Mesenchymal stromal cells as a choice for spinal cord injury treatment

Spinal cord injury (SCI) is a serious clinical problem that affects approximately 17,500 new patients per year in the United States. The main causes of SCI are vehicle collisions, falls, violence (mainly gunshot wounds), and sports/recreational activities. The final severity of the damage results from primary and secondary mechanisms that begin at the time of injury and last for months after trauma. To reduce the extent of damage, several treatments have been proposed. This review summarizes results from several studies that have pointed to cell therapy as the main form of neuroregenerative treatment. Mesenchymal stromal cells (MSCs) are important candidates for tissue regeneration due to the release of bioactive factors, as well as antiapoptotic effects, scar inhibitors, and angiogenic effects. Studies have shown that MSCs act in various ways on injured tissue, such as immunomodulation of the inflamed environment, release of bioactive factors, restoration of axon myelin, prevention of neuronal apoptosis, and neuroregeneration. Current research using MSCs aims to prevent secondary injury, promote regeneration, and replace destroyed spinal cord tissue. This review presents information about the damage from primary and secondary events after SCI, treatments usually used, and pre-clinical and clinical results aiming at the cell therapy using MSCs as a tissue regeneration strategy

Dental pulp-derived Stem Cells: A promising source for regenerative medicine

Submitted by Manoel Barata ([email protected]) on 2019-09-19T17:55:14Z No. of bitstreams: 1 BJSTR.MS.ID.00264.pdf: 417355 bytes, checksum: 0b8f6bf21243e017ad013b397cc0b009 (MD5)Approved for entry into archive by Manoel Barata ([email protected]) on 2019-10-11T19:55:51Z (GMT) No. of bitstreams: 1 BJSTR.MS.ID.00264.pdf: 417355 bytes, checksum: 0b8f6bf21243e017ad013b397cc0b009 (MD5)Made available in DSpace on 2019-10-11T19:55:51Z (GMT). No. of bitstreams: 1 BJSTR.MS.ID.00264.pdf: 417355 bytes, checksum: 0b8f6bf21243e017ad013b397cc0b009 (MD5) Previous issue date: 2019Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Carlos Chagas. Laboratório de Biologia Básica de Células Tronco. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Regenerative medicine has the potential to replace tissues and organs damaged using stem cells (SCs) and their derivatives. Mesenchymal stromal cells (MSCs) are adult cells, multipotent and functionally defined as having self-renewal capacity, differentiation capacity in several cell lines and extensive paracrine and immunomodulatory activity. It has been described the presence of different types of MSCs populations in teeth. Dental pulp-derived SCs (DPSCs) are an important source of SC for cell therapy, can easily be collected and presents a low risk for the patient besides the great availability of material (e.g.: third molar). DPSCs have differentiation potential in multiple lineages like MSCs obtained from other tissues. The use of MSCs on a large scale in the research depends on the quality of the cells that will promote the regeneration of an injured tissue. An important aspect of standardization is the isolation method. The isolated and cultivated DPSCs present fibroblast-like morphology, plastic adherence and demonstrate the immunophenotypic profile suggested by International Society for Cellular Therapy. DPSCs derive from neural crest and have the potential to differentiate into neurons and promote the survival of neuronal cells from the injured site. The characteristics (differentiation and surface markers) presented by DPSCs are very similar with stem cells of other tissues. However, for DPSC to be used on a large scale in regenerative medicine it is necessary to standardize the isolation methodology

Proliferation and differentiation of stem cells in contact with eluate from fibrin-rich plasma membrane

