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Steroid synthesis in vitro by a hilar cell tumor
Steroid synthesis in vitro by a hilar cell tumor S.
L.
JEFFCOATE,
F.
T.
G.
London,
M.B.
PRUNTY,
M.D.,
F.
R.
C.
P.
England
A 57-year-old zuoman with a 5 year history of progressive hirsutism and virilization was found at operation to have a hilar cell tumor in one ovary. Slices of the tumor were incubated separately in vitro with JH-pregnenolone and ‘JC-progesterone. A high (7.5 and 7.2, respectively) percentage conuersion of these precursors to testosterone was established. Labeled androstenedione, dehydroepiandrosterone, epitestosterone, and estrone were also found. The As pathway to androstenedione could well be dominant. Evidence for ll,B-hydroxylation was obtained.
T u M o R s o F the ovarian hilar cells, also known as Leydig cell tumors, are rare. Only 50 cases were collected in a recent review of the world 1iterature.l The difficulties in the differentiation of these tumors from those of luteum cell origin have been discussed elsewhere.? They are usually small, benign, occur about the age of the menopause, and cause virilization. Little is known about the steroid metabolism of such tumors. Case
total hysterectomy and bilateral oophorectomy were performed. Following operation the patient experienced menopausal symptoms which were controlled with estrogen therapy. The hirsutism ceased to progress but the patient still (5 years later) shaves daily. One month after operation the 17-oxosteroid excretion was 11 to 13 mg. per day. Pathologic findings. The left ovary was atrophic and had a small Brenner tumor in it. The right ovary similarly was atrophic and the tumor in it was a hilar cell tumor. Crystalloids of Reinke were prominent (Fig. 1) .
report
Mrs. E. A., aged 57, gave a 5 year history of progressive hirsutism and loss of scalp hair. The menopause had occurred at the age of 50 and there had been no vaginal bleeding since then. Examination showed her to have generalized and severe hirsutism; she was also obese (78.6 kilograms). Blood pressure, serum electrolytes and urea, and hemoglobin levels were normal. Resting steroid excretions were : 17-oxosteroids, 13.1 and 12.0 mg. per day; 17-oxogenic steroids, 10.4 and 8.1 mg. per day. These both rose to 48 mg. Per day after ACTH (20 I.U., twice daily). Gynecologic examination revealed an enlarged clitoris
(4 cm.)
and
enlarged
uterus;
the
Methods
The 17-oxosteroids and 17-oxogenic steroids were measured by standard methods.4s ’ Three in vitro incubations were carried out. Slices (0.5 mm. thick) of “normal” tissue (2.4 grams) from the right ovary were incubated with ‘%-progesterone (5.5 x 10G d.p.m.) and slices of the hilar cell tumor were incubated separately (2.3 grams each) with 14C-progesterone (5.5 x IO6 d.p.m.) and 3H-pregnenolone (44 x lo6 d.p.m. ) . The incubations were performed in 50 ml. of Krebs-Ringer bicarbonate buffer with added glucose, in an atmosphere of O,/CO, (95/5) for 2 hours at 37’ C. with gentle shaking. No cofactors were added since this might distort the biosynthetic activity of the tissue. At the end of the incubation the tissue and incubation medium were separated by filtra-
ovaries
were normal to palpation. At laparotomy (Professor P. Rhodes) the right ovary was found to contain a small (3.8 cm.) yellowish tumor. A
From the Department of Metabolic Diseases, St. Thomas’s Hospital, and Department of Chemical Pathology, St. Thomas’s Hospital Medical School.
