Moléculas HLA-DR e co-estimulatórias tem papel central na função imune de leucócitos. Diferentes emulsões lipídicas (EL) podem alterar funções imunes de leucócitos. Para avaliar os efeitos de diferentes EL sobre a expressão de moléculas HLA-DR, CD80 e CD86 presentes na superfície de monócitos/macrófagos (MO/M?) e CD28 e CD152 presentes na superfície de linfócitos T auxiliares (L? CD4) humanos, células mononucleares do sangue periférico de voluntários saudáveis (n=10) foram separadas com uso de Ficoll Hypaque (d=1,007) e incubadas por 48 horas (MO/M?) e 72 horas (L?) em meio RPMI 1640 acrescidas ou não (controle negativo) de diferentes EL comerciais ou misturas experimentais na concentração de 1mg/mL. De acordo com o tipo da emulsão lipídica adicionada ao meio de cultura, as células foram divididas em seis grupos experimentais: a) Controle negativo - células mononucleares cultivadas sem o acréscimo de EL b) TCLn-6 - células mononucleares cultivadas com EL a base de óleo de soja rica em ácidos graxos poliinsaturados tipo n-6 (AGPI n-6), c) TCLn-6/TCLn-3 - células mononucleares cultivadas com mistura experimental contendo 80% da EL a base de óleo de soja e 20% de EL a base de óleo de peixe rica em AGPI tipo n-3, d) TCM/TCLn6 - células mononucleares cultivadas com EL composta por 50% óleo de coco, rico em triglicérides de cadeia média e 50% de óleo de soja, e) TCM/TCLn-3 - células mononucleares cultivadas com mistura experimental contendo 80% de EL composta por 50% óleo de coco e 50% de óleo de soja e 20% de EL a base de óleo de peixe, f) SMOF - células mononucleares cultivadas com a nova EL contendo 30% de óleo de soja, 30% de triglicérides de cadeia média, 25% de óleo de oliva e 15% de óleo de peixe. As células mononucleares foram ativadas pelo uso de 10?g/mL de fitohemaglutinina. A expressão das moléculas de superfície foi analisada por citometria de fluxo. A porcentagem de fluorescência, que indica o número de células expressando as moléculas em estudo e a intensidade de fluorescência, que indica de forma indireta o número de moléculas expressas por células, foram medidas. Os resultados obtidos foram submetidos à teste estatístico Friedman e pós-teste Student-Newman-Keuls, adotando-se nível de significância de p<0,05. Devido às diferenças na expressão basal dos doadores, os resultados de intensidade de fluorescência foram transformados em porcentagem relativa ao controle basal (Basal=100). Nos grupos TCLn-6, TCLn-6/TCLn-3, TCM/TCLn-6, TCM/TCLn-3 e SMOF, a intensidade de fluorescência de moléculas HLA-DR expressas na superfície de monócitos/macrófagos diminuiu (medianas = 87,6; 84,0; 81,0; 85,0 e 80,0 respectivamente) em relação ao controle negativo (CN) (mediana=100,0) p=0,01. Todos os grupos tratados com EL aumentaram o número de linfócitos T auxiliares expressando moléculas CD28 (medianas = 90,9; 90,4; 91,5; 92,6 e 90,1 respectivamente) em relação ao CN (mediana=82,8) p=0,001 e também o número de moléculas CD152 expressas por células na superfície de linfócitos T auxiliares (medianas = 120,6; 108,8; 127,7; 114,6 e 121,3 respectivamente) em relação ao CN (mediana=100,0), p=0,03. Não foram encontradas diferenças estatísticas na expressão de moléculas CD80 e CD86 na superfície de monócitos/macrófagos cultivados com diferentes EL. Ainda não foram encontradas diferenças no número de linfócitos T auxiliares expressando CD152. Finalmente a expressão por células de moléculas CD28 na superfície de linfócitos T auxiliares também não mostrou alteração significante com as diferentes emulsões lipídicas. Conclusão: Emulsões lipídicas parenterais in vitro, diminuem a expressão de moléculas HLA-DR na superfície de monócitos/macrófagos e aumentam a expressão de moléculas CD28 e CD152 na superfície de linfócitos T auxiliares humanos. Estas alterações podem ser um dos mecanismos pelos quais as EL modulam funções de células imunes / HLA-DR and co-stimulatory molecules play a central role on leucocytes immune function. Different lipid emulsions (LE) may change leucocytes immune function. It is of interest to study the effect of different LE on HLA-DR and costimulatory molecules expression. To access the effect of LE on the HLA-DR, CD80 and CD86 expression on monocytes/macrophages (MO/MØ) surface and CD28 and CD152 (CD80/CD86 co-stimulatory molecules receptor) expression on human T helper lymphocytes (LØ CD4) surface we obtained mononuclear cells from peripheral blood of healthy volunteers (n=10) by using ficoll hypaque (d=1.077). The cells were cultured for 48 hours (MOMØ) and 72 hours (LØ CD4) and incubated with RPMI 1640 medium without (negative control) or added with 1mg/mL of commercial or experimental mixtures of five LE. Groups: a) NC - negative control without LE, b) LCTn-6 - n-6 polyunsaturated fatty acids (PUFA) rich LE ( soybean oil), c) LCTFO - 80% of LCT and 20% of n-3 PUFA rich LE (FO) (fish oil), d) MCT/LCT - LE containing 50% of medium chain triglycerides and 50% of n-6 PUFA rich LE, e) MCT/LCTFO - 80% of MCT/LCT LE and 20% of FO LE and f) SMOF - a new LE containing 30% of soybean oil, 30% of medium chain triglycerides, 25% of olive oil and 15% of fish oil. Mononuclear cells were activated by using 10?g/mL of phytohemagglutinin. Surface molecules expression was measured by flow cytometry. Percentage and intensity of fluorescence were recorded and the data were submitted to Friedman statistical test and Student-Newman-Keuls post test (p<0,05). Due the differences in basal expression between donors, prior to statistical tests, data from intensity of fluorescence were transformed of percentage relative of basal expression (where basal=100). All LE groups LCT, LCTFO, MCT/LCT, MCT/LCTFO and SMOF decreased HLA-DR intensity of fluorescence on monocytes/macrophages (mean= 87.6, 84.0, 81.0, 85.0, and 80.0 respectively) in relation to negative control (NC) (mean=100.0) cultured without LE (p=0,01). All LE groups increased the percentage of lymphocytes expressing CD28 (means=90.9, 90.4, 91.5, 92.6 and 90.1 respectively) in relation to control (mean=82.8) p=0,001 and CD152 intensity of fluorescence on lymphocytes cultured with all different LE (mean=120,6; 108,8; 127,7; 114,6 and 121,3 respectively) in relation to NC (mean=100,0), p=0,03. No significant differences were found on CD80 and CD86 expression on monocytes/macrophages surface, CD28 intensity of fluorescence and the percentage of lymphocytes expressing CD152 on lymphocytes cultured with the different studied LE. Conclusion: In vitro parenteral LE decreased HLA-DR expression on human monocytes/macrophages surface and increase co-stimulatory molecules receptor expression on human lymphocytes surface. These changes could be one of the mechanisms of LE modulation of immune cells functions

Made available in DSpace on 2016-08-10T10:55:55Z (GMT). No. of bitstreams: 1 Fernanda de Oliveira Feitosa de Castro.pdf: 485024 bytes, checksum: 876ee7e1b14c3a6b20a7d9aef9cbe13a (MD5) Previous issue date: 2011-02-25 / Snake s venoms are complexes mixtures of proteins and peptides, aminoacids, nucleotides, lipids and carbohydrates, that showes importants biologicals activities like cytotoxicity, neurotoxicity, systemic myolysis, cardiotoxicity, kidney damage, hematologic disorders, like others. These molecules has shown pharmacological potential components with potent action against bacterias, virus and fungis. This present work aimed assed no cytotoxic effect of the crude venom of the snake Bothrops pauloensis in the culture of peripheral blood mononuclear cells (PBMC). After standardization of obtaining PBMC and evaluation of ideal concentration of fitohemagglutinin (PHA) and interleukin-2 (IL-2) for the cell culture was performed the evaluation of no citotoxic effect on CMN by different doses of crude venom of Bothrops pauloensis. The CMN was separated by density gradient, posterior being cultured. The evaluation of the no cytotoxic activity of the venom was performer by the visualization of the plate after 24, 48 and 72 hours. The concentration 0,05 mg/ml did not appear citotoxic activity, suggesting the possibility of using in testing with CMN infected by viruses, bacteria and fungi. / Os venenos de serpentes são misturas complexas compostas por proteínas e peptídeos, aminoácidos, nucleotídeos, lipídios e carboidratos que apresentam importantes atividades biológicas como citotoxicidade, neurotoxicidade, miólise sistêmica, cardiotoxicidade, danos renais, desordens hematológicas entre outras. Essas moléculas têm-se mostrado como potenciais componentes farmacológicos com potente ação contra a atividade de bactérias, vírus e fungos. O presente trabalho visou avaliar a atividade não citotóxica do veneno bruto da cobra Bothrops pauloensis em células mononucleares (CMN) do sangue periférico humano. Após a padronização da obtenção de CMN do sangue periférico e da concentração ideal de fitohemaglutinina(PHA) e interleucina-2 (IL- 2) para cultivo celular, foi realizada a avaliação do efeito não citotóxico de diferentes concentrações do veneno bruto da Bothrops pauloensis no cultivo celular. As células mononucleares do sangue periférico foram separadas por meio gradiente de densidade e cultivadas em meio de cultura. A avaliação da atividade citotóxica do veneno foi realizada pela visualização da placa após 24, 48 e 72 horas. A concentração 0,05 &#956;g/ml não apresentou atividade citotóxica, sugerindo a possibilidade de utilização em ensaio com CMN infectadas por vírus, bactérias e fungos.