Submitted by Manoel Barata ([email protected]) on 2018-02-09T12:37:17Z No. of bitstreams: 1 AguiarProliferat.pdf: 2378251 bytes, checksum: a4efca8744d7c251c7205603761e7fa0 (MD5)Approved for entry into archive by Manoel Barata ([email protected]) on 2018-02-28T12:24:31Z (GMT) No. of bitstreams: 1 AguiarProliferat.pdf: 2378251 bytes, checksum: a4efca8744d7c251c7205603761e7fa0 (MD5)Made available in DSpace on 2018-02-28T12:24:31Z (GMT). No. of bitstreams: 1 AguiarProliferat.pdf: 2378251 bytes, checksum: a4efca8744d7c251c7205603761e7fa0 (MD5) Previous issue date: 2018Pontifícia Universidade Católica do Paraná. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Curitiba, PR, Brasil.To evaluate the ability of the eluate from fibrin-rich plasma (FRP) membrane to induce proliferation and differentiation of isolated human adipose-derived stem cells (ASCs) into chondrocytes. How method: FRP membranes were obtained by centrifugation of peripheral blood from two healthy donors, cut, and maintained in culture plate wells for 48h to prepare the fibrin eluate. The SCATh were isolated from adipose tissue by collagenase digestion solution, and expanded in vitro. Cells were expanded and treated with DMEM-F12 culture, a commercial media for chondrogenic differentiation, and eluate from FRP membrane for three days, and labeled with BrdU for quantitative assessment of cell proliferation using the HighContent Operetta® imaging system. For the chondrogenic differentiation assay, the SCATh were grown in micromass for 21 days and stained with toluidine blue and aggrecan for qualitative evaluation by light microscopy. The statistical analysis was performed using ANOVA and Tukey’s test. The results were that there was a greater proliferation of cells treated with the eluate from FRP membrane compared to the other two treatments, where the ANOVA test showed significance (p < 0.001). The differentiation into chondrocytes was visualized by the presence of mucopolysaccharide in the matrix of the cells marked in blue toluidine and aggrecan. This way concluded in by treatment with eluate from FRP membrane stimulated cell proliferation and induced differentiation of the stem cells into chondrocytes, suggesting a potential application of FRP membranes in hyaline cartilage regeneration therapies

Mesenchymal stromal cells derived from exfoliated deciduous teeth express neuronal markers before differentiation induction

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&lt;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

Proliferation and differentiation of stem cells in contact with eluate from fibrin-rich plasma membrane

ABSTRACT Objective: To evaluate the ability of the eluate from fibrin-rich plasma (FRP) membrane to induce proliferation and differentiation of isolated human adipose-derived stem cells (ASCs) into chondrocytes. Method: FRP membranes were obtained by centrifugation of peripheral blood from two healthy donors, cut, and maintained in culture plate wells for 48 h to prepare the fibrin eluate. The SCATh were isolated from adipose tissue by collagenase digestion solution, and expanded in vitro. Cells were expanded and treated with DMEM-F12 culture, a commercial media for chondrogenic differentiation, and eluate from FRP membrane for three days, and labeled with BrdU for quantitative assessment of cell proliferation using the High-Content Operetta® imaging system. For the chondrogenic differentiation assay, the SCATh were grown in micromass for 21 days and stained with toluidine blue and aggrecan for qualitative evaluation by light microscopy. The statistical analysis was performed using ANOVA and Tukey's test. Results: There was a greater proliferation of cells treated with the eluate from FRP membrane compared to the other two treatments, where the ANOVA test showed significance (p < 0.001). The differentiation into chondrocytes was visualized by the presence of mucopolysaccharide in the matrix of the cells marked in blue toluidine and aggrecan. Conclusions: Treatment with eluate from FRP membrane stimulated cell proliferation and induced differentiation of the stem cells into chondrocytes, suggesting a potential application of FRP membranes in hyaline cartilage regeneration therapies

HLA-G and CD152 Expression Levels Encourage the Use of Umbilical Cord Tissue-Derived Mesenchymal Stromal Cells as an Alternative for Immunosuppressive Therapy

Mesenchymal stromal cells (MSCs) have been used in immunosuppressive therapy due to their therapeutic effects, with the HLA-G molecule seeming to play a fundamental role. This work evaluated alternative MSC sources to bone marrow (BM), namely, umbilical cord tissue (UC), adipose tissue (AD) and dental pulp tissue (DP), and the influence of interferon-γ (IFN-γ) and hypoxia on the cultivation of these cells for use in immunosuppression therapies. Expression of costimulatory markers CD40, CD80 and CD86 and immunosuppressive molecules CD152 and HLA-G was analyzed. Lymphocyte inhibition assays were also performed. Sequencing of the HLA-G gene from exons 1 to 5 was performed using next-generation sequencing to determine the presence of alleles. UC-derived MSCs (UCMSCs) expressed higher CD152 and HLA-G1 under standard cultivation. UCMSCs and DP-derived MSCs (DPSCs) secreted similar levels of HLA-G5. All MSC sources inhibited the proliferation of peripheral blood mononuclear cells (PBMCs); growth under regular versus hypoxic conditions resulted in similar levels of inhibition. When IFN-γ was added, PBMC growth was inhibited to a lesser extent by UCMSCs. The HLA-G*01:04:01:01 allele appears to generate a more efficient MSC response in inhibiting lymphocyte proliferation. However, the strength of this conclusion was limited by the small sample size. UCMSCs are an excellent alternative to BM in immunosuppressive therapy: they express high concentrations of inhibitory molecules and can be cultivated without stimuli, which minimizes cost

