Made available in DSpace on 2014-12-17T15:32:16Z (GMT). No. of bitstreams: 1 PedroPAS.pdf: 6126862 bytes, checksum: 3ae3ec2483d9e09d3a05fcd73782e862 (MD5) Previous issue date: 2008-02-27 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The giant cell fibroma is a benign neoplasm characterized by the presence of mono, bi or multinucleate cells, which can have a connection to the presence of mast cells. This research aims to analyze, descriptively and comparatively, the immunohystochemistry expression of the tryptase in mast cells of the giant cell f ibroma, f ibrous hyperplasia and samples of the normal oral mucosa. Thirty cases of giant cell fibroma, ten cases of fibrous hyperplasia and ten cases of normal oral mucosa were selected for the analysis of the immunohistochemistry expression, determination of the number of present mast cells, as well as their location and shape. It could be stated that there was a statistically signif icant difference (p<0,001) in relation to the quantity of mast cells among other samples analyzed where the giant cell f ibroma presented lesser quantity of mast cell and the hyperplasia showed higher concentration of this cellular type. Although the oral mucosa has presented a higher quantity of mast cells when compared to the giant cells fibroma, these were found in usual locations in the connective tissue in normal tissues. There could be noticed a statistically significant difference in relation to the number of non-granulated mast cells (p<0,001). On the areas of fibrosis, we could observe a statistically signif icant difference (p<0,006) among the samples. In relation to the present mast cells in perivascular location, no statistically signif icant difference was found. On the morphological analysis there was a predominance of oval mast cells. It was concluded that despite of the fact there was a lesser quantity of mast cells present in cases of giant cell f ibroma, they appeared to have a stronger relation to the present giant fibroblasts in this lesions, around 59,62%, being also evidenced a strong relation between these cells and the fibrosis areas in both cases of giant cell f ibroma and f ibrous hyperplasias and samples of normal oral mucosa, used as control group in our study, confirming, this way, the role of the mast cells as fibrinogenous inductor / O fibroma de c?lulas gigantes constitui-se de uma neoplasia benigna, caracterizada pela presen?a de c?lulas gigantes, mono, bi ou multinucleadas, c?lulas estas que podem guardar rela??o com a presen?a de mast?citos. O prop?sito desta pesquisa consistiu em analisar descritiva e comparativamente a express?o imuno-histoqu?mica da triptase em mast?citos de fibroma de c?lulas gigantes, hiperplasia fibrosa e esp?cimes de mucosa oral normal. Foram selecionados 30 casos de fibroma de c?lulas gigantes, 10 casos de hiperplasia fibrosa e 10 casos de mucosa oral normal, para a an?lise da express?o imuno-histoqu?mica, determina??o do n?mero de mast?citos presentes, bem como a sua forma e localiza??o. Constatou-se diferen?a estatisticamente significativa (p<0,001) em rela??o a quantidade de mast?citos entre os esp?cimes analisados, onde o fibroma de c?lulas gigantes apresentou a menor quantidade de mast?citos e a hiperplasia exibiu a maior concentra??o deste tipo celular. Embora a mucosa oral tenha apresentado uma maior quantidade de mast?citos quando comparado com os casos de f ibroma de c?lulas gigantes, estes se encontravam em localiza??es usuais no tecido conjuntivo em tecidos normais. Verif icou-se, diferen?a estatisticamente significativa, no que diz respeito ao n?mero de mast?citos n?o degranulados (p<0,001). Nas ?reas de fibrose, observamos diferen?a estatisticamente signif icativa (p<0,006) entre os esp?cimes. Com rela??o aos mast?citos presentes em localiza??o perivascular n?o se observou diferen?a estatisticamente significativa. Na an?lise morfol?gica verif icou-se uma predomin?ncia de mast?citos ovais. Concluiu-se que embora uma menor quantidade de mast?citos estivesse presente nos casos de fibroma de c?lulas gigantes, estes exibiam maior rela??o com os fibroblastos gigantes presentes nestas les?es em torno de 59,62%, sendo evidenciada tamb?m uma forte rela??o entre estas c?lulas e ?reas de fibrose tanto nos casos de fibroma de c?lulas gigantes como de hiperplasias fibrosas e esp?cimes de mucosa oral normal, utilizados como controle em nosso estudo, confirmando desta forma, o papel dos mast?citos como indutor fibrinog?nico

