Functional Integrity and Gene Expression Profiles of Human Cord Blood-Derived Hematopoietic Stem and Progenitor Cells Generated In Vitro

Abstract

To date, different experimental strategies have been developed for the ex vivo expansion of human hematopoietic stem (HSCs) and progenitor (HPCs) cells. This has resulted in significant advances on the use of such expanded cells in transplantation settings. To this day, however, it is still unclear to what extent those stem and progenitor cells generated in vitro retain the functional and genomic integrity of their freshly isolated counterparts. In trying to contribute to the solving of this issue, in the present study we have selected and purified three different hematopoietic cell populations: HSCs (CD34+ CD38‐ CD45RA‐ CD71‐ Lin‐ cells), myeloid progenitor cells (CD34+ CD38+ CD45RA+ CD71‐ Lin‐ cells), and erythroid progenitor cells (CD34+ CD38+ CD45RA‐ CD71+ Lin‐ cells), obtained directly from fresh human umbilical cord blood (UCB) units or generated in vitro under particular culture conditions. We, then, compared their functional integrity in vitro and their gene expression profiles. Our results indicate that in spite of being immunophenotipically similar, fresh and in vitro generated cells showed significant differences, both in functional and genetic terms. As compared to their fresh counterparts, those HSCs generated in our culture system showed a deficient content of long‐term culture‐initiating cells, and a marked differentiation bias toward the myeloid lineage. In addition, in vitro generated HSCs and HPCs showed a limited expansion potential. Such functional alterations correlated with differences in their gene expression profiles. These observations are relevant in terms of HSC biology and may have implications in UCB expansion and transplantation. Stem Cells Translational Medicine 2018;7:602–614

Identification of Differentially Expressed Hematopoiesis-associated Genes in Term Low Birth Weight Newborns by Systems Genomics Approach

“…Two microarray datasets associated with gene expression profiling of hematopoietic stem cells GSE64888 (reference dataset I) [23] and GSE107497 (reference dataset II) [24] were retrieved from the GEO database (https:// www.ncbi.nlm.nih.gov/geo/) for comparative analysis of our microarray dataset GSE29807 (test dataset) on differential gene expression in term LBW-NBW newborns [18]. The reference datasets were chosen primarily as they were related to expression profiling of human hematopoietic stem/ progenitor cells and their early myeloid and erythroid proge-ny, using RNA samples derived from human cord blood from healthy newborns same as that for the test dataset.…”

“…As our objective was to study the status of DEHGs in low birth weight (LBW) newborns so, using gene expression information from similar sample source (cord blood) was necessary. The gene expression profiling of the datasets was performed using Affymetrix Human Genome U133 Plus 2.0 Array [18,23] or Affymetrix Human Transcriptome Array 2.0 [24]. A comparative analysis of the reference datasets with the test dataset was done to identify DEHGs between LBW and NBW newborns.…”

“…To segregate the hematopoiesis associated genes present in our test dataset, it was compared with two reference da-tasets (GSE64888, and GSE107497) of hematopoietic stem progenitor cell and their committed erythroid and myeloid progeny [23,24]. The comparison of our test dataset revealed that there were overall 16,589 genes present in our dataset, that are associated with hematopoiesis (D+E+G, Fig.…”

“…An E-value score of the docked complex (LYZ: GM-CSF) was -826.5. Nine hydrogen bonds involved in this interaction are ARG 23 -ASP 49 , ARG 24 -GLN 58 , ARG 62 -VAL 116 , ARG 62 -ILE 117 , ARG 23 -TYR 63 , ARG 98 -GLN 11 , ASN 17 -GLN 104 , ARG 113 -GLU 21 and LYS 72 -ASN 114 . Interacting interface residues are essential for recognition and binding to other proteins.…”

Autologous Cord Blood in Children with Cerebral Palsy: A Review

“…A technique of the ex vivo expansion of CB-derived stem cells has been developed, and its clinical efficacy has been positively verified, despite initial problems [114][115][116][117][118]. However, the cells expanded ex vivo undergo changes in genetic expression, which is an issue that requires further studies [119]. Additionally, the parameters of CB units may be ameliorated by improving the CB thawing procedure [120], using DMSO dextrose instead of DMSO [121], and adding new substances before cryopreservation, such as quercetin [122].…”