Short-term exposure of umbilical cord blood CD34+ cells to granulocyte–macrophage colony-stimulating factor early in culture improves ex vivo expansion of neutrophils

Marturana, Flavia; Timmins, Nicholas; Nielsen, Lars K.

doi:10.3109/14653249.2010.518610

Cited by 13 publications

(10 citation statements)

References 60 publications

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“…Control cultures containing more common high cytokine conditions (100 ng/mL of SCF, 100 ng/mL TPO peptide 32 [referred to as TPO hereafter; Auspep Pty] and 50 ng/mL Flt-3 ligand) were carried out in parallel. The high cytokine condition utilized early acting cytokines 33 and is similar to that used in a recent 3D MSC coculture study. 12 MSC seeding density was varied in low cytokine cultures to determine if increasing levels of MSCs correlated with improvement in CD34 + expansion.…”

Section: Methodsmentioning

confidence: 99%

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly ImprovesIn VitroHematopoietic Stem/Progenitor Cell Expansion

Futrega

Atkinson

Lott

et al. 2017

Tissue Engineering Part C: Methods

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While two-dimensional (2D) monolayers of mesenchymal stem/stromal cells (MSCs) have been shown to enhance hematopoietic stem/progenitor cell (HSPC) expansion in vitro, expanded cells do not engraft long term in human recipients. This outcome is attributed to the failure of 2D culture to recapitulate the bone marrow (BM) niche signal milieu. Herein, we evaluated the capacity of a novel three-dimensional (3D) coculture system to support HSPC expansion in vitro. A high-throughput polydimethylsiloxane (PDMS) microwell platform was used to manufacture thousands of uniform 3D multicellular coculture spheroids. Relative gene expression in 3D spheroid versus 2D adherent BM-derived MSC cultures was characterized and compared with literature reports. We evaluated coculture spheroids, each containing 25–400 MSCs and 10 umbilical cord blood (CB)-derived CD34+ progenitor cells. At low exogenous cytokine concentrations, 2D and 3D MSC coculture modestly improved overall hematopoietic cell and CD34+ cell expansion outcomes. By contrast, a substantial increase in CD34+CD38− cell yield was observed in PDMS microwell cultures, regardless of the presence or absence of MSCs. This outcome indicated that CD34+CD38− cell culture yield could be increased using the microwell platform alone, even without MSC coculture support. We found that the increase in CD34+CD38− cell yield observed in PDMS microwell cultures did not translate to enhanced engraftment in NOD/SCID gamma (NSG) mice or a modification in the relative human hematopoietic lineages established in engrafted mice. In summary, there was no statistical difference in CD34+ cell yield from 2D or 3D cocultures, and MSC coculture support provided only modest benefit in either geometry. While the high-throughput 3D microwell platform may provide a useful model system for studying cells in coculture, further optimization will be required to generate HSPC yields suitable for use in clinical applications.

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Section: Methodsmentioning

confidence: 99%

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly ImprovesIn VitroHematopoietic Stem/Progenitor Cell Expansion

Futrega

Atkinson

Lott

et al. 2017

Tissue Engineering Part C: Methods

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“…Human primary cell culture was performed as previously described [29]. Erythroid and non-erythroid (G/M) differentiation of CD34 + umbilical cord blood was performed as previously described [29-31]. Flow cytometric analyses were performed for erythroid markers (CD45 FITC, GlyA PE, and CD71 APC) and GM markers (CD15 FITC, CD11b PE, and CD45 APC).…”

Section: Methodsmentioning

confidence: 99%

The RNA Binding Protein RBM38 (RNPC1) Regulates Splicing during Late Erythroid Differentiation

et al. 2013

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Alternative pre-mRNA splicing is a prevalent mechanism in mammals that promotes proteomic diversity, including expression of cell-type specific protein isoforms. We characterized a role for RBM38 (RNPC1) in regulation of alternative splicing during late erythroid differentiation. We used an Affymetrix human exon junction (HJAY) splicing microarray to identify a panel of RBM38-regulated alternatively spliced transcripts. Using microarray databases, we noted high RBM38 expression levels in CD71+ erythroid cells and thus chose to examine RBM38 expression during erythroid differentiation of human hematopoietic stem cells, detecting enhanced RBM38 expression during late erythroid differentiation. In differentiated erythroid cells, we validated a subset of RBM38-regulated splicing events and determined that RBM38 regulates activation of Protein 4.1R (EPB41) exon 16 during late erythroid differentiation. Using Epb41 minigenes, Rbm38 was found to be a robust activator of exon 16 splicing. To further address the mechanism of RBM38-regulated alternative splicing, a novel mammalian protein expression system, followed by SELEX-Seq, was used to identify a GU-rich RBM38 binding motif. Lastly, using a tethering assay, we determined that RBM38 can directly activate splicing when recruited to a downstream intron. Together, our data support the role of RBM38 in regulating alternative splicing during erythroid differentiation.

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“…Precursor cells can be stimulated to differentiate into mature neutrophils through signals from granulocyte macrophage colony stimulating factor (GM-CSF) [61] . Neutrophils are thought to be very important to early immune response.…”

Section: Neutrophil Differentiationmentioning

confidence: 99%

The heterogenic properties of monocytes/macrophages and neutrophils in inflammatory response in diabetes

2014

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Inflammation is a complicated biological process in response to harmful stimuli, which involves the cooperation of immune system and vascular system. Upon pathogen invasion or tissue injury, resident innate immune cells such as macrophages and dendritic cells are activated and release inflammatory mediators, which result in the vasodilation and recruitment of leukocytes, mainly monocytes and neutrophils. As two of the most important inflammation-mediating immune cells, macrophages and neutrophils have long been regarded to have a pro-inflammatory effect. However, increasing evidences suggest the role of macrophage and neutrophil in inflammation is more complicated and diversified than we thought. Differently activated macrophages and neutrophils lead to diverse even opposite activities. Precise understanding of the role of different subpopulations is critical to achieve the effective treatment for inflammatory diseases. In this review, we discuss the two potentially distinct activation routes of macrophages and neutrophils in obesity and diabetes.

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Short-term exposure of umbilical cord blood CD34+ cells to granulocyte–macrophage colony-stimulating factor early in culture improves ex vivo expansion of neutrophils

Cited by 13 publications

References 60 publications

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly ImprovesIn VitroHematopoietic Stem/Progenitor Cell Expansion

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly ImprovesIn VitroHematopoietic Stem/Progenitor Cell Expansion

The RNA Binding Protein RBM38 (RNPC1) Regulates Splicing during Late Erythroid Differentiation

The heterogenic properties of monocytes/macrophages and neutrophils in inflammatory response in diabetes

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