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Pathophysiology of uterine leiomyomas Laval University Hospital Research Center and Department of Obstetrics and Gynecology, Laval University, 2705 Blvd. Laurier, Ste. Foy, Quk., Canada Gl V 4G2 Received September 20, 1991
KOUTSILIERIS, M. 1992. Pathophysiology of uterine leiomyomas. Biochem. Cell Biol. 70: 273-278. Uterine leiomyomas is the most common benign neoplasia in women, one of the most frequent causes of infertility in reproductive years, and the leading cause for hysterectomy. The pathophysiology of uterine leiomyomas is uncertain. Therefore, therapeutic approaches have been primarily empirical. It is now well documented that growth factors control the functional and possibly the histological integrity of several tissues. Recently the presence of growth substances in uterine tissues suggested that the role of sex steroid hormones in the pathophysiology of leiomyomas may be mediated by substances influencing the proliferation of smooth muscle cells and fibroblasts. This report summarizes the data related to the pathophysiology of leiomyomas, which indicate a possible role of growth factors in uterine leiomyomas. Key words: leiomyomas, growth factors, uterus, smooth muscle cells, fibroblasts.
KOUTSILIERIS, M. 1992. Pathophysiology of uterine leiomyomas. Biochem. Cell Biol. 70 : 273-278. Le ltiomyome uttrin est la ntoplasie bknigne la plus commune chez les femmes, l'une des causes les plus frtquentes d'infertilitt au cours des anntes de reproduction et la principale cause d'hysttrectomie. La pathophysiologie des ltiomyomes uttrins est ma1 connue. Les premitres approches thtrapeutiques ont donc t t t empiriques. Nous savons bien maintenant que des facteurs de croissance contr6lent I'integritt fonctionnelle et probablement histologique de plusieurs tissus. Rhmment, la presence de facteurs de croissance dans les tissus utkrins a suggbt que le r6le des hormones sexuelles sttroi'diennes dans la pathophysiologie des ltiomyomes pourrait s'exercer par l'intermkdiaire de substances influen~antla proliftration des cellules musculaires lisses et des fibroblastes. Nous rksumons ici des donntes relatives 5 la pathophysiologie des ltiomyomes qui montrent le r6le possible de facteurs de croissance dans les lkiomyomes uttrins. Mots elks : lleomyomes, facteurs de croissance, utbus, cellules musculaires lisses, fibroblastes. [Traduit par la rtdaction]
Introduction Leiomyomas are common, benign tumors which may occur at any anatomical location containing smooth muscle cells (Marshall et al. 1960). Most of these tumors originate in the female genital tract, and in particular, in the uterus (Marshall et al. 1960; Farman 1975). Approximately 28 cases of leiomyomas of the female urethra (Fry et al. 1988) and 200 cases of leiomyomas of the bladder (Kabalin et al. 1990) have been reported in the world literature. Although uterine leiomyoma is the most common solid tumor in women, very little is known regarding its etiology. It is estimated that 20% of all women over the age 35 have leiomyomas (Novak and Woodruff 1979). Other studies suggested that uterine leiomyomas are more common among black and nulliparous women (Novak and Woodruff 1979). A recent study analysing data from gross serial sectioning at 2-mm intervals in 100 consecutive hysterectomies adjuncted to routine pathology concluded that epidemiological studies may not be valid, if they are based only on clinical diagnosis or routine pathology reports (Cramer and Pate1 1990). Therefore, the real incidence of uterine leiomyomas remains unknown. Despite this, leiomyomas arise most often during the third and fourth decades of life and have an important impact upon reproductive performance. The social trend in recent dehydrogenase; ABBREVIATIONS:G-6-PD, glucose-6-~hos~hate 17b-HSD9 17b-h~dr0x~ster0iddeh~drOgenase; GnRH-A, gonadotropin-releasing hormone analogs; LH, luteinizing hormone; kDa, kilodalton(s); EGF, epidermal growth factor; IGF. insulin-like growth factor; IGFBP-I, IGF-binding protein-l; PDGF-A, plateletderived growth factor A; bFGF, basic fibroblast growth factor. Printed in Canada / Imprime au Canada
years where first pregnancies are delayed beyond age 30 could increase the relative frequency of leiomyomas (Buttram 1986). Leiomyomas themselves are relatively avascular and tightly compacted tumors of smooth muscle cells and fibroblasts. Usually they are surrounded by a pseudocapsule of alveolar tissue and have two large vessels to nourish the tumor (Buttram 1986). In certain cases, extensive extramedullary haemopoietic sites in degenerating leiomyomas (Schmid et al. 1990) and histiocytes in leiomyomas in densities higher than in adjacent normal myometrium (Adany et al. 1990) were described. The potential significance of these observations for the pathophysiology of the disease is at the present time unclear. Leiomyomas tend to be multiple and slow growing (Buttram 1986). Most patients who have small leiomyomas are asyrnptomatic, but the leiomyomas range in size and can extend to giant tumor masses that fill the pelvic and abdominal cavities. The intramucosal location and submucosal varieties are associated with the greatest influence upon reproductive capabilities. Since all leiomyoma cells are of identical G-6-PD electrophoretic type, the tumor is considered of unicellular origin, although the (3-6-PD type may vary from one tumor to another within the same uterus (Townsend et al. 1970). ~ecentl;leiomyomas have been reported to bear tumorspecific chromosome aberations, although the majority of them (50-80%) are of normal karyotype (Nibert and Heim 1990). There have been four cfiogenetically subgroups identified as rearrangements of 6p, del(7) (q21.2 q31.2), + 12, and t(12;14) (q14-15; q23-24). Few cfiogenetic similarities so far have been detected between leiomyomas
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and malignant smooth muscle tumors (leiomyomas and rhabdomyosarcomas), but the leiomyoma cytogenic profile has been strikingly similar to lipoma. Similarities existing between leiomyomas and pleomorphic adenomas of the salivary gland and with myxoid liposarcomas have been reported (Nibert and Heim 1990). Leiomyoma is rarely associated with benign-appearing smooth muscle tumors in sites such as lungs and lymph nodes. In the gynecologic literature this was referred to as benign metastasizing leiomyomas (Hartz and Hugenholtz 1942; Clark and Weed 1977). It was suggested that either the primary lesion is a low-grade sarcoma with malignant potential or this is the "end-stage" of intravenous leiomyomatosis (Spiro and McPeak 1966; Wolff et al. 1979). The multifocal origin of these tumors was proposed also (Cho et al. 1989). In pathology, mitotic indices have proven to be highly effective in predicting the courses of smooth muscle tumors of the uterus, but metastasizing smooth muscle tumors with low mitotic indices and histologic appearance of ordinary leiomyomas were reported (Hartz and Hugenholtz 1942). In addition, mitotic activity of leiomyomas was reported to be significantly influenced by the hormonal milieu suggesting that (i) the growth of leiomyomas is affected by sex steroid hormones, and (ii) mitotic indices must be evaluated within the context of the phase of the menstrual cycle before it can be determined whether a smooth muscle cell tumor is malignant (Kawaguchi et al. 1989). Pathogenesis of uterine leiomyomas Enzymatic implications Cytochrome P-450 serves a vital role as the terminal oxidase in the microsomal monooxygenase hydroxylation reaction of a wide variety of xenobiotics including drugs and carcinogens, as well as endogenous substrates such as steroid hormones. Recently sensitive spectrophotometric assays have detected significantly higher cytochrome P-450 activity in leiomyoma than adjacent myometrium. This was proposed to be part of the pathogenetic mechanisms involved in the pathophysiology of leiomyomas (Senler et al. 1985a, 19856). In addition, the 170-HSD activity in leiomyomas did not follow the significant increase of enzymatic activity detected after ovulation as normal myometrium of the same uterus (Eiletz et al. 1980). The moderate increase of the 170-HSD levels after ovulation and the low progesterone receptor binding sites detected in leiomyomas was suggestive for a possible multiple defect of sex steroid hormone metabolism which, in turn, may be causatively linked to the pathogenesis of the disease. Other studies (Herrmann et al. 1987) reported that the specific activity (activity per unit tissue mass) of enzymes involved in the carbohydrate metabolism (hexokinase, phosphofructokinase, lactase dehydrogenase, and glucogen phosphorylase) is not different in leiomyoma tissues when compared with normal adjacent myometrium. The same was true for the specific activity of enzymes involved in tricarboxylic acid cycle (succinate dehydrogenase), but significant differences were detected in the specific activity of enzymes involved in fatty acid oxidation (hydroxyacetyl-CoA dehydrogenase). The latter observation suggested that increased fatty acid utilisation by leiomyomas may be related to increase growth potential of this tissue (Herrmann et al. 1987).
