Comparison of miRNA transcriptome of exosomes in three categories of somatic cells with derived iPSCs

Abstract

Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) through epigenetic manipulation. While the essential role of miRNA in reprogramming and maintaining pluripotency is well studied, little is known about the functions of miRNA from exosomes in this context. To fill this research gap,we comprehensively obtained the 17 sets of cellular mRNA transcriptomic data with 3.93 × 10¹⁰ bp raw reads and 18 sets of exosomal miRNA transcriptomic data with 2.83 × 10⁷ bp raw reads from three categories of human somatic cells: peripheral blood mononuclear cells (PBMCs), skin fibroblasts(SFs) and urine cells (UCs), along with their derived iPSCs. Additionally, differentially expressed molecules of each category were identified and used to perform gene set enrichment analysis. Our study provides sets of comparative transcriptomic data of cellular mRNA and exosomal miRNA from three categories of human tissue with three individual biological controls in studies of iPSCs generation, which will contribute to a better understanding of donor cell variation in functional epigenetic regulation and differentiation bias in iPSCs.

Fig. 1

Schematic workflow of this investigation. (a) Exosome miRNA and total mRNA were collected from three categories of somatic cell and their derived iPSCs, along with identifying hiPSCs characteristics. (b) An overview of the analysis flow of miRNA and mRNA data.

Fig. 2

Characteristics of hiPSCs. (a) The karyotype of hiPSCs derived from three somatic cell types with three samples. (b) The presence of exogenous episomal DNA in hiPSC was identified by agarose gel electrophoresis. Somatic cells transfected by episomal DNA served as the positive control, while the H1 cell line and somatic cells served as the negative controls; GAPDH served as the internal reference. B: PBMCs, F: SFs, U: UCs, i: hiPSCs. The specimens Fi1, Bi2 and Ui3 were chosen to show the results. (c) The expression levels of NANOG OCT4, and SOX2 in somatic cells, their derived hiPSCs and H1 cells were evaluated by qRT-PCR. The gene expression level in each sample was detected triple times, and its mean expression was used as its expression value. Each point represents a sample value. The P-value was calculated by Student’s t-test. *P < 0.05, **P < 0.01, ***P < 0.001. (d) The detection of OCT4, SOX2, SSEA4, TRA-1-60 and TRA-1-81 by immunostaining. scale bar: 200 um. (e) The histology of teratomas induced from hiPSCs derived from different somatic cells. The hiPSCs generated from sample 2 were chosen to show as an example. Teratomas were stained with H&E.

Fig. 3

Gene expression among somatic cells and hiPSCs. (a) Principal components analysis of different somatic cells and their derived hiPSCs. (b) Correltion analysis of different somatic cells and their derived hiPSCs. (c) The mRNA expression levels of the pluripotency and differentiation related genes. (d) Venn plot of DEGs among the three groups. Bips_B: PBMCs-derived hiPSCs Vs. PBMCs, Fips_F: SFs-derived hiPSCs Vs. SFs, Uips_U: UCs-derived hiPSCs Vs. UCs. (e,f) Gene set enrichment analysis of DEGs based on biological process GO and KEGG databases, respectively. The top 20 enriched pathways of each group were displayed.

Fig. 4

Expression of miRNA in exosomes from somatic cells and hiPSCs. (a) Principal components analysis applied to different somatic cells and their derived hiPSCs. (b) Correlation analysis of different somatic cells and their derived hiPSCs. (c) Venn plot of differentially expressed miRNA (DEMs) among the three groups, Bips_B: PBMCs-derived hiPSCs Vs. PBMCs, Fips_F: SFs-derived hiPSCs Vs. SFs, Uips_U: UCs-derived hiPSCs Vs. UCs. (d) The expression profile of the shared DEMs (e,f) Gene set enrichment analysis of DEGs targeted by DEMs based on GO and KEGG databases, respectively. The top 20 enriched pathways of each group were shown.