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Estrone sulfatase activity in human uterine leiomyoma
GYNECOLOGICONCOLOGY37, 315-318 (l!%l)
Estrone Sulfatase Activity in Human Uterine Leiomyoma TAKARA YAMAMOTO, M.D.,’ MAMORU URABE, M.D., KAZUO NAITOH, M.D., Jo KITAWAKI,
M.D., HIDEO HONJO,
M.D., AND HIROJI OKADA, M.D. Department
of Obstetrics
and Gynecology,
Kyoto Prefectural
University
of Medicine,
Kyoto 602, Japan
Received June 19, 1989
Human uterine leiomyoma is a benign tumor and its development is closely related to estrogen. In this study, e&one sulfatase (E$F) activity and concentrations of e&one (E,) and esbone sulfate (E,S) were measured in endometrial, leiomyoma, and myometrial tissues of the same human uterus (n = 11) with a leiomyoma. EISF activity in endometrial tissue overlying a leiomyoma was 2.62 f 0.29 nmole/hr/mg protein (mean f SD), this activity being significantly higher (P < 0.01) compared with that in normal endometrial tissue (2.0 f 0.24 mnole/hr/mg protein). EISF activity in normal endometrial tissue was significantly higher (P < 0.001) compared with that in leiomyoma tissue (0.49 * 0.82 nmole/hr/mg protein) or myometrial tissue (0.76 -C 0.10 nmole/hr/mg protein). We also found a significant difference (P < 0.05) in EISF activity between leiomyoma tissue and myometrial tissue. On the other hand, the E, concentration in endometrial tissue overlying a leiomyoma (10.9 f 8.9 pg/mg protein) proved to be higher than that in endometrial tissue overlying normal myometrium (1.23 + 1.97 pg/mg protein). E,S concentrations in these tissues, however, were 631 f 339 and 902 -C 482 pg/mg protein, respectively, these values showing a trend opposite that of the El concentration data. Thus, these results suggest that high EISF activity and high E, concentration in the endometrium overlying a leiomyoma may be related to estrogen supply to a uterine leiomyoma node. ~1 1990 Academic mess, Inc.
INTRODUCTION While uterine leiomyomas rarely appear in young females before the onset of menstruation, they frequently occur in sexually mature women with menstrual cycles; the frequency generally decreases after menopause. Thus, their appearance and development seem to be closely related to ovarian hormones (i.e., estrogen and progesterone) [l]. Uterine leiomyomas are benign tumors, though they have a tendency to be found in combination with other estrogen-dependent tumors (breast ’ To whom reprint requests should be addressed.
cancer, uterine endometrial cancer, and so on) [ll. Because of this relationship with estrogen, we examined estrogen biosynthetase (aromatase) activity, which converts androgens to estrogens, in leiomyoma and myometrial tissues of the human uterus [2]. Results indicated that this enzyme activity is significantly higher in leiomyoma tissues than in surrounding myometrial tissues. Thus, we suggested the possibility that the estrone produced by the action of aromatase in the leiomyoma is closely related to tumor development and enlargement. On the other hand, the level of 17 p-estradiol dehydrogenase activity in uterine leiomyoma tissues has been found to be higher than [3], lower than [4], or the same as [5] that in uterine tissues surrounding the leiomyoma. Thus, it is of interest to determine how estrogens in the local uterus influence the development of uterine leiomyomas. Moreover, it has been reported that there is a high frequency of endometrial hyperplasia in association with uterine leiomyoma [61. This pathological finding supports the theory that local hyperestrogenism occurs in uterine leiomyoma rather than the general degeneration resulting from mechanical pressure on the leiomyoma. In this study, therefore, we examined estrone sulfatase (E,SF) activity, which converts estrone sulfate (E,S) to estrone (E,) in leiomyoma, myometrial, and endometrial tissues of the same uterus. MATERIALS AND METHODS Steriods
[6,7-3H]Estrone-3-sulfate (E,S; sp act 42.0 Ci/mmole) and [4-‘4Clestrone (E,; sp act 56.4 mCi/mmole) were purchased from NEN Research Products (Boston, MA). Nonlabeled E$ and El were purchased from Sigma Chemical Company (St. Louis, MO). Before use, the radiochemical purity of labeled E,S was checked using thin-layer chromatography.