BIOL. VOL. 70, 1992 TABLE1. Pathophysiology of leiomyomas Enzymatic involvement Higher cytochrome P-450 activity (Senler et al. 1 9 8 5 ~ 19856) . Lower tissue levels of glutathionine (Basu et al. 1988) Lower levels of p-carotene (Palan et al. 1988) Lower tissue aromatase activity (Folkerd et al. 1984) Higher levels of hydroxyacyl-COA dehydrogenase (Herrmann et al. 1987)
Also decreased tissue levels of reduced glutathione (Basu et al. 1988) and 0-carotene (Palan et al. 1988), as well as decreased aromatase activity (Folkerd et al. 1984), was reported in leiomyoma tissue. These findings were compatible with a direct or indirect enzymatic involvement with the metabolic defects underlining the presence of leiomyomas (Table 1). The question is whether these findings are causatively linked with leiomyomas or are the direct result of such pathology. Hormonal implications Cell division, differentiation, and growth of uterine tissues are based on the well-orchestrated secretion of sex steroid hormones by the ovaries. The biological activity of these hormones depends on their specific interaction with specific binding proteins inside the cell. These proteins are referred to as receptors. The tendency of uterine leiomyomas to grow during the reproductive years and to regress postmenopausally suggested that sex steroid hormones may be implicated in the pathophysiology of the disease. This observation led to the initial hypothesis that women bearing such tumors would either have altered serum sex steroid hormone levels or increased sex steroid hormone activity in the tumor. However, serum levels of sex steroid hormones in women with leiomyomas were similar to those of normal women (Buttram 1986). The concentration of tissue levels of estradiol, estrone, and progesterone in normal myometrium was studied during menstrual cycle and was found to be similar to serum concentrations, except for higher levels of tissue estrone in the luteal phase and of 170-HSD activity during the early secretory phase (Eiletz et al. 1980). Other studies detected an increase in intramyoma estrogen concentration (estradiol levels), resulting presumably from inadequate conversion of estradiol to estrone (Pollow et al. 1978). This was explained by a moderate increase of the 170-HSD activity in leiomyoma tissues, as compared with normal myometrium, which was also compatible with the lower levels of estrone reported in leiomyoma tissues (Eiletz et al. 1980). The assessment of estrogen receptor content in leiomyomas with respect to normal myometrium revealed conflicting data, such as (i) altered but not significantly higher content (Otsuka et al. 1989), (ii) not different in content (Sadan et al. 1988), (iii) not different in content and moreover similar regulation of the estrogen receptor by cyclic changes during the menstrual cycle (Soules and McCarty 1982), and (iv) significantly higher content (Tamaya et al. 1985). At this point the progesterone receptor (which is induced by estrogens in uterine tissues) was implicated in the pathogenesis of the disease. Again the number and (or) the regulation of progesterone receptor content throughout the menstrual cycle in leiomyomas, as compared with normal myometrium, revealed conflicting data; i.e., (i) normal (Buchi and Keller
TABLE2. The effect of GnRH-analogs on leiomyomas Uterine leiomyomas shrink up to 50% of their initial volume within 3-4 months of commencing therapy (Healy 1991) Tumors re-enlarge back to their original size 6-12 months after commencement of therapy (Healy 1991) There is a significant reduction in cellularity but there are no significant changes in fibrosis, edema, and mitotic activity (Upadhyaya et al. 1990) Estrogen deprivation is causing hypoplasia but not cell death of smooth muscle cell (Healy 1991) Estradiol, estrone, and estradiol plus estrone sulfate tissue levels are decreased (Pasqualini et al. 1990) Progesterone receptor content is decreased (Pasqualini et al. 1990) Short-term recurrence of leiomyomas is increased in women receiving GnRH-A therapy before myomectomy (Fedele et al. 1990) TABLE3. The possible role of growth factors and growth factor receptors in the physiology and pathophysiology of uterus Animal tissues Estromedins (Sirbasku 1978; Biro 1986) Metastatic-stimulating activity from mouse uterus (Pantazis and Howard 1987; Maharajan et al. 1988) Rat-, rabbit-, bovine-, and porcine-derived growth factors (Simmen et al. 1988; Beck and Garner 1989; Milner et al. 1989) Estrogen-induced EGF in rat uterus (Gonzalez et al. 1984) Human tissues EGF receptors in human leiomyomas (Lumsden et al. 1988) Increased expression of mRNA for the PDGF-A chain in human myometrium during pregnancy (Mendoza et al. 1990) Human myometrium-derived (MDGE) and leiomyoma-derived growth factors (LDGF) (Koutsilieris 1989, 1990)
1983; Soules and McCarty 1982; Sadan et al. 1990), (ii) higher (Tamaya et al. 1985; Chrapusta et al. 1990; Vij et al. 1990; Pasqualini et al. 1990; Benagiano et al. 1990), and (iii) lower and not changing during the menstrual cycle (Eiletz et al. 1980; Folkerd et al. 1984). Estrogen and progesterone receptor content was higher in Caucasian than in Negroid patients (Sadan et al. 1988). This was suggestive of a genetic predisposition and the fact that the estrogen and progesterone receptor content could participate in the pathophysiology of the disease (Sadan et al. 1990). In addition, prostaglandin receptor content (E and F2a) was reported to be lower in leiomyomas than in normal myometrium (Hoffmann et al. 1984), suggesting relative insensitivity to the biological actions of prostaglandins. The prostaglandin insensitivity and estrogen hyperreactivity were proposed to be possible causes for the pathogenesis of the disease (Hoffmann et al. 1984). Recently specific binding sites for GnRH-A were documented in leiomyomas, suggestive of a possible direct effect of GnRH-A on tumor growth (Wiznitzer et al. 1988). GnRH-As have effected regression of uterine leiomyomas through a mechanism thought to be mediated by the inhibition of LH release and steroid synthesis (Maheux and Lemay 1988). Histopathologically, treatment with GnRH-A correlated with a significant reduction in cellularity of leiomyomas, although no significant changes in fibrosis, edema, or mitotic activity were reported (Upadhyaya et al. 1990). There is now agreement that uterine leiomyomas will shrink to 50% of their initial volume within 3-4 months of commencing treatment and that tumor will then re-enlarge back to their original size anywhere from 6 to 12 months after commencement of therapy (Table 2). The shrinkage