684
In vitro
Fig. 1. Hilar cell tumor of the ovary. Crystalloids
tion and appropriate carrier steroids (200 pg ainounts) added. Full details of the incubation technique and the purification of the la.beled products (except in the case of llhydroxyandrostenedione) by chromatography, derivative formation, and recrystallization (after addition of more carrier) are described elsewhere.” The general outline is as follows. Neutral fraction. Four regions were eluted from an overnight Savard (ligroin : propylene glycol) paper chromatogram and further purified. The numbers of the subsequent stages correspond to those in Tables I to III. Androstenedione. Acetylation, thin-layer chromatogram (TLC) ( 1) ; Bush A paper chromatogram (2) ; reduction, Bush A (3) ; TLC (4) ; acetylation, TLC (5) ; recrystallization as testosterone acetate. Delzydroepiandrosterone. TLC (1) ; Bush A (2 ) ; acetylation, TLC (3). I I -Hydroxyandrostenedione. TLC (chloroform : ethyl acetate, 1: 1) ( 1) ; Bush A (2) ; oxidation with 0.4 per cent potassium dichromate in acetic acid at 20° for 5 minutes, TLC (chloroform : ethyl acetate, 2 : 1) (3) ;
steroid
synthesis
by hilar
of Reinke are prominent.
cell
tumor
685
(x240.)
; recrystallization as adrenoBush A (4) sterone. Testosterone, epitestosterone, 17-1lyd1-oxyprogesterone. TLC (separates 1 ‘I-hydroxyprogesterone), Savard (separates testostrrone and epitestosterone which were analyzed separately from this stage) ( 1) ; Bush A (2) ; acetylation, TLC (3) ; recrystallization as testosterone acetate or epitestosterone acetate. Phenolic fraction. Estrone and estradiol were separated on a Bush B, chromatogram. Specific activities were measured after the following stages: Estyadiol. Glass-fiber paper chromatogram, methylation (1) ; TLC (2) ; acetylation, ‘TLC (3)
’
Estrone. Glass fiber paper chromatogram, methylation (1) ; TLC (2) ; reduction, TLC (3) ; acetylation, TLC (4) ; recrystallization as estradiol-3-methyl ether, 1 ‘I-acetate. Results
Testosterone was the major product of the in vitro incubation of slices of the hilar cell tumor with both pregnenolone and pro-
686
Jeffcoate
and
Prunty
Table I. Specific activities of testosterone and epitestosterone in vitro of slices of the “normal” ovarian tissue and a hilar Specific Normal
activity
Hilar
cell
tumor
Progesterone
Epitestosterone
Testosterone
I 2 3 Recrystallization Before Crystals Mother liquors Per cent conversion of substrates
(d.p.m./wg)
tissue
Progesterone Stage
from incubation cell tumor
32.8 30.5 29.4
Testosterone
8.3 4.4 3.9
0.180 0.173 0.179
-
0.11
co.015
Pregnenolone
Epitestosterone
2,400 2,160 2,270
Testosterone
65.1 9.4 6.7
18.9 la.1 16.9
18,500 17,400 17,100
-.
7.2
<0.02
Epitestosterone
1
653 44 43
89.0 91.8 85.1
0.320 0.302 0.313
7.5
0.024
Table II. Specific activities of androstenedione, 11 -hydroxyandrostenedione, and dehydroepiandrosterone from incubation in vitro of slices of “normal” ovarian tissue and a hilar cell tumor Specific Normal
activity
tissue
Hilar
: 3 4 5 Recrystallization Before Crystals Mother liquors Per
cent conversion of substrates
Table III. Specific activities of slices of a hilar cell tumor
11 -Hydroxyandrostenedione
Androstenedione
cell
144 123 119 111 113
Androstenedione
94.8 5.7 3.2 0.3
Pregnenolone
11 -Hydroxyandrostenedione
11,320 .oao 929 888 887
Androstenedione
166 16.3 10.3 9.5
11,500 13,700 12,300 12,400
-
-
5.10 5.59 5.74
0.404 0.390 0.94
0.37
2.7
0.03
5.2
incubations
in vitro
of estrone
and estradiol
from
Specific
activity
1 2 3 4 Recrystallization Before Cxystals Mother liquors Per cent conversion of substrates
Dehydroepiandrosterone 8,720 9,070 9,200 4.1
(d.p.m./bg)
Proeesterone Stage
tumor
Progesterone
Progesterone
Stage
(d.p.m./pg)
Preenenolone
Estrone
Esstradiol
Estrone
Estradiol
16.2 a.3 a.4
20.7 8.2 5.1 -
1,120 76.5 56.7 56.1
193 61.8 13.8
-
--
0.35 0.32 0.43
<0.02
0.03
[0.03]
co.01
In vitro
