colaboram com a vaso-oclusão observada em indivíduos com a doença. Estudos apontam que a alta atividade dos neutrófilos em processos infecciosos nestes pacientes, induz a formação de armadilhas extracelulares de neutrófilos (NETS), influenciando na iniciação e evolução de trombos, bem como a ativação de TLR2 e TLR4. A ativação desses dois receptores é capaz de produzir citocinas e moléculas de adesão importantes na inflamação, mas que de maneira exacerbada podem levar a vaso-oclusão. No presente estudo, foi observada maior expressão de netose, TLR2 e TLR4 nas amostras estimuladas pelo PMA. A literatura relata que o PMA induz notavelmente vias semelhantes a outros estímulos, sendo considerado um bom estímulo artificial para estudos com neutrófilos, conforme demonstrado no presente estudo. Conclusão: Os resultados desse estudo sugerem que o PMA é eficaz no papel de estimulador de netose, TLR2 e TLR4.

Objetivos: Atualmente, o emprego de inibidores de tirosina quinase (TKIs) constitui uma importante opção terapêutica para neoplasias hematológicas. O TKI-258 é um inibidor multi-alvos em fase de estudos que apresenta potencial como quimioterápico para diversos tipos de neoplasias. O índice de seletividade (IS) é um parâmetro bem estabelecido para avaliar a janela entre a citotoxicidade e o efeito antineoplásico de diferentes drogas. O objetivo deste estudo é determinar os índices de citotoxicidade do TKI-258 em diferentes linhagens celulares oncohematológicas e avaliar a seletividade da droga às células tumorais em relação a linfócitos de indivíduos saudáveis. Material e métodos: Foram utilizadas células das linhagens K-562 de leucemia mieloide crônica (LMC), Jurkat de leucemia linfoide aguda (LLA) e Raji de linfoma de Burkitt, bem como células mononucleares de sangue periférico (PBMCs) separadas a centrifugação em gradiente de concentração por Ficoll-Paque™. As células foram mantidas em incubadora úmida a 37°C em meio RPMI-1640 contendo 10% de soro fetal bovino, penicilina e estreptomicina 100 UI/mL e, então, 1x105 células controle e tratadas com TKI-258, em concentrações de 0,25 μM, 0,5 μM, 1 μM, 2 μM e 4 μM, foram semeadas por poço e mantidas por 36 horas. Os índices de citoxicidade (IC50) foram determinados por citometria de fluxo por meio da marcação com iodeto de propídio, marcador de morte celular, e calculados a partir de testes de regressão não linear. O programa GraphPad Prism 9.0.0 foi utilizado para as análises estatísticas. Os IS's foram obtidos a partir da razão entre a citotoxicidade para PBMCs (CC50) e para células neoplásicas (IC50). Resultados: O CC50 do TKI-258 em PBMCs foi de 1,702 (r = 0,9696). Em células K562, o IC50 foi de 2,229 (r = 0,8286); em células Jurkat, de 1,850 (r = 0,8867); e em células Raji, de 0,7623 (r = 0,8246). O índice de seletividade da droga para as diferentes linhagens foi de 0,76, 0,92 e 2,23, respectivamente. Discussão: Visto que as drogas disponíveis para tratamento de neoplasias hematológicas são passíveis de falha, resistência medicamentosa e efeitos adversos importantes, o desenvolvimento de novas terapias é fundamental. Os TKIs alvo-específicos constituem terapia padrão ouro para a LMC, porém poucos estudos visam avaliar a viabilidade da inibição dos receptores de tirosinaquinase como estratégia terapêutica para outros cânceres hematológicos, como LLA e Linfoma de Burkitt. O TKI-258 é um inibidor multi-alvos que apresentou, neste estudo, baixa seletividade para as linhagens de LMC e LLA, porém foi significativamente seletivo para as células de Linfoma de Burkitt, apresentando efeito antioncogênico maior que o efeito citotóxico. Conclusão: O estudo demonstrou que o TKI-258 é eficaz para indução de morte celular de forma seletiva para células Raji de Linfoma de Burkitt, embora a droga não tenha tido bom desempenho para as outras neoplasias testadas. O bom índice de seletividade da droga na linhagem de Burkitt nos testes in vitro pode indicar uma menor incidência de efeitos colaterais relacionados à imunossupressão com o uso in vivo. Desse modo, novos estudos devem ser considerados para avaliar o potencial terapêutico do TKI-258 para o Linfoma de Burkitt.

