Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations

Abstract Background Human Mesenchymal stromal cells (hMSCs) from various tissue sources are widely investigated in clinical trials. These MSCs are often administered to patients immediately after thawing the cryopreserved product (out-of-thaw), yet little is known about the single-cell transcriptomi...

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Autores principales:	Camila Medrano-Trochez, Paramita Chatterjee, Pallab Pradhan, Hazel Y. Stevens, Molly E. Ogle, Edward A. Botchwey, Joanne Kurtzberg, Carolyn Yeago, Greg Gibson, Krishnendu Roy
Formato:	article
Lenguaje:	EN
Publicado:	BMC 2021
Materias:	MSC Cell therapy Bone marrow derived MSC (BM-MSC) Cord tissue derived MSC (CT-MSC) scRNA-seq Medicine (General) R5-920 Biochemistry QD415-436
Acceso en línea:	https://doaj.org/article/6fea1886cd2e42f6a0a90995bb79307f
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spelling	oai:doaj.org-article:6fea1886cd2e42f6a0a90995bb79307f2021-11-07T12:07:35ZSingle-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations10.1186/s13287-021-02627-91757-6512https://doaj.org/article/6fea1886cd2e42f6a0a90995bb79307f2021-11-01T00:00:00Zhttps://doi.org/10.1186/s13287-021-02627-9https://doaj.org/toc/1757-6512Abstract Background Human Mesenchymal stromal cells (hMSCs) from various tissue sources are widely investigated in clinical trials. These MSCs are often administered to patients immediately after thawing the cryopreserved product (out-of-thaw), yet little is known about the single-cell transcriptomic landscape and tissue-specific differences of out-of-thaw human MSCs. Methods 13 hMSC samples derived from 10 “healthy” donors were used to assess donor variability and tissue-of-origin differences in single-cell gene expression profiles. hMSCs derived and expanded from the bone marrow (BM) or cord tissue (CT) underwent controlled-rate freezing for 24 h. Cells were then transferred to the vapor phase of liquid nitrogen for cryopreservation. hMSCs cryopreserved for at least one week, were characterized immediately after thawing using a droplet-based single-cell RNA sequencing method. Data analysis was performed with SC3 and SEURAT pipelines followed by gene ontology analysis. Results scRNA-seq analysis of the hMSCs revealed two major clusters of donor profiles, which differ in immune-signaling, cell surface properties, abundance of cell-cycle related transcripts, and metabolic pathways of interest. Within-sample transcriptomic heterogeneity is low. We identified numerous differentially expressed genes (DEGs) that are associated with various cellular functions, such as cytokine signaling, cell proliferation, cell adhesion, cholesterol/steroid biosynthesis, and regulation of apoptosis. Gene-set enrichment analyses indicated different functional pathways in BM vs. CT hMSCs. In addition, MSC-batches showed significant variations in cell cycle status, suggesting different proliferative vs. immunomodulatory potential. Several potential transcript-markers for tissue source differences were identified for further investigation in future studies. In functional assays, both BM and CT MSCs suppressed macrophage TNFα secretion upon interferon stimulation. However, differences between donors, tissue-of-origin, and cell cycle are evident in both TNF suppression and cytokine secretion. Conclusions This study shows that donor differences in hMSC transcriptome are minor relative to the intrinsic differences in tissue-of-origin. hMSCs with different transcriptomic profiles showed potential differences in functional characteristics. These findings contribute to our understanding of tissue origin-based differences in out-of-thaw therapeutic hMSC products and assist in the identification of cells with immune-regulatory or survival potential from a heterogeneous MSC population. Our results form the basis of future studies in correlating single-cell transcriptomic markers with immunomodulatory functions.Camila Medrano-TrochezParamita ChatterjeePallab PradhanHazel Y. StevensMolly E. OgleEdward A. BotchweyJoanne KurtzbergCarolyn YeagoGreg GibsonKrishnendu RoyBMCarticleMSCCell therapyBone marrow derived MSC (BM-MSC)Cord tissue derived MSC (CT-MSC)scRNA-seqMedicine (General)R5-920BiochemistryQD415-436ENStem Cell Research & Therapy, Vol 12, Iss 1, Pp 1-14 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	MSC Cell therapy Bone marrow derived MSC (BM-MSC) Cord tissue derived MSC (CT-MSC) scRNA-seq Medicine (General) R5-920 Biochemistry QD415-436
