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Molecular remodelling of gene expression profile during differentiation of equine bone marrow-derived MSCs towards adipocytes and chondrocytes

ABSTRACT: Background - The mesenchymal stem cells (MSCs) have a unique self-renewal and developmental potential resulting from their multipotency – the ability to differentiate into particular cell types. It has been confirmed that MSCs have a huge potential and outstanding therapeutic effects in variety of clinical treatments due to the their abilities to differentiate towards ectodermal, endodermal, and mesodermal cell lines. Objectives - the project aims to identify changes in MSCs transcriptome profile during the process of losing multipotency and acquiring the ability to differentiate cells into adipocytes (ADM) and chondrocytes (CDM) equine cell lines. Methods - Bone marrow-derived MSCs were differentiated towards adipocytes and chondrocytes using different procedures. The specificity of obtained cell lines was checked using qPCR, cytometry, and staining approaches. The transcriptome analysis was performed using NGS transcript sequencing on the Illumina HiSeq 3000 platform. Results - The differential expression analysis allowed us to identify the 2325 DEGs between MSC and ADM groups, while 803 DEGs were detected between MCS and CDM cell lines. According to the comparison of both DEG sets the expression of 320 genes was significantly modified despite the direction of differentiation. The direct related to the transformation of MSCs into adipocytes seems to be fat cell differentiation represented by 47 DEGs and included additional GOs as regulation of fat cell differentiation, regulation of cell differentiation, brown fat cell differentiation, and adipocyte fat development. The results indicated the top DEGs involved in differentiation into adipocytes and previously established as related to adipogenesis: down-regulated - MEOX1, CYYR1, and up-regulated – CYP24A1; LEP; TBX5; FABP4 and GPER1. The obtained data indicated molecular mechanism altered during equine cell remodelling towards chondrocyte differentiation. One of identified GOs was ossification process represented by 35 DEGs with the highest up regulation of two genes was observed - IHH (6.46) and PHOSPHO1 (5.46) involved in chondrocyte proliferation and matrix mineralisation; respectively. The highest down regulation was detected for BMPR1B gene, critical for bone and cartilage formation (-5.98) and together with BMP2 and BMP4 (2.26 and 2.48; respectively) triggered BMP signaling. Conclusions - The obtained picture of transcriptome differences between primary donor cells and reprogrammed cell lines revealed a range of transcripts important for molecular alterations resulting from substantial phenotypic change of reprogramed cells under special ex-vivo procedures. The study showed the effects of the radical change of cell fate decisions on the molecular level and reinforced further work on efficient procedures to produce MSC-derived reprogrammed cells that have the potential for regenerative medicine of hard connective tissue in horses.

ORGANISM(S): Equus caballus

PROVIDER: GSE271419 | GEO | 2024/11/30

REPOSITORIES: GEO

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Project description:The utility of human pluripotent stem cells as a tool for understanding disease and as a renewable source of cells for transplantation therapies is dependent on efficient differentiation protocols that convert these cells into relevant adult cell types. Here we report the robust and efficient differentiation of human pluripotent stem cells into adipocytes. We found that inducible expression of PPARG2 in pluripotent stem cell-derived mesenchymal progenitor cells programmed their development towards an adipocyte cell fate. Using this approach, multiple human pluripotent cell lines were differentiated into adipocytes with efficiencies of 85% to 90%. These pluripotent stem cell-derived adipocytes retained their identity independent of transgene expression, could be maintained in culture for several weeks, expressed mature markers, and exhibited mature functional properties such as lipid catabolism in response to a beta-adrenergic stimulus. Global transcriptional and lipid metabolomic analyses further confirmed the identity and maturity of these pluripotent stem cell-derived adipocytes. Mesenchymal progenitor cells (MPCs) derived from human embryonic stem cells hESCs and induced pluripotent stem cells (iPSCs) along with adipose-derived stromal vascular cells (ADSVCs) were subjected to induction of PPAR2 and compared to primary fat samples. Overall 2 ADSVC (ADSVC 24 nd 49) lines, 1 hESC (HUES9) line and 1 iPSC (BJRiPS) line were differentiated into MPCs, PPAR2 programmed, and compared to untreated MPCs and primary fat samples from 2 individuals. Each condition is either represented in duplicate or triplicate on affymetrix HuGene-1_0-st arrays. MPCs derived from the hESC lines HUES2 and HUES8, ADSVCs, and BJRiPS were also run on a separate platform (HG-U133_Plus_2) with more GEO presence to facilitate analysis (34 samples, two platforms total). Supplementary file(s): GeneSymbol-collapsed data represent the final normalized data used for analyses in the manuscript.

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Molecular remodelling of gene expression profile during differentiation of equine bone marrow-derived MSCs towards adipocytes and chondrocytes

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