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Prolactin receptor-associated protein (PRAP) originally cloned in our laboratory was shown to be a novel, luteal isoform of 17β hydroxysteroid dehydrogenase 7 (17βHSD7). In this study, we cloned the promoter region of rat PRAP/17βHSD7 and investigated the mechanisms regulating both basal activity and LH-induced repression of this promoter. Truncated and site-specific mutants of PRAP/17βHSD7 promoter identified two enhancer regions that contained highly conserved Sp1 binding site and bound Sp1 from nuclear extracts of both corpora lutea and a rat luteal cell line. Repression of PRAP/17βHSD7 expression and promoter activity by human chorionic gonadotropin/forskolin was localized to a −52-bp proximal segment of the promoter. This region contained a conserved CCAAT site and bound nuclear factor Y; binding of this transcription factor was inhibited by human chorionic gonadotropin in vivo. Furthermore, mutation of the nuclear factor Y site in the −52-bp promoter-reporter construct abolished forskolin-mediated inhibition of the promoter in a rat luteal cell line. In summary, we have identified the promoter elements involved in the basal expression of PRAP/17βHSD7. We have also found that LH-mediated repression of this gene is at the level of transcription and involves inhibition of nuclear factor YA binding to the CCAAT site within the proximal promoter.

Successful pregnancy requires profound differentiation and reorganization of the uterine tissues including, as pregnancy progresses, extensive apoptosis of decidual tissue to accommodate the developing conceptus. We have previously shown a positive correlation between expression of activin A and apoptosis in the decidua and have also shown that expression of activin A occurs at the time when prolactin (PRL) receptors disappear from decidual cells. The goals of this study were to examine whether activin A plays a role in decidual apoptosis and whether expression of activin A in the decidua is regulated by PRL and placental lactogens. Studies were carried out using primary rat decidual cells, a decidual cell line (GG-AD), and PRL null mice. Treatment of decidual cells with activin A significantly increased DNA degradation, caspase 3 activity, and caspase 3 mRNA expression. However, this effect was observed only in the absence of endogenous activin production by these cells. Addition of follistatin to decidual cells that were producing activin A decreased both caspase 3 activity and mRNA expression. Similarly, addition of activin-blocking antibodies to cultures of GG-AD cells, which also produce activin A, caused a reduction in both DNA degradation and caspase 3 activity. PRL and placental lactogens caused an inhibition of activin A mRNA expression in primary decidual cells. Even more convincingly, decidua of PRL null mice expressed abundant activin A at a time when no expression of this hormone is detected in wild-type mice and treatment of PRL null mice with PRL caused a profound inhibition of activin A mRNA expression. In summary, our investigations into the role and regulation of decidual activin have revealed that activin A can induce cell death in the decidua and that its expression is under tight regulation by PRL and placental lactogens. activin, apoptosis, decidua, prolactin

We investigated whether prolactin (PRL) treatments resembling the intermittent PRL surges of estrous cycles could induce luteal regression in hypophysectomized rats. Immature female rats were stimulated to ovulate and form corpora lutea with exogenous gonadotropins, and were hypophysectomized following ovulation. A single s.c. injection of either vehicle (VEH) or PRL was administered to each rat on post-hypophysectomy Day 8 and again on Day 11. The four resulting treatment groups consisted of rats that received two injections of VEH, VEH followed by PRL, PRL followed by VEH, or two injections of PRL. Rats were killed 24 or 72 h following the second injection. Plasma 20alpha-dihydroprogesterone, luteal weight, and total luteal protein were determined. One ovary was sectioned for immunohistochemistry for monocytes/macrophages, apoptotic nuclei, and major histocompatibility class II (MHC II) molecules. No effect of time (following injection) was observed on any endpoint, indicating that PRL does not have an ongoing regressive action. Time groups from within each treatment group were therefore pooled for analysis. Significant declines (P: < 0.05) in plasma concentrations of 20alpha-dihydroprogesterone, luteal weight, and protein per corpus luteum occurred only after two injections of PRL. Numbers of luteal monocytes/macrophages, apoptotic nuclei, and MHC II-positive cells were low in all groups; numbers of luteal monocytes/macrophages increased following two injections of PRL (P: < 0.05). We conclude that PRL has a cumulative regressive effect on the corpus luteum of the hypophysectomized rat. Drawing a parallel with the estrous cycle, we suggest that continued exposure to PRL, over several cycles, is necessary to induce full luteal regression.

