scholarly journals Promotion of Differentiating Bone Marrow Mesenchymal Stromal Cells (BMSCs) into Cardiomyocytes via HCN2 and HCN4 Cotransfection

2021 ◽  
Vol 2021 ◽  
pp. 1-8
Author(s):  
Xue Luo ◽  
Hongxiao Li ◽  
Xiaolin Sun ◽  
Qisheng Zuo ◽  
Bichun Li ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Western Blot ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ionic Current ◽  
Immunofluorescence Staining ◽  
Patch Clamp Technique ◽  
Marker Proteins ◽  
Myocardial Marker

Aim. Investigation of the influences HCN2 and HCN4 has on bone marrow mesenchymal stromal cells (BMSCs) on cardiomyocyte differentiation. Methods. Miniature adult pigs were used for bone marrow extraction and isolation of BMSCs. The identification of these BMSCs was done by using flow cytometry for the detection of expressed surface antigens CD45, CD11B, CD44, and CD90. Using HCN2 and HCN4 genes cotransfected into BMSCs as group HCN2+HCN4 while myocardial induction solution was used to induced BMSC differentiation in the BMSC induction group. Myocardial marker proteins α-actin and cTnT were detected by immunofluorescence staining, while α-actin, cTnT, and Desmin myocardial marker proteins expressed were detected by Western blot. The whole-cell patch-clamp technique was used to identify and detect cellular HCN2 channels, HCN4 channel current activation curve, and the inhibitory effect of CsCl on heterologous expression currents. Results. Flow cytometry results showed that CD45 and CD11B were expressed negatively while CD90 and CD44 were positive. Post HCN2 and HCN4 gene transfection, immunofluorescence staining, and Western blot showed significantly increased HCN2, HCN4, α-actin, and cTnT expressed in group HCN2+HCN4 were, which could be compared to the expression levels in the BMSC-induced group. The HCN2+HCN4 group was able to document cell membrane channel ion currents that were similar to If properties. Conclusion. HCN2 and HCN4 overexpression can considerably enhance the MSC ability to differentiate into cardiomyocytes in vitro and restore the ionic current.

Mesenchymal Stromal Cells From Myelodysplastic Syndrome Patients Show Lower DICER1 and DROSHA Expression, as Well as Decreased Mir-155 and Mir-181a MicroRNA Expression, When Compared with Healthy Controls

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 397-397
Author(s):  
Carlos Santamaría ◽  
Olga López-VIllar ◽  
Sandra Muntión ◽  
Belén Blanco ◽  
Soraya Carrancio ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Protein Expression ◽  
Myelodysplastic Syndrome ◽  
Western Blot ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Microrna Expression ◽  
Healthy Controls ◽  
Lower Expression

