Human blood and marrow side population stem cell and Stro-1 positive bone marrow stromal cell numbers decline with age, with an increase in quality of surviving stem cells: correlation with cytokines

Abstract

Hematological deficiencies increase with aging leading to anemias, reduced hematopoietic stress responses and myelodysplasias. This study tested the hypothesis that side population hematopoietic stem cells (SP-HSC) would decrease with aging, correlating with IGF-1 and IL-6 levels and increases in bone marrow fat. Marrow was obtained from the femoral head and trochanteric region of the femur at surgery for total hip replacement (N=100). Whole trabecular marrow samples were ground in a sterile mortar and pestle and cellularity and fat content determined. Marrow and blood mononuclear cells were stained with Hoechst dye and the SP-HSC profiles acquired. Marrow stromal cells (MSC) were enumerated flow cytometrically employing the Stro-1 antibody, and clonally in the colony forming unit fibroblast (CFU-F) assay. Plasma levels of IGF-1 (ng/ml) and IL-6 (pg/ml) were measured by ELISA. SP-HSC in blood and bone marrow decreased with age but the quality of the surviving stem cells increased. MSC decreased non-significantly. IGF-1 levels (mean=30.7, SEM=2) decreased and IL-6 levels (mean=4.4, SEM=1) increased with age as did marrow fat (mean=1.2mmfat/g, SEM=0.04). There were no significant correlations between cytokine levels or fat and SP-HSC numbers. Stem cells appear to be progressively lost with aging and only the highest quality stem cells survive.

Fig. 1

SP stem cells/100,000 bone marrow mononuclear cells (MNC), analyzed by flow cytometry, are plotted against age cohorts.

Fig. 2

SP stem cells/100,000 peripheral blood mononuclear cells (MNC), analyzed by flow cytometry, are plotted against age cohorts.

Fig. 3

The ratio of lower side population (LSP) to upper side population (USP), which is a measure of stem cells quality (see Robinson et al., 2005), is plotted against age cohorts.

Fig. 4

Fibroblast colony forming cells (FCFC) in bone marrow/5 × 10⁵ MNC plated, representing clonal mesenchymal stromal cell precursors, are plotted against age cohorts.

Fig. 5

The quantity of fat (mm/g specimen weight) in bone marrow samples is plotted versus age cohorts.

Fig. 6

The levels (ng/ml) of plasma insulin-like growth factor 1 (IGF-1) as determined by ELISA plotted versus age cohorts.

Fig. 7

The levels (pg/ml) of plasma interleukin 6 (IL-6) levels as determined by ELISA assay plotted against age cohorts.