Anticoagulants on yield of bone marrow-derived mononuclear cells harvested from dogs / Anticoagulantes no rendimento de células mononucleares da medula óssea de cães

Cell therapy with bone marrow-derived mononuclear cells is an alternative to therapy with mesenchymal stem cell cultures. The aim of the present research was the comparison of the yield of bone marrow-derived mononuclear cells harvested from dogs with two different anticoagulants. Bone marrow was harvested from the iliac crest of five healthy dogs aged between 15 and 30 months, and the effect of two anticoagulant solutions, CPDA-1 (citrate phosphate dextrose adenine-1) and heparin, on the isolation of mononuclear cells was compared. Mononuclear cells were isolated in a density gradient and stained for CD9 and CD44 for characterization by flow cytometry. Means were compared using Student's paired t-test. Samples harvested with CPDA-1 yielded an average of 5.16x106 (±1.76x106) to 20.20x106 (±1.55x106) mononuclear cells/mL, whereas the yield of samples harvested with heparin varied between 4.56x106(±0.69x106) and 24.30x106 (±2.12x106) mononuclear cells mL-1. By flow cytometry, mean percentage of double-stained cells varied from 1.96% (±0.64%) to 5.01% (±0.73%) for CPDA-1 and from 2.23% (±0.70%) to 7.27% (±0.97%) for heparin. No significant statistical differences were observed on yield or CD9 and CD44 expression. Further studies are recommended to assess efficacy of CPDA on mononuclear cell isolation.

A terapia com células mononucleares de medula óssea é uma alternativa ao cultivo de células-tronco mesenquimais. O objetivo deste trabalho foi comparar o rendimento de células mononucleares derivadas da medula óssea de cães, colhidas com dois anticoagulantes diferentes. Foram coletadas medulas ósseas de cinco cães hígidos, com idades variando entre 15 e 30 meses, por punção na crista ilíaca. Foi comparado o efeito da solução anticoagulante no isolamento das células mononucleares, utilizando-se CPDA-1 (citrato, fosfato, dextrose, adenina) ou heparina como soluções anticoagulantes. As células mononucleares foram isoladas em gradiente de densidade e caracterizadas fenotipicamente em citometria de fluxo. Os resultados foram submetidos ao Teste t pareado para comparação de médias. Nas amostras coletadas com CPDA-1, o rendimento médio variou entre 5,16 x 106 (±1,76x106) a 20,20x106 (±1,55x106) células mononucleares mL-1, enquanto que, nas amostras coletadas com heparina, o rendimento variou entre 4,56x106 (±0,69x106) a 24,30x106(±2,12x106) células mononucleares/mL. Na citometria de fluxo, a média de células duplo-marcadas variou de 1,96% (±0,64%) a 5,01% (±0,73%) para CPDA-1 e de 2,23% (±0,70%) a 7,27% (±0,97%) para heparina. Não foram observadas diferenças estatísticas no rendimento ou na expressão de CD9 e CD44. Recomendam-se estudos adicionais para avaliar melhor a eficácia do CPDA no isolamento de células mononucleares.

Role of cytokines in the differentiation of iPS cells to hepatocytes / 国际外科学杂志

Induced pluripotent stem cell (iPS cell) opened a new way to stem cell research,iPS cells can differentiate into hepatocytes in vitro gradually,this breakthrough makes the iPS cells have become a promising clinical application source.During the process of iPS cells to hepatocytes,a variety of cytokines regulate gene expression by coordinating various liver cell growth signal to promote iPS cells into hepatocytes differentiation and maturation.This article summarizes the relevant cytokines and their role in iPS cells in hepatocytes differentiation.

Morphology and morphometry of feline bone marrow-derived mesenchymal stem cells in culture / Morfologia e morfometria das células-tronco mesenquimais isoladas de medula óssea de gato