of the tumor appears to be estrogen mediated, causing hypoplasia of smooth muscle, but not cell death (Healy 1991). GnRH-A treatment provoked a significant decrease in tissue estradiol and in particular, estrone-S and estradiol-S levels. Progesterone receptor content of leiomyomas was decreased significantly after 3-4 months of GnRH-A treatment (Pasqualini et al. 1990). The mean leiomyoma estrogen content in women treated with GnRH-A was significantly higher than in women receiving placebo. Clinically, the significant increase in leiomyoma estrogen receptor may account for the rapid regrowth of leiomyomas observed after the cessation of GnRH-A therapy (Rein et al. 1990b). This may also explain recent findings showing higher short-term recurrence of uterine leiomyomas in women receiving GnRH-A before myomectomy, thus limiting the efficacy of surgery (Fedele et al. 1990). Therefore, the initial enthusiasm for the potential use of GnRH-As in the therapy of leiomyomas has subsided. Despite this, GnRH-As have nevertheless provided further evidence for the implication of sex steroids in uterine pathophysiology. Although the influence of sex steroid hormones on the growth of uterine leiomyomas is accepted, a good molecular basis for the pathogenesis of the disease is lacking. It is also unclear whether the preceding evidence of a steroid hormone component in the pathophysiology of the disease is related with the initiation of this pathology or would only promote the growth of leiomyomas, which may have been initiated by another mechanism.
Growth factor implications The remarkable increase of tissue by in vivo treatment with estrogens is a well-known response of estrogen-target
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TABLE 4. IGF-1, IGF-11, IGFS receptors, and IGFBPs in uterine physiology and pathophysiology Estrogen-induced IGF-I and -11 mRNA is expressed in rat uterus (Murphy and Friesen 1988; Murphy et al. 1988; Murphy and Ghahary 1990) Estrogen-induced IGF-I and -11 mRNA is expressed in pig uterus (Simmen et al. 1990; Ogasawara et al. 1989) Type I IGF receptor is expressed in rat uterus (Ghahary and Murphy 1989) Diminishing volumes of leiomyomas in women treated with GnRH-As correlated well with decreasing IGF-I and -11 levels in explant cultures of these tissues (Rein et al. 1990a) IGFBP mRNAs are expressed in uterine tissues (Croze et al. 1990) organs. A 5- to 10-fold increase of pituitary gland and a 2- to 5-fold enlargement of mammary and uterus tissues are normal consequences of estrogen treatment. Depending on the dose and duration of estrogen administration, uterine mass may be increased by hypertrophy (increase in cell mass), hyperplasia (increase in cell number), or both. Estrogens stimulate protein synthesis in vivo and in vitro, but a clear demonstration of their effects in vitro is very limited compared with their in vivo mitogenic effects (Biro 1986). Therefore, it is conceivable local regulators could indirectly mediate steroid hormone actions to be implicated (Pollard 1990). Estromedins are the best example of this concept (Sirbasku 1978; Biro 1986), and have been proposed as (i) mediators of estrogen action on target organs and (ii) specific stimulators of estrogen-dependent cells or tissues in an endocrine, paracrine, or autocrine manner. This concept gave rise to the identification of several biologically active peptides from uterine tissues (Table 3) such as (i) metastasis-stimulating factors from mouse uterus (Pantazis and Howard 1987; Maharajan et al. 1988); (ii) rat, rabbit, and bovine uterus-derived growth factors that were acid stable, heat labile, reduced by trypsin, and eluted at molecular mass of 10-30 kDa by gel filtration (Beck and Garner 1989); (iii) a 17-kDa heparin-binding growth factor (HBGF-8) purified from bovine uterus similar to bFGF (Milner et al. 1989); (iv) mitogens distinct from EGF, derived from porcine uterus (Simmen et al. 1988); and (v) heparinbinding growth factors derived from porcine uterus (Bridstock et al. 1990). Of special interest, in this context, is the group of IGFs (Table 4). The inductive effect of estrogens on uterine IGF-I mRNA in rat uterus was reported (Murphy et al. 1988; Murphy and Friesen 1988; Norstedt et al. 1989) and was related to authentic type I IGF receptor which was localized in rat myometrial smooth muscle cells. The expression of mRNA for type I receptor was also regulated by estrogens (Ghahary and Murphy 1989), although similar effects of steroid hormones on uterine IGF-I receptor expression in the pig were not observed (Hofig et al. 1991). The expression and the regulation of the IGF-I gene by estrogens and progesterone were reported also in pig uterus (Simrnen et al. 1990), as well a truncated form of IGF-I was purified from porcine uterus (Ogasawara et al. 1989). This truncated IGF-I form was of high biologic activity for mesenchymal cell types. In addition, decreasing levels of IGF-I and IGF-I1 were detected in explant cultures of leiomyoma and myometrial tissues that were associated with diminishing leiomyoma tumor volumes after treatment by GnRH-A (Rein et al. 1990a). Interestingly, the expression of IGFBP-1 was detected in rat uterus (Croze et al. 1990). The capacity
of this binding protein either to increase or to inhibit the action of IGF-I in various bioassay systems suggested that IGFs and IGFBPs could be implicated in various pathophysiological processes in the uterus (Murphy and Ghahary 1990). The possible involvement of growth factors in the pathophysiology of uterus was further advanced by studies reporting increased expression of mRNA for the PDGF-A chain gene. This was detected in smooth muscle cells of human uterus during physiologic hypertrophy, which occurs in the course of pregnancy (Mendoza et al. 1990; Boehm et al. 1990). In addition, the presence of EGF receptor was detected in mouse uteri (Brown et al. 1989) and human uterine leiomyoma cells (Fayed et al. 1989). Furthermore, estradiol-induced EGF in immature mice uteri (Gonzalez et al. 1984) and binding sites of EGF were detected in the human uterus (Lumsden et al. 1988). Leiomyoma receptors for EGF in leiomyomas treated with GnRH-A were decreased significantly compared with those in leiomyomas of nontreated women (Lumsden et al. 1988). Human uterine extracts were also documented to contain mitogens and inhibitors of skin fibroblast growth (Koutsilieris 1989); normal rat osteoblasts and osteoblast-derived osteosarcoma cells (Koutsilieris 1989; Koutsilieris and Michaud 1990); and myoblasts and smooth muscle cells (Koutsilieris 1990; Koutsilieris et al. 1990). Preferential growth factors for fibroblasts and smooth muscle were also detected in leiomyoma extracts. These mitogen(s) were undetectable in myometrial and endometrial extracts treated and processed identically (Koutsilieris et al. 1990). These data provided additional evidence for the validity of the concept involving locally acting growth factors in the regulation of human myometrium and have supported the argument that locally acting growth substances may be involved in human uterine pathophysiology. Finally, it appears that it will be important to assess the role of certain members of the cell-cycle regulating genes (cyclin family), one of which (PRADI cyclin-like gene) was documented to be overexpressed in a benign parathyroid tumor. This was the result of a chromosome rearrangement that had placed cyclin gene under the control of a new promoter (Motokura et al. 1991). Certainly this type of approach could provide new insights in the pathophysiology of benign tumors, suggesting new avenues of research.