Objetivos: O linfoma de Burkitt é caracterizado por ser uma forma agressiva de linfoma de células B e seu tratamento é realizado por meio de quimioterapia, mas não utiliza inibidores de receptores tirosinaquinase. O TKI-258 é um inibidor de múltiplos receptores tirosinaquinase, ainda em fase de estudos, já testado em diferentes tumores sólidos. O objetivo deste estudo foi avaliar os índices de morte celular em co-culturas de células mononucleares do sangue periférico (PBMCs) e células de linhagem tumoral de Linfoma de Burkitt (RAJI) após tratamento com interleucina-2 (IL-2), TKI-258 e IL-2 em conjunto com TKI-258. Materiais e métodos: Foram utilizadas PBMCs, extraídas por meio da centrifugação com Ficoll-Paque™, e células imortalizadas da linhagem RAJI de Linfoma de Burkitt. As células foram preservadas em incubadora úmida em meio RPMI-1640 a 37 °C, contendo penicilina/estreptomicina e 10% de soro fetal bovino. O índice de citotoxicidade (IC50) do TKI-258 foi determinado previamente por meio citometria de fluxo e calculado pelo teste de regressão não linear, sendo este de 1,5 μM. Em placa de 96 poços foi realizada a co-cultura das PBMCs com as células RAJI, na proporção de 1×106 para 1×105 células por poço, respectivamente. Em seguida, as co-culturas foram divididas em: não tratadas, tratadas com IL-2 1000UI; TKI-258 1,5 μM e TKI-258 1,5 μM mais IL-2 1000UI, e mantidas em incubadora por 4 horas. O percentual de morte celular das co-culturas foi avaliado por citometria de fluxo com marcação com iodeto de propídeo. A análise estatística foi realizada pelo programa GraphPad Prism 9.0.0 sendo considerado estatisticamente significativo quando p < 0,05. Resultados: Foram encontradas diferenças significativas entre o grupo não tratado e os demais (RM one-way ANOVA, F(3,39) = 35.04; p < 0.0001). Não foram observadas diferenças entre os grupos não tratado e tratado com IL-2 (p = 0.9293). O grupo tratado com TKI-258 apresentou um aumento na taxa de mortalidade das células neoplásicas comparado aos grupos anteriores (NTxTKI p < 0.0001; IL-2xTKI p < 0.0001). A maior taxa de mortalidade de células neoplásicas foi encontrada nas amostras tratadas com TKI-258 em associação com IL-2 (NTxIL-2+TKI p < 0.0001; IL-2xIL-2+TKI p < 0.0001; TKIxIL-2+TKI p < 0.0017). Discussão: A associação do TKI-258 com IL-2 apresentou resultados superiores na morte de células neoplásicas e na sobrevida das PBMCs quando comparado aos demais grupos analisados. Diante disso, a IL-2 parece favorecer a atuação desse fármaco, e permitindo a sua atuação em diversas vias simultaneamente, uma vez que, este é um inibidor multi-alvos. Conclusão: O estudo evidenciou que o TKI-258 em associação com a IL-2 foi capaz de produzir uma maior taxa de mortalidade de células da linhagem RAJI quando comparadas com os demais grupos. Novos ensaios com este inibidor em associação ou não à IL-2, para o tratamento para Linfoma de Burkitt são necessários, uma vez que na atualidade, os inibidores de receptores tirosina quinase não são utilizados e os nossos resultados evidenciam o seu potencial terapêutico para o tratamento desta neoplasia.

