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

Abstract The aim of this study was to examine the expression and regulation of type 1 17β-hydroxysteroid dehydrogenase (type 1 17-HSD) enzyme protein and mRNA, and 17-HSD activity in human granulosa cells. The cells were obtained from patients taking part in an in vitro fertilization programme. The cells from each patient were divided into two groups: cells obtained from preovulatory follicles (LGC=granulosa cells from large follicles ≥ 18 mm in diameter), and cells from other visible follicles (SGC=granulosa cells from small follicles, less than 15 mm in diameter). The identity of 17-HSD enzyme protein expressed in human granulosa cells with placental cytosolic 17-HSD (type 1 17-HSD) was assessed by immunoblot analysis using polyclonal antibodies, and the enzyme was immunolocalized in the cytoplasm of granulosa cells. Type 1 17-HSD protein concentration, 17-HSD and cytochrome P450 aromatase (P450arom) activities and oestradiol (OE2) production in cells from LGC were significantly lower than the corresponding values obtained in SGC in the same patient (paired t-test). The type 1 17-HSD protein concentration, 17-HSD activity and P450arom activity were 140±16% (mean±s.e.m.), 121±22% and 113±26% higher in cells from SGC, which was also reflected in a 70±12% higher OE2 production in these cells. In freshly isolated cells from LGC or SGC, a high correlation between 17-HSD and P450arom activities was observed (r=0·93, P<0·001). In long-term cultured cells, type 1 17-HSD was stably expressed at least until day 9, while P450arom expression decreased. In addition, treatments with gonadotrophins did not affect type 1 17-HSD protein concentration and 17-HSD activity. In contrast to this, both P450arom activity and OE2 production were significantly increased (P<0·05). The data, therefore, suggest that type 1 17-HSD and P450arom are expressed in parallel during the latest stages of follicular maturation but, in cultured granulosa-luteal cells, the enzymes are regulated by distinct mechanisms. Journal of Endocrinology (1994) 143, 139–150

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Effects of tri-iodothyronine on alternative splicing events in the coding region of cytochrome P450 aromatase in immature rat Sertoli cells

Journal of Endocrinology ◽

10.1677/joe.0.1700381 ◽

2001 ◽

Vol 170 (2) ◽

pp. 381-393 ◽

Cited By ~ 15

Author(s):

V Pezzi ◽

ML Panno ◽

R Sirianni ◽

P Forastieri ◽

I Casaburi ◽

...

Keyword(s):

Cytochrome P450 ◽

Enzymatic Activity ◽

Protein Content ◽

Sertoli Cells ◽

Male Rats ◽

Strong Increase ◽

Aromatase Activity ◽

P450 Aromatase ◽

Cytochrome P450 Aromatase ◽

Aromatase Mrna

Transient postnatal hypothyroidism in male rats induces a prolonged proliferation of immature Sertoli cells. This change in Sertoli cell replication at young ages is coincident with enhanced and prolonged aromatase activity that leads to a marked increase in the conversion of androgens into estrogens. Both events are drastically inhibited by tri-iodothyronine (T(3)) replacement either in vivo or in vitro. This study, after the immunolocalization of aromatase in cultured rat Sertoli cells, examined the effects elicited by T(3) on this enzyme, by simultaneously investigating three functional levels of aromatase: mRNA expression, protein content, and enzymatic activity. The immunolocalization of cytochrome P450 aromatase (P450 arom) was shown in the cytoplasm of cultured Sertoli cells from 15- and 21-day-old rats. Western blot analysis revealed an enhancement of aromatase protein content upon stimulation with N(6),2'-O-dibutyryladenosine-3':5'-cyclic monophosphate ((Bu)(2)cAMP) that was clearly down-regulated by T(3). The presence of a functional P450 arom protein in purified Sertoli cells was confirmed by the measurement of [(3)H]H(2)O released after incubation with [1 beta-(3)H]androst-4-ene-3,17-dione. With 100 nM T3, a decrease in both P450 arom mRNA levels and aromatase activity was observed. The aromatase enzymatic activity was strongly stimulated by (Bu)(2)cAMP and markedly down-regulated by T(3). In contrast, the strong increase in aromatase mRNA upon (Bu)(2)cAMP stimulation was apparently unaffected by T(3) administration. This paper shows how the identification of an altered transcript induced by T(3) coding for putative truncated and inactive aromatase protein might explain such a decrease in aromatase activity in T(3)-treated cells. On the basis of these results, it is concluded that at least two mechanisms could be involved in the down-regulatory effect of T(3) on aromatase activity in prepuberal Sertoli cells. The first mechanism is linked to a possible direct modulatory role for T(3) in the regulation of the aromatase promoter, whilst the second one is represented by the induction of altered transcripts coding for truncated and inactive aromatase proteins.

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