Conclusions and future directions It is now apparent that the uterus is a rich source of growth factors involved in complex autocrine-paracrine regulatory circuits, which in turn interact with sex steroid hormones. Detailed studies of the appearance of these
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growth factors and their receptors in leiomyomas and myometrium during pregnancy and under different hormonal regimens have yet to be performed. It is presently unclear whether estrogens or growth factors would initiate the growth of leiomyomas, or would promote the growth of these lesions initiated by another mechanism. Although no causative role has been yet directly established for any of the growth factors in uterine biology and pathophysiology, current data provide strong evidence to support such a pathophysiologically important function. Therefore, the study of the growth factor expression in leiomyoma tissues and of their estrogen responsiveness is of fundamental importance in advancing our current knowledge for the pathophysiology of uterine leiomyomas.
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In Advances in gynecological endocrinology. Edited by A.R. Genazzani, S. Betaglia, A. Volti, and F. Faccinnetti. Parthenon Publishing, Park Ridge, N. J. pp. 227-24 1. Marshall, F.C., Uson, A.C., and Melicow, M.M. 1960. Neoplasms and caruncles of the female urethra. Surg. Gynecol. Obstet. 110: 727-732. Mendoza, A.E., Young, R., Orkin, S.H., and Collins, T. 1990. Increased platelet-derived growth factor A-chain expression in human uterine smooth muscle cells during the physiologic hypertrophy of pregnancy. Proc. Natl. Acad. Sci. U.S.A. 87: 2177-2181. Milner, P.G., Li, Y.-S., Hoffman, R.M., et al. 1989. A novel 17 kD heparin-binding growth factor (HBGF-8) in bovine uterus: purification and N-terminal amino acid sequence. Biochem. Biophys. Res. Commun. 165: 1096-1 103. Motokura, T., Bloom, T., Kim, H.G., et al. 1991. A novel cyclin encoded by a bc/l-linked candidate oncogene. Nature (London), 350: 512-515. Murphy, L.J., and Friesen, H.G. 1988. Differential effects of estrogen and growth hormone on uterine and hepatic insulinlike growth factor-I (IGF-I) gene expression in the ovariectomized rat. Endocrinology (Baltimore), 122: 325-334. Murphy, L.J., and Ghahary, A. 1990. Uterine insulin-like growth factor-1: regulation of expression and its role in estrogen-induced uterine proliferation. Endocrinol. Rev. 11: 443-453. Murphy, L.J., Murphy, L.C., and Friesen, H.G. 1988. Estrogen induces insulin-like growth factor-I expression in rat uterus. Mol. Endocrinol. 1: 445-452. Nibert, M., and Heim, S. 1990. Uterine leiomyoma cytogenetics. Genes Chromosomes Cancer 2: 3-13. Norstedt, G., Levinovitz, A., and Eriksson, H. 1989. Regulation of uterine insulin-like growth factor I mRNA and insulin-like growth factor I1 mRNA by estrogen in rat. Acta Endocrinol. 120: 466-472. Novak, E.R., and Woodruff, V.D. 1979. Uterine leiomyomas. In Gynecologic and obstetric pathology. 8 ed. W.B. Saunders, Philadelphia, Pa. pp. 260-279. Ogasawara. M., Karey, K.P., Marquardt, H., and Sirbasku, D.A. 1989. Identification and purification of truncated insulin-like growth factor I from porcine uterus: evidence for high biological potency. Biochemistry, 28: 2710-2721. Otsuka, H., Shinohara, M., Kashiomura, M., et al. 1989. A comparative study of the estrogen receptor ratio in myometrium and uterine leiomyomas. Int. J. Gynaecol. Obstet. 29: 189-194. Palan, P.R., Michail, M., Basu, J., et al. 1988. Decreased P-carotene levels in leiomyomas of the uterus. In The program of The 35th annual meeting of the Society for Gynecologic Investigation, Baltimore, Md. Abstr. no. 557. Pantazis, C.G., and Howard, E.F. 1987. Metastatic-stimulating activity in mouse uterus. Biol. Reprod. 36: 701-708. Pasqualini, J.R., Cornier, E., Grenier, J., et al. 1990. Effects of decapeptyl, an agonistic analog of gonadotropin-releasing hormone on estrogens, estrogen sulphates, and progesterone receptors in leiomyoma and myometrium. Fertil. Steril. 53: 1012-1017. Pollard, J.W. 1990. Regulation of polypeptide growth factor synthesis and growth factor-related gene expression in the rat and mouse uterus before and after implantation. J. Reprod. Fertil. 88: 721-731. Pollow, K., Sinnecker, G., Boquoi, E., and Pollow, B. 1978. In vitro conversion of estradiol-17P into estrone in normal human myometrium and leiomyoma. J. Clin. Chem. Clin. Biochem. 16: 493-502. Rein, M.S., Friedman, A.J., Pandjian, M.R., and Heffner, L.J. 1990a. The secretion of insulin-like growth factors I and I1 by
