Impact of adipocytes on the maintenance of HPCs in vitro. Without frequent media changes and at high cell density, GZL, an established MSC cell line, differentiates preferentially into adipocytes (GZL/Adi). The number of adipocytes is greatly reduced when the cells are propagated under more favorable conditions (GZL). Both GZL and GZL/Adi were used along with primary SCs to sustain LSK cells for 7 days. (A) Cells were harvested on day 7, counted, and used in clonogenic assays. (B) SC, GZL, and GZL/Adi were assayed by quantitative RT-PCR for the expression of adiponectin and FABP4 as indicators of adipogenic differentiation. Data were normalized to primary SCs. (C) SCs, GZL, and GZL/Adi were separated from LSK progeny on day 7 by cell sorting and assayed by quantitative RT-PCR for the expression of neuropilin-1. Expression of Np1 was normalized to primary SCs. P < .05 between SC and GZL/Adi groups in panel A.