Background: This study evaluated the effect of treatment with TKI-258 on apoptosis, involving Rho GTPases and their effectors in SCC-4 cells of oral squamous cell carcinoma., Methods: Markers of cell death and apoptosis were analyzed in control and TKI-258-treated SCC-4 cells by flow cytometry. The involvement of Rho GTPases and effectors in the induction of apoptosis by TKI-258 was evaluated by quantification of cleaved PARP. Also, gene expression analysis of those proteins was performed., Results: The treatment with TKI-258 led to a significant increase in cell death (7-AAD) and apoptosis (annexin V and cleaved PARP). When Rho GTPases were stimulated with LPA and inhibited with toxin A Clostridium difficile, the percentage of apoptotic cells increased and decreased, respectively. A similar effect was found when the treatment was with TKI-258 combined with LPA and toxin A. Treatment with TKI-258 significantly increased RhoA gene expression, while RhoB, RhoC, Rac1, and Cdc42 decreased significantly. ROCKs inhibitors (Y-27632 and HA-1077) reduced apoptosis compared with control. TKI-258 combined with Y-27632 or HA-1077 led to an increase in apoptosis compared with inhibitors only. Treatment with TKI-258 led to an increase in ROCK1 and ROCK2 gene expression, and a decrease in PAK1 and PAK2 gene expression., Conclusions: TKI-258 stimulates apoptosis in SCC-4 cells of oral squamous cell carcinoma. Possibly, RhoA GTPase and their effectors ROCKs participate in the signaling pathway inhibited by TKI-258., Clinical Relevance: Therapies with multi-target inhibitors, such as TKI-258, may be promising alternatives for the clinical treatment of oral squamous cell carcinoma., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

This study aimed to evaluate the effects of radiofrequency (RF) on patellar ligament repair through the analysis of type I and III collagens and immunostaining for TGF-β3. To evaluate the effect of RF on patellar ligament repair of Wistar rats, cross-sectional incision (60% of the width - grade I) was performed in patellar ligaments of the groups: lesion (L, n = 7), treated with RF on the 5-day (5RF, n = 7) and 7-day (7RF, n = 7) post injury were compared to control group (C, n = 7). Histological evaluation, immunohistochemistry, morphometry and statistical analysis were performed. At 10 days post injury, ligament rupture were observed only in L. Active fibroblasts, type 3 collagen and TGF-β3 in L, 5RF and 7RF was significantly (p < 0.05) higher than control (C). Type 1 collagen was significantly (p < 0.05) higher in C than L, 5RF and 7RF. A positive correlation (p < 0.05) was observed: TGF-β3 vs active fibroblasts and TGF-β3 vs type 3 collagen; otherwise, negative correlation (p < 0.05): type I collagen vs TGF-β3. These results suggest that RF seemed to accelerate the wound healing process of the patellar ligament and may be used as a non-invasive treatment of partial ligament injuries., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Background: Identification of the antitumor role of tyrosine kinase inhibitors, such as TKI-258, may lead to novel therapeutics for Oral Squamous Cell Carcinoma (OSCC), which has high mortality rates. TKI-258 blocks Fibroblast Growth Factor Receptors (FGFRs), Platelet-Derived Growth Factor Receptors (PDGFRs), and Endothelial Growth Factor Receptor (VEGFRs)., Objective: This study aimed to evaluate the effect of TKI-258 treatment on cell proliferation in SCC-4 cells of OSCC., Methods: BrdU and KI-67 assays were performed by using SCC-4 cells. Control was compared to 1, 5 and 10μM TKI-258 treatment. Control vehicle was compared to: 60μM LY294002 (LY), 2μM Wortmannin (WTN) and LY+WNT. Moreover, TKI 5μM treatment was compared to: TKI 5μM+LY; TKI 5 μM+WTN; TKI 5μM+LY+WTN. After 6h of treatments, immunofluorescence stained BrdU and KI-67 positive cells. Morphometry of proliferative cells was analyzed considering significance of p<0.05., Results: BrdU and KI-67 assays results were similar for all experiments. TKI-258 treatment leads to an important reduction in proliferation rate in SCC-4 cells in a concentration dependent manner. As expected, there was a significant reduction in the percentage of proliferative cells that had PI3K inhibited. When compared with TKI 5 treatment, proliferating cells were significantly lower with simultaneous PI3K inhibition., Conclusion: This study demonstrated that TKI-258 plays an anti-proliferative role on SCC-4 cells of OSCC. It could be interesting to block multiples pathways such as FGFRs, PDGFRs and VEGFRs. Therefore, TKI-258 is a promising option for novel therapeutics for OSCC, especially if associated with PI3K inhibition., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Objective: This study evaluated the involvement of Rho GTPases proteins in the regulation of cytodifferentiation of the SCC-4 human oral squamous cell carcinoma cell line., Methods: Cytokeratin and vimentin immunofluorescence and F-actin staining, assays were performed with control cells and Clostridium difficile 1, 2 and 4 μg/mL Toxin A (Rho GTPases inhibitor) treated SCC-4 cells on three-dimensional MatrigelTM for 24 h. Samples were analyzed by using confocal laser microscopy. Significances were p.