gesterone. The percentage yield was 7.5 and 7.2. respectively (Table I), and was only 0.1 1 in the adjacent “normal” tissue. With pregnenolone as a substrate there was a low (0.024 per cent) conversion to epitestosterone by the tumor tissue. Conversion to androstenedione was also big-h (Table II) ~ and pregnenolone appeared to be a better precursor than progesterone. There was a high conversion also of pregnenolone to dehydroepiandrosterone. The systhesis from progesterone of 11 -hydroxyandrostenedione, albeit in only 0.03 per cent yield. is shown by the constancy of specific activity of the product but there was no evidence for its synthesis in the normal tissue. This fraction was not investigated in the pregncnolone incubation. Aromatization to estrone but not estradiol from pre:nenolone was also demonstrated in the tumor (Table III), but with progesterone as a substrate aromatization was not definitel>. established. Comment The steroid metabolism of hilar cell tumors has received little attention in the past and there is only one other report of an in vitro study on such a tumor.6 Reinke’s crystalloids, considered by some to be an important featllre distinguishing tumors of hilar cell cells from those arising from lutein cells or adrenal cell rests,’ were absent in the cace of Corral-Gallardo and associate9 but present in our case (Fig. 1). With minces of tissur and an excess of cofactors, CorralGallardo and associate@ demonstrated in vitro a 5.6 per cent conversion of progesterone to androstenedione and 0.3 per cent to testosterone. The testosterone production rate was 6.7 mg. per day, well above the normal female range. The in vitro conversion to testosterone from both pregnenolone and progesterone was very high in the present case with sliced tissue and no colartors. Although nothing is known of the function of normal hilar cells, these studies suggest that they are at least a potential source of ovarian testosterone.
steroid
synthesis
by
hilar
cell
tumor
687
Within the limitations of the technique used, since there was a high conversion ot’ pregnenolone to dehydroepiandrosterone and to androstenedione, the pathway pregnrnolone + 17-hydroxypregnenolone -+ tlehydroepiandrosterone --+ androstenedioncs could be dominant in this tumor. Thus. it is of interest that this has been found to be the case in normal theta tissue, as opposed to granulosa tissue,s and in one in vitro stlld) of an arrhenoblastoma.!’ The finding of a small conversion of progesterone to 1 lp-hydroxyandrostenedione is of great interest: this was about 1 per c,ent of the conversion to androstenedione in the same incubation. There are two nthrr rcports of 1 I-hydroxylation occ-urring in ovarian tumors. One was a “gynandroblastoma,“‘” the other the hilus cell tumor of Corral-Gallardo and associates,” which, under their conditions, converted androstenedione to 1 l/3-hydroxyandrostenedione in a 1 l)cr cent yield. In the latter, with prqqestrrone as a substrate, however, no 1 I-hydroxyl~tion could be demonstrated. It has not been demonstrated that 11 -hydroxylation occurs in the normal ovary and it could not he fwnd in an arrhenoblastoma’l or a masculitlo\,oIt has, however, likewise been blastoma.‘? demonstrated in testicular tumors.‘.‘. ’ hut its significance proves doubtful in attemptill: to distinguish tumors of adrenal rest origin from those of testicular cell origin. It ieelns also that observations of 1 1-hydroxylat~on in tumors of ovarian origin cast doubt on the validity of this criterion in distinguishing these from adrenal rests. In addition, the tumor converted pregnenolone to epitestosterone in a IOM. yield, in fact lower than has heen foluld in incubations under the same conditions of slices of both normal and polycystic ovaries.,’ -4s well as evidence for the high r;ttv of androgen synthesis, there was some fl.br the synthesis of estrogens. Thus it is intel,esting to note that although hilar cell turncrrs arr usually virilizing, one estrogenic‘ tulnllr has been rep0rted.l”
688
Jeffcoate
and
Prunty
REFERENCES
1. 2. 3.
4. 5.
6.
7.
8.
