Análise ultra-estrutural das glândulas localizadas na parede da fístula congênita de lábio inferior de pacientes com a síndrome de Van der Woude

The objective of the present study was to evaluate the glands of wall of congenital fistulae of the lower lip with the transmission electron microscope in order to characterize their microstructural pattern. Thin section of Araldite resin embedded congenital fistulae of the lower lip of four patients with Van der Woude syndrome from the Hospital de Reabilitação de Anomalias Craniofaciais da Universidade de São Paulo, Bauru, SP, were analyzed with a transmission electron microscope. The results showed that the glands were mostly made by typical mucous acini exhibiting, with certain frequency, myoepithelial cells surrounding them. In some of lobules, a few acini smaller than the typical mucous, showed granules of moderate electron density or containing a dense core or exhibiting small dense spherule and predominance granular material. These granules resemble to described recently by others in various human minor salivary glands. We concluded that glands associated with congenital fistula of lower lip of patients with Van der Woude syndrome, in spite of being located in vermilion border of the lip, showed at the transmission electron microscope characteristics of labial minor salivary gland, i.e, are mostly mucous with a few seromucous units, while typical seromucous demilunes are not present.O objetivo do presente estudo foi avaliar a ultraestrutura de glândulas da parede de fístula congênita de lábio inferior ao microscópio eletrônico de transmissão para caracterizar seu padrão microestrutural. Deste modo, as fístulas congênitas de 4 pacientes com a síndrome de Van der Woude do Hospital de Reabilitação de Anomalias Craniofaciais da Universidade de Sâo Paulo, Bauru, SP, foram processadas para inclusão em resina Araldite e os cortes finos foram analisados no microscópio eletrônico de transmissão. Os resultados mostraram que as glândulas estavam constituídas por ácinos mucosos típicos exibindo com certa freqüência células mioepiteliais ao seu redor. Em alguns lóbulos, foram observados em pequeno número, ácinos menores que o mucoso típico, exibindo células com grânulos de moderada eletron-densidade contendo um corpo denso ou uma pequena esférula densa no interior de um material predominante granular. Estes grânulos lembravam os descritos recentemente em glândulas salivares labiais humanas. Em vista dos resultados obtidos concluímos que as glândulas associadas com a fístula congênita de lábio inferior de pacientes com a síndrome de Van der Woude, apesar de estarem localizadas no vermelhão do lábio, mostraram ao microscópio eletrônico de transmissão características de glândula salivar labial, i.e., são predominantemente mucosos com poucas unidades seromucosas, mas semiluas seromucosas típicas não estão presentes

Antibody-drug conjugates (ADC) against cancer stem-like cells (CSC) - Is there still room for optimism?

Cancer stem-like cells (CSC) represent a subpopulation of tumor cells with peculiar functionalities that distinguish them from the bulk of tumor cells, most notably their tumor-initiating potential and drug resistance. Given these properties, it appears logical that CSCs have become an important target for many pharma companies. Antibody-drug conjugates (ADC) have emerged over the last decade as one of the most promising new tools for the selective ablation of tumor cells. Three ADCs have already received regulatory approval and many others are in different phases of clinical development. Not surprisingly, also a considerable number of anti-CSC ADCs have been described in the literature and some of these have entered clinical development. Several of these ADCs, however, have yielded disappointing results in clinical studies. This is similar to the results obtained with other anti-CSC drug candidates, including native antibodies, that have been investigated in the clinic. In this article we review the anti-CSC ADCs that have been described in the literature and, in the following, we discuss reasons that may underlie the failures in clinical trials that have been observed. Possible reasons relate to the biology of CSCs themselves, including their heterogeneity, the lack of strictly CSC-specific markers, and the capacity to interconvert between CSCs and non-CSCs; second, inherent limitations of some classes of cytotoxins that have been used for the construction of ADCs; third, the inadequacy of animal models in predicting efficacy in humans. We conclude suggesting some possibilities to address these limitations

How Tumor Cells Choose Between Epithelial-Mesenchymal Transition and Autophagy to Resist Stress-Therapeutic Implications