315 C@O-8258/!30 $1.50
Copyright 0 1990 by Academic Press, Inc. All rights of reproduction in any form resewed.
316
YAMAMOTO
Tissue Preparation
Uterine tissues were obtained through hysterectomy from patients (n = 11) with uterine leiomyoma. The locations of resected uterine tissues which contributed to this experiment are shown in Fig, 1. Four different samples were resected from each uterus and prepared for analysis. Two of the eleven cases were at the ovarian follicular phase, while the remaining nine cases were at the ovarian luteal phase. Patients ranged from 35 to 46 years in age. Assay of Estrone Sulfatase Activity Concentrations in Tissue
ET AL.
determination of ErSF activity under conditions linear with respect to incubation time and to the amount of enzyme source used. Measurement of E, and E,S concentrations. One milliliter of tissue homogenate was used for the measurement of estrogen concentration. Tissue homogenate was extracted three times using ethyl ether, after which the ether extract was submitted to radioimmunoassay (E, test set from Travenol, Tokyo, Japan) for E, while the water-soluble fraction was submitted to radioimmunoassay for E,S [7].
and Estrogen
Tissue samples from endometrium, uterine leiomyoma, and uterine muscle were minced and homogenized in 5 ml of 0.01 M Tris-HCI buffer solution (pH 7.4). Each homogenate was divided into two parts: one was submitted for assay of estrone sulfatase activity and the other for measurement of estrogen concentration. Assay ofEJF activity. Homogenates were centrifuged at 8OOgfor 10 min, after which [6,7-3H]E,S (2 ,uCi) was added to the supernatant fraction (2.5 ml) obtained, and the mixture was incubated at 37°C for 30 min in air. The enzyme reaction was terminated through the addition of 3 vol of ethyl ether. Next, [4-14C]E, (5000 dpm) and nonlabeled E, (500 pg) were added to the incubated sample. The extraction of nonconjugated steroids using ethyl ether was repeated twice. After the ether extract was evaporated and dissolved in 0.5 ml of methanol, the products were further developed by thin-layer chromatography (TLC, two solvent systems: chloroform-ethyl acetate 4: 1, benzene-ethyl acetate 1: 1, v/v) to isolate the E, formed. The amount of El formed was calculated from the final 3H/‘4C ratio of the El area on the chromatogram. The amount of protein in the 8OOgsupernatant fraction was measured using a Bio-Rad Protein Assay Kit (BioRad Laboratories, Richmond, CA). This assay enabled
RESULTS The EISF activities of various uterine tissues are shown graphically in Fig. 2. EISF activity was 2.62 + 0.29 nmole/hr/mg protein (n = 11, mean + SD) in uterine endometrial tissue overlying the leiomyoma (location C) and 2.0 t 0.24 nmole/hr/mg protein in endometrial tissue overlying the normal myometrium opposite the leiomyoma (location D). Hence, E,SF activity was significantly higher (P < 0.001, t test) in location C than in location D. On the other hand, EISF activity in leiomyoma tissue (location A; 0.49 + 0.82 nmole/hr/mg protein) was lower (P < 0.05) than that in myometrial tissue (location B; 0.76 ? 0.10 nmole/hr/mg protein). Thus, E,SF activity in endometrial tissue was significantly higher (P < 0.01) than that in myometrial tissue or leiomyoma tissue. E, and E,S concentrations were (n = 6, mean + SD) 4.32 & 2.30 and 1004 +- 331, pg/mg protein respectively, in uterine leiomyoma tissue (location A) and 38.7 ? 20.8 and 910 r 352 pg/mg protein, respectively, in myometrial tissue (location B) opposite the leiomyoma. Thus, the El concentration in leiomyoma tissue was significantly lower (P < 0.01) than that in myometrial tissue. 7
P
, r-
FIG. 1. Location of the uterine tissues used in this experiment: (A) uterine leiomyoma, (B) normal myometrium, (C) endometrium overlying leiomyoma, (D) endometrium overlying normal myometrium.
P
7
FIG. 2. Estrone sulfatase (E,SF) activity in various uterine tissues obtained from patients (n = 11) with uterine leiomyoma.
