Abstract Objective: This study aimed to evaluate neuronal markers in stromal cells from human exfoliated deciduous teeth (SHED) and standardize the isolation and characterization of those cells. Methodology: Healthy primary teeth were collected from children. The cells were isolated by enzymatic digestion with collagenase. By following the International Society for Cell and Gene Therapy (ISCT) guidelines, SHED were characterized by flow cytometry and differentiated into osteogenic, adipogenic, and chondrogenic lineages. Colony-forming unit-fibroblasts (CFU-F) were performed to assess these cells’ potential and efficiency. To clarify the neuronal potential of SHED, the expression of nestin and βIII-tubulin were examined by immunofluorescence and SOX1, SOX2, GFAP, and doublecortin (DCX), nestin, CD56, and CD146 by flow cytometry. Results: SHED showed mesenchymal stromal cells characteristics, such as adhesion to plastic, positive immunophenotypic profile for CD29, CD44, CD73, CD90, CD105, and CD166 markers, reduced expression for CD14, CD19, CD34, CD45, HLA-DR, and differentiation in three lineages confirmed by staining and gene expression for adipogenic differentiation. The average efficiency of colony formation was 16.69%. SHED expressed the neuronal markers nestin and βIII-tubulin; the fluorescent signal intensity was significantly higher in βIII-tubulin (p<0.0001) compared to nestin. Moreover, SHED expressed DCX, GFAP, nestin, SOX1, SOX2, CD56, CD146, and CD271. Therefore, SHED had a potential for neuronal lineage even without induction with culture medium and specific factors. Conclusion: SHEDs may be a new therapeutic strategy for regenerating and repairing neuronal cells and tissues.

Abstract: Asthma is a chronic respiratory disease affecting 300 million people worldwide. It results in several structural changes in the airways, which are minimally accessible in clinical practice. Cell therapy using mesenchymal stromal cells (MSCs) is a promising strategy for treating asthma due to the paracrine activity of MSCs, which influences tissue regeneration and modulates the immune response. Studies using extracellular vesicles (EV) released by MSCs have demonstrated their regenerative properties in animal models. The aim of this study was to evaluate the potential of EVs isolated from human bone marrow MSCs (hBM-MSCs) to control lung tissue remodeling in ovalbumin-induced allergic asthma in Balb/c mice. We isolated hBM-MSCs from a single donor, expanded and characterized them, and then isolated EVs. Asthma was induced in 43 male Balb/c mice, divided into four groups: control, asthmatic (AS), asthmatic plus systemic EVs (EV-S), and asthmatic plus intratracheal EVs (EV-IT). Upon completion of asthma induction, animals were treated with EVs either locally (EV-IT) or intravenously (EV-S). Seven days after, we performed bronchoalveolar lavage (BAL) and the total nuclear cells were counted. The animals were euthanized, and the lungs were collected for histopathological analysis of the airways. The EV-S group showed improvement in only the total BAL cell count compared with the AS group, while the EV-IT group showed significant improvement in almost all evaluated criteria. Therefore, we demonstrate that the local application of EVs derived from hBM-MSCs may be a potential treatment in controlling asthma.

ABSTRACT Objective: Adipose tissue-derived stromal/stem cells (ASCs) and vitamin D have immunomodulatory actions that could be useful for type 1 diabetes (T1D). We aimed in this study to investigate the safety and efficacy of ASCs + daily cholecalciferol (VIT D) for 6 months in patients with recent-onset T1D. Materials and methods: In this prospective, dual-center, open trial, patients with recent onset T1D received one dose of allogenic ASC (1 x 106 cells/kg) and cholecalciferol 2,000 UI/day for 6 months (group 1). They were compared to patients who received chol-ecalciferol (group 2) and standard treatment (group 3). Adverse events were recorded; C-peptide (CP), insulin dose and HbA1c were measured at baseline (T0), after 3 (T3) and 6 months (T6). Results: In group 1 (n = 7), adverse events included transient headache (all), mild local reactions (all), tachycardia (n = 4), abdominal cramps (n = 1), thrombophlebitis (n = 4), scotomas (n = 2), and central retinal vein occlusion at T3 (n = 1, resolution at T6). Group 1 had an increase in basal CP (p = 0.018; mean: 40.41+/-40.79 %), without changes in stimulated CP after mixed meal (p = 0.62), from T0 to T6. Basal CP remained stable in groups 2 and 3 (p = 0.58 and p = 0.116, respectively). Group 1 had small insulin requirements (0.31+/- 0.26 UI/kg) without changes at T6 (p = 0.44) and HbA1c decline (p = 0.01). At T6, all patients (100%; n = 7) in group 1 were in honeymoon vs 75% (n = 3/4) and 50% (n = 3/6) in groups 2 and 3, p = 0.01. Conclusions: Allogenic ASC + VIT D without immunosuppression was safe and might have a role in the preservation of β-cells in patients with recent-onset T1D. ClinicalTrials.gov: NCT03920397.