explant cultures of fibroids and myometrium from women treated with a gonadotropin-releasinghormone agonist. Obstet. Gynecol. 76: 388-394. Rein, M.S., Friedman, A.J., Stuart, J.M., and MacLaughlin, D.T. 1990b. Fibroid and myometrial steroid receptors in women treated with gonadotropin-releasing hormone agonist leuprolide acetate. Fertil. Steril. 53: 1018-1023. Sadan, O., van Iddekinge, B., Savage, N., et al. 1988. Ethnic variation in the estrogen and progesterone receptor concentration in leiomyoma and normal myometrium. Gynecol. Endocrinol. 2: 275-282. Sadan, O., van Iddekinge, B., Savage, N., et al. 1990. Endocrine profile associated with estrogen and progesterone receptors in leiomyoma and normal myometrium. Gynecol. Endocrinol. 4: 33-42. Schmid, C., Beharn, A., and Kratochvil, P. 1990. Haematopoietis in a degenerating uterine leiomyoma. Arch. Gynecol. Obstet. 248: 81-86. Senler, T.I., Dean, W .L., Pierce, W .M., and Wittliff, J.L. 1985a. Quantification of cytochrome P-450 dependent cyclohexane hydroxylase activity in normal and neoplastic reproductive tissues. Biochem. J. 227: 379-382. Senler, T.I., Hofmann, G.E., Sanfilippo, J.S., et al. 1985b. Cytochrome P-450 activity in human leiomyoma and normal myometrium. Am. J. Obstet. Gynecol. 153: 551-555. Simmen, R.C.M., KO, Y., Liu, X.H., et al. 1988. A uterine cell mitogen distinct from epidermal growth factor in porcine uterine fluids: characterization and partial purification. Biol. Reprod. 38: 551-561. Simmen, R.C.M., Simrnen, F. A., Hofig, A., et al. 1990. Hormonal regulation of insulin-like growth factor gene expression in pig uterus. Endocrinology (Baltimore), 127: 2166-2174. Sirbasku, D.A. 1978. Estrogen induction of growth factor specific for hormone-responsive mammary, pituitary, and kidney tumor cells. Proc. Natl. Acad. Sci. U.S.A. 75: 3786-3790. Soules, M.R., and McCarty, K.S., Jr. 1982. Leiomyomas: steroid receptor content, variation within normal menstrual cycles. Am. J. Obstet. Gynecol. 143: 6-11. Spiro, R.H., and McPeak, C. J. 1966. On the so-called metastasizing leiomyoma. Cancer (Philadelphia), 19: 544-548. Tamaya, T., Fujimoto, J., and Okada, H. 1985. Comparison of cellular levels of receptors in uterine leiomyoma and myometrium. Acta Obstet. Gynecol. Scand. 64: 307-309. Townsend, D.E., Sparkes, R.S., and McClelland, M.D. 1970. Unicellular histogenesis of uterine leiomyomas as detected by electrophoresis of glucose-6-phosphate dehydrogenase. Am. J. Obstet. Gynecol. 107: 1168-1 173. Upadhyaya, N.B., Doody, M.C., and Googe, P.B. 1990. Histological changes in leiomyomata treated with leuprolide acetate. Fertil. Steril. 54: 811-814. Vij, U., Murugesan, K., Laumas, K.R., and Farook, A. 1990. Progestin and antiprogestin interactions with progesterone receptors in human myomas. Int. J. Gynaecol. Obstet. 31: 347-353. Wiznitzer, A., Marbach, M., Hazum, E., et al. 1988. Gonadotropin-releasing hormone specific binding sites in uterine leiomyomata. Biochem. Biophys. Res. Commun. 152: 1326-1331. Wolff, A., Kaye, G., and Silva, F. 1979. Pulmonary metastases (with admixed epithelial elements) from smooth muscle neoplasms: report of nine cases, including three males. Am. J. Surg. Pathol. 3: 325-342.
Pathophysiology of uterine leiomyomas Laval University Hospital Research Center and Department of Obstetrics and Gynecology, Laval University, 2705 Blvd. Laurier, Ste. Foy, Quk., Canada Gl V 4G2 Received September 20, 1991
KOUTSILIERIS, M. 1992. Pathophysiology of uterine leiomyomas. Biochem. Cell Biol. 70: 273-278. Uterine leiomyomas is the most common benign neoplasia in women, one of the most frequent causes of infertility in reproductive years, and the leading cause for hysterectomy. The pathophysiology of uterine leiomyomas is uncertain. Therefore, therapeutic approaches have been primarily empirical. It is now well documented that growth factors control the functional and possibly the histological integrity of several tissues. Recently the presence of growth substances in uterine tissues suggested that the role of sex steroid hormones in the pathophysiology of leiomyomas may be mediated by substances influencing the proliferation of smooth muscle cells and fibroblasts. This report summarizes the data related to the pathophysiology of leiomyomas, which indicate a possible role of growth factors in uterine leiomyomas. Key words: leiomyomas, growth factors, uterus, smooth muscle cells, fibroblasts.