ESTRONE SULFATASE
ACTIVITY
E, and E,S concentrations in the endometrial tissue overlying a leiomyoma were 10.9 +- 8.9 and 631 2 339 pg/mg protein, respectively, and those in normal endometrial tissue were 1.23 -r 1.97 and 902 2 482 pg/mg protein, respectively. Thus, the El concentration in the endometrial tissue overlying leiomyoma appeared to be higher than that in the endometrial tissue overlying normal myometrium. ES concentration, on the other hand, was lower in endometrial tissue overlying leiomyoma than in normal endometrial tissue overlying nonleiomyoma myometrium. (See Table 1.) DISCUSSION It is well known that the genesis and development of uterine leiomyoma are closely related to estrogen. No significant differences, however, have been found in the estrogen levels in peripheral blood between patients with uterine leiomyoma and those without. We suspect, therefore, that local hyperestrogenism in the uterine leiomyoma, as opposed to general hyperestrogenism throughout the body, is what is related to tumor enlargement. Estrone sulfatase, estrone sulfotransferase, and estradiol dehydrogenase generally regulate estrogen metabolism in the uterus (mainly in uterine endometrium). Thus, in this study, we examined E,SF activity in leiomyoma, myometrium, and endometrial tissues of uteri with leiomyomas. We also measured E, and E,S concentrations in these tissues. Results showed E,SF activity in endometrial tissue to be significantly high compared with that in leiomyoma tissue or myometrial tissue of the same uterus. Furthermore, E,SF activity in the endometrial tissue overlying leiomyoma was significantly higher than that in the endometrial tissue overlying myometrium with no myoma node. These results seem consistent with the corresponding El and E,S concentrations in these tissues as measured in this study. Platia et al. [8] suggested that E,SF activity in the endometrium is higher than that in the myometrium of the uterus. Our present results agree with this assessment. Eiletz et al. [9] reported that the E, concentration (1.5-2.0 pg/mg protein, n = 25) in a uterine leiomyoma is significantly lower than that in the uterine myometrium (7-18 pg/mg protein, it = 50). Our data support this claim that El concentration was higher in myometrial tissue compared with leiomyoma tissue, the results of the assays used being statistically significant (P < 0.01). On the basis of the present results, E,SF activity in the endometrium overlying a leiomyoma may contribute to local hyperestrogenism in the leiomyoma and may assist in the genesis or enlargement of the tumor. EISF activity in the endometrial tissue was similar to that observed by Adessi et al. [lo]. They demonstrated E,SF activity to be 23.13 f. 8.44 and 62.81 2 21.97 pmole E,/min/mg protein (mean f SD) in normal en-
317
IN UTERINE LEIOMYOMA
TABLE 1 Estrone(E,) and E&one Sulfate(E,S) Concentrationsin Various Uterine TissuesObtainedfrom Patientswith Uterine Leiomyoma(n = 6) Concentration (pg/mg protein) Location
W
El
Leiomyoma Normal myometrium Endometrium overlying leiomyoma Endometrium overlying normal myometrium
4.32 + 2.30* 38.7 +- 20.8 10.9 * 1.23 ”
1004 ? 331 910 k 352
8.9
631 f 339
1.97
902 f 482
’ Mean ? SD. * P < 0.01 for the value of location B.
dometrial tissue and mild endometrial hyperplasia tissue, respectively (P < 0.01). In general, hyperplasia of the uterine endometrium is often found in association with uterine leiomyoma [l 11. Deligdish and Loewenthal [l l] reported that cystic glandular hyperplasia was frequently found in the endometrium at the margin of a leiomyoma. Thus, our results would appear to be reasonable. Warren and French [121 reported that EISF activity in uterine endometrium fluctuates during the menstrual cycle. Gurpide et al. [13] and Prost and Adessi [14], however, suggest the contrary, that is, EISF activity in the uterine endometrium does not fluctuate during the menstrual cycle. In support of the latter hypothesis, Urabe er al. [151:demonstrated that EISF activity does not change significantly between proliferative and secretory endometrium. Besides E, sulfatase, DHEA sulfatase also exists in the human uterine endometrium. According to a report by Prost and Adessi [141, the K,,, of E, sulfatase in uterine endometrium is 3.1 PM, whereas that of DHEA sulfatase is 5.7 PM, thus suggesting that the sulfatases for these substrates are entirely different. Carlstrom et al. [16] have speculated that formation of estradiol-17P (E2) from E,S in endometrium may be of importance in the genesis of endometrial disorders in postmenopausal women. As demonstrated in this study, an increase in El sulfatase activity in the local endometrium of the uterus may be closely related to the enlargement of uterine leiomyoma. ACKNOWLEDGMENT We thank Miss Etsuko Murata for her technical assistance in the E,S assay and for typing the manuscript.