Mesenchymal stem cells (MSC) are increasingly being proposed as a therapeutic option for treatment of a variety of different diseases in human and veterinary medicine. Stem cells have been isolated from feline bone marrow, however, very few data exist about the morphology of these cells and no data were found about the morphometry of feline bone marrow-derived MSCs (BM-MSCs). The objectives of this study were the isolation, growth evaluation, differentiation potential and characterization of feline BM-MSCs by their morphological and morphometric characteristics. in vitro differentiation assays were conducted to confirm the multipotency of feline MSC, as assessed by their ability to differentiate into three cell lineages (osteoblasts, chondrocytes, and adipocytes). To evaluate morphological and morphometric characteristics the cells are maintained in culture. Cells were observed with light microscope, with association of dyes, and they were measured at 24, 48, 72 and 120h of culture (P1 and P3). The non-parametric ANOVA test for independent samples was performed and the means were compared by Tukey's test. On average, the number of mononuclear cells obtained was 12.29 (±6.05x106) cells/mL of bone marrow. Morphologically, BM-MSCs were long and fusiforms, and squamous with abundant cytoplasm. In the morphometric study of the cells, it was observed a significant increase in average length of cells during the first passage. The cell lengths were 106.97±38.16µm and 177.91±71.61µm, respectively, at first and third passages (24 h). The cell widths were 30.79±16.75 µm and 40.18±20.46µm, respectively, at first and third passages (24 h).The nucleus length of the feline BM-MSCs at P1 increased from 16.28µm (24h) to 21.29µm (120h). However, at P3, the nucleus length was 26.35µm (24h) and 25.22µm (120h). This information could be important for future application and use of feline BM-MSCs.(AU)

As células tronco mesenquimais são utilizadas na terapia de várias doenças na medicina humana e veterinária. As células tronco foram isoladas da medula óssea de gato, entretanto, existem poucos dados referentes a morfologia e não existem informações sobre a morfometria das células tronco isoladas da medula óssea. Os objetivos do presente estudo foram o isolamento, avaliação do crescimento, potencial de diferenciação e caracterização morfológica e morfométrica das células mesenquimais de gato isoladas de medula óssea. A diferenciação in vitro foi realizada para confirmar a multipotencialidade das células mesenquimais de gato (diferenciação em osteoblastos, condrócitos, adipócitos). As células mesenquimais foram mantidas em cultivo para avaliações morfológica e morfométrica. As células foram coradas e observadas em microscopia ótica. As mensurações foram realizadas com 24, 48, 72 e 120h de cultura (primeira e terceira passagens). O teste não paramétrico ANOVA foi utilizado e as médias foram comparadas pelo teste de Tukey. O número médio de células mononucleares obtido foi de 12,29 (±6,05x106) células/mL de medula óssea. As células mesenquimais são longas e fusiformes, e escamosas com citoplasma abundante. No estudo morfométrico, observou-se aumento no comprimento médio das células durante a primeira passagem. As medidas de comprimento das células foram: 106,97±38,16µm e 177,91±71,61µm, respectivamente, na primeira e terceira passagens (24 horas). As medidas de largura das células foram: 30,79±16,75 µm e 40,18±20,46 µm, respectivamente, na primeira e terceira passagens (24 horas). O comprimento do núcleo na primeira passagem aumentou de 16,28µm (24h) para 21,29µm (120h) e na terceira passagem foi de 26,35µm (24h) para 25,22µm (120h). As informações são importantes para futuras aplicações e uso da célula mesenquimal de gato.(AU)

Enhancement of distribution of dermal multipotent stem cells to bone marrow in rats of total body irradiation by platelet-derived growth factor-AA treatment / 中华放射医学与防护杂志

Objective To observe whether dermal multipotent stem cells (dMSCs) treated with platelet-derived growth factor-AA ( PDGF-AA )could distribute more frequently to the bone marrow in rats of total body irradiation (TBI).Methods Male dMSCs were isolated and 10 μg/L PDGF-AA was added to the culture medium and further cultured for 2 h.Then the expression of tenascin-C were examined by Western blot, and the migration ability of dMSCs was assessed in transwell chamber.The pre-treated dMSCs were transplanted by tail vein injection into female rats administered with total body irradiation, and 2 weeks after transplantation, real-time PCR was employed to measure the amount of dMSCs in bone marrow.Non-treated dMSCs served as control.Results PDGF-AA treatment increased the expression of tenascin-C in dMSCs, made (1.79 ± 0.13) × 105 cells migrate to the lower chamber under the effect of bone marrow extract, and distributed to bone marrow in TBI rats, significantly more than ( 1.24 ± 0.09) ×105 in non-treated dMSCs (t = 8.833, P ＜ 0.0l ).Conclusions PDGF-AA treatment could enhance the migration ability of dMSCs and increase the amount of dMSCs in bone marrow of TBI rats after transplantation.