Cetuximab is a chimeric monoclonal antibody that acts as a competitive antagonist, by binding to EGFR. This cell signalling pathways regulates tumor progression. The oral squamous cell carcinoma undergoes to regional spreading and distant metastasis. This study aimed to evaluate the effect of treatment with Cetuximab on cell migration and invasion in OSCC cells, by using the SCC-4 cell line. Cell migration and cell invasion assay were performed and actin cytoskeleton of control and treated with Cetuximab cells were evaluated. Differences were considered significant when p&lt;0.05.Cetuximab inhibited the migration of SCC-4 cells at three concentrations: 1 µg/mL, 50 µg/mL and 100 µg/mL (p&lt;0.0001) in a dose-dependent manner. The number of SCC-4 treated cells with 1 μg/mL that migrated through the membrane was statistically different from 50 μg/mL (p&lt;0.001) and 100 μg/mL (p&lt;0.0001), and between 50 μg/mL and 100 μg/mL (p&lt;0.01). Cetuximab 50 μg/mL inhibited cell invasion through the MatrigelTM compared with SCC-4 control cells (p&lt;0.01). Cetuximab 50 μg/mL affected the organization of the actin cytoskeleton. Cetuximab has an inhibitory effect on actin cytoskeleton organization, cell migration and invasion, suggesting that Cetuximab treatment can be important to avoid oral squamous cell carcinoma metastasis.