During the estrous cycle, secretion of prolactin is largely restricted to a surge on proestrus. We investigated whether this proestrous prolactin surge initiates regression of the corpora lutea of the preceding cycle. Adult rats were killed prior to the prolactin surge (Proestrus group), following the prolactin surge (Estrus group), after chemical blockade of the prolactin surge with bromocryptine (Estrus1BRC group), and after blockade of the prolactin surge and administration of prolactin (Estrus1BRC1PRL group). Corpora lutea of the current (proestrus) or preceding (estrus) cycle were dissected out, weighed, and sectioned for immunohistochemistry or cultured for examination of in vitro progestin production. Numbers of luteal monocytes/macrophages, differentiated macrophages, and apoptotic nuclei per high-power field were greater for Estrus and Estrus1BRC1PRL than for Estrus1BRC, which in turn had greater numbers than Proestrus (P , 0.05). In contrast, BRC completely reversed the decline in luteal weight observed between Proestrus and Estrus (P , 0.05). Number of major histocompatibility complex II-positive cells was not different between groups (P . 0.05). Finally, progestin production by corpora lutea in vitro was lower for Proestrus than for the other groups (P , 0.05). The results indicate that the prolactin surge alone is not responsible for initiation of apoptosis or immune cell infiltration in regressing corpora lutea of the estrous cycle, although prolactin increases these markers of regression. Prolactin does cause a decline in luteal weight; however, the corpora lutea retain the capacity for steroidogenesis. We conclude that although prolactin has a role in luteal regression, it is not solely responsible for the initiation of this process.

We investigated the physiological basis for the trophic effect of glucocorticoids in rat corpora lutea in the absence of pituitary gonadotropins. Immature (Day 29) Sprague-Dawley rats were given eCG and hCG to induce the development of corpora lutea and were hypophysectomized on Day 32. Beginning on Day 40, rats received twice-daily s.c. injections of either dexamethasone (dex; 200 microg/rat/day) or vehicle (controls) and then were killed on Day 44. Plasma 20alpha-dihydroprogesterone, a major steroid produced by the corpora lutea, was higher (p 2-fold of plasma 20alpha-dihydroprogesterone concentration compared to controls. Glucocorticoid receptor protein (about 92 kDa) was detected in both luteal and nonluteal ovarian tissues in this animal model. These effects of glucocorticoids and the presence of the glucocorticoid receptor raise the possibility of a physiological role for glucocorticoids in the rat corpus luteum.

In hypophysectomized rats, prolactin induces regression of the corpora lutea. Luteal regression is accompanied by infiltration of monocytes/macrophages, declines in luteal mass and plasma progestins, and increased staining for monocyte chemoattractant protein-1 (MCP-1). We investigated whether similar events are induced during the estrous cycle, after the proestrous prolactin surge. Rats were killed on proestrus or on estrus, and one ovary was frozen for immunohistochemical detection of MCP-1, monocytes/macrophages (ED1-positive), and differentiated macrophages (ED2-positive) and for in situ detection of apoptotic nuclei. Corpora lutea of the current (proestrus) or preceding (estrus) cycle were dissected from the ovaries of additional rats and frozen for the same analyses and for determination of total protein content. In sections of whole ovaries, intensity and distribution of MCP-1 staining were increased in corpora lutea of multiple ages on estrus as compared to proestrus, as were numbers of differentiated macrophages and apoptotic nuclei per high-power field. Sections of isolated corpora lutea showed these increases on estrus, and the number of monocytes/macrophages per high-power field was also significantly increased. Accompanying these inflammatory/immune events, the corpora lutea on estrus showed decreased weight and total protein per corpus luteum, as compared to corpora lutea on proestrus. These changes are consistent with a proposed role for prolactin in the initiation of luteal apoptosis and of a sequence of inflammatory/immune events that accompany regression of the rat corpus luteum during the normal estrous cycle.