Abstract Abstract 397 Mesenchymal stromal cells (MSC) are closely related to the regulation of hematopoietic stem cell niche. Recently, Raaijmakers et al (Nature, 2010), published that deletion of Dicer1, a RNase III enzyme involved in microRNA biogenesis, in murine MSC-derived osteoprogenitors triggered peripherical blood cytopenias, myelodysplasia and subsequent AML, showing that molecular alterations in bone marrow microenvironment could result in clonal impaired haematopoiesis. Here, we have investigated whether MSC from myelodysplastic syndrome (MDS) patients show differences in DICER1 and DROSHA, another RNA III endonuclease, in comparison to healthy MSC. In addition, we have analyzed several hematopoietic-related microRNAs in these same samples. Bone marrow MSC from MDS patients (n=35; 10 5q- syndrome, 4 RA, 5 RARS, 10 RCMD, 3 RAEB, 2 MDS-U and 1 hypocellular MDS) and healthy donors (HD, n=20) were isolated and in vitro expanded following standard procedures until the third passage. Additionally, paired mononuclear cells (MNC) from 13 MDS and 8 HD were obtained. Total RNA was isolated using TRIzol reagent (Invitrogen). DICER1 and DROSHA relative gene expressions were assessed by quantitative PCR (Q-PCR) using commercial TaqMan® assay (Applied Biosystems®) with GAPDH as control gene. DICER1 and DROSHA (Abcam) protein expression were evaluated in whole cell lysates by western blot, using calnexin (Stressgen) as control. Several microRNAs with known role in hematopoiesis and immune system regulation were analyzed in 25 MDS and 12 HD by Q-PCR using commercial TaqMan® MicroRNA assay (Applied Biosystems®) with RNU43 as control microRNA. MSC from MDS showed significant lower DICER1 (0.0035±0.0020 vs. 0.0076±0.0092; p=0.044) and DROSHA (0.0070±0.0028 vs. 0.0135±0.0176; p=0.019) gene expression levels than healthy controls. Moreover, MSC from MDS showed lower protein expression of both DICER1 and DROSHA by western blot analysis, confirming Q-PCR findings. By contrast, no difference in either DICER1 (0.0197±0.0151 vs. 0.0173±0.0112; p=0.9) or DROSHA (0.0089±0.0023 vs. 0.0067±0.0037; p=0.09) gene expression were observed between MNC from MDS and HD. As far as microRNA expression, we observed a lower expression of mir-155 (0.63±0.92 vs. 0.94±0.49; p=0.007) and mir-181a (1.30±0.95 vs. 2.02±1.05; p=0.041) in MSC from MDS in comparison to healthy controls. Mir-155 and mir-181a are involved in T-cell and B-cell differentiation, while mir-155 are also related to erythroid and megakarycytic differentiation. We conclude that MSC from MDS patients show lower expression of DICER and DROSHA, two relevant RNA-III endonucleases involved in the microRNA biogenesis, confirming recent findings in murine models. Moreover, the expression of some microRNA is impaired in these cells, raising the possibility that these microenvironmental alterations could be involved in the MDS pathophysiology. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Optimization of a flow cytometry-based protocol for detection and phenotypic characterization of multipotent mesenchymal stromal cells from human bone marrow

2006 ◽  
Vol 70B (6) ◽  
pp. 391-399 ◽  
Author(s):  
Elena A. Jones ◽  
Anne English ◽  
Sally E. Kinsey ◽  
Liz Straszynski ◽  
Paul Emery ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Multipotent Mesenchymal Stromal Cells ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Phenotypic Characterization

scholarly journals Clumping and Viability of Bone Marrow Derived Mesenchymal Stromal Cells under Different Preparation Procedures: A Flow Cytometry-Based In Vitro Study

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Li-li Cui ◽  
Tuure Kinnunen ◽  
Johannes Boltze ◽  
Johanna Nystedt ◽  
Jukka Jolkkonen
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Normal Saline ◽  
Stromal Cells ◽  
Pulse Width ◽  
In Vitro Study ◽  
High Viability ◽  
Storage Solutions

Complications of microocclusions have been reported after intra-arterial delivery of mesenchymal stromal cells. Hence, quantification and efficient limitation of cell clumps in suspension before transplantation is important to reduce the risk. We used a flow cytometry-based pulse-width assay to assess the effects of different cell suspension concentrations (0.2–2.0 × 106/mL), storage solutions (complete growth medium, Dulbecco’s phosphate-buffered saline, and normal saline), storage time in suspension (0–9 h), and freeze-thawing procedure on the clumping of rat bone marrow derived mesenchymal stromal cells (BMMSCs) and also evaluated cell viability at the same time. Surprisingly, increasing the cell concentration did not result in more cell clumps in vitro. Freshly harvested (fresh) cells in normal saline had significantly fewer cell clumps and also displayed high viability (>90%). A time-dependent reduction in viability was observed for cells in all three storage solutions, without any significant change in the clumping tendency except for cells in medium. Fresh cells were more viable than their frozen-thawed counterparts, and fresh cells in normal saline had fewer cell clumps. In conclusion, cell clumping and viability could be affected by different cell preparation procedures, and quantification of cell clumping can be conducted using the flow cytometry-based pulse-width assay before intra-arterial cell delivery.

