Comparison of Gene Expression in Human Embryonic Stem Cells, hESC-Derived Mesenchymal Stem Cells and Human Mesenchymal Stem Cells

Abstract

We present a strategy to identify developmental/differentiation and plasma membrane marker genes of the most primitive human Mesenchymal Stem Cells (hMSCs). Using sensitive and quantitative TaqMan Low Density Arrays (TLDA) methodology, we compared the expression of 381 genes in human Embryonic Stem Cells (hESCs), hESC-derived MSCs (hES-MSCs), and hMSCs. Analysis of differentiation genes indicated that hES-MSCs express the sarcomeric muscle lineage in addition to the classical mesenchymal lineages, suggesting they are more primitive than hMSCs. Transcript analysis of membrane antigens suggests that IL1R1(low), BMPR1B(low), FLT4(low), LRRC32(low), and CD34 may be good candidates for the detection and isolation of the most primitive hMSCs. The expression in hMSCs of cytokine genes, such as IL6, IL8, or FLT3LG, without expression of the corresponding receptor, suggests a role for these cytokines in the paracrine control of stem cell niches. Our database may be shared with other laboratories in order to explore the considerable clinical potential of hES-MSCs, which appear to represent an intermediate developmental stage between hESCs and hMSCs.

Figure 1

Hierarchical clustering of normalized Ct of 380 genes on hESCs, hES-MSCs, or hMSCs using individual samples and genes. Genes are classified using StatMiner Software and the Pearson test.

Figure 2

Differentiation gene expression levels in hESC (blue), hES-MSC (Red), or hMSC (green) populations. The differentiation genes are classified as follow: (a) neuroectodermic genes; (b) endodermic genes; (c) mesodermic genes; (d) and (e) mesenchymal genes including transcription factors, cell adhesion molecules, and protease genes (d) and Extra-cellular matrix genes (e). y-axis values correspond to 2^−NCt  as described in Material and Methods. The black line represents the limit of detection (LOD: > 35 Ct). Asterisks (*) indicate significant expression variation with P < 0.05.

Figure 3

Transcriptomic expression analyses of potential markers for primitive hMSC selection. Relative gene expression of hESC-derived hMSC (hES-MSC) or hMSC populations compared to hESC calibrator group. Messenger RNA levels were determined using QRT-PCR, and relative quantification (RQ) values were calculated by the 2^−ΔΔCt method with GAPDH as internal control and an arbitrary threshold at 35 cycles. Asterisks (*) indicate significant expression variation with P value < 0.05. As compared to hESC calibrator group, arrows pointing downwards (↓) indicate gene extinction (>35 Ct) and arrows pointing upwards (↑) indicate gene appearance (<35 Ct).