Background: Oral squamous cell carcinoma is extremely invasive, and this behavior is regulated by binding of extracellular molecules to the cell membrane receptors. The TKI-258 inhibits phosphorylation of FGFRs VEGFRs and PDGFRs. Our aim was to analyze the effect of TKI-258 treatment in cell movement using SCC-4 cell line from human oral squamous cell carcinoma., Methods: F-actin was stained with rhodamine phalloidin, and confocal analysis was performed. The migration and invasion (membrane covered with Matrigel ™ ) three-dimensional assays were performed, and control and cells treated with TKI-258 that migrated through the membrane were counted after 24 h., Results: Control cells presented abundant cytoplasm with F-actin wide distributed and evident cell cortex; however, treated (1, 5 and 10 μM TKI-258) cells showed round morphology, scanty cytoplasm, F-actin disorganized and preserved cell cortex. TKI-258 (1, 5, and 10 μM) treatment inhibits migrating cells (ANOVA, F = 97.749, d.f. = 3, 10; P < 0.0001), and it was concentration dependent. Invading cell treated with 5 μM TKI-258 was significantly lower (t = 6.708, d.f. = 5, P < 0.001)., Conclusions: These results suggest that the tyrosine kinase inhibitor TKI-258 has an inhibitory effect on cell motility, affecting F-actin, cell migration, and cell invasion, and probably, these processes are regulated by signaling pathways FGFRs and/or PDGFRs and/or VEGFRs., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Depression is a mood disorder that is more prevalent in women and has been closely associated with chronic stress. Many models of depression have been suggested that consider different forms of stress. In fact, stress is present in the life of every human being, but only a few develop depression. Accordingly, it seems wrong to consider all stressed animals to be depressed, emphasizing the importance of predisposition for this mood disorder. Based on this finding, we evaluated a predisposition to depressive behavior of female rats on the forced swim test (FST), and the more immobile the animal was during the FST, the more predisposed to depression it was considered to be. Then, animals were subjected to the stress of social isolation for 21 days and were re-evaluated by the FST. The Predisposed/Isolated rats presented higher immobility times. Once all the rats had prior experience in the FST, we calculated an Index of Increase by Isolation, confirming the previous results. Based on this result, we considered the Predisposed/Isolated group as presenting depressive behavior ('Depressed') and the Nonpredisposed/Nonisolated group as the control group ('Nondepressed'). The animals were distributed into 4 new groups: Nondepressed/Vehicle, Nondepressed/Amitriptyline, Depressed/Vehicle, Depressed/Amitriptyline. After 21 days of treatment, only the Depressed/Vehicle group differed from the other 3 groups, demonstrating the efficacy of amitriptyline in treating the depressive behavior of the Depressed animals, validating the model. This study shows that conducting an FST prior to any manipulation can predict predisposition to depressive behavior in female rats and that the social isolation of predisposed animals for 21 days is effective in inducing depressive behavior. This behavior can be considered real depressive behavior because it takes into account predisposition, chronic mild stress, and the prevalent gender., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Oral squamous cell carcinoma (OSCC) is the most malignant lesion occurring in the head and neck. The Rho-kinases (ROCKs), effectors of Rho proteins, are involved in actin cytoskeletal organization, cell migration, and maintenance cortex. The HA-1077 inhibits the ROCKs. This study aimed to evaluate the effect of treatment with HA-1077 on cell motility in SCC-4 cells, a cell line originating from human OSCC. F-actin of SCC-4 cells treated or not with HA-1077 (1, 50 and 100 μmol/l), and also HA-1077 50 μmol/l and/or inhibitors Y-27632 30 μmol/l was stained with rhodamine-conjugated phalloidin and analyzed by confocal microscopy. Approximately 1×10 cells/well, control and treated with HA-1077 (25, 50, and 100 μmol/l) were added to the migration plate assay. In addition, 1×10 cells/well, control and treated with HA-1077 50 μmol/l, were tested by invasion assays (plate coated with Matrigel). The inhibition of ROCKs with HA-1077 and/or Y-27632 leads to morphological changes, affecting the organization of the actin. The inhibitory effect of HA-1077 (P<0.0001) was dose dependent as the number of cells migrated at 100 μmol/l was statistically different: 25 μmol/l (P<0.0001) and 50 μmol/l (P<0.01). The number of cells treated with HA-1077 50 μmol/l decreased compared with control cells that invaded through Matrigel (P<0.0001). This study shows an inhibitory effect of HA-1077 on cell migration and invasion, suggesting that the use of HA-1077 can be a potential therapy for OSCC.