KOUTSILIERIS, M. 1992. Pathophysiology of uterine leiomyomas. Biochem. Cell Biol. 70 : 273-278. Le ltiomyome uttrin est la ntoplasie bknigne la plus commune chez les femmes, l'une des causes les plus frtquentes d'infertilitt au cours des anntes de reproduction et la principale cause d'hysttrectomie. La pathophysiologie des ltiomyomes uttrins est ma1 connue. Les premitres approches thtrapeutiques ont donc t t t empiriques. Nous savons bien maintenant que des facteurs de croissance contr6lent I'integritt fonctionnelle et probablement histologique de plusieurs tissus. Rhmment, la presence de facteurs de croissance dans les tissus utkrins a suggbt que le r6le des hormones sexuelles sttroi'diennes dans la pathophysiologie des ltiomyomes pourrait s'exercer par l'intermkdiaire de substances influen~antla proliftration des cellules musculaires lisses et des fibroblastes. Nous rksumons ici des donntes relatives 5 la pathophysiologie des ltiomyomes qui montrent le r6le possible de facteurs de croissance dans les lkiomyomes uttrins. Mots elks : lleomyomes, facteurs de croissance, utbus, cellules musculaires lisses, fibroblastes. [Traduit par la rtdaction]
Introduction Leiomyomas are common, benign tumors which may occur at any anatomical location containing smooth muscle cells (Marshall et al. 1960). Most of these tumors originate in the female genital tract, and in particular, in the uterus (Marshall et al. 1960; Farman 1975). Approximately 28 cases of leiomyomas of the female urethra (Fry et al. 1988) and 200 cases of leiomyomas of the bladder (Kabalin et al. 1990) have been reported in the world literature. Although uterine leiomyoma is the most common solid tumor in women, very little is known regarding its etiology. It is estimated that 20% of all women over the age 35 have leiomyomas (Novak and Woodruff 1979). Other studies suggested that uterine leiomyomas are more common among black and nulliparous women (Novak and Woodruff 1979). A recent study analysing data from gross serial sectioning at 2-mm intervals in 100 consecutive hysterectomies adjuncted to routine pathology concluded that epidemiological studies may not be valid, if they are based only on clinical diagnosis or routine pathology reports (Cramer and Pate1 1990). Therefore, the real incidence of uterine leiomyomas remains unknown. Despite this, leiomyomas arise most often during the third and fourth decades of life and have an important impact upon reproductive performance. The social trend in recent dehydrogenase; ABBREVIATIONS:G-6-PD, glucose-6-~hos~hate 17b-HSD9 17b-h~dr0x~ster0iddeh~drOgenase; GnRH-A, gonadotropin-releasing hormone analogs; LH, luteinizing hormone; kDa, kilodalton(s); EGF, epidermal growth factor; IGF. insulin-like growth factor; IGFBP-I, IGF-binding protein-l; PDGF-A, plateletderived growth factor A; bFGF, basic fibroblast growth factor. Printed in Canada / Imprime au Canada
years where first pregnancies are delayed beyond age 30 could increase the relative frequency of leiomyomas (Buttram 1986). Leiomyomas themselves are relatively avascular and tightly compacted tumors of smooth muscle cells and fibroblasts. Usually they are surrounded by a pseudocapsule of alveolar tissue and have two large vessels to nourish the tumor (Buttram 1986). In certain cases, extensive extramedullary haemopoietic sites in degenerating leiomyomas (Schmid et al. 1990) and histiocytes in leiomyomas in densities higher than in adjacent normal myometrium (Adany et al. 1990) were described. The potential significance of these observations for the pathophysiology of the disease is at the present time unclear. Leiomyomas tend to be multiple and slow growing (Buttram 1986). Most patients who have small leiomyomas are asyrnptomatic, but the leiomyomas range in size and can extend to giant tumor masses that fill the pelvic and abdominal cavities. The intramucosal location and submucosal varieties are associated with the greatest influence upon reproductive capabilities. Since all leiomyoma cells are of identical G-6-PD electrophoretic type, the tumor is considered of unicellular origin, although the (3-6-PD type may vary from one tumor to another within the same uterus (Townsend et al. 1970). ~ecentl;leiomyomas have been reported to bear tumorspecific chromosome aberations, although the majority of them (50-80%) are of normal karyotype (Nibert and Heim 1990). There have been four cfiogenetically subgroups identified as rearrangements of 6p, del(7) (q21.2 q31.2), + 12, and t(12;14) (q14-15; q23-24). Few cfiogenetic similarities so far have been detected between leiomyomas
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and malignant smooth muscle tumors (leiomyomas and rhabdomyosarcomas), but the leiomyoma cytogenic profile has been strikingly similar to lipoma. Similarities existing between leiomyomas and pleomorphic adenomas of the salivary gland and with myxoid liposarcomas have been reported (Nibert and Heim 1990). Leiomyoma is rarely associated with benign-appearing smooth muscle tumors in sites such as lungs and lymph nodes. In the gynecologic literature this was referred to as benign metastasizing leiomyomas (Hartz and Hugenholtz 1942; Clark and Weed 1977). It was suggested that either the primary lesion is a low-grade sarcoma with malignant potential or this is the "end-stage" of intravenous leiomyomatosis (Spiro and McPeak 1966; Wolff et al. 1979). The multifocal origin of these tumors was proposed also (Cho et al. 1989). In pathology, mitotic indices have proven to be highly effective in predicting the courses of smooth muscle tumors of the uterus, but metastasizing smooth muscle tumors with low mitotic indices and histologic appearance of ordinary leiomyomas were reported (Hartz and Hugenholtz 1942). In addition, mitotic activity of leiomyomas was reported to be significantly influenced by the hormonal milieu suggesting that (i) the growth of leiomyomas is affected by sex steroid hormones, and (ii) mitotic indices must be evaluated within the context of the phase of the menstrual cycle before it can be determined whether a smooth muscle cell tumor is malignant (Kawaguchi et al. 1989). Pathogenesis of uterine leiomyomas Enzymatic implications Cytochrome P-450 serves a vital role as the terminal oxidase in the microsomal monooxygenase hydroxylation reaction of a wide variety of xenobiotics including drugs and carcinogens, as well as endogenous substrates such as steroid hormones. Recently sensitive spectrophotometric assays have detected significantly higher cytochrome P-450 activity in leiomyoma than adjacent myometrium. This was proposed to be part of the pathogenetic mechanisms involved in the pathophysiology of leiomyomas (Senler et al. 1985a, 19856). In addition, the 170-HSD activity in leiomyomas did not follow the significant increase of enzymatic activity detected after ovulation as normal myometrium of the same uterus (Eiletz et al. 1980). The moderate increase of the 170-HSD levels after ovulation and the low progesterone receptor binding sites detected in leiomyomas was suggestive for a possible multiple defect of sex steroid hormone metabolism which, in turn, may be causatively linked to the pathogenesis of the disease. Other studies (Herrmann et al. 1987) reported that the specific activity (activity per unit tissue mass) of enzymes involved in the carbohydrate metabolism (hexokinase, phosphofructokinase, lactase dehydrogenase, and glucogen phosphorylase) is not different in leiomyoma tissues when compared with normal adjacent myometrium. The same was true for the specific activity of enzymes involved in tricarboxylic acid cycle (succinate dehydrogenase), but significant differences were detected in the specific activity of enzymes involved in fatty acid oxidation (hydroxyacetyl-CoA dehydrogenase). The latter observation suggested that increased fatty acid utilisation by leiomyomas may be related to increase growth potential of this tissue (Herrmann et al. 1987).