REFERENCES 1. Kistner, R. W. Gynecology principles and practice, Book Medical Publishers, Chicago (1979).
3rd ed., Year
318
YAMAMOTO
2. Yamamoto, T., Takamori, K., and Okada, H. Estrogen biosynthesis in leiomyoma and myometrium of the uterus, Horm. Metabol. Res. 16, 678-679 (1984). 3. Fujii, S. Pathogenesis and histogenesis of uterine leiomyoma with special emphasis on sex steroids correlation, Acta Obstet. Gynaecol. Japan. 35, 1166-1174 (1983). 4. Pollow, K., Geilfub, J., Boquoi, E., and Pollow, B. Estrogen and progesterone binding proteins in normal human myometrium and leiomyoma tissue, J. Clin. Chem. C/in. Biochem. 16, 503-511 (1978). 5. Gabb, R. G., and Stone, G. M. Uptake and metabolism of tritiated oestradiol and oestrone by human endometrial and myometrial tissue in vitro, J. Endocrinol. 62, 109-123 (1974). 6. Novak, E. Gynecological and Obstetrical Pathologie, Saunders, Philadelphia (1974). Honjo, H., Kitawaki, J., Itoh, M., Yasuda, J., Iwasaku, K., Urabe, M., Naitoh, K., Yamamoto, T., Okada, H., and Ohkubo, T. Serum and urinary estrone sulfate during the menstrual cycle, measured by a direct radioimmunoassay, and fate of exogenously injected estrone sulfate, Horm. Res. 27, 61-68 (1987). Platia, M. P., Fencl, M. D., Elkind-Hirsch, K. E., Canick, J. A., and Tulchinsky, D. Estrone sulfatase activity in the human brain and estrone sulfate levels in the normal menstrual cycle, J. Steroid Biochem. 21, 237-241 (1984). Eifetz, J., Genz, T., Pollow, K., and Schmidt-Gollwitzer, M. Sex steroid levels in serum, myometrium and fibromyomata in corre-
ET AL. lation with cytoplasmic receptors and 17p-HSD activity in different age-groups and phases of the menstrual cycle, Arch. Gynecol. 229, 13-28 (1980). 16. Adessi, G. L., Prost, O., Agnani, G., Petitjean, A., and Burnod, J. Estrone sulfatase activity in normal and abnormal endometrium, Arch Gynecol. 236, 13-18 (1984). ll ’ Deligdish, L., and Loewenthal, M. Endometrial changes associated with myomata of the uterus, J. Clin. Pathol. 23, 676-680 (1970). 12. Warren, J. C., and French, A. P. Distribution of steroid sulfatase in human tissues, J. Clin. Endocrinol. Metab. 25, 278-282 (1965). 13. Gurpide, E., Gusberg, S. B., and Tseng, L. Estradiol binding and metabolism in human endometrial hyperplasia and adenocarcinoma, J. Steroid Biochem. 7, 891-896 (1976). 14. Prost, O., and Adessi, G. L. Estrone and dehydroepiandrosterone sulfatase activities in normal and pathological human endometrium biopsies, J. Clin. Endocrinol. Metab. 56, 653-661 (1983). 15. Urabe, M., Yamamoto, T., Naitoh, K., Honjo, H., and Okada, H. Estrone sulfatase activity in normal and neoplastic endometrial tissues of human uterus, Asia-Oceania J. Obstet. Gynaecol. 15, 101-106 (1989). 16, Carlstrom, K., von Uexkull, A. K., Einhorn, N., Fredricsson, B., Lunell, N. O., and Sundelin, P. Metabolism of estrone sulfate in human endometrium, Acta Obstet. Gynecol. Stand. 62, 519-524 (1983).