Tumor cells undergo epithelial-mesenchymal transition (EMT) or macroautophagy (hereafter autophagy) in response to stressors from the microenvironment. EMT ensues when stressors act on tumor cells in the presence of nutrient sufficiency, and mechanistic target of rapamycin (mTOR) appears to be the crucial signaling node for EMT induction. Autophagy, on the other hand, is induced in the presence of nutrient deprivation and/or stressors from the microenvironment with 5' adenosine monophosphate-activated protein kinase (AMPK) playing an important, but not exclusive role, in autophagy induction. Importantly, mTOR and EMT on one hand, and AMPK and autophagy on the other hand, negatively regulate each other. Such regulation occurs at different levels and suggests that, in many instances, these two stress responses are mutually exclusive. Nevertheless, EMT and autophagy are able to interconvert and we suggest that this may depend on spatiotemporal changes in the tumor microenvironment and/or on duration/intensity of the stressor signal(s). Eventually, we propose a three-pronged therapeutic approach aimed at targeting these three major tumor cell populations. First, cytotoxic drugs that act on differentiated and proliferating tumor cells and which, per se, may promote induction of EMT or autophagy in surviving tumor cells. Second, inhibitors of mTOR in order to prevent EMT induction. Third inducers of autophagic cell death (autosis) in order to deplete tumor cells that are constitutively in an autophagic state or are induced to enter an autophagic state in response to antitumor therapy

Postmortem acinar autolysis in rat sublingual gland: a morphometric study

ABSTRACT OBJECTIVE: To analyze and to quantify morphological acinar postmortem changes in rat sublingual glands (SLG). MATERIAL AND METHODSs: Fifty rats were divided into two groups of 25 animals each. Group I was used for morphological and morphometric evaluations and group II for the determination of gland density and processed gland volume. Acinar autolytic changes were studied at 0 (control group), 3, 6, 12 and 24 h postmortem periods. The morphometric analysis of the volume density (Vv) and total volume (Vt) of intact (ia) and autolyzed (aa) acini was performed under light microscopy using a Zeiss II integration grid with 100 symmetrically distributed points. RESULTS: Morphologically, temporal progressive nuclear alterations and gradual loss of the structural architecture of acinar cells were found. Regarding quantitative results, both the Vvaa and the Vvia showed statistically significant differences among all postmortem periods (p<0.05). Vvaa increased from 0.42% at 0 h to 75.84% at 24 h postmortem and Vvia decreased from 71.16% to 0% over the same period. For Vtaa and Vtia, no statistically significant differences occurred between 12-24 h and 0-3 h (p&gt;0.05), respectively. Vtaa increased from 0.18 mm³ at 0 h to 38.17 mm³ at 12 h, while Vtia showed a decrease from 33.47 mm³ to 0 mm³ between 3-24 h postmortem. Data concerning Vtaa were adjusted by two-variable linear regression, obtaining the equation: y=-3.54 + 3.38x (r²=0.90). The Vtaa growth rate calculated by this equation was 3.38 mm³/h between 0-12 h. CONCLUSION: Acinar autolysis on rat SLG demonstrated the most significant signs during the first 6 h postmortem and was widely spread through the gland at 12 h

Topical administration of a doxorubicin-specific monoclonal antibody prevents drug-induced mouth apoptosis in mice

One of the most severe side effects of anti-tumour chemotherapy is mucositis due to drug toxicity for rapidly dividing cells. We show here that anti-DXR monoclonal antibodies can prevent DXR-induced damage. Indeed, apoptosis, confined to the proliferative compartment of the basal mucosa, observed in the tongue of DXR-treated mice was completely inhibited by topical application of the anti-DXR antibodies. © 2001 Cancer Research Campaign http://www.bjcancer.co

Estudo radiográfico e histológico do reparo de defeito ósseo perene em calvária de rato após o tratamento com material de enxerto orgânico bovino poroso