Oral squamous cell carcinoma (OSCC) is a disease with high mortality and morbidity. Metastasis is a significant prognostic factor of the OSCC patients. The Rho GTPases are signaling proteins that controls important cellular processes in various complex mechanisms involved in carcinogenesis. This study aimed to evaluate the expression pattern of RhoC in OSCC protein by immunohistochemistry in situ. Immunohistochemical reactions were performed for RhoC by the method of avitina-biotin-peroxidase activity in samples OSCC: well differentiated (BD, n=6), moderately differentiated (MD, n=24) and poorly differentiated (PD, n=13). The morphometry was taken by QuickScore (percentage and intensity of staining) and only intensity staining. There was no statistical difference (p>0.05) through none of the modes of morphometric analysis between BD, MD and PD. And the RhoC staining was not associated with the histopathologic grading (χ2 = 4.65, p>0.05). However, the morphological evaluation of immunostained for RhoC in cases BD, MD, PD OSCC, regardless of histopathologic grading. These results suggest that there is no correlation between the RhoC immunoexpression and histopathological grading of OSCC.

Background: Carboxytherapy (CA) refers to the cutaneous and subcutaneous administration of CO(2) for therapeutic purposes. Radiofrequency (RF) is a method that uses electric current for heating layers of the skin. Both techniques are indicated for the treatment of skin laxity., Objective: The aim of this study was to compare the effects of CA and RF on human skin., Methods: After eight patients underwent abdominoplasty, each of them received a single treatment of CA and a single treatment of RF on the right and left infra-umbilical regions, respectively. In the infra-umbilical region, CA was performed on the right and RF was performed on the left side. Untreated skin was used as a control. The sample collection period lasted 120 days. CA was administered at a velocity of 40 mL/min, and the total quantity of CO(2) infused was approximately 20 mL. RF was carried out at a temperature higher than 40°C on the epidermis for 5 min., Results: CA and RF led to collagen remodeling; however, this result was more evident and lasted longer with RF. With CA an increase in elastic fibers was observed, whereas with RF no alteration was observed., Conclusion: Our results suggest that RF is more efficient than CA in stimulating collagen synthesis.

The pleomorphic adenoma of the parotid (PA) is characterized by the high tissues diversity. Rho GTPases participate in signal transduction pathways that regulate several biological processes, including cell differentiation. A quantitative analysis of RhoA and RhoB GTPases immunoexpression was performed in healthy parotids and in 23 PA cases, predominantly epithelial (PE) or mesenchymal (PM), followed by Student's t test. In PE cases, RhoA immunoexpression was higher in sheets and trabeculae (p < 0.05), whereas RhoB only in sheets (p < 0.05). In normal parotids, RhoA and RhoB were not detected in acinar cells. Ducts have expressed RhoA and RhoB in normal parotids and PA. RhoB was detected in myxoid and chondromyxoid cells. Normal parotids do not express RhoA and RhoB proteins in acinar cells, indicating a lack of function in secretory cells. Despite RhoA and RhoB GTPases are different in their biological roles, no significant difference in immunoexpression of the RhoA and RhoB GTPases in epithelial and mesenchymal structures of PA.

Background: Microdermabrasion is a surface treatment, noninvasive, which uses a negative pressure and drives programmable inert microcrystals on the skin, causing an exfoliation., Objective: The aim of this study was to evaluate the effects of application of microdermabrasion in human skin rejuvenation., Methods: Eleven women who were undergoing abdominoplasty were considered. An area of 25 cm² in the umbilicus to the right was conditioned with microcrystals of Al₂O₃ in maximum flow, negative pressure of 200 mmHg and total of 8 past, the left side being used as control. The number of sessions ranged from one to five, with weekly intervals, and timing of sample collection ranged from 0 to 132 days. Samples were fixed in 10% formaldehyde in phosphate buffer and were evaluated histologically., Results: A mild to marked hyperpigmentation was observed and remained for a variable period. Histological findings suggest an improvement in the epidermal layer with increased thickness and reestablishing their interdigitations in the dermis initially observed an increase in collagen synthesis. The analysis showed a late stay of epidermal changes, which did not occur in the dermis., Conclusion: Under the conditions and parameters used in this work, the microdermabrasion had a positive skin structure, showing that a viable resource in promoting skin rejuvenation.

