Extended passaging, but not aldehyde dehydrogenase activity, increases the chondrogenic potential of human adipose-derived adult stem cells

Adipose-derived adult stem (ADAS) cells represent an abundant population of multipotent mesodermal cells residing in various adipose tissue depots. ADAS cell preparations appear heterogeneous, yet at a clonal level, greater than 50% of these cells exhibit multilineage differentiation potential. To date, there have been few attempts to define prospectively a homogenous population of multipotent cells. In this study, we investigated whether aldehyde dehydrogenase (ALDH) can be used to enrich ADAS cells with increased chondrogenic potential. ALDH has been previously used to isolate primitive hematopoietic progenitors and has been implicated in early neurogenesis. Human ADAS cells were purified based on ALDH activity, and the cells were expanded and induced for chondrogenic differentiation using BMP-6 in a 3-D alginate culture. No significant differences in chondrogenic potential were observed in the ALDH-positive cells compared to unsorted controls. In contrast, significant differences were noted between cells assayed at passage 4 (P4) and cells assayed at passage 9 (P9). Following BMP-6 induction, AGC1 gene expression in P9 cells increased 290-fold over P4 cells. Similarly, COL2A1 expression in P9 cells increased fivefold compared to P4 cells, while COL10A1 levels remained unchanged. Immunohistochemical analysis over 28 days revealed consistent findings at the protein level for collagen II, collagen X, and aggrecan. No changes in telomerase activity were detected across passage, suggesting that ADAS cells retain some level of "stemness" in monolayer culture. These findings suggest that the chondrogenic potential of ADAS cells increases with passage number, although ALDH may not be a suitable marker for chondrogenesis.