Estrone Sulfatase Activity in Human Uterine Leiomyoma TAKARA YAMAMOTO, M.D.,’ MAMORU URABE, M.D., KAZUO NAITOH, M.D., Jo KITAWAKI,
M.D., HIDEO HONJO,
M.D., AND HIROJI OKADA, M.D. Department
of Obstetrics
and Gynecology,
Kyoto Prefectural
University
of Medicine,
Kyoto 602, Japan
Received June 19, 1989
Human uterine leiomyoma is a benign tumor and its development is closely related to estrogen. In this study, e&one sulfatase (E$F) activity and concentrations of e&one (E,) and esbone sulfate (E,S) were measured in endometrial, leiomyoma, and myometrial tissues of the same human uterus (n = 11) with a leiomyoma. EISF activity in endometrial tissue overlying a leiomyoma was 2.62 f 0.29 nmole/hr/mg protein (mean f SD), this activity being significantly higher (P < 0.01) compared with that in normal endometrial tissue (2.0 f 0.24 mnole/hr/mg protein). EISF activity in normal endometrial tissue was significantly higher (P < 0.001) compared with that in leiomyoma tissue (0.49 * 0.82 nmole/hr/mg protein) or myometrial tissue (0.76 -C 0.10 nmole/hr/mg protein). We also found a significant difference (P < 0.05) in EISF activity between leiomyoma tissue and myometrial tissue. On the other hand, the E, concentration in endometrial tissue overlying a leiomyoma (10.9 f 8.9 pg/mg protein) proved to be higher than that in endometrial tissue overlying normal myometrium (1.23 + 1.97 pg/mg protein). E,S concentrations in these tissues, however, were 631 f 339 and 902 -C 482 pg/mg protein, respectively, these values showing a trend opposite that of the El concentration data. Thus, these results suggest that high EISF activity and high E, concentration in the endometrium overlying a leiomyoma may be related to estrogen supply to a uterine leiomyoma node. ~1 1990 Academic mess, Inc.
INTRODUCTION While uterine leiomyomas rarely appear in young females before the onset of menstruation, they frequently occur in sexually mature women with menstrual cycles; the frequency generally decreases after menopause. Thus, their appearance and development seem to be closely related to ovarian hormones (i.e., estrogen and progesterone) [l]. Uterine leiomyomas are benign tumors, though they have a tendency to be found in combination with other estrogen-dependent tumors (breast ’ To whom reprint requests should be addressed.
cancer, uterine endometrial cancer, and so on) [ll. Because of this relationship with estrogen, we examined estrogen biosynthetase (aromatase) activity, which converts androgens to estrogens, in leiomyoma and myometrial tissues of the human uterus [2]. Results indicated that this enzyme activity is significantly higher in leiomyoma tissues than in surrounding myometrial tissues. Thus, we suggested the possibility that the estrone produced by the action of aromatase in the leiomyoma is closely related to tumor development and enlargement. On the other hand, the level of 17 p-estradiol dehydrogenase activity in uterine leiomyoma tissues has been found to be higher than [3], lower than [4], or the same as [5] that in uterine tissues surrounding the leiomyoma. Thus, it is of interest to determine how estrogens in the local uterus influence the development of uterine leiomyomas. Moreover, it has been reported that there is a high frequency of endometrial hyperplasia in association with uterine leiomyoma [61. This pathological finding supports the theory that local hyperestrogenism occurs in uterine leiomyoma rather than the general degeneration resulting from mechanical pressure on the leiomyoma. In this study, therefore, we examined estrone sulfatase (E,SF) activity, which converts estrone sulfate (E,S) to estrone (E,) in leiomyoma, myometrial, and endometrial tissues of the same uterus. MATERIALS AND METHODS Steriods
[6,7-3H]Estrone-3-sulfate (E,S; sp act 42.0 Ci/mmole) and [4-‘4Clestrone (E,; sp act 56.4 mCi/mmole) were purchased from NEN Research Products (Boston, MA). Nonlabeled E$ and El were purchased from Sigma Chemical Company (St. Louis, MO). Before use, the radiochemical purity of labeled E,S was checked using thin-layer chromatography.