GD2+ Selected Mesenchymal Stromal Cells (MSCs) Demonstrate a More Robust Proliferation and Differentiation Potential Compared to Unselected Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1919-1919
Author(s):  
Caridad Martinez ◽  
Ted J. Hofmann ◽  
Roberta Marino ◽  
Massimo Dominici ◽  
Edwin M. Horwitz
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Doubling Time ◽  
Blue Staining ◽  
Differentiation Potential ◽  
Proliferation And Differentiation

Abstract Human mesenchymal stromal cells (MSCs) are spindle-shape, plastic-adherent cells with capacity to differentiate to bone, cartilage, and fat. MSCs express fibroblast, endothelial, and lymphocyte antigens, e.g. CD105, CD73, CD90, and CD166 which are the cornerstone of phenotypic characterization of these cells. We recently showed that MSCs are the only bone marrow cell to express GD2, a neural ganglioside. Now, for the first time we show that GD2 may serve as the single, unique, and definitive marker of marrow and adipose derived MSCs that can be used to isolate GD2+ MSCs, which possess important biologic properties justifying prospective isolation. MSCs expression of GD2 is uniformly high on freshly isolated and culture-expanded cells. Using the Miltenyi AutoMACS® device and a monoclonal antibody recognizing GD2 (clone 14.G2A) we prospectively isolated a highly enriched MSC population from bone marrow MNCs. The selected fraction was >98% pure for GD2+ cells determined by flow cytometry. Light microscopy showed that the GD2-selected cells were smaller, thinner, and more spindle-like when attached to plastic compared to unselected MSCs which spread wider along the surface of the culture flask, the so-called “fried egg” appearance. The doubling time of GD2-selected MSCs was 30 hrs compared to 90 hrs for unselected cells representing a 3-fold greater growth rate. Cell cycle analysis by flow cytometry showed ∼80% of cells were in G0/G1 and ∼20% were in S/G2/M phases of the cell cycle in both populations. With the shorter doubling time, this data indicates that GD2-selected MSCs move through the cell cycle more rapidly than unselected cells. In accordance with this finding, electron microscopy showed few organelles in the GD2-selected cells, but increase lamellar bodies indicating overall less complexity, but consistent with a greater membrane turnover rate (cell division) than unselected MSCs. Moreover, flow cytometric analysis revealed an increased expression of receptors for bFGF and EFG, known mitogenic factor receptors for MSCs, compared to unselected MSCs. In vitro differentiation of GD2-selected MSCs showed a more robust osteoid matrix formation (osteoblast) and proteoglycan formation (chondroblast) assayed by semi-quantitative Alizarin Red and Alcian blue staining, respectively. Additionally, more GD2-selected MSCs differentiated to adipocytes than among unselected cells. Surprisingly, GD2 expression persisted on the in vitro human MSC-differentiated osteoblasts, chondroblasts, and adipocytes, in contrast to human bone-derived osteoblasts, adipose tissue, and cartilage which lacked GD2 expression. We conclude that GD2 is a unique, stably expressed surface MSC marker which can be used to prospectively isolate MSCs from marrow, GD2-selcted cells have a more robust in vitro proliferation and differentiation potential which may be valuable for cell therapy, and biologically, in vitro isolated MSCs may not represent the in vivo progenitor for bone, fat, or cartilage.

scholarly journals Infusing Mesenchymal Stromal Cells into Porcine Kidneys during Normothermic Machine Perfusion: Intact MSCs Can Be Traced and Localised to Glomeruli

2019 ◽  
Vol 20 (14) ◽  
pp. 3607 ◽  
Author(s):  
Merel Pool ◽  
Tim Eertman ◽  
Jesus Sierra Parraga ◽  
Nils ’t Hart ◽  
Marieke Roemeling-van Rhijn ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Adipose Tissue ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ex Vivo ◽  
Human Adipose Tissue ◽  
Machine Perfusion ◽  
Normothermic Machine Perfusion ◽  
Glomerular Capillaries

