Prospective Identification and Characterization of Human Mesenchymal Stem Cell Subpopulations.

Abstract Mesenchymal stem cells (MSC) are multipotent cells that readily differentiate into a variety of cell types including osteoblasts, chondrocytes, and adipocytes. They support hematopoiesis (stroma function) and have potent immunomodulatory properties making them promising candidates for clinical use. Although MSC cultures are heterogeneous in morphology, no specific markers are available that would allow for the prospective isolation of distinct subpopulations. Therefore we investigated whether potential MSC subpopulation markers could be identified by gene expression profiling of MSC that were sorted based on functional parameters (i.e. proliferation characteristics). Human MSC were generated in standard cultures and stained with carboxyfluoresceine-succinimidylesther (CFSE) for cell division tracking. FACS analysis of CFSE-stained cells after 10 days culture showed that the majority of cells had undergone more than 3 cell divisions (51.2 ± 1.7%) whereas only 3.5 ± 0.9 % had not divided. Based on these results MSC were sorted by FACS according to their divisional history into non-proliferating cells (NP-MSC) and rapidly-proliferating cells (RP-MSC). Interestingly, the CFU-F frequencies of the NP-MSC were considerably (5-times) lower compared to RP-MSC. Furthermore, RP-MSC with low forward-/side-scatter properties had a higher CFU-F frequency than those with high scatter properties. Comparative oligonucleotide microarray analysis of FACS-sorted NP-MSC and RP-MSC populations identified a total of 112 differentially (≥4-fold) expressed genes. Fifty genes were significantly higher expressed in RP-MSC with 12 of them being cell cycle associated. Sixty-two genes showed a higher expression in NP-MSC and more than 20 of them were related to adhesion molecules, extracellular matrix proteins or cell senescence. Two of the differentially expressed genes (VCAM-1, FMOD) that corresponded to cell surface molecules were further tested for their potential to prospectively identify MSC subpopulations. Remarkably, FACS analysis using anti-VCAM-1 (CD106) and anti-fibromodulin (fmod) antibodies showed that approximately 40% and 3% of MSC were CD106neg/fmodneg and CD106pos/fmodpos, respectively. MSC were then sorted by FACS according to their expression of these markers and assayed for CFU-F frequency. Significant differences in CFU-F frequencies were observed between the double-negative and the double-positive MSC (12.3 ± 2.8 and 4.7 ± 2.1 colonies per 100 cells for CD106neg/fmodneg and CD106pos/fmodpos MSC, respectively). Experiments to test for differentiation potential differences are ongoing. Taken together, the data clearly demonstrate functional differences between MSC subpopulations. Furthermore, cell sorting based on proliferation characteristics and gene expression profiling enabled to identify surface markers that allowed for the first time to prospectively identify MSC subpopulations.

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Comparison of different isolation techniques prior gene expression profiling of blood derived cells: impact on physiological responses, on overall expression and the role of different cell types

The Pharmacogenomics Journal ◽

10.1038/sj.tpj.6500240 ◽

2004 ◽

Vol 4 (3) ◽

pp. 193-207 ◽

Cited By ~ 143

Author(s):

S Debey ◽

U Schoenbeck ◽

M Hellmich ◽

B S Gathof ◽

R Pillai ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Physiological Responses ◽

Cell Types ◽

Isolation Techniques ◽

Different Cell Types

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Nonsense-Mediated mRNA Decay Is Essential for the Hematopietic Compartement.

Blood ◽

10.1182/blood.v110.11.506.506 ◽

2007 ◽

Vol 110 (11) ◽

pp. 506-506

Author(s):

Joachim Weischenfeldt ◽

Inge Damgaard ◽

David Bryder ◽

Claus Nerlov ◽

Bo Porse

Keyword(s):