BIOL. VOL. 70, 1992 TABLE1. Pathophysiology of leiomyomas Enzymatic involvement Higher cytochrome P-450 activity (Senler et al. 1 9 8 5 ~ 19856) . Lower tissue levels of glutathionine (Basu et al. 1988) Lower levels of p-carotene (Palan et al. 1988) Lower tissue aromatase activity (Folkerd et al. 1984) Higher levels of hydroxyacyl-COA dehydrogenase (Herrmann et al. 1987)
Also decreased tissue levels of reduced glutathione (Basu et al. 1988) and 0-carotene (Palan et al. 1988), as well as decreased aromatase activity (Folkerd et al. 1984), was reported in leiomyoma tissue. These findings were compatible with a direct or indirect enzymatic involvement with the metabolic defects underlining the presence of leiomyomas (Table 1). The question is whether these findings are causatively linked with leiomyomas or are the direct result of such pathology. Hormonal implications Cell division, differentiation, and growth of uterine tissues are based on the well-orchestrated secretion of sex steroid hormones by the ovaries. The biological activity of these hormones depends on their specific interaction with specific binding proteins inside the cell. These proteins are referred to as receptors. The tendency of uterine leiomyomas to grow during the reproductive years and to regress postmenopausally suggested that sex steroid hormones may be implicated in the pathophysiology of the disease. This observation led to the initial hypothesis that women bearing such tumors would either have altered serum sex steroid hormone levels or increased sex steroid hormone activity in the tumor. However, serum levels of sex steroid hormones in women with leiomyomas were similar to those of normal women (Buttram 1986). The concentration of tissue levels of estradiol, estrone, and progesterone in normal myometrium was studied during menstrual cycle and was found to be similar to serum concentrations, except for higher levels of tissue estrone in the luteal phase and of 170-HSD activity during the early secretory phase (Eiletz et al. 1980). Other studies detected an increase in intramyoma estrogen concentration (estradiol levels), resulting presumably from inadequate conversion of estradiol to estrone (Pollow et al. 1978). This was explained by a moderate increase of the 170-HSD activity in leiomyoma tissues, as compared with normal myometrium, which was also compatible with the lower levels of estrone reported in leiomyoma tissues (Eiletz et al. 1980). The assessment of estrogen receptor content in leiomyomas with respect to normal myometrium revealed conflicting data, such as (i) altered but not significantly higher content (Otsuka et al. 1989), (ii) not different in content (Sadan et al. 1988), (iii) not different in content and moreover similar regulation of the estrogen receptor by cyclic changes during the menstrual cycle (Soules and McCarty 1982), and (iv) significantly higher content (Tamaya et al. 1985). At this point the progesterone receptor (which is induced by estrogens in uterine tissues) was implicated in the pathogenesis of the disease. Again the number and (or) the regulation of progesterone receptor content throughout the menstrual cycle in leiomyomas, as compared with normal myometrium, revealed conflicting data; i.e., (i) normal (Buchi and Keller
TABLE2. The effect of GnRH-analogs on leiomyomas Uterine leiomyomas shrink up to 50% of their initial volume within 3-4 months of commencing therapy (Healy 1991) Tumors re-enlarge back to their original size 6-12 months after commencement of therapy (Healy 1991) There is a significant reduction in cellularity but there are no significant changes in fibrosis, edema, and mitotic activity (Upadhyaya et al. 1990) Estrogen deprivation is causing hypoplasia but not cell death of smooth muscle cell (Healy 1991) Estradiol, estrone, and estradiol plus estrone sulfate tissue levels are decreased (Pasqualini et al. 1990) Progesterone receptor content is decreased (Pasqualini et al. 1990) Short-term recurrence of leiomyomas is increased in women receiving GnRH-A therapy before myomectomy (Fedele et al. 1990) TABLE3. The possible role of growth factors and growth factor receptors in the physiology and pathophysiology of uterus Animal tissues Estromedins (Sirbasku 1978; Biro 1986) Metastatic-stimulating activity from mouse uterus (Pantazis and Howard 1987; Maharajan et al. 1988) Rat-, rabbit-, bovine-, and porcine-derived growth factors (Simmen et al. 1988; Beck and Garner 1989; Milner et al. 1989) Estrogen-induced EGF in rat uterus (Gonzalez et al. 1984) Human tissues EGF receptors in human leiomyomas (Lumsden et al. 1988) Increased expression of mRNA for the PDGF-A chain in human myometrium during pregnancy (Mendoza et al. 1990) Human myometrium-derived (MDGE) and leiomyoma-derived growth factors (LDGF) (Koutsilieris 1989, 1990)
1983; Soules and McCarty 1982; Sadan et al. 1990), (ii) higher (Tamaya et al. 1985; Chrapusta et al. 1990; Vij et al. 1990; Pasqualini et al. 1990; Benagiano et al. 1990), and (iii) lower and not changing during the menstrual cycle (Eiletz et al. 1980; Folkerd et al. 1984). Estrogen and progesterone receptor content was higher in Caucasian than in Negroid patients (Sadan et al. 1988). This was suggestive of a genetic predisposition and the fact that the estrogen and progesterone receptor content could participate in the pathophysiology of the disease (Sadan et al. 1990). In addition, prostaglandin receptor content (E and F2a) was reported to be lower in leiomyomas than in normal myometrium (Hoffmann et al. 1984), suggesting relative insensitivity to the biological actions of prostaglandins. The prostaglandin insensitivity and estrogen hyperreactivity were proposed to be possible causes for the pathogenesis of the disease (Hoffmann et al. 1984). Recently specific binding sites for GnRH-A were documented in leiomyomas, suggestive of a possible direct effect of GnRH-A on tumor growth (Wiznitzer et al. 1988). GnRH-As have effected regression of uterine leiomyomas through a mechanism thought to be mediated by the inhibition of LH release and steroid synthesis (Maheux and Lemay 1988). Histopathologically, treatment with GnRH-A correlated with a significant reduction in cellularity of leiomyomas, although no significant changes in fibrosis, edema, or mitotic activity were reported (Upadhyaya et al. 1990). There is now agreement that uterine leiomyomas will shrink to 50% of their initial volume within 3-4 months of commencing treatment and that tumor will then re-enlarge back to their original size anywhere from 6 to 12 months after commencement of therapy (Table 2). The shrinkage
of the tumor appears to be estrogen mediated, causing hypoplasia of smooth muscle, but not cell death (Healy 1991). GnRH-A treatment provoked a significant decrease in tissue estradiol and in particular, estrone-S and estradiol-S levels. Progesterone receptor content of leiomyomas was decreased significantly after 3-4 months of GnRH-A treatment (Pasqualini et al. 1990). The mean leiomyoma estrogen content in women treated with GnRH-A was significantly higher than in women receiving placebo. Clinically, the significant increase in leiomyoma estrogen receptor may account for the rapid regrowth of leiomyomas observed after the cessation of GnRH-A therapy (Rein et al. 1990b). This may also explain recent findings showing higher short-term recurrence of uterine leiomyomas in women receiving GnRH-A before myomectomy, thus limiting the efficacy of surgery (Fedele et al. 1990). Therefore, the initial enthusiasm for the potential use of GnRH-As in the therapy of leiomyomas has subsided. Despite this, GnRH-As have nevertheless provided further evidence for the implication of sex steroids in uterine pathophysiology. Although the influence of sex steroid hormones on the growth of uterine leiomyomas is accepted, a good molecular basis for the pathogenesis of the disease is lacking. It is also unclear whether the preceding evidence of a steroid hormone component in the pathophysiology of the disease is related with the initiation of this pathology or would only promote the growth of leiomyomas, which may have been initiated by another mechanism.