Normothermic machine perfusion (NMP) of kidneys offers the opportunity to perform active interventions, such as the addition of mesenchymal stromal cells (MSCs), to an isolated organ prior to transplantation. The purpose of this study was to determine whether administering MSCs to kidneys during NMP is feasible, what the effect of NMP is on MSCs and whether intact MSCs are retained in the kidney and to which structures they home. Viable porcine kidneys were obtained from a slaughterhouse. Kidneys were machine perfused during 7 h at 37 °C. After 1 h of perfusion either 0, 105, 106 or 107 human adipose tissue derived MSCs were added. Additional ex vivo perfusions were conducted with fluorescent pre-labelled bone-marrow derived MSCs to assess localisation and survival of MSCs during NMP. After NMP, intact MSCs were detected by immunohistochemistry in the lumen of glomerular capillaries, but only in the 107 MSC group. The experiments with fluorescent pre-labelled MSCs showed that only a minority of glomeruli were positive for infused MSCs and most of these glomeruli contained multiple MSCs. Flow cytometry showed that the number of infused MSCs in the perfusion circuit steeply declined during NMP to approximately 10%. In conclusion, the number of circulating MSCs in the perfusate decreases rapidly in time and after NMP only a small portion of the MSCs are intact and these appear to be clustered in a minority of glomeruli.

scholarly journals The Metastatic Bone Marrow Niche in Neuroblastoma: Altered Phenotype and Function of Mesenchymal Stromal Cells

Cancers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3231 ◽  
Author(s):  
Caroline Hochheuser ◽  
Lieke M. J. van Zogchel ◽  
Marion Kleijer ◽  
Carlijn Kuijk ◽  
Simon Tol ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Bone Marrow Niche ◽  
Main Site ◽  
Bm Niche ◽  
Promote Tumor Cell ◽  
And Function ◽  
And Control

Background: The bone marrow (BM) is the main site of metastases and relapse in patients with neuroblastoma (NB). BM-residing mesenchymal stromal cells (MSCs) were shown to promote tumor cell survival and chemoresistance. Here we characterize the MSC compartment of the metastatic NB BM niche. Methods: Fresh BM of 62 NB patients (all stages), and control fetal and adult BM were studied by flow cytometry using well-established MSC-markers (CD34−, CD45−, CD90+, CD105+), and CD146 and CD271 subtype-markers. FACS-sorted BM MSCs and tumor cells were validated by qPCR. Moreover, isolated MSCs were tested for multilineage differentiation and Colony-forming-unit-fibroblasts (CFU-Fs) capacity. Results: Metastatic BM contains a higher number of MSCs (p < 0.05) with increased differentiation capacity towards the osteoblast lineage. Diagnostic BM contains a MSC-subtype (CD146+CD271−), only detected in BM of patients with metastatic-NB, determined by flow cytometry. FACS-sorting clearly discriminated MSC(-subtypes) and NB fractions, validated by mRNA and DNA qPCR. Overall, the CD146+CD271− subtype decreased during therapy and was detected again in the majority of patients at relapse. Conclusions: We demonstrate that the neuroblastoma BM-MSC compartment is different in quantity and functionality and contains a metastatic-niche-specific MSC-subtype. Ultimately, the MSCs contribution to tumor progression could provide targets with potential for eradicating resistant metastatic disease.

scholarly journals The Analysis of In Vivo Aging in Human Bone Marrow Mesenchymal Stromal Cells Using Colony-Forming Unit-Fibroblast Assay and the CD45lowCD271+ Phenotype

