Focal adhesion protein abnormalities in myelodysplastic mesenchymal stromal cells

Aanei, Carmen Mariana; Eloae, Florin Zugun; Flandrin-Gresta, Pascale; Tavernier, Emmanuelle; Carasevici, E; Guyotat, Denis; Campos, Lydia

doi:10.1016/j.yexcr.2011.08.007

Cited by 24 publications

(32 citation statements)

References 40 publications

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“…22,24,25 Two studies by Aanei et al demonstrated again that MDS-MSC present intrinsic growth deficiencies that are related to focal adhesion protein abnormalities; the MSC from patients with refractory cytopenia (mostly LR-MDS patients) had reduced proliferation capacity while cells from patients with an excess of blasts (HR-MDS patients) had increased proliferation. 26,27 This differs from our observations and may be explained by differences in the patients' characteristics, cell sorting strategy, and culture conditions. MDS-MSC were positive (>90%) in all cases for CD73, CD90 and CD105; these markers are homogeneously expressed on MSC and useful for robust in vitro assays.…”

Section: Discussioncontrasting

confidence: 99%

Mesenchymal stromal cells from patients with myelodyplastic syndrome display distinct functional alterations that are modulated by lenalidomide

et al. 2013

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ABSTRACTtypes including MDS associated with single del(5q), and characterized their clonogenicity and differentiation potential. Methods Human samplesTwenty MDS patients were studied; their characteristics are detailed in Online Supplementary Table S1. Our control cohort consisted of one male and five female healthy individuals undergoing orthopedic surgery with a median age of 60 years (range, 56-65). MSC were obtained as previously described. 18 Patients were classified for the study as "lower risk" with and without del(5q) (LR, LR-5q ) (<5% bone marrow blasts and IPSS-low/int-1), and as "higher-risk" (HR) (>5% bone marrow blasts and IPSS-int-2/high). Magnetically sorted HSPC were obtained from peripheral blood of healthy individuals after mobilization. Samples were collected after approval by our Institutional Review Board, and written informed consent was obtained. A detailed description is available in the Online Supplementary Material. Lenalidomide treatmentLEN was kindly provided by Celgene (Munich, Germany) and prepared using 10% dimethyl-sulfoxide as a vehicle. The experiments were performed with 0.1 μM and 1 μM of the drug. Characterization of the mesenchymal stromal cellsClonality, self-renewal, generation of single cell-derived colonies (SCD), expression of cell surface and adhesion molecules with flow cytometry, differentiation potential, proliferation, viability, apoptosis, senescence, and cell cycle were studied. Detailed information about the experiments is provided in the Online Supplementary Material. Co-culture of myelodysplastic syndrome-mesenchymal stromal cells with hematopoietic stem and progenitor cells, clonogenicity and apoptosis assayThe capacity of MSC to support HSPC was tested using a 4-week cobblestone area-forming (CAF-C) assay and a 14-day colony-forming unit granulocyte-erythrocytemonocyte/macrophage (CFU-GEMM) assay as described in the Online Supplementary Material. HSPC or the cell line KG1-α were labeled with carboxyfluorescein-diacetate-succinimidyl-ester (CFSE) prior to co-culture with MSC for 72 h, or left unlabeled and co-cultured for 24 h, with or without the addition of 100 nM recombinant tumor necrosis factor alpha (TNF-α, Peprotech). Apoptosis and proliferation were studied by annexin-V staining and CFSE dilution using flow cytometry. Cytokine measurement and transwell migration assaySupernatant from MSC was obtained after culture with or without LEN and used for enzyme-linked immunoabsorbent assay (ELISA) determinations of SDF-1α, angiopoietin-1 (ANG1), and stem cell factor (SCF) levels. Supernatant was also used for a 4-h transwell migration assay with 1x10 5 HSPC, as described previously. 18 The CXCR4 antagonist AMD3100 (Sigma) was used at a concentration of 10 μM.RNA extraction, complementary DNA synthesis and real time polymerase chain reaction mRNA extracted with Trizol reagent (Invitrogen) was transcribed into complementary-DNA and used for real time polymerase chain reaction (RT-PCR) analysis for fatty acid binding protein 4 (FABP4), SDF-1α , ANG1, and SCF. Ab...

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Section: Discussioncontrasting

confidence: 99%

Mesenchymal stromal cells from patients with myelodyplastic syndrome display distinct functional alterations that are modulated by lenalidomide

et al. 2013

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“…Further, cell-to-cell interaction and adhesion among mesenchymal stromal microenvironment niche and HSC are essential to maintain stem cell characteristics and contribute to abnormal HSC growth and maturation [5]. Matsuda-Hashii et al showed that cultures treated with HGF neutralizing antibody inhibited MSCs adhesion [28].…”

Section: Discussionmentioning

confidence: 99%

“…The BM microenvironment is composed of several cell types, including mesenchymal stromal cells (MSCs), which are key components in supporting self-renewal and proliferation of hematopoietic cell progenitors [4]. Numerous studies have demonstrated the morphological and functional alterations in MSCs from MDS patients [5], such as modifications in gene expression and in cytokine secretion [6].…”

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confidence: 99%

Serine Protease Inhibitor Kunitz-Type 2 Is Downregulated in Myelodysplastic Syndromes and Modulates Cell–Cell Adhesion