Abstract Induced pluripotent stem cells (iPSCs) are generated from adult cells that have been reprogrammed to pluripotency. However, in vitro cultivation and genetic reprogramming increase genetic instability, which could result in chromosomal abnormalities. Maintenance of genetic stability after reprogramming is required for possible experimental and clinical applications. The aim of this study was to analyze chromosomal alterations by using the G-banding karyotyping method applied to 97 samples from 38 iPSC cell lines generated from peripheral blood or Wharton’s jelly. Samples from patients with long QT syndrome, Jervell and Lange-Nielsen syndrome and amyotrophic lateral sclerosis and from normal individuals revealed the following chromosomal alterations: acentric fragments, chromosomal fusions, premature centromere divisions, double minutes, radial figures, ring chromosomes, polyploidies, inversions and trisomies. An analysis of two samples generated from Wharton’s jelly before and after reprogramming showed that abnormal clones can emerge or be selected and generate an altered lineage. IPSC lines may show clonal and nonclonal chromosomal aberrations in several passages (from P6 to P34), but these aberrations are more common in later passages. Many important chromosomal aberrations were detected, showing that G-banding is very useful for evaluating genetic instability with important repercussions for the application of iPSC lines.

Abstract Platelet-activating factor (PAF) is a potent proinflammatory mediator that is produced in increased amounts in the lungs of asthmatic humans and horses. The present pilot study, shows that mesenchymal stromal cells can modulate alveolar macrophage function in asthma, interfering in the activity of PAF, being another potential pathway for mesenchymal stromal cells benefits in asthma.

Fundamento:Pacientes portadores de cardiomiopatia dilatada idiopática apresentam alta morbimortalidade, mesmo em tratamento clínico otimizado. A infusão autóloga de células-tronco adultas da medula óssea mostrou resultados clínicos preliminares promissores nesses pacientes.Objetivo:Determinar a eficácia do transplante autólogo de células-tronco adultas da medula óssea sobre as funções sistólica e diastólica, e o grau de insuficiência mitral em pacientes portadores de cardiomiopatia dilatada idiopática em classes funcionais NYHA II e III.Métodos:Infundiram-se 4,54 x 108 ± 0,89 x 108 células-tronco adultas da medula óssea nas artérias coronárias de 24 pacientes com cardiomiopatia dilatada idiopática em classes funcionais NYHA II e III. Após 3 meses, 6 meses e 1 ano, avaliaram-se as mudanças de classe funcional, das funções ventricular esquerda sistólica e diastólica, e do grau da insuficiência mitral.Resultados:No seguimento, seis (25%) pacientes melhoraram sua classe funcional e oito (33,3%) mantiveram sua classe funcional inicial. A fração de ejeção ventricular esquerda aumentou 8,9%, 9,7% e 13,6%, após 3 e 6 meses e 1 ano (p = 0,024; p = 0,017 e p = 0,018), respectivamente. A função diastólica ventricular esquerda e o grau de insuficiência mitral não demonstraram mudanças significativas. Dois pacientes (8,3%) receberam cardioversor e ressincronizador implantável. Ocorreram quatro (16,6%) mortes súbitas e quatro (16,6%) mortes por insuficiência cardíaca terminal. A sobrevida média desses oitos pacientes foi de 2,6 anos.Conclusão:A infusão intracoronariana de células-tronco adultas da medula óssea em pacientes com cardiomiopatia dilatada idiopática promoveu melhora ou estabilização da classe funcional, e melhora da fração de ejeção ventricular esquerda em expressivo número de pacientes, sugerindo eficácia dessa intervenção. Não se observaram mudanças significativas na função diastólica ventricular esquerda e nem no grau de insuficiência mitral.