Purpose: This study evaluated the healing process of teeth replanted after root treatment and intracanal dressing with indomethacin alone or indomethacin with calcium hydroxide (Ca[OH]2)., Materials and Methods: Through a case-control study, 24 teeth of 6 adult dogs were extracted, dried, and divided into 4 groups according to the root surface treatment protocols performed before replantation and the intracanal medication used after replantation. In group 1 (negative control), root surfaces were treated by immersion in a 0.9% saline solution and then replanted. In the other groups, the roots were immersed for 10 minutes in Ca(OH)2 (group 2), indomethacin (group 3), or a solution of indomethacin and Ca(OH)2 (group 4). After 2 weeks, group 1 teeth were subjected to single-visit root canal treatment and obturation with gutta-percha and sealer consisting of zinc oxide and eugenol. The teeth in the other groups were subjected to intracanal dressing with the same material used for immersion. After an additional period of 28 weeks, the animals were euthanized and the jaws containing the replanted teeth were processed for histologic analysis. Histometric values were statistically analyzed, with significance set at a P value less than or equal to .05., Results: Group 1 exhibited significantly more normal periodontium than group 4 (P = .02). Total resorption was greater in group 4 than in group 1 (P = .02). No statistically significant difference in the percentage of surface resorption or in total inactive resorption was observed between the groups., Conclusions: The findings of this study suggest that intracanal dressing and topical root treatment with Ca(OH)2 with or without indomethacin is not recommended for teeth dried for 50 minutes, but the use of indomethacin alone as root surface treatment for delayed tooth replantation deserves further study using longer drying periods. In addition, the present results suggest that a single-visit root canal, performed up to 2 weeks after replantation, might be indicated for teeth dried for up to 50 minutes., (Copyright © 2014 American Association of Oral and Maxillofacial Surgeons. Published by Elsevier Inc. All rights reserved.)

Purpose of Investigation: To evaluate the distribution of GTPases RhoA, RhoB, and Cdc42 in cervical intraepithelial neoplasias (CIN) and invasive neoplasias of the uterine cervix., Materials and Methods: samples of neoplastic lesions of the uterine cervix of 44 patients were classified in: CIN I (n = 10), CIN II (n = 10), CIN III (n = 09), and invasive carcinoma (n = 15). Antibodies anti-RhoA, anti-RhoB, and anti-Cdc42 were used and staining was classified as: negative, mild, moderate, and intense positive., Results: When compared with dysplastic cells, superficial cells showed: higher expression of RhoB in CIN I (p = 0.0018), and lower expression of Cdc42 in CIN I (p = 0.0225). The authors observed higher expression of RhoA (p = 0.0002) and RhoB (p = 0.0046) in CIN dysplastic cells when compared with invasive carcinoma cells., Conclusions: GTPases Rho may be involved with the regulation of biological processes, important to the progression of cervical neoplasias. Probably, RhoA is important for maintenance of cell differentiation and RhoB protects cells from malignant cervical neoplasia.

Rho GTPases are proteins that regulate cell cycle, shape, polarization, invasion, migration, and apoptosis, which are important characteristics of normal and neoplastic cells. Rho GTPases expression has been reported in normal tooth germ and several pathologies; however, it has not been evaluated in ameloblastomas. The aim of this study was to analyze the expression and distribution of RhoA, RhoB, Rac1, and Cdc42 Rho GTPases in solid and unicystic ameloblastomas. Three-micrometer sections from paraffin-embedded specimens were evaluated by using an avidin-biotin immunohistochemical method with antibodies against the proteins mentioned above. RhoA and RhoB staining was observed in a high number of cells (P < 0.05) and greater intensity in non-polarized ones. Rac1 was not observed, and Cdc42 did not show any statistical differences between the number of non-polarized and basal positive cells (P > 0.05). Upon comparing the studied ameloblastomas, a higher number of positive cells in the unicystic variant was observed than that in the solid one (P < 0,05). The results obtained suggest that these GTPases could play a role in the ameloblastoma neoplastic epithelial cell phenotype determination (polarized or non-polarized), as well as in variant (solid or unicystic) and subtype (follicular or plexiform) determination. Furthermore, they could participate in solid ameloblastoma invasion mechanisms., (© 2011 John Wiley & Sons A/S.)

