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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function

ABSTRACT: Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function

PROVIDER: PRJNA800085 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR17723000.fastq.gz	Fastqsanger.gz
	SRR17723001.fastq.gz	Fastqsanger.gz
	SRR17723002.fastq.gz	Fastqsanger.gz
	SRR17723003.fastq.gz	Fastqsanger.gz
	SRR17723004.fastq.gz	Fastqsanger.gz

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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [RNA-seq]

Project description:Donor variation is a prominent critical issue limiting applicability of cell-based therapies. We asked whether reference protocols for pre-clinical bone marrow stromal cell (BMSC) propagation using human platelet lysate (hPL) to replace xenogeneic fetal bovine serum (FBS) might also reduce donor variability for osteo-chondral regeneration. We therefore investigated the impact of donor variability, hPL- vs FBS driven propagation and exhaustive proliferation, on BMSC epigenome, transcriptome, phenotype, coagulation risk and osteochondral regenerative function. Polychromatic flow cytometry revealed maintained canonical fibroblastic phenotype in all cells tested. We confirmed significantly declining proliferative potential in FBS-expanded BMSC after proliferative challenge. Notably, BMSC propagation under the aegis of hPL, compared to FBS, significantly increased BMSC proliferation, created significantly different gene expression trajectories and diverging surface marker signatures, already after just one culture passage. We observed limited but measurable effects on DNA methylation irrespective of culture duration. Moreover, expansion under xenogenic serum conditions resulted in significant loss of function during 3D cartilage organoid-like disk formation and significantly increased clotting risk. Superior chondrogenic function under hPL conditions was maintained over culture duration. As an additional observation, platelet blood group and isoagglutinin levels showed phenotypic changes but only minor impact on BMSC chondrogenesis and clotting behaviour.

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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [MethylCap-seq]

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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [MethylCap-seq]

Project description:Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [MethylCap-seq]

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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [RNA-seq]

Project description:Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function [RNA-seq]

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Batch Effects during Human Bone Marrow Stromal Cell Propagation Prevail Donor Variation and Culture Duration: Impact on Genotype, Phenotype and Function.

Project description:Donor variation is a prominent critical issue limiting the applicability of cell-based therapies. We hypothesized that batch effects during propagation of bone marrow stromal cells (BMSCs) in human platelet lysate (hPL), replacing fetal bovine serum (FBS), can affect phenotypic and functional variability. We therefore investigated the impact of donor variation, hPL- vs. FBS-driven propagation and exhaustive proliferation, on BMSC epigenome, transcriptome, phenotype, coagulation risk and osteochondral regenerative function. Notably, propagation in hPL significantly increased BMSC proliferation, created significantly different gene expression trajectories and distinct surface marker signatures, already after just one passage. We confirmed significantly declining proliferative potential in FBS-expanded BMSC after proliferative challenge. Flow cytometry verified the canonical fibroblastic phenotype in culture-expanded BMSCs. We observed limited effects on DNA methylation, preferentially in FBS-driven cultures, irrespective of culture duration. The clotting risk increased over culture time. Moreover, expansion in xenogenic serum resulted in significant loss of function during 3D cartilage disk formation and significantly increased clotting risk. Superior chondrogenic function under hPL-conditions was maintained over culture. The platelet blood group and isoagglutinins had minor impact on BMSC function. These data demonstrate pronounced batch effects on BMSC transcriptome, phenotype and function due to serum factors, partly outcompeting donor variation after just one culture passage.

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Batch effect during human bone marrow stromal cell propagation prevails donor variation and culture duration impact on phenotype, transcriptome and function

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