Background:Morbimortality in patients with dilated idiopathic cardiomyopathy is high, even under optimal medical treatment. Autologous infusion of bone marrow adult stem cells has shown promising preliminary results in these patients.Objective:Determine the effectiveness of autologous transplantation of bone marrow adult stem cells on systolic and diastolic left ventricular function, and on the degree of mitral regurgitation in patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III.Methods:We administered 4,54 x 108 ± 0,89 x 108 bone marrow adult stem cells into the coronary arteries of 24 patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III. Changes in functional class, systolic and diastolic left ventricular function and degree of mitral regurgitation were assessed after 3 months, 6 months and 1 year.Results:During follow-up, six patients (25%) improved functional class and eight (33.3%) kept stable. Left ventricular ejection fraction improved 8.9%, 9.7% e 13.6%, after 3, 6 and 12 months (p = 0.024; 0.017 and 0.018), respectively. There were no significant changes neither in diastolic left ventricular function nor in mitral regurgitation degree. A combined cardiac resynchronization and implantable cardioversion defibrillation was implanted in two patients (8.3%). Four patients (16.6%) had sudden death and four patients died due to terminal cardiac failure. Average survival of these eight patients was 2.6 years.Conclusion:Intracoronary infusion of bone marrow adult stem cells was associated with an improvement or stabilization of functional class and an improvement in left ventricular ejection fraction, suggesting the efficacy of this intervention. There were no significant changes neither in left ventricular diastolic function nor in the degree of mitral regurgitation.

Objetivo: Realizar uma avaliação comparativa de duas técnicas cirúrgicas que são usadas para criar um infarto agudo do miocárdio pela oclusão da artéria coronária descendente anterior esquerda, a fim de gerar ratos com uma fração de ejeção ventricular esquerda inferior a 40%. Métodos: O estudo foi completamente randomizado e composto por 89 ratos anestesiados com halotano, que foram divididos dentro de três grupos. O grupo controle (SHAM) composto por 14 ratos, cuja artéria coronária descendente anterior esquerda não foi ocluída. Grupo 1 (G1): composto por 35 ratos intubados endotraquealmente e ventilados mecanicamente, cuja artéria coronária descendente anterior esquerda foi ocluída. Grupo 2 (G2): constituído por 40 ratos sendo ventilados manualmente utilizando um respirador nasal, cuja artéria coronária descendente anterior esquerda foi ocluída. Outras diferenças entre as duas técnicas incluem o método de realizar a toracotomia e remover o pericárdio, a fim de expor o coração, e o uso de diferentes métodos e tipos de sutura para fechar o tórax. Sete dias após a cirurgia, a função cardíaca de todos os ratos sobreviventes foi determinada por ecocardiografia. Resultados: Nenhum rato do grupo SHAM foi a óbito ou teve fração de ejeção ventricular esquerda menor que 40%. Nove dos 16 ratos sobreviventes do G1 (56,3%) e seis dos 20 ratos sobreviventes do G2 (30%) tiveram uma fração de ejeção ventricular esquerda inferior a 40%. Conclusão: Os resultados indicam uma tendência da técnica utilizada no G1 ser melhor do que a do G2. Esta melhora deve-se provavelmente à maior duração do tórax aberto, o que reduz a pressão de tempo sobre o cirurgião, permitindo uma oclusão da artéria coronária descendente anterior esquerda com maior acurácia.

Objective: To perform a comparative assessment of two surgical techniques that are used creating an acute myocardial infarc by occluding the left anterior descending coronary artery in order to generate rats with a left ventricular ejection fraction of less than 40%. Methods: The study was completely randomized and comprised 89 halothane-anaesthetised rats, which were divided into three groups. The control group (SHAM) comprised fourteen rats, whose left anterior descending coronary artery was not occluded. Group 1 (G1): comprised by 35 endotracheally intubated and mechanically ventilated rats, whose left anterior descending coronary artery was occluded. Group 2 (G2): comprised 40 rats being manually ventilated using a nasal respirator whose left anterior descending coronary artery was occluded. Other differences between the two techniques include the method of performing the thoracotomy and removing the pericardium in order to expose the heart, and the use of different methods and suture types for closing the thorax. Seven days after surgery, the cardiac function of all surviving rats was determined by echocardiography. Results: No rats SHAM group had progressed to death or had left ventricular ejection fraction less than 40%. Nine of the 16 surviving G1 rats (56.3%) and six of the 20 surviving G2 rats (30%) had a left ventricular ejection fraction of less than 40%. Conclusion: The results indicate a tendency of the technique used in G1 to be better than in G2. This improvement is probably due to the greater duration of the open thorax, which reduces the pressure over time from the surgeon, allowing occlusion of left anterior descending coronary artery with higher accuracy.

