Efeitos da ciclosporina a sobre a função renal de cães da raça Golden Retriever normais ou afetados pela distrofia muscular

The muscular dystrophy of Golden Retriever is a degenerative miopaty caused by the absence of dystrophy and it is genetically homologue of the Duchenne muscular dystrophy in humans, so, these dogs are considerably experimental models for studies on cellular therapy. Their successful depends of the adequate immunosuppression. Cyclosporin A is indicated for that and the monitoring of blood concentration and adverse effects are essential to viabilise the therapy. It was studied GRMD dogs, and normal dogs from the same breed, submitted for therapy with CsA, associated, on GRMD, of cell transplantation. It was evaluated the possible effects of the drug on renal functions. It has been considerate the clinic manifestations, urinalisis, testis of glomerular function and blood concentrations of urea, cretinine, sodium and potassium. In our results we found a discrete increase of blood urea on booth groups; increased levels of urine's cylinders and protein and also increase of urinary density on GRMD group. CsA therapy could make acute lesions on renal tubules, especially on GRMD. These dogs also have different reactions than normal dogs on relation of ions homeostasis and renal function. However, earlier diagnosis and adequate treatment could prevent the development of renal diseases.

Análise das dosagens e concentrações séricas da ciclosporina A em cães da raça Golden Retriever normais ou afetados pela distrofia muscular

The muscular dystrophy of Golden Retriever (GRMD) is a degenerative miopaty caused by the absence of dystrophy and it is genetically homologue of the Duchenne muscular dystrophy in humans, so, these dogs are considerably experimental models for studies on cellular therapy. Their successful depends of the adequate immunosuppression. Cyclosporin A (CsA) is indicated for that and the monitoring of the blood concentration and adverse effects are essential to viabilise the therapy. It was studied GRMD dogs, and normal dogs from the same breed, submitted for therapy with CsA, associated, on GRMD, of cell transplantation. It was evaluated blood concentration of the drug, between two or tree days using the method of FPIA. In our results we found that the CsA blood concentrations oscillated too much on six than eight of our animals. We concluded that the doses varieties individually and the correct dosage as to important as the evaluation of the blood concentration of the drug and became viable for cell therapy.

Platelet-Rich Plasma And Chronic Wounds: Remaining Fibronectin May Influence Matrix Remodeling And Regeneration Success

Background: Platelet-rich plasma has been largely used as a therapeutic option for the treatment of chronic wounds of different etiologies. The enhanced regeneration observed after the use of platelet-rich plasma has been systematically attributed to the growth factors that are present inside platelets' granules. Aim: We hypothesize that the remaining plasma and platelet-bound fibronectin may act as a further bioactive protein in platelet-rich plasma preparations. Methods: Recent reports were analyzed and presented as direct evidences of this hypotheses. Results: Fibronectin may directly influence the extracellular matrix remodeling during wound repair. This effect is probably through matrix metalloproteinase expression, thus exerting an extra effect on chronic wound regeneration. Conclusions: Physicians should be well aware of the possible fibronectin-induced effects in their future endeavors with PRP in chronic wound treatment. © 2013 International Society for Cellular Therapy.

Implante autólogo de células mesenquimais no tratamento de tendinites induzidas em eqüinos: avaliação clínica, ultrasonográfica, histopatológica e imunoistoquímica

Pós-graduação em Medicina Veterinária - FMVZ

Isolamento, caracterização e diferenciação de células tronco embrionárias e mesenquimais de equinos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Proposta de um protocolo de terapia celular para o tratamento da doença pulmonar obstrutiva crônica

The main feature of pulmonary emphysema is airflow obstruction resulting from the destruction of the alveolar walls distal to the terminal bronchioles. Existing clinical approaches have improved and extended the quality of life of emphysema patients. However, no treatment currently exists that can change the disease course and cure the patient. The different therapeutic approaches that are available aim to increase survival and/or enhance the quality of life of emphysema patients. In this context, cell therapy is a promising therapeutic approach with great potential for degenerative pulmonary diseases. In this protocol proposition, all patients will be submitted to laboratory tests, such as evaluation of heart and lung function and routine examinations. Stem cells will be harvested by means of 10 punctures on each anterior iliac crest, collecting a total volume of 200 mL bone marrow. After preparation, separation, counting and labeling (optional) of the mononuclear cells, the patients will receive an intravenous infusion from the pool of Bone Marrow Mononuclear Cells (BMMC). This article proposes a rational and safe clinical cellular therapy protocol which has the potential for developing new projects and can serve as a methodological reference for formulating clinical application protocols related to the use of cellular therapy in COPD. This study protocol was submitted and approved by the Brazilian National Committee of Ethics in Research (CONEP - Brazil) registration number 14764. It is also registered in ClinicalTrials.gov (NCT01110252). (c) 2013 Sociedade Portuguesa de Pneumologia. Published by Elsevier Espana, S.L. All rights reserved.