Over the last few years, various bone graft materials of bovine origin to be used in oromaxillofacial surgeries have entered the market. In the present study, we determined the capacity of a block organic bone graft material (Gen-ox, Baumer SA, Brazil) prepared from bovine cancellous bone to promote the repair of critical size bone injuries in rat calvaria. A transosseous defect measuring approximately 8mm in diameter was performed with a surgical trephine in the parietal bone of 25 rats. In 15 animals, the defects were filled with a block of graft material measuring 8mm in diameter and soaked in the animal's own blood, and in the other 10 animals the defects were only filled with blood clots. The calvariae of rats receiving the material were collected 1, 3 and 6 months after surgery, and those of animals receiving the blood clots were collected immediately and 6 months after surgery. During surgery, the graft material was found to be of easy handling and to adapt perfectly to the receptor bed after soaking in blood. The results showed that, in most animals treated, the material was slowly resorbed and served as a space filling and maintenance material, favoring angiogenesis, cell migration and adhesion, and bone neoformation from the borders of the lesion. However, a foreign body-type granulomatous reaction, with the presence of numerous giant cells preventing local bone neoformation, was observed in two animals of the 1-month subgroup and in one animal of the 3-month subgroup. These cases were interpreted as resulting from the absence of demineralization and the lack of removal of potential antigen factors during production of the biomaterial. We conclude that, with improvement in the quality control of the material production, block organic bone matrix will become a good alternative for bone defect repair in the oromaxillofacial region due to its high osteoconductive capacity.Nos últimos anos, vários materiais de origem bovina para enxerto ósseo têm sido lançados no comércio para serem utilizados em cirurgias bucomaxilofaciais. Na presente pesquisa avaliamos a capacidade de um material de enxerto ósseo orgânico em bloco (Gen-ox, Baumer SA - Brasil) preparado à partir de osso medular bovino, em promover a reparação de lesões ósseas de tamanho crítico em calvária de ratos. Uma lesão trans-óssea de aproximadamente 8mm de diâmetro foi realizada com trefina cirúrgica nos ossos parietais de 25 ratos, sendo que em 15 os defeitos foram preenchidos com bloco de 8mm de diâmetro do material de enxerto, embebida com sangue do próprio animal e em 10 somente com coágulo sanguíneo. As calvárias dos ratos tratados com o material foram coletadas após 1, 3 e 6 meses após as cirurgias e dos tratados com coágulo sanguíneo, imediatamente após as cirurgias e decorridos 6 meses. Durante as cirurgias verificamos que esse material enxertado é de fácil manuseio e se adapta perfeitamente ao leito receptor após sua embebição com sangue. Os resultados obtidos mostraram que na maioria dos casos tratados, o material foi reabsorvido lentamente e serviu como material de preenchimento e mantenedora de espaço, favorecendo a angiogênese, migração e adesão celular e a neoformação óssea à partir das bordas da lesão. No entanto, em 2 casos no subgrupo tratado - 1 mês e 1 caso no subgrupo tratado - 3 meses, ocorreu a presença de reação granulomatosa tipo corpo estranho com a presença de inúmeras células gigantes, que inibiu a neoformação óssea local. Esses casos foram interpretados como decorrentes de falhas na desmineralização e na retirada de potenciais fatores antigênicos durante a produção do biomaterial. Concluímos que com um aprimoramento do controle de qualidade na linha de produção do material, a matriz orgânica medular em bloco poderá ser uma boa alternativa para ser usada no reparo de defeitos ósseos na região buco-maxilofacial, devido a sua alta capacidade osteocondutora

Critical Role of TLR9 in Acute Graft-versus-Host Disease

Abstract Graft-vs-host disease (GVHD) is a major complication after allogeneic bone marrow transplantation. Different studies have demonstrated that intestinal bacterial breakdown products and loss of gastrointestinal tract integrity, both induced by conditioning regiments, are critical in the pathogenesis of acute GVHD. Using C57BL/6 knockout mice, we evaluated the role of TLR4 and TLR9, which recognize bacterial LPS and DNA, respectively, in the GVHD associated with allogeneic bone marrow transplantation. When myeloablative-irradiated TLR9 knockout (TLR9−/−) mice were used as graft recipients, survival and clinical score of acute GVHD were improved as compared with the wild-type recipient mice (18/30 vs 1/31 mice still alive at day 70 in a total of four experiments); while no differences were observed using recipient TLR4 knockout (TLR4−/−) mice. The reduced mortality and morbidity in TLR9−/− mice related with reduced stimulatory activity of TLR9−/− spleen APCs after conditioning and reduced proliferation of allogeneic donor T cells. Experiments using TLR9+/+ into TLR9−/− and TLR9−/− into TLR9+/+ chimeric mice as recipients indicated a critical role for nonhematopoietic TLR9+/+ cells interacting with bacterial breakdown products released in myeloablated mice. Altogether these data reveal a novel important role of TLR9 in GVHD, a finding that might provide tools to reduce this complication of allogeneic transplantation