2019 ◽  
Vol 2019 ◽  
pp. 1-14 ◽  
Author(s):  
Payal Ganguly ◽  
Jehan J. El-Jawhari ◽  
Agata N. Burska ◽  
Frederique Ponchel ◽  
Peter V. Giannoudis ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Human Serum ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Connexin 43 ◽  
Colony Forming Unit ◽  
Age Related ◽  
Old Donor

Uncultured mesenchymal stromal cells (MSCs) are increasingly used in therapies; however, the effects of donor age on their biological characteristics and gene expression remain unclear. The aim of this study was to investigate age-related changes in bone marrow (BM) MSCs following minimal or no culture manipulation. Iliac crest BM was aspirated from 67 healthy donors (19-89 years old) and directly used for the colony-forming unit-fibroblast (CFU-F) assay or CD45lowCD271+ cell enumeration. The colonies were analysed for colony area and integrated density (ID) when grown in standard MSC media or media supplemented with human serum from young (YS) or old (OS) donors. There was a notable age-related decline in the number of MSCs per millilitre of BM aspirate revealed by the CFU-F assay (r=−0.527, p<0.0001) or flow cytometry (r=−0.307, p=0.0116). Compared to young donors (19-40 years old), colony IDs were significantly lower in older donors (61-89 years old), particularly for smaller-sized colonies (42% lower, p<0.01). When cultured in media supplemented with OS, young and old donor MSCs formed colonies with lower IDs, by 21%, p<0.0001, and 27%, p<0.05, respectively, indicating the formation of smaller sparser colonies. No significant differences in the expression of selected adipogenic, osteogenic, stromal, and bone remodelling genes as well as CD295, CD146, CD106, and connexin 43 surface molecules were found in sorted CD45lowCD271+ MSCs from young and old donors (n=8 donors each). Altogether, these results show similar trends for age-related decline in BM MSC numbers measured by the CFU-F assay and flow cytometry and reveal age-related effects of human serum on MSC colony formation. No significant differences in selected gene expression in uncultured CD45lowCD271+ MSCs suggest that old donor MSCs may not be inferior in regard to their multipotential functions. Due to large donor-to-donor variation in all donor groups, our data indicate that an individual’s chronological age is not a reliable predictor of their MSC number or potency.

224 CHARACTERIZATION OF EQUINE ENDOMETRIAL-DERIVED MESENCHYMAL STROMAL CELLS

2016 ◽  
Vol 28 (2) ◽  
pp. 243
Author(s):  
E. Rink ◽  
H. French ◽  
E. Watson ◽  
C. Aurich ◽  
F. X. Donadeu
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Cell Lines ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Magnetic Beads ◽  
Type I ◽  
Trilineage Differentiation ◽  
Significant Difference ◽  
Cell Fractions

Equine mesenchymal stromal cells (MSC) are mainly harvested from bone marrow and adipose tissue, requiring surgical procedures. Although human endometrium is known to harbor mesenchymal precursor cells, the presence of MSC in equine endometrium, a dynamic tissue, has not been investigated. This study reports for the first time the culture and characterisation of MSC from equine endometrium compared with equine bone marrow (BM)-derived MSC. Samples of equine endometrium (n = 6) and BM (n = 3) were collected postmortem. Endometrial tissue was digested using a dissociation medium containing collagenase I and DNase type I, and CD227 (mucin-1)-bound magnetic beads were utilised to separate epithelial (CD227+) from stromal (CD227–) cell fractions. Red blood cells from BM samples were excluded using a density gradient. All cell fractions were cultured in DMEM/F-12 containing 10% fetal bovine serum. After expansion, colony-forming unit (CFU) assay at passage 2, trilineage differentiation (adipogenic, chondrogenic, osteogenic), and flow cytometry analysis at passage 3/4 were performed for CD227– fractions and BM-MSC. Descriptive statistical analysis and 2-tailed t-test was performed with IBM SPSS Statistics 22 (SPSS Inc./IBM, Chicago, IL, USA). Both isolated cell fractions were plastic adherent and grew well under standard MSC culture conditions, although endometrial CD227– cells attached quicker to culture plasticware than did BM-MSC. The CFU assay at passage 2 showed no significant difference in cloning efficiency (CE) between BM-MSC (20.78 ± 2.86%) and CD227– (24.89 ± 3.04%) cell lines (P = 0.36). Flow cytometry showed the expression of MSC markers (CD29, CD44, CD90, CD105) and perivascular markers (CD146, NG2) but almost no expression of haematopoietic markers (CD34, CD45) in both cell lines (Table 1). No statistically relevant difference was seen except for the higher expression of NG2 in BM-MSC (P = 0.054). Trilineage differentiation was successfully induced in both cell lines. In conclusion, we showed the presence of putative MSC in equine endometrium. We successfully isolated and cultured these cells, which display comparable characteristics in MSC criteria as well-established BM-derived MSC. These endometrial-derived MSC may provide a convenient source for veterinary regenerative therapies in equine reproduction. Table 1.Flow cytometry marker expression (mean ± standard error) at passage 4