The interest in evaluating the effect of the phytochemical constituents in food and herb teas in attenuating the oxidative stress associated with high intensity physical exercise has increased in the last years. In this context, leaf tea of plants of the Passiflora genus can inhibit the cell damage because of the oxidative stress due to the presence of phenolic compounds. In the present study, the phytochemical profile was investigated by colorimetric or precipitation methods, while the total phenol content was quantified by the Folin-Ciocalteu method. To study the acute and chronic effect of the extract on the oxidative stress, Wistar rats (250-300 g) were divided randomly into groups (n=8 and n=6, respectively) that were treated with the extract and were submitted or not to high intensity physical exercises. The oxidative stress marker levels were analyzed. Because of the high content of phenolic constituents (430.63 ± 46.71 μg EAG.g-1 extract), it was verified a scavenger activity of the free radicals DPPH● and ABTS●+ up of 97%. The extract inhibited also levels of lipid peroxidation induced by AAPH (above 95 %), FeSO4 (up 40.82%) and H2O2 (up62.50%), suggesting the the extract is active against peroxil and hydroxyl radical. The acute administration of the extract protected rats from lipoperoxidation with reductions in levels in plasmatic (70.41%), hepatic (64.1%) and cardiac (20.0 %) samples, and protected liver against protein oxidation (94.08%). Sulfhydryl levels in plasmatic (264.32%), hepatic (261.86%) and cardiac (478.8%) tissues were high. The research showed that physical training used was able to increase lipid oxidation in the urinary tract (201%), blood tissue (123%) and liver (161%), and the protein oxidation rate (226%) rats exercised compared to sedentary control group. In contrast, treatment with the extract , after 8 weeks of treating the rats submitted to high intensity physical exercises with the extract, there was a chronic prevention of the lipoperoxidation in structures of the urinary (50.96%), plasmatic (39.91%), hepatic (83.64%) and cardiac (15.82%) tissues, as well as, oxidative lesion of hepatic proteins (9.62%). Therefore, these results suggest benefits of P. cincinnata leaf ethanol extract to health. / Nos últimos anos, têm aumentado o interesse em avaliar o efeito de constituintes fitoquímicos presentes em alimentos e chás de ervas na atenuação do estresse oxidativo associado ao exercício físico de alta intensidade. Nesse contexto, o chá das folhas de plantas do gênero Passiflora pode inibir o dano celular causado pelo estresse oxidativo devido à presença de componentes fenólicos. No presente estudo, o perfil fitoquímico do extrato etanólico das folhas de P. cincinnata Mast foi investigado através de reações colorimétricas ou de precipitação, enquanto o teor de fenóis totais foi quantificado usando o método de Folin-Ciocalteu. Para estudar o efeito agudo e crônico do extrato sobre o estresse oxidativo, ratos Wistar (250-300g) foram divididos aleatoriamente em grupos (n=8 e n=6, respectivamente) que foram tratados com o extrato (200 mg.kg-1) e submetidos ou não a exercícios físicos de alta intensidade. Os níveis de marcadores do estresse oxidativo foram analisados. Devido ao elevador teor fenólico (430,63 ± 46,71 μg GAEq.g-1 extrato), foi constatada atividade seqüestradora dos radicais DPPH● e ABTS●+ acima de 97%. O extrato inibiu, também, valores elevados da lipoperoxidação in vitro induzida por AAPH (acima 95 %), FeSO4 (até 40,82 %) e H2O2 (até 62,50 %), sugerindo o ação ativo contra radicais peroxil e hidroxil. A administração aguda do extrato protegeu ratos da lipoperoxidação plasmática (70,41%), hepática (64,1%) e cardíaca (20,0%), além de proteger a oxidação protéica (94,08%) hepática. Os níveis de sulfidrila plasmático (264,32%), hepático (261,86%) e cardíaco (478,8%) foram elevados. Constatamos, também, que o treinamento físico utilizado foi capaz de aumentar à oxidação lipídica no aparelho urinário (201 %), tecido sanguíneo (123 %) e hepática (161 %), além de oxidação a proteínas cardíaca (226 %) os animais exercitados (GT) em comparação ao controle sedentário (GC) . Em contrapartida, o tratamento com o extrato, após 8 semanas de tratamento, foi capaz de prevenir a lipoperoxidação de estruturas do aparelho urinário (50,96%), sanguínea (39,91%), hepático (83,64%) e cardíaca (15,82%), bem como, lesões oxidativas a proteínas hepáticas (9,62%) induzidas pelo exercício físico. Desta forma, os resultados sugerem que o consumo diário do extrato etanólico da P. cincinnata pode trazer benefícios à saúde.

