Aromatization of Androstenedione by Human Adipose Tissue Stromal Cells in Monolayer Culture*

Stromal cells and adipocytes were separated after collagenase treatment of adipose tissue obtained from women undergoing elective surgery, and these cells were used to study aromatization of 3Handrostenedione in vitro. Aromatization activity was estimated either 1) by determining the incorporation of tritium from 1-3Handrostenedione into 3Hwater or else 2) by determining the formation of 3Hestrone (E1) and 3Hestradiol (E2) from 1,2,6,7-3Handrostenedione. It was established that only 13% of the aromatase activity of adipose tissue resided in the adipocyte fraction, whereas 87% of the aromatase activity was in the stromal/vascular fraction. Subsequent studies of aromatization were conducted utilizing stromal/vascular cells grown to confluence in monolayer culture. In such cells, the formation of 3HE2 was slower initially but increased with time, and after 48 h of incubation, the amount of 3HE2 produced exceeded that of 3HE1. The rate of 3HE1 formation, as a function of 3Handrostenedione concentration, followed Michaelis-Menten kinetics. The Vmax ranged from 0.8-3.0 pmol and ranged from 0.16-0.67 pmol mg-1 cell protein 6 h-1 in cells from omental adipose tissue. The apparent Km for 3Handrostenedione in stromal cells derived from both omental and sc tissue was the same, i.e. about 25 nM. We conclude that the ability of human adipose tissue to form estrogen is not a function primarily of the adipocytes but rather resides principally in the cells of the stroma.