Dunnihoo, D. R., Grieme, D. L., and Woolf, R. B.: Obst. & Gynoc. 27: 703, 1966. Pruntv. F. T. G.: T. Endocrinol: 38: 85. 1967. Jeffcoate, S. L., Biooks, R. V., London,‘D. R., Prunty, F. T. G., and Rhodes, P.: J. Endocrinol. Submitted. Gibson, G., and Norymberski, J. K.: Ann. Rheum. Dis. 13: 59, 1954. Appleby, J. I., Gibson, G., Norymberski, J. K., and Stubbs, R. D.: Biochem. J. 60: 453, 1955. Corral-Gallardo, J., Acevedo, H. A., de Salazar, J. L. P., Loria, M., and Goldzieher, J. W.: Acta endocrinol. 52: 425, 1966. Scully, R. E.: In Grady, H. G., and Smith, D. E., editors. The Ovary, Baltimore, 1963, The Williams & Wilkins Comnanv. D. 143. Ryan, K. J., and Petro, Z.: J. Chin. Endocrin01. 26: 46, 1966.
9.
10.
11.
12.
13.
14.
15.
Kase, N., and Conrad, S. H.: AM. J. ORST. 8 GYNEC. 90: 1251, 1964. Nocke, W.: In Proc. II International Congress of Endocrirolcgy, Amsterdam, 1965, Excerpta Medica Foundation, p. 1304. Savard, K.. Gut, M., Dorfman, R. I., Gabrilove, J. L., and Soffer, L. V.: J. Clin. Endocrinol. 21: 165, 1961. Bryson, M. J., Dominguez, 0. V., Kaiser, I. H, Samuels, L. T., and Sweat, M. L.:. J. Clin. Endocrinol. 22: 773. 1962. Prunty, F. T. G.: Chemistry and Treatment of Adrenocortical Diseases, Springfield, Illinois, 1964, Charles C Thomas, Publisher, p. 241. Engel, L. L., Lanman, G., Scully, R. E., and Villee, D. B.: J. Clin. Endocrinol. 26: 381, 1966. Plate, W. P.: Acta endocrinol. 26: 489, 1957.
L.
JEFFCOATE,
F.
T.
G.
London,
M.B.
PRUNTY,
M.D.,
F.
R.
C.
P.
England
A 57-year-old zuoman with a 5 year history of progressive hirsutism and virilization was found at operation to have a hilar cell tumor in one ovary. Slices of the tumor were incubated separately in vitro with JH-pregnenolone and ‘JC-progesterone. A high (7.5 and 7.2, respectively) percentage conuersion of these precursors to testosterone was established. Labeled androstenedione, dehydroepiandrosterone, epitestosterone, and estrone were also found. The As pathway to androstenedione could well be dominant. Evidence for ll,B-hydroxylation was obtained.
T u M o R s o F the ovarian hilar cells, also known as Leydig cell tumors, are rare. Only 50 cases were collected in a recent review of the world 1iterature.l The difficulties in the differentiation of these tumors from those of luteum cell origin have been discussed elsewhere.? They are usually small, benign, occur about the age of the menopause, and cause virilization. Little is known about the steroid metabolism of such tumors. Case
total hysterectomy and bilateral oophorectomy were performed. Following operation the patient experienced menopausal symptoms which were controlled with estrogen therapy. The hirsutism ceased to progress but the patient still (5 years later) shaves daily. One month after operation the 17-oxosteroid excretion was 11 to 13 mg. per day. Pathologic findings. The left ovary was atrophic and had a small Brenner tumor in it. The right ovary similarly was atrophic and the tumor in it was a hilar cell tumor. Crystalloids of Reinke were prominent (Fig. 1) .
report
Mrs. E. A., aged 57, gave a 5 year history of progressive hirsutism and loss of scalp hair. The menopause had occurred at the age of 50 and there had been no vaginal bleeding since then. Examination showed her to have generalized and severe hirsutism; she was also obese (78.6 kilograms). Blood pressure, serum electrolytes and urea, and hemoglobin levels were normal. Resting steroid excretions were : 17-oxosteroids, 13.1 and 12.0 mg. per day; 17-oxogenic steroids, 10.4 and 8.1 mg. per day. These both rose to 48 mg. Per day after ACTH (20 I.U., twice daily). Gynecologic examination revealed an enlarged clitoris
(4 cm.)