Stem cell-based regenerative medicine is one of the most intensively researched medical issues. Pre-clinical studies in a large-animal model, especially in swine or miniature pigs, are highly relevant to human applications. Mesenchymal stem cells (MSCs) have been isolated and expanded from different sources. Objective: This study aimed at isolating and characterizing, for the first time, bone marrow-derived MSCs (BM-MSCs) from a Brazilian minipig (BR1). Also, this aimed to validate a new large-animal model for stem cell-based tissue engineering. Material and Methods: Bone marrow (BM) was aspirated from the posterior iliac crest of twelve adult male BR1 under general anesthesia. MSCs were selected by plastic-adherence as originally described by Friedenstein. Cell morphology, surface marker expression, and cellular differentiation were examined. The immunophenotypic profile was determined by flow cytometry. The differentiation potential was assessed by cytological staining and by RT-PCR. Results: MSCs were present in all minipig BM samples. These cells showed fibroblastic morphology and were positive for the surface markers CD90 (88.6%), CD29 (89.8%), CD44 (86.9%) and negative for CD34 (1.61%), CD45 (1.83%), CD14 (1.77%) and MHC-II (2.69%). MSCs were differentiated into adipocytes, osteoblasts, and chondroblasts as demonstrated by the presence of lipidic-rich vacuoles, the mineralized extracellular matrix, and the great presence of glycosaminoglycans, respectively. The higher gene expression of adipocyte fatty-acid binding protein (AP2), alkaline phosphatase (ALP) and collagen type 2 (COLII) also confirmed the trilineage differentiation (p<0.001, p<0.001, p=0.031; respectively). Conclusions: The isolation, cultivation, and differentiation of BM-MSCs from BR1 makes this animal eligible as a useful large-animal model for stem cell-based studies in Brazil.

OBJECTIVE: To present the initial results of first three years of implementation of a genetic evaluation test for bone marrow-derived mesenchymal stem cells in a Cell Technology Center.METHODS: A retrospective study was carried out of 21 candidates for cell therapy. After the isolation of bone marrow mononuclear cells by density gradient, mesenchymal stem cells were cultivated and expanded at least until the second passage. Cytogenetic analyses were performed before and after cell expansion (62 samples) using G-banded karyotyping.RESULTS: All the samples analyzed, before and after cell expansion, had normal karyotypes, showing no clonal chromosomal changes. Signs of chromosomal instability were observed in 11 out of 21 patients (52%). From a total of 910 analyzed metaphases, five chromatid gaps, six chromatid breaks and 14 tetraploid cells were detected giving as total of 25 metaphases with chromosome damage (2.75%).CONCLUSION: The absence of clonal chromosomal aberrations in our results for G-banded karyotyping shows the maintenance of chromosomal stability of bone marrow-derived mesenchymal stem cells until the second passage; however, signs of chromosomal instability such as chromatid gaps, chromosome breaks and tetraploidy indicate that the long-term cultivation of these cells can provide an intermediate step for tumorigenesis.