Effects of lithium on the proliferation and migration of neuroblasts in the rostral migratory stream in adult mice

The discovery of neurogenesis in adult brains opened the possibility of cellular therapy strategies for the treatment of neurodegenerative diseases, such as Alzheimer’s disease. Neurogenesis in the adult brain occurs in two areas: subgranular zone of the hippocampus and subventricular zone (SVZ) of the lateral ventricles. Neurons that originate from the SVZ migrate to the olfactory bulb (OB) through the rostral migratory stream (RMS). In Alzheimer’s disease, there is a progressive neuronal dysfunction and degeneration, resulting in brain atrophy and cognitive impairments including olfactory dysfunction. Several studies have demonstrated that pharmacological treatment with lithium exerts positive effects on adult neurogenesis, and one pathway seems to be the modulation of factors that regulate the migration of neuroblasts. The objective of this study was to investigate whether treatment with lithium promotes the increase of migratory neuroblasts using as parameter the RMS. Adult male C57BL/6 mice were divided into control and lithium-treated groups. The animals were treated for 6 weeks and, at four different time points, i.e., 10 days, 7 days, 3 days and 1 day before the end of treatments, they received an injection of BrdU (cell proliferation marker). The animals were sacrificed by perfusion fixation and the brains were immunohistochemically labeled for BrdU for analysis of migrating neuroblasts in the RMS. The results showed that the number of BrdU+ cells in the RMS was not significantly different between the two groups, suggesting that lithium, alone, is not capable of increasing the number of neuroblasts migrating from the SVZ to the OB

Estudo da capacidade imunomoduladora in vivo de células-tronco mesenquimais obtidas da matriz do cordão umbilical equino

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Isolamento e caracterização de células-tronco mesenquimais de filtros reutilizáveis e descartáveis de medula óssea

OBJETIVO: Comparar as células-tronco mesenquimais humanas obtidas de filtros de coleta reutilizáveis àquelas coletadas em filtros descartáveis e caracterizá-las utilizando os critérios da International Society for Cellular Therapy. MÉTODOS: Foram isoladas células-tronco mesenquimais humanas de kits de coleta de medula óssea reutilizáveis e descartáveis, pela lavagem dos filtros com meio de cultura. As células isoladas foram caracterizadas de acordo com os critérios estabelecidos pela International Society for Cellular Therapy, por meio das técnicas de citometria de fluxo, diferenciação in vitro e citoquímica. RESULTADOS: As amostras foram obtidas de filtro descartável (n=3) e reutilizável (n=3). Todas as amostras obtidas de filtros descartáveis produziram células-tronco mesenquimais, e todas as células-tronco mesenquimais humanas derivadas de medula óssea preencheram os critérios estabelecidos pela International Society for Cellular Therapy. CONCLUSÃO: Este estudo mostrou que as células-tronco mesenquimais também podem ser obtidas de kits de coleta reutilizáveis (que permanecem em uso em vários centros, no mundo inteiro), para serem empregadas em pesquisa como uma fonte alternativa e ética.

In vitro characterisation and expansion of human regulatory T cells for their in vivo application in the induction of tolerance in haematopoietic stem cell and solid organ transplantation