Indiana University-Purdue University Indianapolis (IUPUI) / Platelet-activating Factor (PAF) stimulates various cell types by the activation of the G-protein coupled PAF-receptor (PAFR). Systemic PAFR activation induces an acute pro-inflammatory response, as well as delayed systemic immunosuppressive effects in vivo. De novo enzymatic PAF synthesis and degradation are closely regulated, but oxidative stressors, such as UVB, and cigarette smoke, can generate PAF-like species via the oxidation of membrane lipids in an unregulated process. Mast cells (MCs) and the PAFR have been shown to be necessary to mediate the resulting systemic immune suppression from oxidative stressors. The work herein implicates pro-oxidative chemotherapeutics, such as melphalan and etoposide, in mediating augmentation in tumor growth by inducing the generation of PAFR agonists via the oxidation of membrane lipids. This work also demonstrates the role of MCs and MC-released mediators in PAFR systemic immunosuppression. Through a contact hypersensitivity (CHS) model, the MC PAFR was found to be necessary and sufficient for PAF to mediate systemic immunosuppression. Additionally, activation of the MC PAFR seems to induce MC histamine and prostaglandin E2 release. Furthermore, by transplanting histamine- or COX-2-deficient MCs into MC-deficient mice, MC-derived histamine and prostaglandin release were found to be necessary for PAF to induce systemic immunosuppression. Lastly, we have evidence to suggest that prostaglandin release modulates MC migration to draining lymph nodes, a process necessary to promote immunosuppression. These studies fit with the hypothesis that MC PAFR activation mediates PAFR systemic immunosuppression in part by histamine and prostaglandin release.

Indiana University-Purdue University Indianapolis (IUPUI) / Neurofibromatosis type 1 is a genetic disease that results from either heritable or spontaneous autosomal dominant mutations in the NF1 gene, which encodes a protein serving, at least in part, to accelerate the intrinsic hydrolysis of active Ras-GTP to inactive Ras-GDP. A second-hit NF1 mutation precedes predominant NF1 neoplasms, including juvenile myelomoncytic leukemia (JMML) and plexiform neurofibroma formation, potentially fatal conditions with no medical therapy. While NF1 loss of heterozygosity (LOH) in myeloid progenitor cells sufficiently engenders leukemogenesis, plexiform neurofibroma formation depends on LOH in Schwann cells and Nf1 heterozygosity in the hematopoietic system. Specifically, recruited Nf1+/- mast cells accelerate tumorigenesis through secreted cytokines and growth factors. Nf1+/- mast cells depend upon deregulated signaling in c-kit pathways, a receptor system conserved in hematopoietic stem cells (HSCs). Accordingly, Nf1-/- myeloid progenitor cells, which can induce a JMML-like disease in mice, also demonstrate deregulated c-kit receptor signaling. C-kit-activated Nf1+/- mast cells and Nf1-/- myeloid progenitors both show increased latency and potency of active Erk1 and Erk2, the principal cytosolic-to-nuclear effectors of canonical Ras-Raf-Mek signaling. Thus, Erk represents a potential regulator of leukemogenesis and tumor-associated inflammation. However, single and combined Erk1 and Erk2 roles in HSC function, myelopoiesis, and mature mast cell physiology remain unknown, and recent hematopoietic studies relying on chemical Mek-Erk inhibitors have produced conflicting results. Here, we show that hematopoietic stability, myelopoiesis, and mast cell generation require Erk1 or Erk2, but individual isoforms are largely dispensable. Principally, Erk-disrupted hematopoietic stem cells incorporate BrdU but are incapable of dividing, a novel and cell type-specific Erk function. Similarly, mast cell proliferation requires Erk but cytokine production proceeds through other pathways, elucidating molecule-specific functions within the c-kit cascade. Based on these findings, we have reduced tumor mast cell infiltration by treating genetically-engineered tumor model mice with PD0325901, a preclinical Mek-Erk inhibitor. Moreover, we have devised a quadruple transgenic HSC transplantation model to examine dual Erk disruption in the context of Nf1 nullizygosity, testing whether diseased hematopoiesis requires Erk. These insights illuminate cell-specific Erk functions in normal and Nf1-deficient hematopoiesis, informing the feasibility of targeting Mek-Erk in NF1-associated disease.