This article addresses the role of GnRH in ovulation in the context of two general models of GnRH action-deterministic and permissive. According to the deterministic model, increased GnRH secretion is required to induce the preovulatory LH surge and thus ovulation. The permissive model, in contrast, holds that GnRH secretion need not increase. Rather, the preovulatory LH surge results from enhanced sensitivity of the pituitary gland to GnRH. Studies in rodents and rabbits support the deterministic model whereas evidence in primates suggests that GnRH is permissive. Three lines of evidence are presented to support the conclusion that GnRH plays a deterministic role in sheep. First, a large GnRH surge is secreted together with the preovulatory LH surge. Second, the follicular phase increase in circulating estradiol concentration stimulates this GnRH surge by a positive feedback effect. Third, initiation of the LH surge requires an abrupt increase in GnRH, and maintenance of the LH surge requires continued GnRH support. Collectively, these observations document the fundamental importance of a GnRH surge to ovulation and generation of the estrous cycle of sheep.

Decreased suppressors of cytokine signaling (SOCS) activity in human gestational tissues may play a part in the onset/progression of term labor. Since SOCS proteins negatively regulate cytokine-mediated inflammatory processes, we hypothesized that SOCS proteins are elevated in gestational tissues from spontaneous preterm deliveries with intrauterine infection. SOCS1, −2 and −3 mRNAs and proteins were detectable by RT-PCR and immunoblotting respectively, in preterm amnion, choriodecidua and placenta, irrespective of infection status. Immunoperoxidase staining localized SOCS1, −2 and −3 to all cell types of the gestational membranes, with infiltrating leukocytes reacting strongly in infected tissues. In villous placenta, SOCS was immunolocalized to the syncytiotrophoblast with marked staining of round mesenchymal cells, possibly Hofbauer cells. Nuclear SOCS staining was seen in amnion, chorion and placental syncytiotrophoblasts. SOCS proteins were, in general, significantly more abundant in placenta compared with amnion or choriodecidua. Placental SOCS1 and interleukin-1β concentrations were positively correlated (r2=0.47; P

In the last 10 years many exciting discoveries have been made regarding the development, function, and regression of the corpus luteum (CL) of pregnancy. Techniques such as microarray analysis have enhanced the speed with which new genes of interest are identified, and trans-genic technology has revealed the frequently surprising functions of known and novel factors. The field of angiogenesis has blossomed, and the remarkably intense and rapid angiogenesis that occurs in the CL has made it an organ of choice for studies of this process. The cloning of steroid ogenic acute regulatory protein (StAR) has changed the way we think about steroidogenesis in the CL. Although StAR is no longer a novelty, the mechanism of its action continues to be a controversial topic and an exciting field of research. Similarly, the cloning of 17β-hydrox-ysteroid dehydrogenase type 7 has filled in one of the missing pieces of the steroidogenic puzzle in the CL of the rodent; however, more questions remain unanswered. A luteotropic role for progesterone, proposed long ago, has now been supported by numerous studies, and our knowledge of the mechanisms by which this and other hormones act on the CL continues to expand. In this chapter, we summarize the current body of knowledge, in particular the new information of the last 10 years of research, to present a complete picture of the CL of pregnancy. This fascinating organ, with its rapid formation and destruction and its complex regulation, continues to be a fertile field for research.

Successful pregnancy requires profound differentiation and reorganization of the uterine tissues including, as pregnancy progresses, extensive apoptosis of decidual tissue to accommodate the developing conceptus. We have previously shown a positive correlation between expression of activin A and apoptosis in the decidua and have also shown that expression of activin A occurs at the time when prolactin (PRL) receptors disappear from decidual cells. The goals of this study were to examine whether activin A plays a role in decidual apoptosis and whether expression of activin A in the decidua is regulated by PRL and placental lactogens. Studies were carried out using primary rat decidual cells, a decidual cell line (GG-AD), and PRL null mice. Treatment of decidual cells with activin A significantly increased DNA degradation, caspase 3 activity, and caspase 3 mRNA expression. However, this effect was observed only in the absence of endogenous activin production by these cells. Addition of follistatin to decidual cells that were producing activin A decreased both caspase 3 activity and mRNA expression. Similarly, addition of activin-blocking antibodies to cultures of GG-AD cells, which also produce activin A, caused a reduction in both DNA degradation and caspase 3 activity. PRL and placental lactogens caused an inhibition of activin A mRNA expression in primary decidual cells. Even more convincingly, decidua of PRL null mice expressed abundant activin A at a time when no expression of this hormone is detected in wild-type mice and treatment of PRL null mice with PRL caused a profound inhibition of activin A mRNA expression. In summary, our investigations into the role and regulation of decidual activin have revealed that activin A can induce cell death in the decidua and that its expression is under tight regulation by PRL and placental lactogens.

