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

Abstract

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 [3H]androstenedione in vitro. Aromatization activity was estimated either 1) by determining the incorporation of tritium from [1-3H]androstenedione into [3H]water or else 2) by determining the formation of [3H]estrone (E1) and [3H]estradiol (E2) from [1,2,6,7-3H]androstenedione. 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 [3H]E2 was slower initially but increased with time, and after 48 h of incubation, the amount of [3H]E2 produced exceeded that of [3H]E1. The rate of [3H]E1 formation, as a function of [3H]androstenedione 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 [3H]androstenedione 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.

Melatonin inhibits aromatase promoter expression by regulating cyclooxygenases expression and activity in breast cancer cells

“…Ovaries are the main sites of oestrogen synthesis in premenopausal non-pregnant women. However, after menopause, local synthesis of oestrogens in some tissues, including mammary tissue, acquires a special importance in mammary carcinogenesis (Ackerman et al, 1981;Pasqualini, 2004;Pasqualini and Chetrite, 2005;Suzuki et al, 2005). The local synthesis of oestrogens depends on the activity of different enzyme families (aromatase, sulfatases, etc) to transform androgens into oestrogens, as well as compounds of weak oestrogenic activity into more active forms.…”

Inhibitory effects of pharmacological doses of melatonin on aromatase activity and expression in rat glioma cells

“…Aromatase activity in C6 glioma cells was measured by the tritiated water release assay, based on the formation of tritiated water during aromatisation of an androgenic substrate such as [1b-3 H(N)]-androst-4-ene-3,17-dione (Ackerman et al, 1981). Glioma cells were seeded onto 60 Â 15 mm tissue culture dishes (2 Â 10 6 cells per dish) in DMEM/HAM F12 mixture supplemented with 10% FBS, 20 U ml À1 penicillin and 20 mg ml À1 streptomycin.…”

The effect of obesity on the ratio of type 3 17β-hydroxysteroid dehydrogenase mRNA to cytochrome P450 aromatase mRNA in subcutaneous abdominal and intra-abdominal adipose tissue of women

“…3 These conversions, catalysed by 17b-hydroxysteroid dehydrogenase (17b-HSD) and cytochrome P450 arom (aromatase) respectively, have been demonstrated to occur in adipose tissue. 4,5 In adipose tissue, aromatase activity and mRNA is localised predominantly in preadipocytes. 5,6 Killinger et al 7 reported that there are regional differences in preadipocyte sex steroid production.…”

“…4,5 In adipose tissue, aromatase activity and mRNA is localised predominantly in preadipocytes. 5,6 Killinger et al 7 reported that there are regional differences in preadipocyte sex steroid production. In cultured preadipocytes derived from human subcutaneous abdominal and omental adipose tissue, the formation of androgens from androstenedione was 10-fold greater than the formation of estrone, whereas in adipose tissue from the buttock and thigh, the production of androgens and estrone from androstenedione was equivalent.…”