Solid organ transplantation (SOT) is considered the treatment of choice for many end-stage organ diseases. Thus far, short term results are excellent, with patient survival rates greater than 90% one year post-surgery, but there are several problems with the long term acceptance and use of immunosuppressive drugs. Hematopoietic Stem Cells Transplantation (HSCT) concerns the infusion of haematopoietic stem cells to re-establish acquired and congenital disorders of the hematopoietic system. The main side effect is the Graft versus Host Disease (GvHD) where donor T cells can cause pathology involving the damage of host tissues. Patients undergoing acute or chronic GvHD receive immunosuppressive regimen that is responsible for several side effects. The use of immunosuppressive drugs in the setting of SOT and GvHD has markedly reduced the incidence of acute rejection and the tissue damage in GvHD however, the numerous adverse side effects observed boost the development of alternative strategies to improve the long-term outcome. To this effect, the use of CD4+CD25+FOXP3+ regulatory T cells (Treg) as a cellular therapy is an attractive approach for autoimmunity disease, GvHD and limiting immune responses to allograft after transplantation. Treg have a pivotal role in maintaining peripheral immunological tolerance, by preventing autoimmunity and chronic inflammation. Results of my thesis provide the characterization and cell processing of Tregs from healthy controls and patients in waiting list for liver transplantation, followed by the development of an efficient expansion-protocol and the investigation of the impact of the main immunosuppressive drugs on viability, proliferative capacity and function of expanded cells after expansion. The conclusion is that ex vivo expansion is necessary to infuse a high Treg dose and although many other factors in vivo can contribute to the success of Treg therapy, the infusion of Tregs during the administration of the highest dose of immunosuppressants should be carefully considered.

Células madre mesenquimales orales: estado del arte en Odontología

Desde la organogénesis y hasta estadios adultos, las células madre mesenquimales participan activamente dando origen y manteniéndola homeostasis del organismo. En la cavidad oral han sido aisladas desde variadas estructuras del órgano dental tales como el ligamento periodontal, pulpa dental, tejido gingival, folículo dental y papila apical significando una prometedora fuente de células madre mesenquimales las que pueden ser caracterizadas de acuerdo a los criterios mínimos establecidos por "The International Society for Cellular Therapy" que son: a) La adherencia al plástico; b) La expresión de marcadores CD73, CD90, CD105 y la carencia de CD34, CD45, CD14, CD11, CD79, CD19 y HLA-DR (clase II); c) Capacidad multipotencial de diferenciación hacia linaje osteogénico, condrogénico y adipogénico. El objetivo de esta revisión consiste en realizar un levantamiento de la situación actual de este tema efectuando una revisión comprensiva de la literatura en los campos de; identificación a través demarcadores de superficie, aislamiento por medio de mecanismos de digestión enzimática o explante, almacenamiento atendiendo a la necesidad de suprimir el uso de suero fetal bovino como medio de cultivo en un esfuerzo por avanzar hacia aplicaciones terapéuticas, banca o criopreservación destacando nuevas experiencia en este campo como lo es la criopreservación de piezas dentales completas gracias a la tecnología láser Nd:YAG. Y, finalmente, las aplicaciones clínicas que promete este grupo de células a través de la medicina regenerativa y la ingeniería tisular tanto en el campo de la odontología como la medicina general.

Cellular immune responses to HCV core increase and HCV RNA levels decrease during successful antiretroviral therapy.

BACKGROUND: Hepatitis C virus (HCV) infection is a major cause of morbidity in HIV infected individuals. Coinfection with HIV is associated with diminished HCV-specific immune responses and higher HCV RNA levels. AIMS: To investigate whether long-term combination antiretroviral therapy (cART) restores HCV-specific T cell responses and improves the control of HCV replication. METHODS: T cell responses were evaluated longitudinally in 80 HIV/HCV coinfected individuals by ex vivo interferon-gamma-ELISpot responses to HCV core peptides, that predominantly stimulate CD4(+) T cells. HCV RNA levels were assessed by real-time PCR in 114 individuals. RESULTS: The proportion of individuals with detectable T cell responses to HCV core peptides was 19% before starting cART, 24% in the first year on cART and increased significantly to 45% and 49% after 33 and 70 months on cART (p=0.001). HCV-specific immune responses increased in individuals with chronic (+31%) and spontaneously cleared HCV infection (+30%). Median HCV RNA levels before starting cART were 6.5 log(10) IU/ml. During long-term cART, median HCV-RNA levels slightly decreased compared to pre-cART levels (-0.3 log10 IU/ml, p=0.02). CONCLUSIONS: Successful cART is associated with increasing cellular immune responses to HCV core peptides and with a slight long-term decrease in HCV RNA levels. These findings are in line with the favourable clinical effects of cART on the natural history of hepatitis C and with the current recommendation to start cART earlier in HCV/HIV coinfected individuals.

Cell Therapy Assistance in Reconstructive Surgery for Musculoskeletal Tissues Following Burn and Trauma: Swiss Cellular Transplantation Platform

The sterol compositions of three oceanic jellyfish have been determined using gas chromatographic mass spectrometric techniques involving the use of two separate gas chromatographic column systems. The components in overlapping peaks have been identified by comparison of the mass spectra of peaks in the two column systems using subtractive techniques. A mid-water animal, Periphylla periphylla, was found to contain a very complex and unusual sterol profile including rare 5alpha-stanols, whereas two other oceanic jellyfish Pelagia noctiluca and Atolla wyvillei contained similar mixtures of delta5 sterols to those previously isolated from coastal species.