OBJECTIVE: Prolactin is capable of both trophic and lytic actions in rat corpora lutea. In corpora lutea responding to a trophic prolactin signal, the long form of the prolactin receptor is the dominant form and is upregulated by prolactin. We investigated whether mRNA for the short form of the prolactin receptor was dominant in corpora lutea responding to a lytic prolactin signal, and whether the relative concentrations of the mRNAs for both forms of the prolactin receptor were changed during this response. DESIGN AND METHODS: Immature rats were ovulated by injection of 5 IU equine chorionic gonadotrophin and 5 IU human chorionic gonadotrophin, and were hypophysectomized shortly after ovulation. Nine days after hypophysectomy, rats were injected with prolactin (500 microg/day) or vehicle for 24 (n=6, n=6) or 72 h (n=13, n=5). Total RNA was isolated from corpora lutea and mRNA for both types of prolactin receptor were analyzed by semiquantitative RT-PCR using the ribosomal protein S16 as the internal control. RESULTS: The intensities of the long- and short-form prolactin receptor signals were normalized to the S16 internal control and expressed as relative densitometric units. The normalized values at 24h for prolactin-treated vs vehicle-treated rats were 0.23 +/- 0.05 vs 0.49 +/- 0.15 (P>0.05) for the short form and 4.04 +/- 0.8 vs 4.23 +/- 0. 6 (P>0.05) for the long form. The values for 72 h were 0.30 +/- 0.05 vs 0.24 +/- 0.05 (P>0.05) for the short form and 2.76 +/- 0.4 vs 5. 53 +/- 0.3 (P

The preovulatory LH surge in the ewe is stimulated by a large sustained surge of GnRH. We have previously demonstrated that the duration of this GnRH signal exceeds that necessary to initiate and sustain the LH surge. The objective of the present study was to determine whether a similar excess exists for amplitude of the GnRH surge. Experiments were performed using an animal model in which GnRH secretion was blocked by progesterone, which in itself does not block the pituitary response to GnRH. To assess the amplitude of the GnRH surge needed to induce the LH surge, we introduced artificial GnRH surges of normal contour and duration but varying amplitudes. Twelve ewes were run through 3 successive artificial follicular phases (total of 36). Six of these artificial follicular phases were positive controls, in which progesterone was removed, the estradiol stimulus was provided, and vehicle was infused. In these control cycles, animals generated endogenous LH surges. In the remaining artificial follicular phases, progesterone was not withdrawn, the estradiol stimulus was provided, and either vehicle (negative control) or GnRH solutions of varying concentrations (experimental) were infused. The circulating GnRH concentrations achieved by infusion were monitored. No LH surges were observed in negative controls, whereas LH surges were induced in experimental cycles provided a sufficient dose of GnRH was infused. A highly significant dose-response relationship was observed between the amplitude of the GnRH surge and both the amplitude of the LH surge and the area under the curve describing the LH response, but no such relationship existed between the amplitude of the GnRH surge and the duration of the LH response. In numerous cases, LH surges similar to those in the positive control animals resulted from infusion of amounts of GnRH estimated to be considerably less than those delivered to the pituitary during the endogenously generated GnRH/LH surge. These findings indicate that, in the ewe, increased GnRH secretion drives the preovulatory LH surge in a dose-dependent fashion, and they provide evidence that the amplitude of the GnRH surge may exceed that needed to generate the LH surge.

