Gene expression profiling suggests a pathological role of human bone marrow-derived mesenchymal stem cells in aging-related skeletal diseases

Abstract

Aging is associated with bone loss and degenerative joint diseases, in which the aging of bone marrow-derived mesenchymal stem cell (bmMSC)[1] may play an important role. In this study, we analyzed the gene expression profiles of bmMSC from 14 donors between 36 and 74 years old, and obtained age-associated genes (in the background of osteoarthritis) and osteoarthritis-associated genes (in the background of old age). Pathway analysis of these genes suggests that alterations in glycobiology might play an important role in the aging of human bmMSC. On the other hand, antigen presentation and signaling of immune cells were the top pathways enriched by osteoarthritis-associated genes, suggesting that alteration in immunology of bmMSC might be involved in the pathogenesis of osteoarthritis. Most intriguingly, we found significant age-associated differential expression of HEXA, HEXB, CTSK, SULF1, ADAMTS5, SPP1, COL8A2, GPNMB, TNFAIP6, and RPL29; those genes have been implicated in the bone loss and the pathology of osteoporosis and osteoarthritis in aging. Collectively, our results suggest a pathological role of bmMSC in aging-related skeletal diseases, and suggest the possibility that alteration in the immunology of bmMSC might also play an important role in the etiology of adult-onset osteoarthritis.

Figure 1. Growth rate of human bone marrow-derived plastic-adherent cells.

(A) Plastic-adherent cells harvested separately from bone marrows of 14 donors were seeded at a density of 1.3 × 10³ cells/cm² (passage 1). Cells were counted after 3 days (left panel) and 6 days (right panel) post-seeding using a hemocytometer, and the fold increase in cell number was calculated. Donors are color-coded as shown in Table I. (B) Cells harvested 6 days post-seeding were subjected to flow cytometric analyses. A representative result is shown.

Figure 2. Representative plots of donor age versus normalized mRNA expression level for selective age-associated genes.

Each solid dot represented a bone marrow donor. The regression lines (solid line) and correlation coefficients (r) showed trend of change in gene expression with increasing donor age. Donors are color-coded as shown in Table I.

Figure 3. N-glycan degradation pathway enriched by pathway analysis.

Enzymes (in EC number) with their names of coding genes and the corresponding sites of actions in the modification of N-glycan are indicated. Genes (n=9) that are differentially expressed with age and enriched by IPA analysis are marked in red.

Figure 4. DNA synthesis measured by BrdU incorporation assays.

BmMSC from donors 1, 3, 10, 12, and 13 were seeded into 96-well culture plate (1.2 × 10² cells/well). Cells were either left untreated or treated with 4, 8, 12, and 16 mM Na₂SO₄ (Merck, Darmstadt, Germany) for 48 h. Then, either BrdU or PBS was added in medium, and cells were incubated for 24 h. Subsequently, cells were subjected to detection of the BrdU signals. The fold increase in DNA synthesis was calculated by comparing the BrdU signals of Na₂SO₄ treated cells to that of untreated cells (to which a value of 1 was assigned). The gender and age of donors are as indicated in the figure. Data represent the mean ± S.D. from three duplicate analyses. **, p<0.05; *, p=0.051 (comparison of individual aged sample to the average of adult samples, t-test).