Células-tronco/progenitoras frequentemente não estão disponíveis em quantidade suficiente para restauração de órgãos e tecidos danificados, sendo necessária sua expansão in vitro. Instalações físicas adequadas, pessoal técnico qualificado, reagentes de grau clínico e protocolos bem definidos de acordo com as condições de boas práticas de fabricação são imprescindíveis para assegurar a qualidade e segurança das células infundidas no paciente. A medula óssea e o sangue de cordão umbilical ainda são as fontes de células mais utilizadas em terapias. Protocolos bem sucedidos de expansão utilizando células-tronco hematopoéticas, células-tronco mesenquimais e células progenitoras endoteliais já têm sido empregados em estudos pré-clínicos e clínicos. A escolha do tipo celular adequado deve ser direcionada pelo tamanho da lesão ou natureza do tecido tratado e pelo efeito terapêutico desejado. Estudos recentes têm demonstrado que propriedades de diferentes células expandidas in vitro podem ser combinadas para obtenção de um resultado melhor no tratamento de algumas doenças. Células em culturas de longo termo precisam ser acompanhadas por meio de diversas técnicas de citogenética clássica e molecular para demonstrar que não há evidências de transformação espontânea ou sinais de imortalização. Ensaios utilizando a infusão de células expandidas através da barreira alogeneica e xenogeneica, apresentaram melhora funcional e foram alcançados sem imunossupressão e sem evidências de infiltrados celulares que indicariam resposta imune. Porém, mais estudos precisam ser realizados para avaliar a imunogenicidade destas células e garantir a segurança da terapia celular alogênica permitindo sua consolidação no uso clínico. Aqui apresentamos uma atualização sobre expansão celular associada com seu uso clínico.

Stem/progenitor cells are not frequently available in large enough amounts to repair damaged tissues and organs and so in vitro expansion is necessary. Appropriate facilities, qualified technicians, clinical-grade reagents and well defined protocols relating to good manufacturing products are essential to assure the quality and security of the cells injected in the patient. Bone marrow and human umbilical cord blood are still the best sources of cells for therapies. Successful expansion protocols using hematopoietic stem cells, mesenchymal stem cells and endothelial progenitor cells have already been used in clinical and pre-clinical trials. Adequate cell choice should consider the extent of injury or nature of the damaged tissue and the desired therapeutic effect. Recent studies have demonstrated that properties of different in vitro expanded cells can be combined aiming to improve the outcome of the treatment of some diseases. Long-term cell cultures need to be followed up by classical and molecular cytogenetic techniques to demonstrate that there is no evidence of spontaneous transformation or signs of immortalization. Assays using expanded cell infusions across both xenogeneic and allogeneic transplant barriers showed functional improvement and were achieved without immunosuppression and without evidence of a cellular infiltrate that would indicate an immune response. However, more research needs to be performed to evaluate the immunogenicity of these cells and to guarantee the safety of allogeneic cell therapy, allowing consolidation of their clinical use. Here, we present an update regarding cellular expansion associated with their clinical use.

OBJETIVO: As células progenitoras endoteliais (CPE), caracterizadas pelo marcador CD133+, contribuem para a neovascularização, e o aumento no número dessas células pode ser uma ferramenta terapêutica promissora. O sangue de cordão umbilical humano contém um número significante de CPE, sugerindo a possibilidade do uso destas células para a revascularização de tecidos isquêmicos. O objetivo desse trabalho foi analisar a funcionalidade das células CD133+ diferenciadas in vitro. MÉTODOS: As células diferenciadas foram caracterizadas por citometria de fluxo; a expressão do mRNA de VEGF foi avaliada por RT-PCR e a funcionalidade, por meio de ensaios de formação de túbulos capilares. RESULTADOS: As células diferenciadas perderam os marcadores de CPE, mantiveram em níveis baixos os marcadores das linhagens hematopoética e monocíticas e aumentaram a expressão dos marcadores de células endoteliais adultas. As células diferenciadas apresentaram transcritos no mRNA de VEGF e mostraram-se capazes de formar túbulos capilares in vitro. CONCLUSÃO: As células CD133+ diferenciadas in vitro em células endoteliais demonstraram serem funcionalmente ativas, abrindo perspectiva para seu uso futuro em aplicações terapêuticas.

OBJECTIVE: Endothelial progenitor cells (EPC) caracterized by the CD133+ marker, contribute to the neovascularization. Increasing EPC number in vitro could be a promising therapeutic tool. Human umbilical cord blood maintains a significant number of EPC, suggesting the possibility to use these cells to induce the revascularization of ischemic tissues. The aim of this study was to analize the in vitro function of differentiated CD133+ cells. METHODS: Cells were characterized by flow cytometry, VEGF mRNA expression was evaluated by the RT-PCR analysis and the functionally by essays of capillary tubes formation. RESULTS: Differentiated cells lost EPC markers, maintained low levels of markers for hematopoietic and monocytic cell lines and increased the expression of adult endothelial cell markers. Differentiated cells expressed VEGF mRNA and were capable to induce in vitro capillary tubules formation. CONCLUSION: CD133+ cells differentiated into endothelial cells in vitro are functionally active initiating the possibility of their use in future therapeutic applications.