and
enlarged
uterus;
the
Methods
The 17-oxosteroids and 17-oxogenic steroids were measured by standard methods.4s ’ Three in vitro incubations were carried out. Slices (0.5 mm. thick) of “normal” tissue (2.4 grams) from the right ovary were incubated with ‘%-progesterone (5.5 x 10G d.p.m.) and slices of the hilar cell tumor were incubated separately (2.3 grams each) with 14C-progesterone (5.5 x IO6 d.p.m.) and 3H-pregnenolone (44 x lo6 d.p.m. ) . The incubations were performed in 50 ml. of Krebs-Ringer bicarbonate buffer with added glucose, in an atmosphere of O,/CO, (95/5) for 2 hours at 37’ C. with gentle shaking. No cofactors were added since this might distort the biosynthetic activity of the tissue. At the end of the incubation the tissue and incubation medium were separated by filtra-
ovaries
were normal to palpation. At laparotomy (Professor P. Rhodes) the right ovary was found to contain a small (3.8 cm.) yellowish tumor. A
From the Department of Metabolic Diseases, St. Thomas’s Hospital, and Department of Chemical Pathology, St. Thomas’s Hospital Medical School.
684
In vitro
Fig. 1. Hilar cell tumor of the ovary. Crystalloids
tion and appropriate carrier steroids (200 pg ainounts) added. Full details of the incubation technique and the purification of the la.beled products (except in the case of llhydroxyandrostenedione) by chromatography, derivative formation, and recrystallization (after addition of more carrier) are described elsewhere.” The general outline is as follows. Neutral fraction. Four regions were eluted from an overnight Savard (ligroin : propylene glycol) paper chromatogram and further purified. The numbers of the subsequent stages correspond to those in Tables I to III. Androstenedione. Acetylation, thin-layer chromatogram (TLC) ( 1) ; Bush A paper chromatogram (2) ; reduction, Bush A (3) ; TLC (4) ; acetylation, TLC (5) ; recrystallization as testosterone acetate. Delzydroepiandrosterone. TLC (1) ; Bush A (2 ) ; acetylation, TLC (3). I I -Hydroxyandrostenedione. TLC (chloroform : ethyl acetate, 1: 1) ( 1) ; Bush A (2) ; oxidation with 0.4 per cent potassium dichromate in acetic acid at 20° for 5 minutes, TLC (chloroform : ethyl acetate, 2 : 1) (3) ;
steroid
synthesis
by hilar
of Reinke are prominent.
cell
tumor
685
(x240.)
; recrystallization as adrenoBush A (4) sterone. Testosterone, epitestosterone, 17-1lyd1-oxyprogesterone. TLC (separates 1 ‘I-hydroxyprogesterone), Savard (separates testostrrone and epitestosterone which were analyzed separately from this stage) ( 1) ; Bush A (2) ; acetylation, TLC (3) ; recrystallization as testosterone acetate or epitestosterone acetate. Phenolic fraction. Estrone and estradiol were separated on a Bush B, chromatogram. Specific activities were measured after the following stages: Estyadiol. Glass-fiber paper chromatogram, methylation (1) ; TLC (2) ; acetylation, ‘TLC (3)
’
Estrone. Glass fiber paper chromatogram, methylation (1) ; TLC (2) ; reduction, TLC (3) ; acetylation, TLC (4) ; recrystallization as estradiol-3-methyl ether, 1 ‘I-acetate. Results
Testosterone was the major product of the in vitro incubation of slices of the hilar cell tumor with both pregnenolone and pro-
686
Jeffcoate
and
Prunty
Table I. Specific activities of testosterone and epitestosterone in vitro of slices of the “normal” ovarian tissue and a hilar Specific Normal
activity
Hilar
cell
tumor
Progesterone
Epitestosterone
Testosterone
I 2 3 Recrystallization Before Crystals Mother liquors Per cent conversion of substrates
(d.p.m./wg)
tissue
Progesterone Stage
from incubation cell tumor
32.8 30.5 29.4
Testosterone
8.3 4.4 3.9
0.180 0.173 0.179
-
0.11
co.015
Pregnenolone
Epitestosterone
2,400 2,160 2,270
Testosterone
65.1 9.4 6.7
18.9 la.1 16.9
18,500 17,400 17,100
-.