Avaliação sensitiva, motora e cistométrica de cães com lesão medular crônica, submetidos ao transplante de células-tronco mesenquimais derivadas de tecido adiposo

RESUMO: A utilização de células-tronco na reparação de lesões tem sido extensivamente investigada. Neste estudo, examinamos os efeitos terapêuticos de dois transplantes (12x106 céls/transplante) de células-tronco mesenquimais alogênicas derivadas do tecido adiposo (CTDAs) em 11 cães com lesões crônicas traumáticas toracolombares da medula espinhal. As CTDAs foram foram cultivadas in vitro, a proliferação e a viabilidade foram avaliadas. As suspensões foram expandidas e administradas no espaço intradural com intervalo de uma semana entre transplantes. Os cães foram submetidos à avaliações clínicas, laboratoriais, radiográficas, tomográficas, sensitivas, motoras e cistométricas. A maioria dos animais não tinha raça definida (63,63%), mesma proporção para o acometimento de fêmeas e foi observada predominância de fratura com subluxação vertebral (81,81%). Na comparação dos cães pré e pós-transplante não foram observadas alterações hematológicas e três animais (27,27%) apresentaram cistite bacteriana. Em relação a sensibilidade, motricidade e cistometria, também não houve alterações significativas dos índices antes e pós transplantes, sendo observado a ausência nociceptiva na maioria dos animais (72,73%), paraplegia e incontinência urinária na mesma proporção. Neste estudo concluiu-se que o protocolo utilizado de transplante de CTDAs, demonstrou ser um tratamento seguro para cães com lesão medular crônica, com melhora discreta da funcionalidade vesical, porém sem melhora clínica significativa

Padronização e avaliação histológica de um modelo experimental de lesão medular

A lesão medular é incapacitante, irreversível e de custo econômico e social elevado. Neste estudo, objetivou-se padronizar um modelo de lesão medular, que produza paraplegia, com o uso de cateter e avaliar histologicamente a efetividade da lesão para estudos com terapia celular. Foram realizadas as lesões medulares em ratos Wistar, utilizando-se o cateter Fogarty n.3 e compressão na região toracolombar (T8-T9) durante 5 minutos. Foram estudados três grupos: grupo A, animais controle sem lesão medular; grupo B, animais submetidos à lesão, utilizando-se 50µL de compressão; grupo C, animais submetidos à lesão, utilizando-se 80µL de compressão. Foi realizada avaliação motora pela aplicação da escala BBB, antes da compressão, após recuperação anestésica, 24 e 72 horas depois da compressão e sete dias após a compressão. Após o sétimo dia da lesão, os animais foram submetidos à eutanásia, foi feita a retirada da medula espinhal, fígado e rins e realizada a análise histológica com a coloração hematoxilina-eosina. A mortalidade variou entre os grupos, com 0% no grupo A, 38,5% no B e 48% no C. Nesses dois últimos grupos, a causa da morte foi edema pulmonar neurogênico, confirmado clínica e histologicamente. As medulas espinhais histologicamente apresentaram diferentes graus de edema, congestão vascular e hemorragia, enquanto que os fígados e os rins apresentaram diferentes graus de congestão vascular e necrose. Em relação à recuperação dos movimentos, no grupo A, verificou-se 100% de escore 21; no B, 25% de escore 21; 37,5% de escore 11; e 37,5% de escore 0; enquanto, no grupo C, verificou-se 100% de escore 0. Conclui-se que o procedimento realizado utilizando-se 80µL de solução salina para preencher o balão do cateter foi mais eficiente, apesar de maior mortalidade, pois apresentou maior porcentagem de animais com lesão completa (paraplegia)