INTRODUÇÃO: O transplante de células no miocárdio tem se mostrado tecnicamente reprodutível, entretanto, existem dúvidas em relação a melhor fração das células da medula óssea a ser utilizada. Desta forma, o objetivo deste estudo é analisar o resultado do transplante de células tronco mononucleares (MO) e mesenquimais (ME) no infarto do miocárdio. MÉTODO: Quarenta e dois ratos Wistar foram induzidos ao infarto do miocárdio. Após uma semana, os animais foram submetidos à ecocardiografia para avaliação da fração de ejeção (FE) e dos volumes diastólico (VDFVE) e sistólico (VSFVE) finais do ventrículo esquerdo. Após dois dias, os animais foram reoperados e divididos em grupos: 1) controle (n=21), que recebeu 0,15 ml de meio de cultura, 2) MO (n=8) e 3) ME (n=13), que receberam 3x10(6) células mononucleares e mesenquimais, respectivamente, no infarto. As MO foram obtidas a partir de uma punção da medula e isoladas pelo método Ficoll-Hypaque, as ME, após o mesmo processo, foram cultivadas por 14 dias. Os animais foram submetidos à ecocardiografia após um mês. A histologia foi realizada pelo método Tricrômio de Gomery. RESULTADOS: Não houve diferença entre os grupos na ecocardiografia de base. Entretanto, um mês após o transplante, observou-se uma diminuição da FE no grupo controle e uma estabilização nos demais grupos. Os três grupos apresentaram dilatação ventricular. A análise histológica do infarto identificou, no grupo ME, células endoteliais e musculares lisas, no grupo MO, apenas células endoteliais. CONCLUSÕES: Tanto o grupo MO, como o ME, apresentaram uma estabilização da FE após o infarto do miocárdio, uma regeneração vascular, entretanto, com remodelamento ventricular.

BACKGROUND: Bone marrow stem cell (SC) transplantation into failing myocardium has emerged as a novel therapeutic option for the treatment of ventricular dysfunction. Both mononuclear (MoSC) and mesenchymal (MeSC) stem cells have been proposed as ideal cell types to this goal. The objective of this study is to compare the efficacy of these cells in improving ventricular function in a rat model of post-infarct ventricular dysfunction. METHOD: Myocardial infarction was induced in Wistar rats by left coronary occlusion. After 1 week, 42 animals with resulting ejection fractions (EF) lower than 30% were included in the study. MoSC and MeSC were obtained from bone marrow aspirates and separated by the Ficoll-Hypaque method. MeSC were cultured for 14 days before injection. Nine days after infarction, rats received intramyocardial injections of MoSC (n=8), MeSC (n=13) or culture medium as a control (n=21). Echocardiographic evaluation was performed at baseline and after one month. RESULTS: There were no significant differences in the baseline ejection fractions or the left ventricular end diastolic volumes (LVEDV) between all groups. After 1 month, ejection fraction decreased in the Control Group and remained unchanged in MoSC and MeSC Groups. In all three groups ventricular dilation was observed. Histopathology of the infarcted area where injections were performed identified new smooth muscle cells and endothelial cells in the MeSC Group and only new endothelial cells in MoSC Group CONCLUSIONS: Both MoSC and MeSC provided stabilization in the ejection fraction in this post-infarction ventricular dysfunction model however, no therapy prevented ventricular dilation.