Estudo estereológico comparativo entre os ameloblastos de secreção e de maturação em incisivos de ratos

Ameloblastos são células de origem epitelial, que apresentam funções de síntese, secreção e maturação do esmalte dental. Assim sendo, ameloblastos passam por mudanças morfológicas durante o seu curso de desenvolvimento. Desse modo, cinco ratos adultos Wistar foram utilizados para avaliar a morfologia de ameloblastos da fase secretora e em maturação, usando de métodos estereológicos em microscopia eletrônica de transmissão. Os dados foram analisados pela análise de variância a um critério (ANOVA). Os resultados mostraram que da fase de secreção para a de maturação da amelogênese: a) ocorreu uma redução de 23% no volume absoluto nuclear e o volume absoluto citoplasmático não mostrou diferença estatisticamente significante (p>;0.05); b) o volume total e a superfície total do retículo endoplasmático rugoso (RER) diminuíram 74% e 90%, respectivamente; c) o volume total e a superfície total das mitocôndrias aumentaram 742% e 384%, respectivamente; d) a relação superfície-volume do RER e das mitocôndrias diminuíram 59% e 42%, respectivamente; e) os pequenos grânulos de secreção, dispostos principalmente na área apical, desapareceram junto com o Processo de Tomes e apareceram os lisossomos e vacúolos digestivos principalmente na área supranuclear. Portanto, apesar de ocorrer uma significante adaptação morfológica durante a modulação da fase secretora para a de maturação do desenvolvimento do esmalte, o volume citoplasmático do ameloblasto permanece inalterado.The ameloblasts both in secretion phase and in smooth-ended ameloblasts in maturation phase were studied using stereologic methods in transmission electron microscopy (TEM). From secretion to maturation phase of amelogenesis, the nucleus volume decreased 23% and cytoplasm volume did not show significant changes; the total volume and surface of the rough endoplasmic reticulum (RER) decreased 74% and 90%, respectively, and of the mitochondria increased 742% and 384%, respectively; the surface-to-volume ratio for RER and mitochondria decrease 59% and 42%, respectively; and the predominantly apical secretory granules disappeared joined at Tomes process and lysosomes and phagic vacuoles have appeared principly in supranuclear cytoplasm. Although significant morphologic variation occurs from the secretory to the maturation phase of the ameloblasts, their cytoplasm volume remains unaltered

Development of a direct ESI-MS method for measuring the tannin precipitation effect of proline-rich peptides and in silico studies on the proline role in tannin-protein interactions

Tannins are a heterogeneous class of polyphenols that are present in several plants and foods. Their ability to interact and precipitate proline-rich proteins leads to different effects such as astringency or antidiarrheal activity. Thus, evaluation of the tannin content in plant extracts plays a key role in understanding their potential use as pharmaceuticals and nutraceuticals. Several methods have been proposed to study tannin-protein interactions but few of them are focused on quantification. The purpose of the present work is to set up a suitable and time efficient method able to quantify the extent of tannin protein precipitation. Bradykinin, chosen as a model, was incubated with increasing concentrations of 1,2,3,4,6-penta-O-galloyl-\u3b2-D-glucose and tannic acid selected as reference of tannic compounds. Bradykinin not precipitated was determined by a mass spectrometer TSQ Quantum Ultra Triple Quadrupole (direct infusion analysis). The results were expressed as PC 50 , which is the concentration able to precipitate 50% of the protein. The type of tannin-protein interaction was evaluated also after precipitate solubilisation. The involvement of proline residues in tannin-protein interactions was confirmed by repeating the experiment using a synthesized peptide (RR-9) characterized by the same bradykinin sequence, but having proline residues replaced by glycine residues: no interaction occurred between the peptide and the tannins. Moreover, modelling studies on PGG-BK and PGG-RR-9 were performed to deeply investigate the involvement of prolines: a balance of hydrophobic and H-bond contacts stabilizes the PGG-BK cluster and the proline residues exert a crucial role thus allowing the PGG molecules to elicit a sticking effect