Growth factor implications The remarkable increase of tissue by in vivo treatment with estrogens is a well-known response of estrogen-target
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TABLE 4. IGF-1, IGF-11, IGFS receptors, and IGFBPs in uterine physiology and pathophysiology Estrogen-induced IGF-I and -11 mRNA is expressed in rat uterus (Murphy and Friesen 1988; Murphy et al. 1988; Murphy and Ghahary 1990) Estrogen-induced IGF-I and -11 mRNA is expressed in pig uterus (Simmen et al. 1990; Ogasawara et al. 1989) Type I IGF receptor is expressed in rat uterus (Ghahary and Murphy 1989) Diminishing volumes of leiomyomas in women treated with GnRH-As correlated well with decreasing IGF-I and -11 levels in explant cultures of these tissues (Rein et al. 1990a) IGFBP mRNAs are expressed in uterine tissues (Croze et al. 1990) organs. A 5- to 10-fold increase of pituitary gland and a 2- to 5-fold enlargement of mammary and uterus tissues are normal consequences of estrogen treatment. Depending on the dose and duration of estrogen administration, uterine mass may be increased by hypertrophy (increase in cell mass), hyperplasia (increase in cell number), or both. Estrogens stimulate protein synthesis in vivo and in vitro, but a clear demonstration of their effects in vitro is very limited compared with their in vivo mitogenic effects (Biro 1986). Therefore, it is conceivable local regulators could indirectly mediate steroid hormone actions to be implicated (Pollard 1990). Estromedins are the best example of this concept (Sirbasku 1978; Biro 1986), and have been proposed as (i) mediators of estrogen action on target organs and (ii) specific stimulators of estrogen-dependent cells or tissues in an endocrine, paracrine, or autocrine manner. This concept gave rise to the identification of several biologically active peptides from uterine tissues (Table 3) such as (i) metastasis-stimulating factors from mouse uterus (Pantazis and Howard 1987; Maharajan et al. 1988); (ii) rat, rabbit, and bovine uterus-derived growth factors that were acid stable, heat labile, reduced by trypsin, and eluted at molecular mass of 10-30 kDa by gel filtration (Beck and Garner 1989); (iii) a 17-kDa heparin-binding growth factor (HBGF-8) purified from bovine uterus similar to bFGF (Milner et al. 1989); (iv) mitogens distinct from EGF, derived from porcine uterus (Simmen et al. 1988); and (v) heparinbinding growth factors derived from porcine uterus (Bridstock et al. 1990). Of special interest, in this context, is the group of IGFs (Table 4). The inductive effect of estrogens on uterine IGF-I mRNA in rat uterus was reported (Murphy et al. 1988; Murphy and Friesen 1988; Norstedt et al. 1989) and was related to authentic type I IGF receptor which was localized in rat myometrial smooth muscle cells. The expression of mRNA for type I receptor was also regulated by estrogens (Ghahary and Murphy 1989), although similar effects of steroid hormones on uterine IGF-I receptor expression in the pig were not observed (Hofig et al. 1991). The expression and the regulation of the IGF-I gene by estrogens and progesterone were reported also in pig uterus (Simrnen et al. 1990), as well a truncated form of IGF-I was purified from porcine uterus (Ogasawara et al. 1989). This truncated IGF-I form was of high biologic activity for mesenchymal cell types. In addition, decreasing levels of IGF-I and IGF-I1 were detected in explant cultures of leiomyoma and myometrial tissues that were associated with diminishing leiomyoma tumor volumes after treatment by GnRH-A (Rein et al. 1990a). Interestingly, the expression of IGFBP-1 was detected in rat uterus (Croze et al. 1990). The capacity
of this binding protein either to increase or to inhibit the action of IGF-I in various bioassay systems suggested that IGFs and IGFBPs could be implicated in various pathophysiological processes in the uterus (Murphy and Ghahary 1990). The possible involvement of growth factors in the pathophysiology of uterus was further advanced by studies reporting increased expression of mRNA for the PDGF-A chain gene. This was detected in smooth muscle cells of human uterus during physiologic hypertrophy, which occurs in the course of pregnancy (Mendoza et al. 1990; Boehm et al. 1990). In addition, the presence of EGF receptor was detected in mouse uteri (Brown et al. 1989) and human uterine leiomyoma cells (Fayed et al. 1989). Furthermore, estradiol-induced EGF in immature mice uteri (Gonzalez et al. 1984) and binding sites of EGF were detected in the human uterus (Lumsden et al. 1988). Leiomyoma receptors for EGF in leiomyomas treated with GnRH-A were decreased significantly compared with those in leiomyomas of nontreated women (Lumsden et al. 1988). Human uterine extracts were also documented to contain mitogens and inhibitors of skin fibroblast growth (Koutsilieris 1989); normal rat osteoblasts and osteoblast-derived osteosarcoma cells (Koutsilieris 1989; Koutsilieris and Michaud 1990); and myoblasts and smooth muscle cells (Koutsilieris 1990; Koutsilieris et al. 1990). Preferential growth factors for fibroblasts and smooth muscle were also detected in leiomyoma extracts. These mitogen(s) were undetectable in myometrial and endometrial extracts treated and processed identically (Koutsilieris et al. 1990). These data provided additional evidence for the validity of the concept involving locally acting growth factors in the regulation of human myometrium and have supported the argument that locally acting growth substances may be involved in human uterine pathophysiology. Finally, it appears that it will be important to assess the role of certain members of the cell-cycle regulating genes (cyclin family), one of which (PRADI cyclin-like gene) was documented to be overexpressed in a benign parathyroid tumor. This was the result of a chromosome rearrangement that had placed cyclin gene under the control of a new promoter (Motokura et al. 1991). Certainly this type of approach could provide new insights in the pathophysiology of benign tumors, suggesting new avenues of research.
Conclusions and future directions It is now apparent that the uterus is a rich source of growth factors involved in complex autocrine-paracrine regulatory circuits, which in turn interact with sex steroid hormones. Detailed studies of the appearance of these
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growth factors and their receptors in leiomyomas and myometrium during pregnancy and under different hormonal regimens have yet to be performed. It is presently unclear whether estrogens or growth factors would initiate the growth of leiomyomas, or would promote the growth of these lesions initiated by another mechanism. Although no causative role has been yet directly established for any of the growth factors in uterine biology and pathophysiology, current data provide strong evidence to support such a pathophysiologically important function. Therefore, the study of the growth factor expression in leiomyoma tissues and of their estrogen responsiveness is of fundamental importance in advancing our current knowledge for the pathophysiology of uterine leiomyomas.
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