7.2
<0.02
Epitestosterone
1
653 44 43
89.0 91.8 85.1
0.320 0.302 0.313
7.5
0.024
Table II. Specific activities of androstenedione, 11 -hydroxyandrostenedione, and dehydroepiandrosterone from incubation in vitro of slices of “normal” ovarian tissue and a hilar cell tumor Specific Normal
activity
tissue
Hilar
: 3 4 5 Recrystallization Before Crystals Mother liquors Per
cent conversion of substrates
Table III. Specific activities of slices of a hilar cell tumor
11 -Hydroxyandrostenedione
Androstenedione
cell
144 123 119 111 113
Androstenedione
94.8 5.7 3.2 0.3
Pregnenolone
11 -Hydroxyandrostenedione
11,320 .oao 929 888 887
Androstenedione
166 16.3 10.3 9.5
11,500 13,700 12,300 12,400
-
-
5.10 5.59 5.74
0.404 0.390 0.94
0.37
2.7
0.03
5.2
incubations
in vitro
of estrone
and estradiol
from
Specific
activity
1 2 3 4 Recrystallization Before Cxystals Mother liquors Per cent conversion of substrates
Dehydroepiandrosterone 8,720 9,070 9,200 4.1
(d.p.m./bg)
Proeesterone Stage
tumor
Progesterone
Progesterone
Stage
(d.p.m./pg)
Preenenolone
Estrone
Esstradiol
Estrone
Estradiol
16.2 a.3 a.4
20.7 8.2 5.1 -
1,120 76.5 56.7 56.1
193 61.8 13.8
-
--
0.35 0.32 0.43
<0.02
0.03
[0.03]
co.01
In vitro
gesterone. The percentage yield was 7.5 and 7.2. respectively (Table I), and was only 0.1 1 in the adjacent “normal” tissue. With pregnenolone as a substrate there was a low (0.024 per cent) conversion to epitestosterone by the tumor tissue. Conversion to androstenedione was also big-h (Table II) ~ and pregnenolone appeared to be a better precursor than progesterone. There was a high conversion also of pregnenolone to dehydroepiandrosterone. The systhesis from progesterone of 11 -hydroxyandrostenedione, albeit in only 0.03 per cent yield. is shown by the constancy of specific activity of the product but there was no evidence for its synthesis in the normal tissue. This fraction was not investigated in the pregncnolone incubation. Aromatization to estrone but not estradiol from pre:nenolone was also demonstrated in the tumor (Table III), but with progesterone as a substrate aromatization was not definitel>. established. Comment The steroid metabolism of hilar cell tumors has received little attention in the past and there is only one other report of an in vitro study on such a tumor.6 Reinke’s crystalloids, considered by some to be an important featllre distinguishing tumors of hilar cell cells from those arising from lutein cells or adrenal cell rests,’ were absent in the cace of Corral-Gallardo and associate9 but present in our case (Fig. 1). With minces of tissur and an excess of cofactors, CorralGallardo and associate@ demonstrated in vitro a 5.6 per cent conversion of progesterone to androstenedione and 0.3 per cent to testosterone. The testosterone production rate was 6.7 mg. per day, well above the normal female range. The in vitro conversion to testosterone from both pregnenolone and progesterone was very high in the present case with sliced tissue and no colartors. Although nothing is known of the function of normal hilar cells, these studies suggest that they are at least a potential source of ovarian testosterone.