Roversi

Lopes²,

Machado-Neto³

et al. 2014

Stem Cells and Development

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Myelodysplastic syndromes (MDS) are clonal disorders involving hematopoietic stem cells (HSC) characterized by ineffective hematopoiesis. In addition to HSC defects, a defective hematopoiesis supporting capacity of mesenchymal stromal cells (MSCs) in the microenvironment niche has been implicated in MDS pathophysiology. The interaction between the dysfunctional MSCs MDS and HSC regulates diverse adhesion-related processes, such as progenitor cell survival, proliferation, differentiation, and self-renewal. As previously reported, a microarray analysis identified serine protease inhibitor kunitz-type 2 (SPINT2), an inhibitor of hepatocyte growth factor (HGF) activation, to be downregulated in MSCs from MDS patients. To define the role of SPINT2 in MDS hematopoietic microenvironment, an analysis of the effect of SPINT2 silencing in MSCs was carried out. We herein reported significantly lower levels of SPINT2 whereas HGF was expressed at higher levels in MSCs from MDS patients compared with healthy controls. SPINT2 underexpression results in an increased expression, production, and secretion of HGF and stromal cell-derived factor 1 (SDF-1) by MSCs. An increased adhesion of normal HSC or malignant cells onto MSCs silenced for SPINT2 was also observed. The altered MSCs adhesion in SPINT2-knockdown cells was correlated with increased CD49b and CD49d expression and with a decrease in CD49e expression. Our results suggest that the SPINT2 underexpression in the MSC from MDS patients is probably involved in the adhesion of progenitors to the bone marrow niche, through an increased HGF and SDF-1 signaling pathway.

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“…[6][7][8][9][10] However, other authors report results indicating the opposite. [11][12][13] The discrepancies between the results of the studies might be attributed to the use of different MDS subtypes, different passage numbers and even dissimilar cell populations.…”

Section: Introductionmentioning

confidence: 96%

Down-regulation of Dicer1 promotes cellular senescence and decreases the differentiation and stem cell-supporting capacities of mesenchymal stromal cells in patients with myelodysplastic syndrome

Zhao¹,

Wu²,

Fei³

et al. 2014

Haematologica

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ABSTRACTdence suggests that microRNA (miRNA, miR) are functionally involved in MSC senescence. [18][19][20] The ablation of Dicer1 (an RNAse III endonuclease essential for miRNA biogenesis) and the loss of mature miRNA in embryonic fibroblasts and endothelial cells inhibited cell proliferation and induced a premature senescence phenotype. 21,22 Although recent striking findings indicate that Dicer1 deletion can induce an MDS phenotype in mice 23 and that Dicer1 expression is reduced in MSC from MDS patients, 24 the precise mechanism by which the Dicer1 gene contributes to the pathogenesis of MDS remains elusive. We inferred that disordered expression of Dicer1 could contribute to abnormal MSC senescence and impaired stromal support in MDS.The aims of this study were, therefore, to evaluate the senescent features of BM-MSC from patients with MDS and to explore the role of Dicer1 in the senescence of MSC and the disturbed stromal function observed in patients with MDS. Methods Patients and control samplesA total of 77 patients with MDS were included in this study. Their characteristics are detailed in Online Supplementary Table S1. Patients were diagnosed with MDS according to the minimum diagnostic criteria established by the Conference on MDS. 25Patients were classified for the study as "lower-risk" (LR) [International Prognostic Scoring System (IPSS) low/int-1] and as "higher-risk" (HR) (IPSS-int-2/high).26 Twenty-two healthy volunteers were used as controls (median age 62 years; age range, 43-78 years) and were matched for gender and age. All study participants signed an informed consent form. The research was approved by the ethics committee of the Sixth Hospital affiliated with Shanghai Jiao Tong University, and all patient-relevant research strictly abided by the Declaration of Helsinki. Mesenchymal stromal cell characterizationBM-MSC were isolated and cultured; their immunophenotype, clonogenic and proliferative potential, and differentiation were studied and SA-β-Gal assay were performed. Detailed information about the experiments is provided in the Online Supplementary Material. RNA isolation and real-time polymerase chain reaction analysisTotal RNA was isolated using TRIzol (Invitrogen, Paisley, Scotland) according to the manufacturer's instructions. For mRNA detection, the RNA was reverse transcribed using the RevertAid First Strand cDNA Synthesis Kit (Fermentas, Burlington, Canada) according to the manufacturer's protocol, and real-time polymerase chain reaction (RT-PCR) was performed using RealMasterMix (Takara, Dalian, China). The process used was described in detail in our previous study. 27 The primers are listed in Online Supplementary Table S2. RT-PCR for miRNA was performed using the PrimeScript ™ miRNA qPCR Starter Kit Ver. 2.0 (Takara, Dalian, China). RNU6B was used as the housekeeping gene. Fold change was calculated using the DDCT method of relative quantification. Isolation of CD34 + cells and CD271 + cellsMononuclear cells were obtained from BM aspirates by density gradient separation and...

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Focal adhesion protein abnormalities in myelodysplastic mesenchymal stromal cells

Cited by 24 publications

References 40 publications

Mesenchymal stromal cells from patients with myelodyplastic syndrome display distinct functional alterations that are modulated by lenalidomide

Mesenchymal stromal cells from patients with myelodyplastic syndrome display distinct functional alterations that are modulated by lenalidomide

Serine Protease Inhibitor Kunitz-Type 2 Is Downregulated in Myelodysplastic Syndromes and Modulates Cell–Cell Adhesion

Down-regulation of Dicer1 promotes cellular senescence and decreases the differentiation and stem cell-supporting capacities of mesenchymal stromal cells in patients with myelodysplastic syndrome

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