<p>Mast cells (MCs) are increasingly recognized as powerful sentinel cells responsible for modulating the early immune responses to a wide range of infectious agents. This protective role is attributable in part to their preponderance at the host-environment interface and their innate capacity to rapidly release modulators of immune cell trafficking which promotes the early recruitment of pathogen-clearing immune cells from the blood. However, host-adapted pathogens had been a critical threat to human for a long time because they have evolved mechanisms directed at overcoming protective immunity. </p><p>In this work, we outline <italic>Salmonella enterica</italic> serovar Typhimurium has evolved a novel mechanism to inactivate peripheral MCs resulting in limited neutrophil responses at infection sites in early stage of infection. Because of the delay in bacterial clearance at the point of entry, <italic>Salmonella</italic> are able to multiply and rapidly disseminate to distal sites. Suppression of local MCs' degranulation restricted outflow of vascular contents into infection sites, thus facilitating bacterial spread. </p><p>We discover MC suppression is mediated by the Salmonella Protein Tyrosine Phosphatase (SptP), which shares structural homology with <italic>Yersinia</italic> YopH. Interestingly, SptP, not only shares homology with phosphatases found in MCs, they are also homologous to YopH an effector protein expressed by plague causing <italic>Yersinia pestis</italic>. We show that YopH had MC suppressing abilities as SptP suggesting that this activity is shared among some of the more virulent bacterial pathogens. The functionally relevant domain in SptP is its enzymatic site and that it works by dephosphorylating the vesicle fusion protein N-ethylmalemide-sensitive factor (NSF) and by blocking phosphorylation of Syk, which is located in downstream and upstream of tyrosine phosphorylation signaling pathway in MCs. </p><p>Without SptP, orally challenged <italic>S.</italic> Typhimurium failed to suppress MC degranulation and exhibited limited colonization of the mesenteric lymph nodes. Administration of SptP to sites of Escherichia coli infection markedly enhanced its virulence. Thus, SptP-mediated inactivation of local MCs is a powerful mechanism utilized by <italic>S.</italic> Typhimurium to impede early innate immunity. This finding provides a logical explanation for why previous attempts by others to demonstrate a protective role for MCs against <italic>Salmonella</italic> infections have resulted in equivocal results. </p><p>Taken together, this work highlights an overlooked virulence mechanism possessed by certain host adapted pathogens to avoid the host's innate immune system. Additionally, this innate immune-quelling property of SptP may hold future promise in tempering harmful inflammatory disorders in the body of an immune competent host.</p> / Dissertation

La peau est un « avant poste » fascinant du système immunitaire. Elle forme une barrière entre l'environnement extérieur et l’organisme. Elle est aussi le point d'entrée pour les agents pathogènes, contre lesquels le système immunitaire organise des réponses adaptatives. Les acteurs de l'immunité innée de la peau contrôlent l'invasion des pathogènes et perçoivent également des changements environnementaux physiques et chimiques directs. Plusieurs composants du système immunitaire, tels que des cellules dendritiques (DCs), les macrophages (MΦ) et les mastocytes (MCs), participent à l'éradication des pathogènes et à l'initiation des réponses mémoires adaptatives. Ce qui permet une mobilisation rapide des cellules T effectrices ainsi que la sécrétion des anticorps par les cellules B à la suite d’une seconde exposition aux agents pathogènes. Les MCs qui sont des cellules résidentes du derme, jouent un rôle déterminant dans la libération de signaux d’alertes et sont classiquement considérés comme des cellules effectrices de la réaction allergique cutanée liée à l'IgE. Plusieurs observations récentes indiquent que les MCs seraient aussi impliqués dans les processus immunorégulateurs lors de l'initiation des réponses immunitaires adaptatives, dans le maintien de la tolérance périphérique aux composants de la peau et dans la régénération de la peau au cours des processus de cicatrisation. Cependant, les interactions entre les MCs et d'autres cellules immunitaires innées et adaptatives recrutées dans des conditions inflammatoires cutanées n'ont pas été élucidées en détail. Dans ce travail, nous décrivons l'utilisation d'une nouvelle souris possédant des MCs fluorescents (RMB), dans laquelle nous avons marqué les MCs FcεRI+ avec un marqueur fluorescent rouge tomato (TdT) et avec un système d'ablation conditionnelle basé sur l'expression concurrente du récepteur de la toxine diphtérique (DTR). Avec ces souris RMB, nous avons visualisé la dynamique des MCs et nous avons suivi les interactions entre les MCs et les lymphocytes T régulateurs (Tregs) après l'activation des MCs par l'IgE, dans une réaction inflammatoire typique de l'anaphylaxie cutanée passive (PCA). Dans un second volet d’étude, nous avons évalué le rôle des MCs lors d'un modèle expérimental de la greffe de peau de l'oreille, afin de révéler leur influence dans la cinétique de rejet ou prise de greffe du transplant. Nous avons constaté que 1) l'activation et la dégranulation des MCs induites par le pontage du récepteur FcεRI via des IgE couplées à un antigène multivalent sont les seules responsables de la réaction de PCA, et induisent le recrutement de Tregs ayant une grande motilité sur le site de l'inflammation. Nous avons constaté dans ces conditions, que les MCs restent immobiles, et que les Tregs établissent des contacts dynamiques avec les MCs dans le derme. 