The mechanisms of idiopathic severe aplastic anemia (SAA) in children are not completely understood. Insufficiency of the bone marrow microenvironment, in which mesenchymal stem cells (MSCs) are an important element, can be a potential factor associated with hematopoietic impairment. In the current study, we studied whether aberrant gene expression could be found in MSCs from children with SAA. Using microarray analysis, two different patterns of global gene expression were detected in the SAA MSCs. Fourteen genes (POLE2, HGF, KIF20A, TK1, IL18R1, KITLG, FGF18, RRM2, TTK, CXCL12, DLG7, TOP2A, NUF2, and TYMS), which are related to DNA synthesis, cytokines, or growth factors, were significantly downregulated. Further, knockdown of gene expression was performed using the small hairpin RNA (shRNA)-containing lentivirus method. We found that knockdown of CXCL12, HGF, IL-18R1, FGF18, or RRM2 expression compelled MSCs from the controls to behave like those from the SAA children, with decreased survival and differentiation potential. Among them, inhibition of CXCL12 gene expression had the most profound effects on the behavior of MSCs. Further experiments regarding re-introduction of the CXCL12 gene could largely recover the survival and differentiation potential in MSCs with inhibition of CXCL12 expression. Our findings suggest that MSCs from children with SAA exhibit aberrant gene expression profiles and downregulation of CXCL12 gene may be associated with alterations in the bone marrow microenvironment.