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Transcriptomic Insights into the effect of Mesenchymal Stromal Cells Culture Medium on the Hematopoietic Stem/Progenitor Cells Co-culture

ABSTRACT: To investigate the molecular mechanisms underlying the altered hematopoietic support of mesenchymal stromal cells (MSC) expanded in human platelet lysate (hPL)-supplemented medium, we performed a comprehensive transcriptomic analysis comparing hPL-MSC and fetal bovine serum (FBS)-expanded MSC. MSC from three bone marrow donors were cultured under different adaptation regimens and co-cultured with hematopoietic stem and progenitor cells (HSPC). Transcriptomic analysis revealed distinct gene expression profiles, with 13% of genes differentially expressed between hPL-MSC and FBS-MSC, impacting cell signaling, extracellular matrix, and chemotaxis. Key pathways, including TGF-beta, PI3K-Akt, and Wnt signaling, were altered, affecting MSC-mediated HSPC expansion. hPL-MSC exhibited increased expression of inhibitory factors and reduced expression of promoters of HSPC proliferation, influencing Notch, TGF-β, JAK/STAT, and Wnt regulation. These findings highlight critical molecular interactions in MSC-HSPC co-culture and provide insights for optimizing xeno-free MSC expansion for hematopoietic applications.

ORGANISM(S): Homo sapiens

PROVIDER: GSE294580 | GEO | 2026/01/14

REPOSITORIES: GEO

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Project description:Purpose: The population of muscle-derived stem cells called MuStem cells is presented as promising candidate for cell-based therapy of muscle diseases. To validate if this agent can be really presented as therapeutic product and so to be eligible to a future clinical use, it is now required to demonstrate beforehand an efficacy with cells prepared in compliance with good manufacturing practices (GMPs). The aim of the current study was to evaluate the use of two xeno-free blood derivatives corresponding to human serum (HS) and human platelet lysate (hPL) as alternatives to controverted but until now used fetal bovine serum (FBS) for isolation and expansion of human MuStem (hMuStem) cells. Methods: A comparative study was performed with hMuStem cells isolated and in vitro expanded by using commercially available HS and hPL to determine its impact on their proliferation rates, clonogenicity, myogenic commitment level and oligopotency with regard to results obtained under FBS-based medium. Also, their respective phenotype and global gene expression patterns were investigated by flow cytometry and high throughput 3’ digital gene expression RNA-sequencing in order to define a possible differential impact of the human nutrients tested. Results: Comparatively to FBS-based medium, use of HS- and hPL-supplemented ones efficiently supported long-term proliferation of hMuStem cells and enhanced clonogenicity, without main modification of their expression profile and allowing besides limiting the supplementation in growth factors. In vitro differentiation assay combined to transforming growth factor β1 (TGF-β1)-depletion experiments showed a lower myogenic commitment level as well as fusion ability of hMuStem cells when cultured with hPL-based medium according to a TGF-β1-independent process. Use of hPL-derived 3D hydrogel or fibrinogen-depleted hPL demonstrated that heparin-free hPL derivatives maintain consequent myogenic differentiation potential. In addition, the reduced myogenicity was shown to be rapidly reversible following replacement of hPL by HS or fibrinogen-depleted hPL. Conclusions: All together, our original findings position HS and hPL as efficient and suitable alternatives to FBS for preparation of hMuStem cell batch in compliance with GMPs.

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Transcriptomic Insights into the effect of Mesenchymal Stromal Cells Culture Medium on the Hematopoietic Stem/Progenitor Cells Co-culture

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