315 C@O-8258/!30 $1.50
Copyright 0 1990 by Academic Press, Inc. All rights of reproduction in any form resewed.
316
YAMAMOTO
Tissue Preparation
Uterine tissues were obtained through hysterectomy from patients (n = 11) with uterine leiomyoma. The locations of resected uterine tissues which contributed to this experiment are shown in Fig, 1. Four different samples were resected from each uterus and prepared for analysis. Two of the eleven cases were at the ovarian follicular phase, while the remaining nine cases were at the ovarian luteal phase. Patients ranged from 35 to 46 years in age. Assay of Estrone Sulfatase Activity Concentrations in Tissue
ET AL.
determination of ErSF activity under conditions linear with respect to incubation time and to the amount of enzyme source used. Measurement of E, and E,S concentrations. One milliliter of tissue homogenate was used for the measurement of estrogen concentration. Tissue homogenate was extracted three times using ethyl ether, after which the ether extract was submitted to radioimmunoassay (E, test set from Travenol, Tokyo, Japan) for E, while the water-soluble fraction was submitted to radioimmunoassay for E,S [7].
and Estrogen
Tissue samples from endometrium, uterine leiomyoma, and uterine muscle were minced and homogenized in 5 ml of 0.01 M Tris-HCI buffer solution (pH 7.4). Each homogenate was divided into two parts: one was submitted for assay of estrone sulfatase activity and the other for measurement of estrogen concentration. Assay ofEJF activity. Homogenates were centrifuged at 8OOgfor 10 min, after which [6,7-3H]E,S (2 ,uCi) was added to the supernatant fraction (2.5 ml) obtained, and the mixture was incubated at 37°C for 30 min in air. The enzyme reaction was terminated through the addition of 3 vol of ethyl ether. Next, [4-14C]E, (5000 dpm) and nonlabeled E, (500 pg) were added to the incubated sample. The extraction of nonconjugated steroids using ethyl ether was repeated twice. After the ether extract was evaporated and dissolved in 0.5 ml of methanol, the products were further developed by thin-layer chromatography (TLC, two solvent systems: chloroform-ethyl acetate 4: 1, benzene-ethyl acetate 1: 1, v/v) to isolate the E, formed. The amount of El formed was calculated from the final 3H/‘4C ratio of the El area on the chromatogram. The amount of protein in the 8OOgsupernatant fraction was measured using a Bio-Rad Protein Assay Kit (BioRad Laboratories, Richmond, CA). This assay enabled
RESULTS The EISF activities of various uterine tissues are shown graphically in Fig. 2. EISF activity was 2.62 + 0.29 nmole/hr/mg protein (n = 11, mean + SD) in uterine endometrial tissue overlying the leiomyoma (location C) and 2.0 t 0.24 nmole/hr/mg protein in endometrial tissue overlying the normal myometrium opposite the leiomyoma (location D). Hence, E,SF activity was significantly higher (P < 0.001, t test) in location C than in location D. On the other hand, EISF activity in leiomyoma tissue (location A; 0.49 + 0.82 nmole/hr/mg protein) was lower (P < 0.05) than that in myometrial tissue (location B; 0.76 ? 0.10 nmole/hr/mg protein). Thus, E,SF activity in endometrial tissue was significantly higher (P < 0.01) than that in myometrial tissue or leiomyoma tissue. E, and E,S concentrations were (n = 6, mean + SD) 4.32 & 2.30 and 1004 +- 331, pg/mg protein respectively, in uterine leiomyoma tissue (location A) and 38.7 ? 20.8 and 910 r 352 pg/mg protein, respectively, in myometrial tissue (location B) opposite the leiomyoma. Thus, the El concentration in leiomyoma tissue was significantly lower (P < 0.01) than that in myometrial tissue. 7
P
, r-
FIG. 1. Location of the uterine tissues used in this experiment: (A) uterine leiomyoma, (B) normal myometrium, (C) endometrium overlying leiomyoma, (D) endometrium overlying normal myometrium.
P
7
FIG. 2. Estrone sulfatase (E,SF) activity in various uterine tissues obtained from patients (n = 11) with uterine leiomyoma.
