Background and objectives Multipotential stromal cells (MSCs) have considerable regenerative capabilities and their premature senescence in vivo may be responsible for altered bone phenotypes in age-related skeletal diseases including osteoporosis and osteoarthritis. Following minimal culture expansion, MSCs lose their native molecular phenotype (Churchman, Arthritis Rheum, 2012). However it is not known whether this continues to change as MSC reach senescence and whether any molecular markers could be predictive of senescence-related loss of MSC osteogenic capacity.
Materials and methods Bone marrow (BM, n = 7) was obtained from a deliberately diverse age range (2–72 years) of healthy donors. MSCs were selected for by plastic adherence and culture expanded up to senescence. Throughout the expansion period samples were taken for RNA and gDNA extraction as well as osteogenic differentiation. Gene expression profiles were studied by 96-gene Taqman low density array, whilst epigenetic senescence signature analysis was performed on the gDNA.
Results Cultures from older donors reached senescence considerably earlier compared to younger donors (r = -0.703). For all donors combined, MSC osteogenic capacity correlated better with population doublings (PDs) before senescence than with accrued PDs. Regardless of the donor age, the majority (59/78) of the genes were unchanged throughout the expansion period, with exceptions including TNFRSF11B/osteoprotegrin, CDH11/osteogenic cadherin, COL1A1/collagen type I and SPARC/osteonectin (all higher by senescence compared to early passage), and NANOG (lower in senescent cultures). The methylation status of cells correlated equally well with both PD and PD before senescence. Following osteogenic differentiation many genes were upregulated up to 200-fold and TNFRSF11B was less upregulated closer to senescence.
Conclusion Compared to methylation markers, the expression of TNFRSF11B/osteoprotegrin may be a better predictor of BM MSC senescence and related loss of osteogenic capability independent of donor age.
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