Doenças pulmonares intersticiais fibrosantes são doenças que afetam homens, mulheres e crianças, tem prognóstico ruim e os pacientes possuem sobrevida estimada entre 3 e 5 anos após a confirmação diagnóstica, sobretudo os portadores de Fibrose pulmonar idiopática. Estudos recentes demonstram a capacidade das células-tronco em se diferenciar em diferentes linhagens celulares e tecidos, como já comprovado em órgãos como coração, fígado, trato gastrointestinal, sistema nervoso e pulmão. Os objetivos deste trabalho foram os de estabelecer a espécie suína como modelo experimental e utilizar a terapia celular experimentalmente como possibilidade de estudo para tratamento das doenças pulmonares intersticiais fibrosantes. Para estabelecer o modelo experimental e induzir a doença nos dois grupos de animais estudados (grupos tratado e controle) foi utilizado sulfato de bleomicina pela via intratraqueal em procedimento único. Após a instalação da doença, os animais dos grupos tratado e controle foram submetidos a tomografia computadorizada de alta resolução (TCAR); um grupo tratado com terapia celular e após noventa dias os dois grupos reavaliados com TCAR antes da eutanásia, totalizando para os dois grupos, cento e oitenta dias de doença instalada. As análises tomográficas mostraram que o tempo para que a doença intersticial seja estabelecida ocorre até três meses após a infusão de bleomicina. As provas histológicas corroboram a viabilidade do modelo testado e as análises imunohistoquímicas sugerem a migração das células mononucleares de medula óssea para os pulmões, bem como a presença de populações celulares que indicam provável reestruturação do parênquima pulmonar. / Fibrotic Interstitial pulmonary illnesses affect men, women and children, and presents bad prognostic, 3-5 years depending on the diagnostic confirmation, mainly in idiopathic pulmonary fibrosis. Recent studies demonstrate the capacity of the stem cells in differentiating into different cellular lineages and different tissues, as in heart, liver, gastrointestinal, nervous system and lung. The objectives of this study were to investigate the possibility to consider the swine as experimental model of fibrotic pulmonary disease and experimental stem cell therapy. Bleomycin sulphate was injected into the trachea to induce the pulmonary disease in control and treatment groups. High resolution computed scan (TCAR) was carried out in both groups after the confirmation of the disease. The tomographic analyses showed that the interstitial illness was established after three months of the bleomicine infusion. Histologic investigation revealed the viability of the tested model and the imunohistochemical analyses suggest the migration of the mononuclear cells to the lungs, as well as the presence of new cellular populations that would indicate probable reorganization of the pulmonary parenchyma.

