Homing Genes Expression in Fucosyltransferase VI-Treated Umbilical Cord Blood CD133+ Cells which Expanded on Protein-Coated Nanoscaffolds.

Abstract

Umbilical cord blood (UCB)-derived hematopoietic stem cells (HSCs) are considered because of their self-renewing, differentiating, proliferating, and readily available properties. Moreover, HSCs' homing to the hematopoietic microenvironment is an important step in their transplantation process. But low content of progenitor cells in one unit of UCB and defect in the bone marrow (BM) homing limit their applications. Hence, we decided to correct this deficiency with ex vivo incubation of CD133+ cells using fucosyltransferase VI and GDP-fucose. Then C-X-C chemokines receptor-4 (CXCR4), very late activation antigen-4 (VLA4), very late activation antigen-5 (VLA5), lymphocyte function-associated antigen-1 (LFA-1), and E-cadherin (E-cad) genes expressions were investigated with the goal of homing evaluation. The purity of MACS isolated CD133+ cells and confirmation of fucosylation were done by flow cytometry, and the viability of cells seeded on protein-coated poly L-lactic acid (PLLA) scaffold was proven via MTT assay. Scanning electron microscopy (SEM), CFU assays, and expression assays of CXCR4, VLA4, VLA5, LFA-1 and E-cad by real-time PCR were performed, too. Flow cytometry data showed that isolated cells were suitable for fucosyltransferase VI (FT-VI) incubation and expansion on nanoscaffolds. MTT, CFU assays, and SEM micrographs demonstrated fibronectin (FN)-collagen-selectin (FCS)-coated scaffold serve as best environment for viability, clonogenicity, and cell attachment. High levels of homing genes expression were also observed in cells seeded on FCS-coated scaffolds. Also, CXCR4 flow cytometry analysis confirmed real-time data. FCS-PLLA scaffolds provided optimal conditions for viability of FT-VI-treated CD133+ cells, and clonogenicity with the goal of improving homing following UCB-HSCs transplantation.