The corpus luteum is a transient endocrine gland that is specialized for the production of progesterone and that plays a critical role in the establishment and maintenance of pregnancy. The life span of the corpus luteum varies between species and, within a species, can be dramatically altered by events such as mating or pregnancy. Regardless of the duration of its life span, the corpus luteum eventually enters a dynamic regression process during which it loses the capacity to produce progesterone and undergoes structural involution. The overall process of luteal regression has been referred to by a variety of terms over the last several decades. In Table ​Table1,1, we show the results of a cursory MEDLINE/PubMed search on luteal regression performed in November of 2002, which reveals that the most frequently utilized term for this process is "luteolysis," followed by "luteal regression," and then, to a lesser extent, by "functional luteolysis" and "structural luteolysis." Other terms, such as "luteal involution," "functional luteal regression," or "structural luteal regression" lag far behind in usage. The necessity of using several keywords to search for information on luteal regression is illustrated by the fact that in our sample MEDLINE/PubMed search, only 47% of the manuscripts retrieved using the keyword "luteal regression" were also retrieved when using the keyword "luteolysis." This inconsistency in terminology can result in serious under-retrieval and under-citation of papers using less popular keywords. Table 1 Results of a cursory MEDLINE/PubMed search performed on November 2002 using the terms as listed. To quote Rossdale and Cox in their communication entitled Terminology: a mark of scientific progress [1]: "Scientific terminology must be as exact as is possible within the state of knowledge available." As our knowledge of the molecular mechanisms participating in the process of luteal regression evolves, some of the commonly used terms cited above have been rendered obsolete, inappropriate, and even incorrect. The foremost of these is "luteolysis," still one of the most common terms used to define the process of luteal regression, most probably because it is listed among Medical Subject Headings (MeSH), and, therefore, used most frequently. MeSH is the U.S. National Library of Medicine's controlled vocabulary used for indexing articles in PubMed. MeSH terminology provides a consistent way to retrieve information that may use different terminology for the same concepts. In MeSH, "luteolysis" is defined as "degradation of corpus luteum" and further described as "...characterized by the involution and cessation of its endocrine function." From a cell biology point of view, "lysis" means "rupture of cell plasma membrane, leading to the release of cytoplasm and the death of the cell" [2]. If interpreted with strict accordance to the Latin, lysis refers to "disintegration," as recently discussed by Davis and Rueda [3]. Over the last decade, it has become clear that the involution of the corpus luteum is associated with a phenomenon of programmed cell death or apoptosis [4]. Because apoptosis is an organized process that does not involve a major inflammatory response, and because the majority of cells are removed prior to rupture, the term "lysis" when applied to luteal regression is largely inaccurate. Therefore, while the term luteolysis remains very popular – 1251 manuscripts were retrieved using the word "luteolysis" and only 403 were retrieved using the terms "luteal regression" in the MEDLINE/PubMed search (see Table ​Table1)1) – this term no longer accurately reflects the mechanisms involved in luteal regression. In 1999, McCraken et al. [5] stated that "the term luteolysis may be something of a misnomer;" but also referred to the deep entrenchment of this term in the literature. However, the exponential increase in the number of scientific manuscripts published in recent years ensures that a prompt move to replace the older term with a more accurate descriptor will rapidly result in an overturn of the current ranking displayed in Table ​Table11 and that only a minority of papers will continue to be retrieved using the older term. Despite the fact that a strict definition of "luteolysis" appears to regard only the structural disintegration of the corpus luteum, broader interpretations are rife within the literature. Rothchild [6] defined luteolysis as "stopping the secretion of progesterone by the corpus luteum." Two other recent reviews take a middle road. Hoyer [7] refers to the fall in progesterone secretion as functional regression but considers structural regression, or cell death, the "true" regression. Niswender et al. [8], while referring to luteal regression as the structural demise of the corpus luteum, go on to describe the loss of progesterone synthesis and secretion – not a structural event – as part of the regression of the corpus luteum. As the previous paragraph shows, the adjectives "structural" and "functional" cannot be overlooked when addressing terminology in the field of luteal regression. These words have had a consistent presence in literature concerning the regression of the corpus luteum since Malven and Sawyer [9] described the process of luteal regression in the rat as having two components: "1) an initial termination of luteal secretory function and 2) a subsequent morphological regression of the nonfunctioning corpora lutea." A few years later Malven et al. [10] used the term "structural luteolysis" to describe the morphological regression of persistent, nonfunctional corpora lutea in the hypophysectomized rat. The definition of "nonfunctional" provided in this manuscript expanded upon the 1966 description of the first component of luteal regression in the rat by explaining that the corpora lutea "did not secrete enough progesterone to induce deciduoma formation following endometrial traumatization." Thus, the corpora lutea were "nonfunctional" in terms of progesterone production and the potential for establishment of pregnancy. Since the appearance of these early papers, the terms functional and structural regression of the corpus luteum (or the more popular phrasing "functional and structural luteolysis") have gained relative acceptance and usage. The cursory MEDLINE/PubMed search shown in Table ​Table11 turned up 82 references containing the phrase "functional luteolysis," of which 5 were 2001 and 2002 publications, and 72 references containing the phrase "structural luteolysis," of which 14 were 2001 and 2002 publications. The prevalence of the term "structural luteolysis" over "functional luteolysis" in recently published articles may be due in part to a greater disparity in the perceived meaning of the latter term. This stems largely from studies in rats, in which it was revealed that corpora lutea which no longer secreted significant quantities of progesterone retained many functions, including active steroidogenesis [11-13]. It is our contention that these two frequently described components of the process of luteal regression (i.e. cessation of progesterone secretion and disappearance of the structure) merit equal attention. However, the timing and inter-relationship of these two types of regressive changes appears to vary by species. As new information on the molecular steps triggered during luteal regression is obtained, these mechanisms will need to be defined as they relate to these facets of the overall regressive process. Davis and Rueda [3] advised that the terms functional and structural used in the context of luteal regression should be adequately defined by the authors in the context of the particular species and reproductive stage being studied, with which we concur. It is not extreme to predict that with the advancement of knowledge in the process of luteal regression researchers will find increasing difficulties in establishing the limits as to what mechanisms belong to the "functional" facet of luteal regression, and which ones are part of the "structural" facet, as luteal regression is a process of synchronized events. For example, it now appears that progesterone itself is responsible for preventing the onset of apoptosis and structural disintegration [14-17].