spellingShingle	MSC Cell therapy Bone marrow derived MSC (BM-MSC) Cord tissue derived MSC (CT-MSC) scRNA-seq Medicine (General) R5-920 Biochemistry QD415-436 Camila Medrano-Trochez Paramita Chatterjee Pallab Pradhan Hazel Y. Stevens Molly E. Ogle Edward A. Botchwey Joanne Kurtzberg Carolyn Yeago Greg Gibson Krishnendu Roy Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
description	Abstract Background Human Mesenchymal stromal cells (hMSCs) from various tissue sources are widely investigated in clinical trials. These MSCs are often administered to patients immediately after thawing the cryopreserved product (out-of-thaw), yet little is known about the single-cell transcriptomic landscape and tissue-specific differences of out-of-thaw human MSCs. Methods 13 hMSC samples derived from 10 “healthy” donors were used to assess donor variability and tissue-of-origin differences in single-cell gene expression profiles. hMSCs derived and expanded from the bone marrow (BM) or cord tissue (CT) underwent controlled-rate freezing for 24 h. Cells were then transferred to the vapor phase of liquid nitrogen for cryopreservation. hMSCs cryopreserved for at least one week, were characterized immediately after thawing using a droplet-based single-cell RNA sequencing method. Data analysis was performed with SC3 and SEURAT pipelines followed by gene ontology analysis. Results scRNA-seq analysis of the hMSCs revealed two major clusters of donor profiles, which differ in immune-signaling, cell surface properties, abundance of cell-cycle related transcripts, and metabolic pathways of interest. Within-sample transcriptomic heterogeneity is low. We identified numerous differentially expressed genes (DEGs) that are associated with various cellular functions, such as cytokine signaling, cell proliferation, cell adhesion, cholesterol/steroid biosynthesis, and regulation of apoptosis. Gene-set enrichment analyses indicated different functional pathways in BM vs. CT hMSCs. In addition, MSC-batches showed significant variations in cell cycle status, suggesting different proliferative vs. immunomodulatory potential. Several potential transcript-markers for tissue source differences were identified for further investigation in future studies. In functional assays, both BM and CT MSCs suppressed macrophage TNFα secretion upon interferon stimulation. However, differences between donors, tissue-of-origin, and cell cycle are evident in both TNF suppression and cytokine secretion. Conclusions This study shows that donor differences in hMSC transcriptome are minor relative to the intrinsic differences in tissue-of-origin. hMSCs with different transcriptomic profiles showed potential differences in functional characteristics. These findings contribute to our understanding of tissue origin-based differences in out-of-thaw therapeutic hMSC products and assist in the identification of cells with immune-regulatory or survival potential from a heterogeneous MSC population. Our results form the basis of future studies in correlating single-cell transcriptomic markers with immunomodulatory functions.
format	article
author	Camila Medrano-Trochez Paramita Chatterjee Pallab Pradhan Hazel Y. Stevens Molly E. Ogle Edward A. Botchwey Joanne Kurtzberg Carolyn Yeago Greg Gibson Krishnendu Roy
author_facet	Camila Medrano-Trochez Paramita Chatterjee Pallab Pradhan Hazel Y. Stevens Molly E. Ogle Edward A. Botchwey Joanne Kurtzberg Carolyn Yeago Greg Gibson Krishnendu Roy
author_sort	Camila Medrano-Trochez
title	Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
title_short	Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
title_full	Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
title_fullStr	Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
title_full_unstemmed	Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
title_sort	single-cell rna-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations
publisher	BMC
publishDate	2021
url	https://doaj.org/article/6fea1886cd2e42f6a0a90995bb79307f
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Single-cell RNA-seq of out-of-thaw mesenchymal stromal cells shows tissue-of-origin differences and inter-donor cell-cycle variations

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