O Transtorno Depressivo Maior (TDM) é uma condição psiquiátrica que afeta 4,4% da população mundial, exibindo um substancial sofrimento pessoal, incapacidade e custos sociais, e estima-se que ele será a principal causa de incapacidade no mundo em 2030. O surgimento de novas ferramentas e modelos de pesquisa envolvendo o TDM irá auxiliar no entendimento desta doença. Deste modo, o objetivo deste trabalho foi gerar uma coleção de células-tronco pluripotentes induzidas humanas (hiPSCs) de um grupo de indivíduos com TDM. Foram coletadas amostras de células mononucleares (MNCs) de 66 indivíduos afetados, e geradas 6 linhagens de hiPSCs. Através de diversos testes de caracterização, a pluripotência destas células foi confirmada. Além disto, também foi padronizada a diferenciação destas hiPSCs em neurônios serotonérgicos. Neurônios derivados dessas hiPSCs poderão constituir material de estudo para outros grupos de pesquisa interessados no estudo da TDM, e ser utilizados em testes futuros para prever resposta a medicamentos. / Major Depressive Disorder (MDD) is a psychiatric condition that affects 4.4% of the world\'s population, exhibiting substantial personal suffering, disability and social costs, and is estimated to be the leading cause of disability in the world by 2030. Emergence of new tools and research models involving TDM will aid in the understanding of this disease. Thus, the objective of this work was to generate a collection of human induced pluripotent stem cells (hiPSCs) from a group of individuals with MDD. Samples of mononuclear cells (MNCs) of 66 affected individuals were collected, and 6 lines of hiPSCs were generated. Through several characterization tests, the pluripotency of these cells was confirmed. In addition, the differentiation of these hiPSCs into serotonergic neurons was also standardized. Neurons derived from these hiPSCs could constitute study material for other research groups interested in the study of MDD, and be used in future tests to predict drug response.

As células-tronco adultas possuem capacidade de se transformar em certos tipos de tecidos. Surge desta forma, uma nova modalidade de tratamento para doenças cardíacas, assim, uma avaliação na distribuição e quantificação do número de células por diferentes vias de infusão tornam-se importantes, pois o microambiente e interação célula/célula interferem no processo de transdiferenciação e/ou fusão. Foram utilizados nove cães, com peso entre 25 e 30kg e hígidos, no qual foi realizada punção, separação, marcação e infusão de células mononucleares da medula óssea pelas vias intracoronariana, transendocárdica e venosa retrógrada. Pôde-se observar a presença de células por todo tecido cardíaco em todas as vias, contudo o padrão de distribuição das mesmas difere, no qual há um predomínio de células em tecido conjuntivo, principalmente em epicárdio nas vias transendocárdica e venosa retrógrada e um padrão intersticial muscular intracoronariana. Há diferenças significativas na quantidade de células nas regiões apical, basal, medial e apical quando relacionada à via de infusão (P&lt;0,01). / The adult stem cells have a capacity to transform in different kinds of tissue. It is emerges like a new treatment modality of cardiac diseases, so, an evaluation in cell distribution and quantification by different routes of infusion in heart are necessary, because the microenvironment and the interaction cell/cell age in the transdifferentiation and fusion process. It was used 09 healthy mongrel dogs, with 25 -30kg of weight, which was performed a punction of iliacal crest, bone marrow mononuclear cells separation by Ficoll density, stained with Hoechst and infusion by intracoronary, transendocardial and retrograde routes. The marked cells was observed in all cardiac tissue, however, the pattern of distribution of the same ones differs, however in the transendocardial and retrograde route the cells could be founded mainly into the connective tissue - epicardium and sub-epicardic connective tissue and in the intracoronary route, the cells was observed mainly in the muscular interstice. It was observed a significant number of cells in the atrial, basal, medium and apical cardiac region when related with the route of infusion.

