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Matrix metalloproteinases (MMPs) are highly expressed in the human endometrium during menstruation, and these enzymes participate in the cyclic destruction and regeneration characteristic of the primate endometrium. To examine hormonal regulation of MMPs in vivo, we evaluated MMP expression and localization in the endometrium of ovariectomized rhesus macaques under various hormonal conditions. Although all MMPs were up-regulated by progesterone (P4) withdrawal, their expression declined spontaneously after menstruation in the absence of P4. Of 7 MMPs examined, only matrilysin and stromelysin-3 were suppressed any further when P4 levels were experimentally re-elevated. MMP expression was confined to the upper functionalis zone during menstruation, but after menstrual breakdown was complete, matrilysin and the tissue inhibitor of MMPs, TIMP-1, shifted expression from the functionalis to the basalis zone in the absence of both estradiol and P4. The spiral arteries in the functionalis, but not the basalis, were intense foci of MMP and TIMP-1 expression. Menstruation and MMP expression after P4 withdrawal were similar in both the presence and absence of estradiol. In sum, endometrial MMPs in vivo are strongly up-regulated by P4 withdrawal, but zone-specific tissue gradients greatly influence the pattern and degree of MMP expression.

The objective of this study was to detect and characterize a secreted oviduct-specific glycoprotein (OGP) in the rhesus macaque (Macaca mulatta) and to compare the characteristics of this OGP to those previously characterized in baboons and women. Oviducts were obtained from untreated ovariectomized rhesus and from ovariectomized rhesus either treated with estradiol (E2) for 14 days or treated sequentially with E2 for 14 days and then with E2 plus progesterone (P4) for an additional 14 days. Segments of oviducts were either fixed for morphological analysis, cultured for OGP synthesis and release, or frozen for RNA analysis. The proteins present in the culture media were separated by one-dimensional SDS-PAGE, and OGP was detected on Western blots using polyclonal antibodies generated against the reduced form of baboon OGP or a 17-amino acid segment of the baboon core protein. Cross-reacting antigens were present in the 120-kDa region, identical to what was observed for baboon and human OGP. Indirect immunogold localization of OGP on thin sections demonstrated specific clustering of gold particles over the apical secretory granules of the secretory cells of the oviductal epithelium. A cDNA was generated using RT-PCR and 5' and 3' rapid amplification of cDNA ends (RACE), and sequenced. The total transcript was 2237 nucleotides in length plus a poly(A) tail. The largest open reading frame was 624 amino acids, which would produce a protein of 69.3 kDa. The nucleotide sequence was more than 95% identical to the nucleotide sequences of baboon and human OGP. Northern blots revealed a single message at 2.4 kilobases (kb) in oviduct samples obtained from E2-treated rhesus. This message was absent in oviducts obtained from untreated ovariectomized and from sequential E2 plus P4-treated rhesus macaques. In summary, the rhesus oviduct synthesizes and secretes an OGP in the presence of E2 that is immunologically and structurally similar to the baboon and human OGP. The presence of a highly homologous glycoprotein in several primates suggests a similar function for OGP in the reproductive process.

We measured androgen receptors (AR) and 5 alpha-reductase activity (5 alpha RA) in the ductuli efferentes and epididymides from adult rhesus macaques. Tissue samples were either assayed biochemically for AR or stained immunocytochemically (ICC) with a monoclonal antibody against AR. To estimate 5 alpha RA, tissue microsomes were incubated with [1 alpha,2 alpha-3H]testosterone, and the [3H]dihydrotestosterone formed was quantified. We found significant regional differences in the levels of both 5 alpha RA and AR in the excurrent ducts. In general, both enzyme activity and AR levels were higher in the caput and corpus epididymis than in ductuli efferentes and cauda epididymis. With ICC, positive nuclear AR staining was detected in all epithelial cell types, whereas variable numbers of stromal cells were positively stained. Our data demonstrate that there are segmental differences in the concentrations of 5 alpha RA and AR in epididymis and suggest that there may be regional differences in the regulation of epididymal functions by androgen.

Ovarian androgens may act locally to modulate follicular and luteal function in various species. This study examined the distribution of androgen receptors within the primate ovary throughout the menstrual cycle. Ovaries were collected from rhesus and cynomolgus monkeys during the early, mid-, and late (n = 3-5 per stage) follicular and luteal phases of the cycle. The tissues were processed for indirect immunocytochemical localization of androgen receptors with a specific monoclonal antibody against human androgen receptor (AN1-15). In addition, ovaries (n = 3) were collected from rhesus monkeys for biochemical detection of androgen receptor using 3H-androgen and AN1-15. Specific immunocytochemical staining, as determined by comparing adjacent tissue sections incubated with either AN1-15 or a nonspecific control antibody, was exclusively nuclear. Androgen receptor was detected in the germinal epithelium and ovarian stroma at all stages of the cycle. The thecal and granulosa cells of growing follicles, and of many but not all atretic follicles, contained androgen receptors. Luteinizing granulosa cells of the periovulatory follicle and luteal cells from the early and midluteal phase stained intensely for androgen receptor. Regressing corpora lutea of the late luteal phase also stained for androgen receptor; however, fully regressed corpora lutea in the early follicular phase of the next cycle did not exhibit receptor staining. Luteal cells that were androgen receptor-positive also stained histochemically for the presence of 3 beta-hydroxysteroid dehydrogenase. Sucrose gradient analysis with radiolabeled androgen demonstrated a shift in the androgen receptor peak in monkey ovarian tissue upon addition of AN1-15, confirming the presence of androgen receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