OBJETIVO: Apresentar os resultados funcionais, imunocitoquímicos e histopatológicos, in vitro ou em espécimes cardíacas após isolamento, cultura e co-cultura de células tronco mesenquimais, células mioblásticas esqueléticas e transplantadas e co-transplantadas em animais de laboratório com miocardiopatia isquêmica e fração de ejeção do ventrículo esquerdo menor de 40%. MÉTODO: Foram empregados 72 ratos Wistar, divididos em quatro grupos de acordo com o meio de cultura ou das células injetáveis: Grupo controle em que foi injetado apenas o meio de cultura (22 ratos); Grupo de células tronco mesenquimais (17 ratos); Grupo de células mioblásticas esqueléticas (16 ratos) e grupo co-cultura (17 ratos). Nos estudos imunocitoquímicos, as células foram marcadas com anti-vimentina, anti-desmina e anti-miosina. Nos estudos histopatológicos, as lâminas foram coradas com Tricômio de Gomori e identificados neovasos e tecido muscular. Na análise funcional, foi medida a fração de ejeção do ventrículo esquerdo em dois momentos do seguimento, uma semana após o infarto do miocárdio e um mês após a injeção. RESULTADOS: A fração de ejeção do ventrículo esquerdo entre os quatro grupos não apresentou diferença estatística significante (P=0,276), o ecocardiograma de seguimento demonstrou diferença estatística significante (P=0,001). Essa diferença ocorreu entre o grupo controle e o grupo de células mioblásticas esqueléticas (P=0,037), entre o grupo controle e o grupo co-cultura (P<0,001) e o grupo de células tronco mesenquimais e o grupo co-cultura (P=0,025). Quando se compararam as medidas obtidas dos dois ecocardiogramas em cada grupo, encontrou-se diferença no grupo controle (P=0,005) para pior e no grupo co-cultura (P=0,006) para melhor. No estudo imunocitoquímico in vitro, identificou-se células tronco mesenquimais quando marcou-se com anti-vimentina e células musculares, com anti-desmina. Nas espécimes cardíacas, identificou-se tecido muscular marcada com anti-desmina e células mioblásticas esqueléticas marcadas com anti-miosina rápida. No estudo histopatológico, observaram-se novos vasos no grupo de células tronco mesenquimais, no grupo de células mioblásticas esqueléticas, tecido muscular e angiogênese e miogênese no grupo co-cultura. CONCLUSÃO: A fração de ejeção do ventrículo esquerdo melhorou no grupo em que foram injetadas células musculares, mais acentuadamente no grupo co-cultura. Nos achados imunocitoquímicos, na cultura e no co-cultivo encontraram-se as células correspondentes. Nas espécimes cardíacas, foram encontradas células musculares e mioblásticas esqueléticas. Na histopatologia, encontraram-se novos vasos, tecido muscular e ambos quando se injetou células tronco mesenquimais, células mioblásticas esqueléticas e a co-cultura das duas células, respectivamente.

OBJECTIVE: To present the functional, immunocytochemical and histopathologic results (in vitro or in heart specimens) after isolation, culture and co-culture of mesenchymal stem cells and skeletal myoblast cells transplanted and co-transplanted in experimental animals with ischemic heart disease and left ventricular ejection fractions lower than 40%. METHOD: We utilized 72 Wistar rats, divided into four groups according to the culture media or injected cells: control group into which only culture media was injected (22 rats); mesenchymal stem cell group (17 rats); myoblast skeletal cell group (16 rats) and co-culture group (17 rats). In the immunohistochemical studies, the cells were stained with anti-vimentin, anti-desmin and anti-myosin. In the histopathologic analysis, slides were stained with Gomori Trichrome, and neo-vessels and muscle tissues were identified. In the functional analysis the left ventricle ejection fraction was analyzed one week after myocardial infarction and one month after the injection. RESULTS: The initial left ventricle ejection fraction (control echo) was not statistically significant between the four groups (P=0.276), but was significantly different in the follow-up examination (P=0.001). This difference was seen between the control and the myoblast skeletal cells groups (P=0.037), between the control and the co-culture groups (P<0.001), and between the mesenchymal stem cell and co-culture groups (P=0.025). When the initial and final echocardiograms in each group were compared, the control group deteriorated (P=0.005) and the co-culture group improved (P=0.006). With the immunocytochemical in vitro analysis, mesenchymal stem cells were identified when stained with anti-vimentin and muscle cells when stained with anti-desmin. In the heart specimens, muscle tissue, stained with anti-desmin and skeletal myoblasts cells, stained with fast anti-miosin were identified. In the histopathologic analysis, new vessels were observed in the mesenchymal stem cell and skeletal myoblast groups, and muscular tissue, angiogenesis and myogenesis in the co-culture group. CONCLUSION: The left ventricle ejection fraction improved in the group in which muscle cells were injected and more strikingly in the co-culture group. The immunohistochemical findings in the culture and co-culture groups evidenced the corresponding cells. In the heart specimens, muscle and skeletal myoblast cells were found. In the histopathologic examination, new vessels and muscle tissue were found in the mesenchymal stem cell, skeletal myoblast cell and co-culture groups.