Macrophages are leukocytes of the innate immune system that display great phenotypic plasticity to mediate diverse functions. The ontogeny of tissue resident macrophages has been debated in recent decades. It is now recognised that tissue macrophages can be replenished from embryonically-derived precursors, and/or monocyte intermediates in a tissue specific manner. Interferon Regulatory Factor 5 (IRF5) is a transcription factor that promotes a pro-inflammatory phenotype in macrophages in vitro and in vivo. Indeed, IRF5 contributes to the pathogenesis of experimental inflammatory arthritis, lupus, and obesity via recruitment and activation of effector cells. Research described here as part of this thesis, involves the profiling of the intestinal Mononuclear Phagocyte system to investigate the role of IRF5 in the development of monocyte-derived macrophages in the Colonic Lamina Propria (cLP) which are exclusively replenished by adult Ly6C^hi monocytes. Using Mixed Bone Marrow Chimaeras (MBMCs) we showed that in shared environment Wild-Type (WT) cLP macrophages dominated IRF5-deficient (Irf5^-/-) cLP macrophages in both steady state and inflammation. The development of in vitro bone marrow derived macrophages, and the reconstitution of the haematopoietic compartment in bone marrow of MBMCs were not significantly affected by IRF5 deficiency. IRF5 promoted the accumulation of WT monocytes in the cLP of MBMCs in a process possibly dependent on the CCL2/CCR2 axis. Furthermore, IRF5 expression committed Ly6C^hi monocytes to a pro-inflammatory macrophage fate in the inflamed cLP, characterised by protein expression of the cytokines IL1β, and TNFα, and the expression of Ccl4 and Ccl8 transcripts, whilst loss of IRF5 favoured accumulation of CD11b⁺ IRF4-dependent Dendritic Cells. Of significance, IRF5 expression might have prevented further differentiation of inflammatory macrophages into tissue-resident macrophages, thus supporting an inflammatory state. Irf5-/- mice were protected from Helicobacter hepaticus + αIL10R colitis. Intriguingly, protection from colitis may also be conferred by the presence of Irf5-/- haematopoietic cells, evidenced by WT:Irf5-/- MBMCs . Modulation of IRF5 activity may therefore be a viable therapeutic strategy. RNA sequencing identified that C1q, Cd81, and Ccl8 were upregulated in WT macrophages from MBMC, which may prove therapeutic targets.

Les nouveau-nés (enfants, ruminants) sont particulièrement sensibles à l’infection intestinale par le parasite Cryptosporidium parvum car leur système immunitaire est encore en cours de développement. Peu de solutions de contrôle existent à ce jour. Il n’existe pas de vaccin et seule une molécule l’Halocur™ possède une AMM pour les veaux mais l’utilisation du traitement est contraignante et il peut présenter une toxicité pour l’animal. Le développement de nouvelles alternatives immunoprophylactiques requiert de mieux comprendre les mécanismes immunitaires mis en jeux lors de l’infection. L’immunité innée joue un rôle prépondérant pour le contrôle de la phase aigüe de l’infection et nous avions montré au laboratoire que les phagocytes mononucléés CD11c+ sont des acteurs déterminant dans le processus protection. Lors de cette thèse nous avons confirmé le rôle des cellules dendritiques (DC) CD103+ en utilisant des souriceaux BatF3-/- chez qui le développement des deux sous-populations CD103+CD11b+ et CD103+CD11b- est altéré au niveau intestinal ce qui rend les animaux beaucoup plus sensibles à l’infection. / Newborns (children, ruminants) are particularly susceptible to intestinal infection by the parasite Cryptosporidium parvum because their immune system is still developing. To date, parasite control methods are limited. There is no vaccine and the only molecule which possess a marketing authorization for calves, Halocur ™, presents toxicity at 2 times the therapeutic dose. The development of new immunoprophylactic methods requires better understanding of the immune mechanisms occurring during infection. Innate immunity plays a major role in controlling the acute phase of infection and we previously demonstrated in the laboratory that intestinal mononuclear phagocytes CD11c+ are key players in the protection process. In this thesis, we confirmed the role of dendritic cells (DC) CD103+ using mice BatF3-/- in which the development of the two DC subsets CD103+CD11b+ and CD103+CD11b- is altered in the intestine making these animals more susceptible to infection. This high susceptibility can be partially mitigated by preventive administration of IL-12 to Batf3-/- neonatal mice. Batf3-/- adult mice which are only deficient for the CD103+CD11b- DC subset were transiently susceptible to infection in contrast to conventional mice that are highly resistant.