2) En outre, nous avons mis en place un modèle pour identifier les paramètres moléculaires de l'interaction MC-Treg et avons constaté que le complexe de l'antigène avec l'IgE peut être présenté aux Tregs en association avec les molécules du complexe majeur d'histocompatibilité de classe II, permettant la formation des contacts stables MC-Treg. 3) En utilisant un modèle de transplantation de la peau in vivo, nous avons montré que l'ablation conditionnelle des MCs conduit à une accélération du rejet du greffon dans le cas d'une transplantation en présence d’une disparité d’antigènes d’histocompatibilité mineurs depuis une souris mâle sur une souris femelle. Nous avons également constaté un impact inattendu de l'ablation des MCs dans la greffe de peau en l’absence de disparité antigénique d'une souris femelle sur une souris femelle, conduisant à un rejet rapide. Les MCs semblent donc être essentiels pour la cicatrisation et la régénération tissulaire après greffe. (...) / The skin is a fascinating outpost of the immune system. It performs a barrier function between the outside environment and the inner body and is also a port of entry for pathogens against which the immune system mounts adapted responses. The skin innate immune defenses control pathogen invasion and perceive also direct physical and chemical environmental changes. Several component of the immune system such as dendritic cells (DC), macrophages (MΦ) and mast cells (MC) participate in initial pathogen clearance and in initiating adaptive memory responses, allowing rapid mobilization of effector T cells and secretion of B cellderived antibodies after secondary pathogen challenge. MCs residing in the dermis exert a determinant alert function through the liberation of various factors and are classically considered as effector cells in the IgE-mediated cutaneous allergic reaction. As emerging now, MC are also involved in immunoregulatory processes during the initiation of adaptive immune responses, the maintenance of peripheral tolerance to skin components and skin regeneration during wound healing. Yet, the crosstalks between MCs and other innate and adaptive immune cells recruited during cutaneous inflammatory conditions have not been elucidated in detail. Here, we report the use of a novel Mast cell fluorescent reporter mouse (RMB), in which we tagged FcεRI+ MCs, with red fluorescence marker tomato (Tdt) and with a conditional ablation system based on concurrent diphtheria toxin receptor (DTR) expression. Using these RMB mice, we visualized MC dynamics and monitored MC interactions with regulatory T lymphocytes (Tregs) after IgE-mediated activation of MCs, in a typical passive cutaneous anaphylaxis (PCA) inflammatory reaction. Using another setting, we further assessed the role of MC during experimental ear skin grafting to reveal their potential influence in skin grafting and rejection. We found that 1) the activation and degranulation of MCs induced by FcεRI crosslinking by multivalent IgE is solely responsible for the PCA reaction and induces the recruitment of highly motile regulatory T cells (Tregs) to the site of inflammation. In these conditions, we found that MC remain sessile and Tregs establish dynamic contacts with MC in the dermis. 2) Further we set up a model system to reveal the molecular requirement for MC-Treg interaction and found that antigen complexed with IgE were able to be presented to Treg in association with major histocompatibility complex class II molecules allowing the formation of stable MC-Treg contacts. 3) Using in vivo skin transplantation model, we showed that conditional ablation of MCs leads to an acceleration of skin transplant rejection in sex-mismatched model (male skin transplant to female). We also found an unexpected impact of MC conditional ablation in sex-matched skin graft (female skin transplant to female) leading to rapid rejection, implying that MCs are essential for the wound healing reaction and the regeneration of tissue continuity after grafting. The aforementioned results point out to an important immunoregulatory role of MC beyond their classically described activator functions in inflamed tissues. The fact that MC constantly interact with Treg during inflammatory processes suggest that MCs could participate in skin homeostasis by exerting tolerogenic functions. These functions remain to be elucidated at the molecular level as presented in the discussion.