scholarly journals Bone Marrow-Derived Mesenchymal Stromal Cells (MSCs) Modulate the Inflammatory Character of Alveolar Macrophages from Sarcoidosis Patients

2020 ◽  
Vol 9 (1) ◽  
pp. 278 ◽  
Author(s):  
Ian McClain Caldwell ◽  
Christopher Hogden ◽  
Krisztian Nemeth ◽  
Michael Boyajian ◽  
Miklos Krepuska ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Alveolar Macrophages ◽  
Stromal Cells ◽  
Cultured Cells ◽  
Intracellular Cytokines ◽  
Tnf Α ◽  
Inflammatory Phenotype ◽  
Anti Inflammatory

Sarcoidosis is a devastating inflammatory disease affecting many organs, especially the lungs and lymph nodes. Bone marrow-derived mesenchymal stromal cells (MSCs) can “reprogram” various types of macrophages towards an anti-inflammatory phenotype. We wanted to determine whether alveolar macrophages from sarcoidosis subjects behave similarly by mounting an anti-inflammatory response when co-cultured with MSCs. Fifteen sarcoidosis and eight control subjects underwent bronchoscopy and bronchoalveolar lavage (BAL). Unselected BAL cells (70–94% macrophages) were isolated and cultured with and without MSCs from healthy adults. Following stimulation of the cultured cells with lipopolysaccharide, the medium was removed to measure interleukin 10 and tumor necrosis factor alpha (IL-10 and TNF-α). In two additional sarcoidosis subjects, flow cytometry was used to study intracellular cytokines and surface markers associated with alveolar macrophages to confirm the results. Unselected BAL cells from sarcoidosis subjects co-cultured with MSCs showed a reduction in TNF-α (pro-inflammatory M1) and an increase in IL-10 (anti-inflammatory M2) in 9 of 11 samples studied. Control subject samples showed few, if any, differences in cytokine production. Unselected BAL cells from two additional patients analyzed by flow cytometry confirmed a switch towards an anti-inflammatory state (i.e., M1 to M2) after co-culture with MSCs. These results suggest that, similarly to other macrophages, alveolar macrophages also respond to MSC contacts by changing towards an anti-inflammatory phenotype. Based on our results, we hypothesize that mesenchymal stromal cells applied to the airways might alleviate lung inflammation and decrease steroid need in patients with sarcoidosis.

Interference of bone marrow CD56 + mesenchymal stromal cells in minimal residual disease investigation of neuroblastoma and other CD45 − /CD56 + pediatric malignancies using flow cytometry

2019 ◽  
Vol 66 (8) ◽  
Author(s):  
Ioannis Theodorakos ◽  
Georgios Paterakis ◽  
Vassilios Papadakis ◽  
Ales Vicha ◽  
Georgios Topakas ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Minimal Residual Disease ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Residual Disease ◽  
Pediatric Malignancies ◽  
Minimal Residual
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