ESTRONE SULFATASE
ACTIVITY
E, and E,S concentrations in the endometrial tissue overlying a leiomyoma were 10.9 +- 8.9 and 631 2 339 pg/mg protein, respectively, and those in normal endometrial tissue were 1.23 -r 1.97 and 902 2 482 pg/mg protein, respectively. Thus, the El concentration in the endometrial tissue overlying leiomyoma appeared to be higher than that in the endometrial tissue overlying normal myometrium. ES concentration, on the other hand, was lower in endometrial tissue overlying leiomyoma than in normal endometrial tissue overlying nonleiomyoma myometrium. (See Table 1.) DISCUSSION It is well known that the genesis and development of uterine leiomyoma are closely related to estrogen. No significant differences, however, have been found in the estrogen levels in peripheral blood between patients with uterine leiomyoma and those without. We suspect, therefore, that local hyperestrogenism in the uterine leiomyoma, as opposed to general hyperestrogenism throughout the body, is what is related to tumor enlargement. Estrone sulfatase, estrone sulfotransferase, and estradiol dehydrogenase generally regulate estrogen metabolism in the uterus (mainly in uterine endometrium). Thus, in this study, we examined E,SF activity in leiomyoma, myometrium, and endometrial tissues of uteri with leiomyomas. We also measured E, and E,S concentrations in these tissues. Results showed E,SF activity in endometrial tissue to be significantly high compared with that in leiomyoma tissue or myometrial tissue of the same uterus. Furthermore, E,SF activity in the endometrial tissue overlying leiomyoma was significantly higher than that in the endometrial tissue overlying myometrium with no myoma node. These results seem consistent with the corresponding El and E,S concentrations in these tissues as measured in this study. Platia et al. [8] suggested that E,SF activity in the endometrium is higher than that in the myometrium of the uterus. Our present results agree with this assessment. Eiletz et al. [9] reported that the E, concentration (1.5-2.0 pg/mg protein, n = 25) in a uterine leiomyoma is significantly lower than that in the uterine myometrium (7-18 pg/mg protein, it = 50). Our data support this claim that El concentration was higher in myometrial tissue compared with leiomyoma tissue, the results of the assays used being statistically significant (P < 0.01). On the basis of the present results, E,SF activity in the endometrium overlying a leiomyoma may contribute to local hyperestrogenism in the leiomyoma and may assist in the genesis or enlargement of the tumor. EISF activity in the endometrial tissue was similar to that observed by Adessi et al. [lo]. They demonstrated E,SF activity to be 23.13 f. 8.44 and 62.81 2 21.97 pmole E,/min/mg protein (mean f SD) in normal en-
317
IN UTERINE LEIOMYOMA
TABLE 1 Estrone(E,) and E&one Sulfate(E,S) Concentrationsin Various Uterine TissuesObtainedfrom Patientswith Uterine Leiomyoma(n = 6) Concentration (pg/mg protein) Location
W
El
Leiomyoma Normal myometrium Endometrium overlying leiomyoma Endometrium overlying normal myometrium
4.32 + 2.30* 38.7 +- 20.8 10.9 * 1.23 ”
1004 ? 331 910 k 352
8.9
631 f 339
1.97
902 f 482
’ Mean ? SD. * P < 0.01 for the value of location B.
dometrial tissue and mild endometrial hyperplasia tissue, respectively (P < 0.01). In general, hyperplasia of the uterine endometrium is often found in association with uterine leiomyoma [l 11. Deligdish and Loewenthal [l l] reported that cystic glandular hyperplasia was frequently found in the endometrium at the margin of a leiomyoma. Thus, our results would appear to be reasonable. Warren and French [121 reported that EISF activity in uterine endometrium fluctuates during the menstrual cycle. Gurpide et al. [13] and Prost and Adessi [14], however, suggest the contrary, that is, EISF activity in the uterine endometrium does not fluctuate during the menstrual cycle. In support of the latter hypothesis, Urabe er al. [151:demonstrated that EISF activity does not change significantly between proliferative and secretory endometrium. Besides E, sulfatase, DHEA sulfatase also exists in the human uterine endometrium. According to a report by Prost and Adessi [141, the K,,, of E, sulfatase in uterine endometrium is 3.1 PM, whereas that of DHEA sulfatase is 5.7 PM, thus suggesting that the sulfatases for these substrates are entirely different. Carlstrom et al. [16] have speculated that formation of estradiol-17P (E2) from E,S in endometrium may be of importance in the genesis of endometrial disorders in postmenopausal women. As demonstrated in this study, an increase in El sulfatase activity in the local endometrium of the uterus may be closely related to the enlargement of uterine leiomyoma. ACKNOWLEDGMENT We thank Miss Etsuko Murata for her technical assistance in the E,S assay and for typing the manuscript.
