The identity of the cells that form the periosteum during development is controversial with current dogma suggesting these are derived from a Sox9-positive progenitor. Herein, we characterize a newly created Prrx1eGFP reporter transgenic mouse line during limb formation and postnatally. Interestingly, in the embryo Prrx1eGFP-labeled cells become restricted around the Sox9-positive cartilage anlage without themselves becoming Sox9-positive. In the adult, the Prrx1eGFP transgene live labels a subpopulation of cells within the periosteum that are enriched at specific sites, and this population is diminished in aged mice. The green fluorescent protein (GFP)-labeled subpopulation can be isolated using fluorescence-activated cell sorting (FACS) and represents approximately 8% of all isolated periosteal cells. The GFP-labeled subpopulation is significantly more osteogenic than unlabeled, GFP-negative periosteal cells. In addition, the osteogenic and chondrogenic capacity of periosteal cells in vitro can be extended with the addition of fibroblast growth factor (FGF) to the expansion media. We provide evidence to suggest that osteoblasts contributing to cortical bone formation in the embryo originate from Prrx1eGFP-positive cells within the perichondrium, which possibly piggyback on invading vascular cells and secrete new bone matrix. In summary, the Prrx1eGFP mouse is a powerful tool to visualize and isolate periosteal cells and to quantify their properties in the embryo and adult.

The presence of estrogen receptor ␣ (ER␣) in osteocytes was identified immunocytochemically in transverse sections from 560 to 860 m distal to the midshaft of normal neonatal and adult male and female rat ulnas (n ‫؍‬ 3 of each) and from adult male rat ulnas that had been exposed to 10 days of in vivo daily 10-minute periods of cyclic loading producing peak strains of either ؊3000 (n ‫؍‬ 3) or ؊4000 microstrain (n ‫؍‬ 5). Each animal ambulated normally between loading periods, and its contralateral ulna was used as a control. In animals in which limbs were subject to normal locomotor loading alone, 14 ؎1.2% SEM of all osteocytes in each bone section were ER␣ positive. There was no influence of either gender (p ‫؍‬ 0.725) or age (p ‫؍‬ 0.577) and no interaction between them (p ‫؍‬ 0.658). In bones in which normal locomotion was supplemented by short periods of artificial loading, fewer osteocytes expressed ER␣ (7.5 ؎ 0.91% SEM) than in contralateral control limbs, which received locomotor loading alone (14 ؎ 1.68% SEM; p ‫؍‬ 0.01; median difference, 6.43; 95% CI, 2.60, 10.25). The distribution of osteocytes expressing ER␣ was uniform across all sections and thus did not reflect local peak strain magnitude. This suggests that osteocytes respond to strain as a population, rather than as individual strain-responsive cells. These data are consistent with the hypothesis that ER␣ is involved in bone cells' responses to mechanical strain. High strains appear to decrease ER␣ expression. In osteoporotic bone, the high strains assumed to accompany postmenopausal bone loss may reduce ER␣ levels and therefore impair the capacity for appropriate adaptive remodeling. (J Bone Miner Res 2002;17:1646 -1655)

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