Morphogenesis and cytodifferentiation are distinct processes in tooth development. Cell proliferation predominates in morphogenesis; differentiation involves changes in form and gene expression. The cytoskeleton is essential for both processes, being regulated by Rho GTPases. The aim of this study was to verify the expression, distribution, and role of Rho GTPases in ameloblasts and odontoblasts during tooth development in correlation with actin and tubulin arrangements and amelogenin and dentin sialophosphoprotein (DSPP) expression. RhoA, Rac1, and Cdc42 were strongly expressed during morphogenesis; during cytodifferentiation, RhoA was present in ameloblasts and odontoblasts, Rac1 and its effector Pak3 were observed in ameloblasts; and Cdc42 was present in all cells of the tooth germ and mesenchyme. The expression of RhoA mRNA and its effectors RockI and RockII, Rac1 and Pak3, as analyzed by real-time polymerase chain reaction, increased after ameloblast and odontoblast differentiation, according to the mRNA expression of amelogenin and DSPP. The inhibition of all Rho GTPases by Clostridium difficile toxin A completely abolished amelogenin and DSPP expression in tooth germs cultured in anterior eye chamber, whereas the specific inhibition of the Rocks showed only a partial effect. Thus, both GTPases are important during tooth morphogenesis. During cytodifferentiation, Rho proteins are essential for the complete differentiation of ameloblasts and odontoblasts by regulating the expression of amelogenin and DSPP. RhoA and its effector RockI contribute to this role. A specific function for Rac1 in ameloblasts remains to be elucidated; its punctate distribution indicates its possible role in exocytosis/endocytosis.

During the rat submandibular gland (SMG) development, organogenesis and cytodifferentiation depend on the actin cytoskeleton, which is regulated by small Rho GTPases. These proteins link cell surface receptors to pathways that regulate cell motility, polarity, gene expression, vesicular trafficking, proliferation and apoptosis. The aim of this study was to evaluate, by immunohistochemistry, the distribution pattern of RhoA, RhoB, RhoC, Rac1 and Cdc42 during cytodifferentiation of the rat SMG and in male adults. All GTPases were found in epithelial and mesenchymal tissues throughout gland development. Rac1 appeared to be important for parenchyma expansion at the beginning of cytodifferentiation, while RhoC, Cdc42 and the inactive phosphorylated form of Rac1 seemed associated with lumen formation and cell polarization in terminal tubules. RhoA and RhoB labeling was evident throughout development. All GTPases were differentially expressed in the adult gland, suggesting that they play specific roles during differentiation and function of the rat SMG.

Rho GTPases participate in a wide variety of signal transduction pathways regulating the actin cytoskeleton, gene expression, cellular migration and proliferation. The aim of this study was to evaluate the role of Rho GTPases in signal transduction pathways during acinus formation in a human salivary gland (HSG) cell line initiated by extracellular matrix (ECM; Matrigel) alone or in combination with epidermal growth factor, basic fibroblast growth factor and lysophosphatidic acid (LPA). Immunohistochemical and Western blotting analyses showed that HSG cells contained RhoA, RhoB, Rac1 and Cdc42 proteins. All growth factors enhanced the effects of ECM on acinus formation, in a pathway dependent on PI3-kinase and Rho GTPases. The role of ROCK, a major RhoA effector, seemed limited to cortical actin polymerization. LPA stimulated cell migration and acinus formation in a PI3-kinase-independent pathway. The results suggest that Rho proteins are important for epithelial-mesenchymal interactions during salivary gland development.