REFERENCES 1. Kistner, R. W. Gynecology principles and practice, Book Medical Publishers, Chicago (1979).
3rd ed., Year
318
YAMAMOTO
2. Yamamoto, T., Takamori, K., and Okada, H. Estrogen biosynthesis in leiomyoma and myometrium of the uterus, Horm. Metabol. Res. 16, 678-679 (1984). 3. Fujii, S. Pathogenesis and histogenesis of uterine leiomyoma with special emphasis on sex steroids correlation, Acta Obstet. Gynaecol. Japan. 35, 1166-1174 (1983). 4. Pollow, K., Geilfub, J., Boquoi, E., and Pollow, B. Estrogen and progesterone binding proteins in normal human myometrium and leiomyoma tissue, J. Clin. Chem. C/in. Biochem. 16, 503-511 (1978). 5. Gabb, R. G., and Stone, G. M. Uptake and metabolism of tritiated oestradiol and oestrone by human endometrial and myometrial tissue in vitro, J. Endocrinol. 62, 109-123 (1974). 6. Novak, E. Gynecological and Obstetrical Pathologie, Saunders, Philadelphia (1974). Honjo, H., Kitawaki, J., Itoh, M., Yasuda, J., Iwasaku, K., Urabe, M., Naitoh, K., Yamamoto, T., Okada, H., and Ohkubo, T. Serum and urinary estrone sulfate during the menstrual cycle, measured by a direct radioimmunoassay, and fate of exogenously injected estrone sulfate, Horm. Res. 27, 61-68 (1987). Platia, M. P., Fencl, M. D., Elkind-Hirsch, K. E., Canick, J. A., and Tulchinsky, D. Estrone sulfatase activity in the human brain and estrone sulfate levels in the normal menstrual cycle, J. Steroid Biochem. 21, 237-241 (1984). Eifetz, J., Genz, T., Pollow, K., and Schmidt-Gollwitzer, M. Sex steroid levels in serum, myometrium and fibromyomata in corre-
ET AL. lation with cytoplasmic receptors and 17p-HSD activity in different age-groups and phases of the menstrual cycle, Arch. Gynecol. 229, 13-28 (1980). 16. Adessi, G. L., Prost, O., Agnani, G., Petitjean, A., and Burnod, J. Estrone sulfatase activity in normal and abnormal endometrium, Arch Gynecol. 236, 13-18 (1984). ll ’ Deligdish, L., and Loewenthal, M. Endometrial changes associated with myomata of the uterus, J. Clin. Pathol. 23, 676-680 (1970). 12. Warren, J. C., and French, A. P. Distribution of steroid sulfatase in human tissues, J. Clin. Endocrinol. Metab. 25, 278-282 (1965). 13. Gurpide, E., Gusberg, S. B., and Tseng, L. Estradiol binding and metabolism in human endometrial hyperplasia and adenocarcinoma, J. Steroid Biochem. 7, 891-896 (1976). 14. Prost, O., and Adessi, G. L. Estrone and dehydroepiandrosterone sulfatase activities in normal and pathological human endometrium biopsies, J. Clin. Endocrinol. Metab. 56, 653-661 (1983). 15. Urabe, M., Yamamoto, T., Naitoh, K., Honjo, H., and Okada, H. Estrone sulfatase activity in normal and neoplastic endometrial tissues of human uterus, Asia-Oceania J. Obstet. Gynaecol. 15, 101-106 (1989). 16, Carlstrom, K., von Uexkull, A. K., Einhorn, N., Fredricsson, B., Lunell, N. O., and Sundelin, P. Metabolism of estrone sulfate in human endometrium, Acta Obstet. Gynecol. Stand. 62, 519-524 (1983).