A model of cellular dosimetry for macroscopic tumors in radiopharmaceutical therapy.

PURPOSE: In the radiopharmaceutical therapy approach to the fight against cancer, in particular when it comes to translating laboratory results to the clinical setting, modeling has served as an invaluable tool for guidance and for understanding the processes operating at the cellular level and how these relate to macroscopic observables. Tumor control probability (TCP) is the dosimetric end point quantity of choice which relates to experimental and clinical data: it requires knowledge of individual cellular absorbed doses since it depends on the assessment of the treatment's ability to kill each and every cell. Macroscopic tumors, seen in both clinical and experimental studies, contain too many cells to be modeled individually in Monte Carlo simulation; yet, in particular for low ratios of decays to cells, a cell-based model that does not smooth away statistical considerations associated with low activity is a necessity. The authors present here an adaptation of the simple sphere-based model from which cellular level dosimetry for macroscopic tumors and their end point quantities, such as TCP, may be extrapolated more reliably. METHODS: Ten homogenous spheres representing tumors of different sizes were constructed in GEANT4. The radionuclide 131I was randomly allowed to decay for each model size and for seven different ratios of number of decays to number of cells, N(r): 1000, 500, 200, 100, 50, 20, and 10 decays per cell. The deposited energy was collected in radial bins and divided by the bin mass to obtain the average bin absorbed dose. To simulate a cellular model, the number of cells present in each bin was calculated and an absorbed dose attributed to each cell equal to the bin average absorbed dose with a randomly determined adjustment based on a Gaussian probability distribution with a width equal to the statistical uncertainty consistent with the ratio of decays to cells, i.e., equal to Nr-1/2. From dose volume histograms the surviving fraction of cells, equivalent uniform dose (EUD), and TCP for the different scenarios were calculated. Comparably sized spherical models containing individual spherical cells (15 microm diameter) in hexagonal lattices were constructed, and Monte Carlo simulations were executed for all the same previous scenarios. The dosimetric quantities were calculated and compared to the adjusted simple sphere model results. The model was then applied to the Bortezomib-induced enzyme-targeted radiotherapy (BETR) strategy of targeting Epstein-Barr virus (EBV)-expressing cancers. RESULTS: The TCP values were comparable to within 2% between the adjusted simple sphere and full cellular models. Additionally, models were generated for a nonuniform distribution of activity, and results were compared between the adjusted spherical and cellular models with similar comparability. The TCP values from the experimental macroscopic tumor results were consistent with the experimental observations for BETR-treated 1 g EBV-expressing lymphoma tumors in mice. CONCLUSIONS: The adjusted spherical model presented here provides more accurate TCP values than simple spheres, on par with full cellular Monte Carlo simulations while maintaining the simplicity of the simple sphere model. This model provides a basis for complementing and understanding laboratory and clinical results pertaining to radiopharmaceutical therapy.

A colorectal cancer classification system that associates cellular phenotype and responses to therapy.

Colorectal cancer (CRC) is a major cause of cancer mortality. Whereas some patients respond well to therapy, others do not, and thus more precise, individualized treatment strategies are needed. To that end, we analyzed gene expression profiles from 1,290 CRC tumors using consensus-based unsupervised clustering. The resultant clusters were then associated with therapeutic response data to the epidermal growth factor receptor-targeted drug cetuximab in 80 patients. The results of these studies define six clinically relevant CRC subtypes. Each subtype shares similarities to distinct cell types within the normal colon crypt and shows differing degrees of 'stemness' and Wnt signaling. Subtype-specific gene signatures are proposed to identify these subtypes. Three subtypes have markedly better disease-free survival (DFS) after surgical resection, suggesting these patients might be spared from the adverse effects of chemotherapy when they have localized disease. One of these three subtypes, identified by filamin A expression, does not respond to cetuximab but may respond to cMET receptor tyrosine kinase inhibitors in the metastatic setting. Two other subtypes, with poor and intermediate DFS, associate with improved response to the chemotherapy regimen FOLFIRI in adjuvant or metastatic settings. Development of clinically deployable assays for these subtypes and of subtype-specific therapies may contribute to more effective management of this challenging disease.