<p>Mast cells (MC) are potent inflammatory cells that are known primarily for their prominent role in IgE mediated allergies. However, they also provide beneficial functions to the host, e.g. in bacterial and parasitic defence. MCs react rapidly upon stimulation by releasing potent granule-stored mediators, and serine proteases of the chymase or tryptase families are such major granule constituents. </p><p>As a first step towards a better understanding of the biological function of these proteases, we have determined the extended cleavage specificities of four mammalian mast cell chymases, by utilizing a substrate phage display approach. The specificities of these enzymes have then been used to compare their functional characteristics.</p><p>The major mucosal MC chymase in mice, mMCP-1, was found to possess a strict preference in four amino acid positions of the peptide substrate. Using this sequence to search the mouse proteome for potential <i>in vivo</i> substrates led to the identification of several very interesting potential novel substrates. Some of them may explain the increased epithelial permeability provided by this enzyme.</p><p>Human MCs, express only one single α-chymase, and the rodent α-chymases have secondarily gained elastase-like primary cleavage specificity. However, rodents express additional chymases, the β-chymases, and rodent β-chymases may have adopted the function of the α-chymases. The cleavage specificities of the human chymase and two rodent β-chymases were therefore determined (rat rMCP-1 and mouse mMCP-4). N-terminal of the cleaved bond the three chymases showed similar preferences, but C-terminal the human chymase and mMCP-4 shared a high preference for acidic amino acids in the P2´ position and therefore seem to be functional homologues. The molecular interactions mediating the preference for acidic amino acids in position P2´ were further investigated. By site-directed mutagenesis of the human chymase, amino acids Arg143 and Lys192 were concluded to synergistically mediate this preference.</p><p>Our data show that chymases, of different MC subpopulations, display quite different extended cleavage specificities. However mouse do possess a MC chymase with almost identical cleavage specificity as the human MC chymase indicating a strong evolutionary pressure to maintain this enzyme specificity.</p>

Implanted biomaterials often induce inflammation that frequently leads to the foreign body response, fibrosis, and the failure of the implant. Thus, it is important to evaluate how cells interact with materials to promote a more regenerative response. It is critical to determine how to modulate the response of tissue resident innate immune cells, as they are among the first cells to interact with implanted materials. Among tissue resident innate immune cells are mast cells, which are inflammatory sentinels that degranulate and orchestrate the fate of other cell populations, such as monocytes/macrophages and lymphocytes. Mast cells have also been reported to play a vital role in the foreign body response of implanted biomaterials as well as angiogenesis. The goal of this study was to determine how to modulate mast cell responses to electrospun scaffolds by altering scaffold architecture and composition to promote anti-inflammatory and regenerative cell-scaffold interactions. Scaffold architecture was manipulated by changing either fiber diameter or pore diameter and mast cell responses were mediated by endogenous and exogenous DAMPs (i.e. IL-33 and LPS, respectively). Particularly in response to IL-33, scaffolds with increased fiber and pore diameter promoted less inflammatory cytokine and chemokine release while increasing angiogenic cytokine release. Additionally, taking scaffolds that promoted increased inflammatory cytokine expression and increasing the pore diameter alone dampened inflammatory cytokine expression. The next question we wanted to answer was how might the degradative byproducts of scaffolds alter mast cell inflammatory responses. Given the widespread use of polylactic acid, we decided to investigate this question using lactic acid as a degradative byproduct. In the presence of physiologically relevant levels of lactic acid, IL-33- and IgE-mediated inflammatory cytokines and chemokines are suppressed, while angiogenic cytokines are enhanced. This response was shown to be pH- and MCT1-dependent and was recapitulated in primary human skin mast cells as well as in vivo. In summary, scaffold architecture and the presence of select polymer degradative byproducts have the potential of selectively suppressing inflammatory cytokines and enhancing angiogenic cytokines.