We obtained the testes, ductuli efferentes, and epididymides from adult rhesus and cynomolgus macaques and examined these tissues for estrogen receptors (ER) with immunocytochemistry (ICC) and a sucrose gradient assay. Both techniques employed monoclonal antibodies prepared against ER, and both showed that high concentrations of ER were present OFFy in the ductuli efferentes. Moreover, all specific staining was confined to the nuclei of the nonciliated, absorptive epithelial cells. The quantity of salt-extractable ER in the ductuli efferentes (834 +/- 161 [SEM] fmol/mg DNA [n = 8]) did not differ significantly from the amounts measured with the identical assay in oviducts and endometrium of estrogenized female macaques. Testes and epididymides of macaques had no specific staining by ICC and barely detectable amounts by biochemical analysis (7 +/- 4 [n = 3], 8 +/- 2 [n = 5], 33 +/- 16 [n = 3], and 6 +/- 3 [n = 8] fmol/mg DNA for testis and caput, corpus, and cauda epididymis, respectively). The functional significance of the high levels of ER in the ductuli efferentes of macaques remains to be determined.

With the aid of monoclonal antibodies specific to the estrogen and progestin receptors, we have examined the cellular localization of these proteins in the reproductive tract of male and female macaques. Two striking findings have resulted from our work with these new reagents. First, these receptors are detectable only in cell nuclei, regardless of hormonal treatment, and second, they are often detectable in stromal, but not epithelial cells when the epithelial cells undergo various estrogen or progestin-dependent events. The latter observation has led us to conclude that stromal cell-epithelial cell interactions may play previously unappreciated roles in the hormonal control of the primate reproductive tract. The lines of evidence that have drawn us to this conclusion will be reviewed.

We sampled oviducts and endometria of 27 cynomolgus macaques during the menstrual cycle and measured the concentration of nuclear and cytoplasmic estrogen receptors in these tissues by exchange assay. We assessed the stage of the cycle by correlating serum estradiol (E2) and progesterone (P), as measured by radioimmunoassay, with the morphological condition of the ovaries, oviducts and endometrium of each animal. We have previously identified a series of oviductal stages that reflected the orderly sequence of cytological changes in the oviduct during the cycle, and we normalized receptor measurements to these stages. The amounts of nuclear and cytoplasmic estrogen receptor in both the oviduct and the endometrium were approximately twofold greater in the follicular phase than in the luteal phase. In the follicular phase, elevated receptor levels were associated with oviductal proliferation and differentiation, as well as with endometrial proliferation. During the luteal phase, lowered levels were correlated with atrophy and dedifferentiation in the oviduct, but with hypertrophy and progestational development in the endometrium. When the luteal phase of one cycle ends and the follicular phase of the next begins, it is a decline in serum P, not a rise in serum E2, that precedes the elevation in estrogen receptor level and the onset of proliferation in the oviduct and endometrium. Proliferation of the reproductive tract and elevations in nuclear estrogen receptor levels during the early follicular phase can therefore be viewed as consequences of the release of the system from antagonism by P.

We sampled the reproductive tracts of 27 cynomolgus macaques during the menstrual cycle and correlated the cytologic changes in the oviductal epithelium with changes in the serum levels of estradiol (E2) and progesterone (P) and with the histology of the ovaries and the endometria. We identified an orderly sequence of changes in the oviductal epithelium from the early follicular to the late luteal phase, and we classified this sequence into eight stages, named as follows: preciliogenic, ciliogenic, ciliogenic-ciliated, ciliated-ciliogenic, ciliated-secretory, early regression, late regression and full regression. The preciliogenic and ciliogenic phases were coincident with menses and the early follicular phase. The ciliogenic-ciliated, ciliated-ciliogenic and ciliated-secretory phases during which the oviductal epithelium became progressively more differentiated were coincident, respectively, with the midfollicular, late follicular and periovulatory phases of the cycle. The early, late and full regression stages during which the epithelium became progressively more atrophied, deciliated and nonsecretory were coincident, respectively, with the early, mid and late luteal phases of the cycle. The cyclic changes in the endometrium of cynomolgus macaques were similar to those reported for the rhesus macaque.

We compared the morphology of ovaries from fetal rhesus monkeys at two different stages of gestation with the ability of ovarian tissue to synthesize androgens and estrogens in vitro. Ovaries collected between 80 and 104 days of gestation contained single-layered primordial follicles but no multilayered or antral follicles. Within these ovaries we found theca-like interstitial cells which contained abundant smooth endoplasmic reticulum and lipid droplets. These ovaries produced androstenedione and dehydropiandrosterone (but not estradiol) in vitro in the absence of exogenous substrates. Ovaries collected between Days 124 and 153 of gestation, however, contained numerous, well-developed multilayered and antral follicles. The ultrastructural characteristics of the thecal and granulosa cell layers were similar to those of adult ovarian follicles. These ovaries synthesized large amounts of androstenedione, dehydroepiandrosterone, and estradiol in vitro. Identity of the estradiol was confirmed by incubating homogenates of late gestation ovaries with [14C]progesterone, by isolation of estradiol and formation of its derivative, and by recrystallization to constant specific activity. In vitro cyclic AMP synthesis was stimulated by Pergonal (luteinizing hormone [LH] + follicle stimulating hormone [FSH] ) only in ovaries which contained multilayered and antral follicles, an indication that fetal ovaries contain gonadotropin receptors during late stages of development. These data indicate that fetal ovaries of rhesus monkeys attain the capacity for de novo estrogen biosynthesis and to respond to gonadotropins during late gestation, when multilayered and antral follicles have developed.