Expression data from consecutive stages of human early in vitro T-cell differentiation

Human T-cell development is less well studied than its murine counterpart due to the lack of genetic tools and the difficulty of obtaining cells and tissues. However, recent technological advances allow identification of the transcriptional landscape of differentiating human thymocytes. Here we report the gene expression profiles of 11 immature, consecutive T-cell developmental stages. The changes in gene expression of cultured stem cells on OP9-DL1 match those of ex vivo isolated human thymocytes. These analyses led us to define evolutionary conserved gene signatures that represent pre- and post- αβ T-cell commitment stages. We found that loss of CD44 marks T-cell commitment in early CD7+CD5+CD45dim cells, before the acquisition of CD1a surface expression. The CD44-CD1a- post-committed thymocytes have initiated in frame TCR rearrangements and have completely lost the capacity to develop into myeloid, B- and NK-cells, unlike uncommitted CD44+CD1a- thymocytes. Therefore, loss of CD44 represents a previously unrecognized stage that defines the earliest committed T-cell population in the human thymus. We used microarrays to detail the transcriptional program underlying human early T-cell differentiation, from CD34+ HSCs until the CD4+CD8+ DP stage. Throughout the differentiation of human HSC towards the T-cell lineage, we sorted T-cell fractions at 5 time points (days 0, 7, 12, 18, and 27) based on the expression of cell surface markers including CD45, CD34, CD7, CD5, CD1a, CD4 and CD8. Following the sorting of consecutive early T-cell development stages, the samples were then used for RNA extraction and hybridization on Affymetrix microarrays.