Localisé entre l'Epithélium Pigmentaire Rétinien (EPR) et les segments externes des photorécepteurs, l'espace sous-rétinien est une zone immunosuppressive ; régulée par des signaux comme la thrombospondine-1 (TSP-1) ou Fas Ligand (FasL), qui empêchent l'accumulation des phagocytes mononucléés (PMs), en particulier des monocytes inflammatoires. La Dégénérescence Maculaire Liée à l'Age (DMLA) est associée à une rupture de l'immunosuppression de cet espace, et s'accompagne d'une accumulation de PMs ; causant la mort des photorécepteurs, la dédifférenciation de l'EPR et une néovascularisation pathologique. Des études d'associations génétiques ont établi un lien entre la DMLA et un haplotype qui affecte le locus 10q26, qui contient trois gènes : PLEKHA1, ARMS2 et HTRA1. L'haplotype est associé à une augmentation de la transcription de HTRA1 dans les lymphocytes ou les cellules de l'EPR. HTRA1 code pour une sérine protéase qui a une multitude de substrats ; mais le mécanisme par lequel elle pourrait être impliquée dans la pathogenèse de la DMLA reste inconnu. TSP-1 est une glycoprotéine exprimée par l'EPR, les macrophages résidents et inflammatoires. Le domaine C-terminal de TSP-1 contient deux séquences VVM qui peuvent chacune interagir avec un récepteur CD47. Dans cette étude, nous montrons que HTRA1 clive TSP-1 et inhibe l'élimination des PMs régulée par l'interaction entre TSP-1 et CD47 à l'état physiologique, in vitro et in vivo. L'activation pharmacologique de CD47 nous a permis d'annuler les effets pro-inflammatoires de HTRA1 et pourrait représenter un espoir thérapeutique pour le contrôle de la progression de la DMLA chez les patients porteurs de l'haplotype à risque. / Localized between the Retinal Pigment Epithelium (RPE) and the photoreceptors outer segments, the subretinal space is an immunosuppressive zone, mediated by signals such as Thrombospondin-1 (TSP-1), Fas Ligand (FasL) that prevent the accumulation of Mononuclear Phagocytes (MPs) and in particular pathogenic inflammatory monocytes. Age related Macular Degeneration (AMD) is associated with a breakdown of this immunosuppressivity and an accumulation of MPs, which causes photoreceptor degeneration, RPE dedifferentiation and pathological neovascularization. Genome association studies showed a strong link between AMD and a relatively common haplotype of 10q26 locus that contains the PLEKHA1, ARMS2 and HTRA1 genes. The disease haplotype is associated with increased HTRA1 transcription in cell types such as lymphocytes and RPE cells. HTRA1 is a serine protease with a number of substrates, but the mechanism by which it might be involved in AMD pathogenesis is unknown. TSP-1 is a glycoprotein expressed by RPE, resident macrophages and inflammatory macrophages. The C-terminal domain of TSP-1 contains two VVM sequences that can each interact with a CD47 receptor. We show that HTRA1 induced subretinal MP accumulation is dependent on TSP-1 deactivation in an RPE/Mo co-culture model and in a laser induced inflammation model in vivo. This pathogenic effect of HTRA1 was reversible by synthetic CD47 agonists. Our study reveals a comprehensive mechanism how the risk-allele 10q26 participates in the pathogenesis of AMD and opens new therapeutic avenues to restore subretinal immunosuppressivity and inhibit the inflammation-dependent neurodegeneration.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / Most injuries of the tendons in small animals are linked to trauma and the most important injuries are partial or complete section due to the action of sharp or acute objects or laceration associated with automobile accident leading to a functional disability of the member. The treatments offered are different, but effects such as improving the quality and speed of healing are not confirmed by controlled studies. Recently, the medical advances have shown growing interest in the use of cell therapy in treatment of degenerative diseases and also in slow or ineffective healing. This study aimed to evaluate the effects of the transplant autogenous of mononuclear cells of bone marrow with or without MEC in tendon healing. Compared tendons were treated only with mononuclears cells (GI); mononuclears cells embedded in MEC of collagen (GII); only MEC of collagen (GIII) with the control group (GIV), besides the presence of such cells in the new tissue. It was induced Achilles tendon injury trial in common law of the pelvic member in 36 dogs, randomly separated into four groups with nine experimental animals each, and followed by autogenou transplantation of at least 0,6 x 108 and a maximum 7,4 x 108 mononuclear cells. kg-1, with more than 90% viability, marked with nanocristal Q-tracker 655. The animals were evaluated by clinical and hematological parameters and through biopsy cyhtological features. Analyses performed at 7th, 14th and 30th days of evolution through cytological features analyses and direct fluorescent histopathological about the quality of healing tissue as: increasing of the inflammatory infiltrate, presence of extracellular matrix, increased cell proliferation and marked presence of mononuclear cells