A total of 142 seronegative volunteers were given one of the following rubella vaccines: Cendehill, HPV77. DE-5, RA27/3, or a new Japanese vaccine, To-336. To-336 vaccine produced a slightly higher geometric mean antibody titre (G.M.T.) (65.7) than did the HPV77. DE-5 (63.1) or RA27/3 vaccine (61.9) but the G.M.T. induced by Cendehill vaccine was much lower (39.3).Reactions, particularly joint symptoms, occurred least commonly after vaccination with To-336 vaccine. Joint symptoms occurred within seven days of menstruation in 30 out of 37 (81%) vaccines (P

Inflammatory cytokines secreted by the placenta and fetal membranes are believed to play an important role in the initiation of parturition. The suppressor of cytokine signaling (SOCS) proteins regulate signal transduction by several cytokines that have been reported to affect gestational tissues. The presence, distribution and roles of SOCS proteins, however, have not been described in human gestational tissues. Using reverse transcriptase (RT)-PCR and Western blot analysis we investigated the expression of SOCS1, SOCS2, and SOCS3 mRNA and protein, respectively, by human villous placenta, amnion and choriodecidua (n = 3-4). Tissues were obtained from uncomplicated pregnancies at term after either spontaneous labor and vaginal delivery or caesarean section (before labor). Messenger RNAs for SOCS1, SOCS2, and SOCS3 were expressed in all tissue types, irrespective of labor status. SOCS proteins were, however, only detectable in villous placenta and in one case in the choriodecidua. Labor was associated with abrogated expression of SOCS1 and SOCS3 proteins in villous placenta and the choriodecidua sample. Following labor the band for SOCS2 protein increased slightly in size which may indicate post-translational modification of SOCS2. Reduced expression of SOCS proteins in gestational tissues may provide a mechanism by which inflammatory cytokines enter into a positive feedback loop of inflammatory changes leading to delivery.