Gene Expression ◽

Bone Marrow ◽

T Cells ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Mrna Decay ◽

Double Positive ◽

Mrna Pool ◽

Nonsense Mediated Mrna Decay

Abstract Nonsense-mediated mRNA decay (NMD) is a conserved cellular surveillance system that degrades mRNAs with premature termination codons (PTCs). PTC-containing transcripts can arise from faulty events such as erroneous mRNA processing events as well as mutations, and their translation may lead to the synthesis of deleterious proteins. In addition to serving as a genomic protection system, experiments in tissue culture cells have demonstrated that NMD regulates 5% of the normal mRNA pool suggesting that the NMD pathway may have a broader role in gene regulation. Finally, NMD has also been proposed to be important during lymphocyte development as a tool of riding the cells of transcripts resulting from unproductive re-arrangements events of T cell receptor and immunoglobulin genes. Although NMD has been studied extensively at the biochemical level, the actual role and importance of NMD in the mammalian organism has not been investigated. We therefore generated a conditional Upf2 knock-out mouse line (UPF2 being an essential NMD factor) which we crossed to different hematopoietic relevant Cre expressing lines. Full ablation of UPF2 (using the inducible Mx1-Cre deleter) led to complete loss of all nucleated cells in the bone marrow and death of the animals within 10 days. A similar phenotype was observed when Upf2fl/fl; Mx1Cre BM cells were transplanted into lethally irradiated WT recipients and induced with poly-IC, demonstrating the cell autonomous nature of the phenotype. Deletion of UPF2 in the myeloid lineage using the LysM-Cre deleter resulted in efficient ablation of UPF2 and the absence of NMD in reporter transfected bone marrow derived macrophages (BMDMs). However, the steady state levels of myeloid cells appeared unaltered. Finally, deletion of UPF2 in T cells using a Lck-Cre deleter led to a marked reduction of both CD4/CD8 double-positive and single-positive T cells and accumulation of PTC containing transcripts. Gene expression profiling experiments of BMDM and thymocytes from WT and UPF2-ablated animals identified a common core set of 27 up-regulated genes consistent with the role of NMD as a mRNA degrading system. The gene expression profiling data suggest that ablation of NMD leads to accumulation of unfolded proteins. In summary, these studies demonstrate the vital and cell-autonomous role of NMD in the hematopoietic system.

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Abstract 1528: Selective gene-expression profiling of metastasizing breast tumor cell subpopulations complements the predictive power of Mammaprint Dx and Oncotype Dx

10.1158/1538-7445.am2016-1528 ◽

2016 ◽

Author(s):

George S. Karagiannis ◽

Sumanta Goswami ◽

Joan G. Jones ◽

Maja H. Oktay ◽

John S. Condeelis

Keyword(s):

Gene Expression ◽

Tumor Cell ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Breast Tumor ◽

Predictive Power ◽

Oncotype Dx ◽

Breast Tumor Cell ◽

Cell Subpopulations

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Clustering cell types and identifying cell-type specific transcripts by gene expression profiling in single neurons

Nature Genetics ◽

10.1038/14354 ◽

1999 ◽

Vol 23 (S3) ◽

pp. 59-59

Author(s):

F.J. Livesey ◽

S. Gustincich ◽

M. Steffen ◽

V. Cheung ◽

E. Raviola ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Cell Types ◽

Single Neurons ◽

Cell Type ◽

Cell Type Specific

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Gene Expression Profiling Reveals Distinct Cocaine-Responsive Genes in Human Fetal CNS Cell Types

Journal of Addiction Medicine ◽

10.1097/adm.0b013e318199d863 ◽

2009 ◽

Vol 3 (4) ◽

pp. 218-226 ◽

Cited By ~ 16

Author(s):

Chun-Ting Lee ◽

Elin Lehrmann ◽

Teruo Hayashi ◽

Rose Amable ◽

Shang-Yi Tsai ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Cell Types

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Modeling multifunctionality of genes with secondary gene co-expression networks in human brain provides novel disease insights

10.1101/2020.09.29.317305 ◽

2020 ◽

Author(s):

Juan A. Sánchez ◽

Ana L. Gil-Martinez ◽

Alejandro Cisterna ◽

Sonia García-Ruíz ◽

Alicia Gómez ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Analysis ◽

Gene Function ◽

Expression Profiling ◽

Cell Types ◽

Cell Type ◽

Bulk Tissue ◽

Function Cell ◽

Tissue Gene Expression

AbstractMotivationCo-expression networks are a powerful gene expression analysis method to study how genes co-express together in clusters with functional coherence that usually resemble specific cell type behaviour for the genes involved. They can be applied to bulk-tissue gene expression profiling and assign function, and usually cell type specificity, to a high percentage of the gene pool used to construct the network. One of the limitations of this method is that each gene is predicted to play a role in a specific set of coherent functions in a single cell type (i.e. at most we get a single <gene, function, cell type> for each gene). We present here GMSCA (Gene Multifunctionality Secondary Co-expression Analysis), a software tool that exploits the co-expression paradigm to increase the number of functions and cell types ascribed to a gene in bulk-tissue co-expression networks.ResultsWe applied GMSCA to 27 co-expression networks derived from bulk-tissue gene expression profiling of a variety of brain tissues. Neurons and glial cells (microglia, astrocytes and oligodendrocytes) were considered the main cell types. Applying this approach, we increase the overall number of predicted triplets <gene, function, cell type> by 46.73%. Moreover, GMSCA predicts that the SNCA gene, traditionally associated to work mainly in neurons, also plays a relevant function in oligodendrocytes.AvailabilityThe tool is available at GitHub,https://github.com/drlaguna/GMSCA as open source software.ImplementationGSMCA is implemented in R.

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