<p>Akrilamid je supstanca koja se prirodno stvara pečenjem i prženjem hrane bogate skrobom.&nbsp; Cilj rada je bio da se ispita subhronični i akutni uticaj akrilamida na debelo&nbsp;crevo prepubertalnih pacova. Dobijeni rezultati su pokazali da akrilamid ne naru&scaron;ava morfologiju zida creva, ali dovodi do promena u volumenskoj gustini njegovih tunika i lamina. Najizraženije promene&nbsp; su bile na&nbsp; vezivnom tkivu&nbsp; debelog creva.&nbsp; Analizom peharastih ćelija&nbsp; i sadržaja mucina pokazano je da akrilamid utiče i na ugljenohidratnu i na proteinsku komponentu mucina. Subhronični tretman je&nbsp; doveo do smanjenja broja&nbsp;limfocita i&nbsp; eozinofila, a&nbsp; kod akutnog tretmana&nbsp; je primećeno nakupljanje limfocita i eozinofila u kolonu akrilamidom tretiranih jedinki. Broj mastocita je u sva tri eksperimenta bio smanjen kod tretiranih životinja. Duže izlaganje akrilamidu ima imunosupresivno dejstvo kod pacova.</p> / <p>Acrylamide is natural product of cooking (baking, roasting) starchy food.&nbsp; Aim of the study was to evaluate the risk of subchronic and acute acrylamide treatment on juvenile&nbsp; rat colon.&nbsp; Changes&nbsp; in colon wall morphology was detected by stereological methods since histological evaluation reveal normal colon architecture&nbsp; after acrylamide intoxication.&nbsp; The changes was&nbsp; most prominent on&nbsp; connective tissue of rat colon.&nbsp; Acrylamide affected both protein&nbsp; component of mucins and glycans linked to peptide backbone.&nbsp; In subchronic&nbsp; treatment acrylamide caused reduction of lymphocytes&nbsp; and eosinophils &nbsp;number, while acute experiment lead to lymphocytes&nbsp; and&nbsp; eosinophils&nbsp; accumulation in colon tissue. Acrylamide intoxication decreased mast cell number in all experiments. Longer acrylamide exposure had immunosuppressive effect in rats.</p>

La peau est un « avant poste » fascinant du système immunitaire. Elle forme une barrière entre l'environnement extérieur et l’organisme. Elle est aussi le point d'entrée pour les agents pathogènes, contre lesquels le système immunitaire organise des réponses adaptatives. Les acteurs de l'immunité innée de la peau contrôlent l'invasion des pathogènes et perçoivent également des changements environnementaux physiques et chimiques directs. Plusieurs composants du système immunitaire, tels que des cellules dendritiques (DCs), les macrophages (MΦ) et les mastocytes (MCs), participent à l'éradication des pathogènes et à l'initiation des réponses mémoires adaptatives. Ce qui permet une mobilisation rapide des cellules T effectrices ainsi que la sécrétion des anticorps par les cellules B à la suite d’une seconde exposition aux agents pathogènes. Les MCs qui sont des cellules résidentes du derme, jouent un rôle déterminant dans la libération de signaux d’alertes et sont classiquement considérés comme des cellules effectrices de la réaction allergique cutanée liée à l'IgE. Plusieurs observations récentes indiquent que les MCs seraient aussi impliqués dans les processus immunorégulateurs lors de l'initiation des réponses immunitaires adaptatives, dans le maintien de la tolérance périphérique aux composants de la peau et dans la régénération de la peau au cours des processus de cicatrisation. Cependant, les interactions entre les MCs et d'autres cellules immunitaires innées et adaptatives recrutées dans des conditions inflammatoires cutanées n'ont pas été élucidées en détail. Dans ce travail, nous décrivons l'utilisation d'une nouvelle souris possédant des MCs fluorescents (RMB), dans laquelle nous avons marqué les MCs FcεRI+ avec un marqueur fluorescent rouge tomato (TdT) et avec un système d'ablation conditionnelle basé sur l'expression concurrente du récepteur de la toxine diphtérique (DTR). Avec ces souris RMB, nous avons visualisé la dynamique des MCs et nous avons suivi les interactions entre les MCs et les lymphocytes T régulateurs (Tregs) après l'activation des MCs par l'IgE, dans une réaction inflammatoire typique de l'anaphylaxie cutanée passive (PCA). Dans un second volet d’étude, nous avons évalué le rôle des MCs lors d'un modèle expérimental de la greffe de peau de l'oreille, afin de révéler leur influence dans la cinétique de rejet ou prise de greffe du transplant. Nous avons constaté que 1) l'activation et la dégranulation des MCs induites par le pontage du récepteur FcεRI via des IgE couplées à un antigène multivalent sont les seules responsables de la réaction de PCA, et induisent le recrutement de Tregs ayant une grande motilité sur le site de l'inflammation. Nous avons constaté dans ces conditions, que les MCs restent immobiles, et que les Tregs établissent des contacts dynamiques avec les MCs dans le derme. 