in tissue that characterize the improvement of the regeneration of tendineous repair. Statistically, the group treated with the combination of MEC of collagen embedded with mononuclears cells was more significant when compared to other groups. Thus it appears that after examining the results, tendon cell therapy with mononuclear cells improved the organization of proliferative tendon repair between 14th and 30th of days after treatment. / A maioria das lesões tendíneas em pequenos animais está relacionada a traumas, e as lesões mais importantes envolvem secção parcial ou completa, devido à ação de objetos cortantes ou agudos, ou laceração associada com acidente de automóvel levando a uma incapacidade funcional do membro. Os efeitos como a melhora da qualidade ou rapidez da cicatrização não são confirmados por estudos controlados. Recentemente, os avanços médicos têm demonstrado crescente interesse na utilização de terapia celular em tratamentos de doenças degenerativas e também em cicatrização lenta ou ineficaz. Este estudo teve como objetivo avaliar os efeitos do transplante autógeno de células mononucleares (CM) de medula óssea com ou sem MEC na cicatrização tendínea. Foram comparados tendões tratados somente com CM (GI); CM embebidos em MEC de colágeno (GII); somente MEC de colágeno (GIII) com o grupo controle (GIV), além da presença dessas células no tecido neoformado. Foi induzida lesão experimental no tendão calcâneo comum do membro pélvico direito em 36 cães, separados aleatoriamente em quatro grupos experimentais com nove animais cada, seguida por transplante autógeno de no mínimo 0,6 x 108 e máximo 7,4 x 108 CM. kg-1, com viabilidade superior a 90%, marcadas com nanocristal Q-tracker 655. Os animais foram avaliados por parâmetros clínicos e hematológicos e, através de biópsia realizada aos 7º, 14º e 30º dias de evolução por meio de características citológicas, análises fluorescente direta e histopatológicas quanto à qualidade de cicatrização tecidual como: aumento do infiltrado inflamatório, presença de matriz extracelular, aumento da proliferação celular e presença de CM marcadas no tecido que caracterizam uma melhora discreta da regeneração do reparo tendíneo quanto ao quesito inflamação. Estatisticamente, o grupo tratado com a associação de MEC de colágeno embebidos com CM foi mais significativo quando comparados aos outros grupos. Dessa forma conclui-se que após a análise dos resultados, a terapia celular tendínea com CM favoreceu a fase proliferativa do reparo tendíneo entre o 14º e 30º dias após o tratamento e novas pesquisas devem buscar sua utilização como auxílio à regeneração.

Changes to the composition and function of the gut mononuclear phagocyte (MNP) compartment are associated with the development of intestinal inflammation. Much work has focused on the role of MNPs in gut-associated lymphoid tissue in maintaining homeostasis, however little is known regarding the roles of MNPs during colitis. We have investigated MNPs in the large intestinal lamina propria during the steady state and inflammation. One of our primary aims was to determine the contribution of MNP subsets to intestinal pathology. For our studies of inflammation, we focused mainly on the Helicobacter hepaticus infection &plus; anti-IL-10R model, which induces inflammation of the colon and caecum (typhlocolitis). We defined the composition of the MNP compartment alongside intestinal pathology scores throughout Hh &plus; anti-IL-10R typhlocolitis. Peak pathology, 2-3 weeks after induction of colitis, coincided with peak frequencies of CX₃CR1^int Ly6C^&plus; MNPs. Having observed the accumulation of CX₃CR1^int CD64^&plus; monocyte/macrophage MNPs in the inflamed lamina propria, we conducted comparative whole genome microarray analysis of these cells isolated from the large intestine three weeks after Hh &plus; anti-IL-10R treatment. CX₃CR1^int CD64^&plus; MNPs selectively expressed a variety of pro- and anti-inflammatory genes, including a number of genes which individually can both promote and negatively regulate inflammation. IL-23 is essential for Hh &plus; anti-IL-10R-induced intestinal pathology. We investigated the role of MNPs as a source of IL-23 which drives Hh &plus; anti-IL-10R colitis. Unexpectedly, our results indicate that normally hyporesponsive CX₃CR1^hi macrophages may act as the initial source of IL-23, which induces development of colitis. Recruitment of Ly6C^&plus; MHCII^&plus; MNPs to the lamina propria was IL-23-dependent, and these cells also expressed IL-23, which may establish a positive feedback loop of immune cell recruitment, activation and IL-23 production. Finally, we also examined how MNPs might be recruited to the colonic lamina propria during inflammation. Our studies support the conclusion that CCR6 is not required for accumulation of monocyte-derived populations in the inflamed intestine. We cannot rule out a role for CCR2, however preliminary data from the Hh &plus; anti-IL-10R colitis model suggest a potential role for CCR1 or its close relation CCRL2. Such pathways could represent new therapeutic targets in inflammatory bowel disease.