Direct intracardiac injection of umbilical cord-derived stromal cells and umbilical cord blood-derived endothelial cells for the treatment of ischemic cardiomyopathy

Submitted by Luciane Willcox ([email protected]) on 2016-09-05T18:06:03Z No. of bitstreams: 1 Suss et al Direct intracardiac injection.pdf: 1458058 bytes, checksum: f352b414b697cd29ce2061be68ee252b (MD5)Approved for entry into archive by Luciane Willcox ([email protected]) on 2016-09-05T18:16:00Z (GMT) No. of bitstreams: 1 Suss et al Direct intracardiac injection.pdf: 1458058 bytes, checksum: f352b414b697cd29ce2061be68ee252b (MD5)Made available in DSpace on 2016-09-05T18:16:00Z (GMT). No. of bitstreams: 1 Suss et al Direct intracardiac injection.pdf: 1458058 bytes, checksum: f352b414b697cd29ce2061be68ee252b (MD5) Previous issue date: 2015-01-08This work was supported by grants from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) (CT-Saúde/MS/SCTIE/DECIT/MCT/CNPq N 17/2008) and Fundação Araucária (Convênio no. 416/2009).Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Pontifícia Universidade Católica do Paraná. Núcleo de Tecnologia Celular. Curitiba, PR, Brasil.The development of new therapeutic strategies is necessary to reduce the worldwide social and economic impact of cardiovascular disease, which produces high rates of morbidity and mortality. A therapeutic option that has emerged in the last decade is cell therapy. The aim of this study was to compare the effect of transplanting human umbilical cord-derived stromal cells (UCSCs), human umbilical cord blood-derived endothelial cells (UCBECs) or a combination of these two cell types for the treatment of ischemic cardiomyopathy (IC) in a Wistar rat model. IC was induced by left coronary artery ligation, and baseline echocardiography was performed seven days later. Animals with a left ventricular ejection fraction (LVEF) of 40% were selected for the study. On the ninth day after IC was induced, the animals were randomized into the following experimental groups: UCSCs, UCBECs, UCSCs plus UCBECs, or vehicle (control). Thirty days after treatment, an echocardiographic analysis was performed, followed by euthanasia. The animals in all of the cell therapy groups, regardless of the cell type transplanted, had less collagen deposition in their heart tissue and demonstrated a significant improvement in myocardial function after IC. Furthermore, there was a trend of increasing numbers of blood vessels in the infarcted area. The median value of LVEF increased by 7.19% to 11.77%, whereas the control group decreased by 0.24%. These results suggest that UCSCs and UCBECs are promising cells for cellular cardiomyoplasty and can be an effective therapy for improving cardiac function following IC

Treatment of Chronic Kidney Disease with Extracellular Vesicles from Mesenchymal Stem Cells and CD133+ Expanded Cells: A Comparative Preclinical Analysis

Chronic kidney disease (CKD) is characterized by structural abnormalities and the progressive loss of kidney function. Extracellular vesicles (EVs) from human umbilical cord tissue (hUCT)-derived mesenchymal stem cells (MSCs) and expanded human umbilical cord blood (hUCB)-derived CD133+ cells (eCD133+) maintain the characteristics of the parent cells, providing a new form of cell-free treatment. We evaluated the effects of EVs from hUCT-derived MSCs and hUCB-derived CD133+ cells on rats with CDK induced by an adenine-enriched diet. EVs were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis (NTA) and electron microscopy. The animals were randomized and divided into the MSC-EV group, eEPC-EV group and control group. Infusions occurred on the seventh and 14th days after CKD induction. Evaluations of kidney function were carried out by biochemical and histological analyses. Intense labeling of the &alpha;-SMA protein was observed when comparing the control with MSC-EVs. In both groups treated with EVs, a significant increase in serum albumin was observed, and the increase in cystatin C was inhibited. The results indicated improvements in renal function in CKD, demonstrating the therapeutic potential of EVs derived from MSCs and eCD133+ cells and suggesting the possibility that in the future, more than one type of EV will be used concurrently