2) En outre, nous avons mis en place un modèle pour identifier les paramètres moléculaires de l'interaction MC-Treg et avons constaté que le complexe de l'antigène avec l'IgE peut être présenté aux Tregs en association avec les molécules du complexe majeur d'histocompatibilité de classe II, permettant la formation des contacts stables MC-Treg. 3) En utilisant un modèle de transplantation de la peau in vivo, nous avons montré que l'ablation conditionnelle des MCs conduit à une accélération du rejet du greffon dans le cas d'une transplantation en présence d’une disparité d’antigènes d’histocompatibilité mineurs depuis une souris mâle sur une souris femelle. Nous avons également constaté un impact inattendu de l'ablation des MCs dans la greffe de peau en l’absence de disparité antigénique d'une souris femelle sur une souris femelle, conduisant à un rejet rapide. Les MCs semblent donc être essentiels pour la cicatrisation et la régénération tissulaire après greffe. (...) / The skin is a fascinating outpost of the immune system. It performs a barrier function between the outside environment and the inner body and is also a port of entry for pathogens against which the immune system mounts adapted responses. The skin innate immune defenses control pathogen invasion and perceive also direct physical and chemical environmental changes. Several component of the immune system such as dendritic cells (DC), macrophages (MΦ) and mast cells (MC) participate in initial pathogen clearance and in initiating adaptive memory responses, allowing rapid mobilization of effector T cells and secretion of B cellderived antibodies after secondary pathogen challenge. MCs residing in the dermis exert a determinant alert function through the liberation of various factors and are classically considered as effector cells in the IgE-mediated cutaneous allergic reaction. As emerging now, MC are also involved in immunoregulatory processes during the initiation of adaptive immune responses, the maintenance of peripheral tolerance to skin components and skin regeneration during wound healing. Yet, the crosstalks between MCs and other innate and adaptive immune cells recruited during cutaneous inflammatory conditions have not been elucidated in detail. Here, we report the use of a novel Mast cell fluorescent reporter mouse (RMB), in which we tagged FcεRI+ MCs, with red fluorescence marker tomato (Tdt) and with a conditional ablation system based on concurrent diphtheria toxin receptor (DTR) expression. Using these RMB mice, we visualized MC dynamics and monitored MC interactions with regulatory T lymphocytes (Tregs) after IgE-mediated activation of MCs, in a typical passive cutaneous anaphylaxis (PCA) inflammatory reaction. Using another setting, we further assessed the role of MC during experimental ear skin grafting to reveal their potential influence in skin grafting and rejection. We found that 1) the activation and degranulation of MCs induced by FcεRI crosslinking by multivalent IgE is solely responsible for the PCA reaction and induces the recruitment of highly motile regulatory T cells (Tregs) to the site of inflammation. In these conditions, we found that MC remain sessile and Tregs establish dynamic contacts with MC in the dermis. 2) Further we set up a model system to reveal the molecular requirement for MC-Treg interaction and found that antigen complexed with IgE were able to be presented to Treg in association with major histocompatibility complex class II molecules allowing the formation of stable MC-Treg contacts. 3) Using in vivo skin transplantation model, we showed that conditional ablation of MCs leads to an acceleration of skin transplant rejection in sex-mismatched model (male skin transplant to female). We also found an unexpected impact of MC conditional ablation in sex-matched skin graft (female skin transplant to female) leading to rapid rejection, implying that MCs are essential for the wound healing reaction and the regeneration of tissue continuity after grafting. The aforementioned results point out to an important immunoregulatory role of MC beyond their classically described activator functions in inflamed tissues. The fact that MC constantly interact with Treg during inflammatory processes suggest that MCs could participate in skin homeostasis by exerting tolerogenic functions. These functions remain to be elucidated at the molecular level as presented in the discussion.