steroid
synthesis
by
hilar
cell
tumor
687
Within the limitations of the technique used, since there was a high conversion ot’ pregnenolone to dehydroepiandrosterone and to androstenedione, the pathway pregnrnolone + 17-hydroxypregnenolone -+ tlehydroepiandrosterone --+ androstenedioncs could be dominant in this tumor. Thus. it is of interest that this has been found to be the case in normal theta tissue, as opposed to granulosa tissue,s and in one in vitro stlld) of an arrhenoblastoma.!’ The finding of a small conversion of progesterone to 1 lp-hydroxyandrostenedione is of great interest: this was about 1 per c,ent of the conversion to androstenedione in the same incubation. There are two nthrr rcports of 1 I-hydroxylation occ-urring in ovarian tumors. One was a “gynandroblastoma,“‘” the other the hilus cell tumor of Corral-Gallardo and associates,” which, under their conditions, converted androstenedione to 1 l/3-hydroxyandrostenedione in a 1 l)cr cent yield. In the latter, with prqqestrrone as a substrate, however, no 1 I-hydroxyl~tion could be demonstrated. It has not been demonstrated that 11 -hydroxylation occurs in the normal ovary and it could not he fwnd in an arrhenoblastoma’l or a masculitlo\,oIt has, however, likewise been blastoma.‘? demonstrated in testicular tumors.‘.‘. ’ hut its significance proves doubtful in attemptill: to distinguish tumors of adrenal rest origin from those of testicular cell origin. It ieelns also that observations of 1 1-hydroxylat~on in tumors of ovarian origin cast doubt on the validity of this criterion in distinguishing these from adrenal rests. In addition, the tumor converted pregnenolone to epitestosterone in a IOM. yield, in fact lower than has heen foluld in incubations under the same conditions of slices of both normal and polycystic ovaries.,’ -4s well as evidence for the high r;ttv of androgen synthesis, there was some fl.br the synthesis of estrogens. Thus it is intel,esting to note that although hilar cell turncrrs arr usually virilizing, one estrogenic‘ tulnllr has been rep0rted.l”
688
Jeffcoate
and
Prunty
REFERENCES
1. 2. 3.
4. 5.
6.
7.
8.
Dunnihoo, D. R., Grieme, D. L., and Woolf, R. B.: Obst. & Gynoc. 27: 703, 1966. Pruntv. F. T. G.: T. Endocrinol: 38: 85. 1967. Jeffcoate, S. L., Biooks, R. V., London,‘D. R., Prunty, F. T. G., and Rhodes, P.: J. Endocrinol. Submitted. Gibson, G., and Norymberski, J. K.: Ann. Rheum. Dis. 13: 59, 1954. Appleby, J. I., Gibson, G., Norymberski, J. K., and Stubbs, R. D.: Biochem. J. 60: 453, 1955. Corral-Gallardo, J., Acevedo, H. A., de Salazar, J. L. P., Loria, M., and Goldzieher, J. W.: Acta endocrinol. 52: 425, 1966. Scully, R. E.: In Grady, H. G., and Smith, D. E., editors. The Ovary, Baltimore, 1963, The Williams & Wilkins Comnanv. D. 143. Ryan, K. J., and Petro, Z.: J. Chin. Endocrin01. 26: 46, 1966.
9.
10.
11.
12.
13.
14.
15.
Kase, N., and Conrad, S. H.: AM. J. ORST. 8 GYNEC. 90: 1251, 1964. Nocke, W.: In Proc. II International Congress of Endocrirolcgy, Amsterdam, 1965, Excerpta Medica Foundation, p. 1304. Savard, K.. Gut, M., Dorfman, R. I., Gabrilove, J. L., and Soffer, L. V.: J. Clin. Endocrinol. 21: 165, 1961. Bryson, M. J., Dominguez, 0. V., Kaiser, I. H, Samuels, L. T., and Sweat, M. L.:. J. Clin. Endocrinol. 22: 773. 1962. Prunty, F. T. G.: Chemistry and Treatment of Adrenocortical Diseases, Springfield, Illinois, 1964, Charles C Thomas, Publisher, p. 241. Engel, L. L., Lanman, G., Scully, R. E., and Villee, D. B.: J. Clin. Endocrinol. 26: 381, 1966. Plate, W. P.: Acta endocrinol. 26: 489, 1957.