Possible involvement of IFNT in lymphangiogenesis in the corpus luteum during the maternal recognition period in the cow

The corpus luteum (CL), which secretes large amounts of progesterone and is thus essential for establishing pregnancy, contains various types of immune cells that may play essential roles in CL function by generating immune responses. The lymphatic system is the second circulation system and is necessary for immune function, but the lymphatic system of the bovine CL has not been characterized in detail. We collected bovine CLs on days 12 and 16 of the estrous cycle (C12 and C16) and days 16 and 40 of early pregnancy (P16 and P40). Lymphatic endothelial hyaluronan receptor 1 (LYVE1) protein was detected in the CL by immunohistochemistry and western blotting and increased at P40 compared with C16. The mRNA expression levels of lymphangiogenic factors, such as vascular endothelial growth factor-C (VEGFC), VEGFD, and their common receptor VEGFR3, as well as the lymphatic endothelial cell (LyEC) marker podoplanin, increased in P16 and P40 CLs. Thus, it is suggested that the lymphatic system of the bovine CL reconstitutes during early pregnancy. Interferon tau (IFNT) from the conceptus in the uterus is a candidate for activating luteal lymphangiogenesis during the maternal recognition period (MRP). We found that treatment of LyECs isolated from internal iliac lymphatic vessels with IFNT stimulated LyEC proliferation and significantly increased mRNA expression of VEGFC and IFN-stimulated gene 15. Moreover, both IFNT and VEGFC induced LyECs to form capillary-like tubes in vitro. In conclusion, it is suggested that new lymphangiogenesis in the bovine CL begins during the MRP and that IFNT may mediate this novel phenomenon.     

Expression and localization of apelin and its receptor APJ in the bovine corpus luteum during the estrous cycle and prostaglandin F2alpha-induced luteolysis

Angiogenesis, changes in blood flow, and extracellular matrix remodeling are the processes associated with the development and demise of the bovine corpus luteum (CL) during the estrous cycle. APJ (putative receptor protein related to angiotensin type 1 receptor) is a G-protein-coupled receptor, and its ligand, apelin, has been identified as a novel regulator of blood pressure and as an angiogenic factor. We hypothesized that the apelin-APJ system is involved in luteal function. This study investigated whether apelin-APJ exists in bovine CL and determined their expression profiles and localization during luteal phase and prostaglandin F(2)(alpha) (PGF(2)(alpha))-induced luteolysis. During the luteal phase, apelin mRNA expression increased from early to late CL and decreased in regressed CL. APJ mRNA expression increased from early to mid-CL and remained elevated in late and regressed CL. Apelin and APJ proteins were immunohistochemically detected only in the smooth muscle cells of intraluteal arterioles during the luteal phase. PGF(2)(alpha) stimulated apelin and APJ mRNA expression at 0.5-2 and 2 h respectively, and then the mRNA expression of apelin-APJ was inhibited from 4 h during PGF(2)(alpha)-induced luteolysis. Additionally, apelin mRNA and protein were stimulated at 1 h after PGF(2)(alpha) injection only in the periphery of mid- but not early CL. The present study indicated that the apelin-APJ was localized in the smooth muscle cells of intraluteal arterioles, and responded to PGF(2)(alpha) at the periphery of mid-CL in the cow. Thus, the apelin-APJ system may be involved in the maturation of CL and the luteolytic cascade as a regulator of intraluteal arterioles in cow.     

Fibroblast growth factor 2 is a key determinant of vascular sprouting during bovine luteal angiogenesis

Fibroblast growth factor (FGF) 2 and vascular endothelial growth factor (VEGF) A are thought to be key controllers of luteal angiogenesis; however, their precise roles in the regulation and coordination of this complex process remain unknown. Thus, the temporal and spatial patterns of endothelial network formation were determined by culturing mixed cell types from early bovine corpora lutea on fibronectin in the presence of FGF2 and VEGFA (6 h to 9 days). Endothelial cells, as determined by von Willebrand factor immunohistochemistry, initially grew in cell islands (days 0-3), before undergoing a period of vascular sprouting to display a more tubule-like appearance (days 3-6), and after 9 days in culture had formed extensive intricate networks. Mixed populations of luteal cells were treated with SU1498 (VEGF receptor 2 inhibitor) or SU5402 (FGF receptor 1 inhibitor) or control on days 0-3, 3-6 or 6-9 to determine the role of FGF2 and VEGFA during these specific windows. The total area of endothelial cells was unaffected by SU1498 treatment during any window. In contrast, SU5402 treatment caused maximal reduction in the total area of endothelial cell networks on days 3-6 vs controls (mean reduction 81%; P<0.001) during the period of tubule initiation. Moreover, SU5402 treatment on days 3-6 dramatically reduced the total number of branch points (P<0.001) and degree of branching per endothelial cell island (P<0.05) in the absence of changes in mean island area. This suggests that FGF2 is a key determinant of vascular sprouting and hence critical to luteal development.     

The class II major histocompatibility complex molecule BoLA-DR is expressed by endothelial cells of the bovine corpus luteum

Cells expressing class II major histocompatibility complex (MHC) molecules are found within the corpus luteum (CL) of several species. Expression and localization of class II MHC molecules in the bovine CL were examined in the present study. Immunohistochemical evaluation revealed class II MHC molecules on single cells in early CL (days 4 and 5 post-estrus). Two class II MHC-expressing cell types were observed in midcycle CL (days 10-12 post-estrus), single cells similar to those observed in the early CL, and endothelial cells. Not all endothelial cells expressed class II MHC, and further investigation revealed expression of only one type of class II MHC molecule, DR, on endothelial cells. Class II MHC was also localized to endothelial cells in late CL (day 18 post-estrus). Steroidogenic luteal cells were negative for class II MHC throughout the estrous cycle. Quantitative RT-PCR revealed higher (P < 0.05) concentrations of mRNA encoding the alpha-subunit of DR (DRA) in late CL when compared with those in the early CL. DRA mRNA abundance was also measured in cultures of mixed luteal and luteal endothelial (CLENDO) cells, in the presence or absence of tumor necrosis factor-alpha (TNF). No differences were found in the DRA mRNA concentration between mixed luteal and CLENDO cell cultures, and TNF had no effect on DRA mRNA concentration in both cell types. Expression of DR by endothelial cells of the midcycle CL may induce anergy of T lymphocytes, or stimulate them to secrete products that enhance normal luteal function.     

Expression and regulation of oestrogen receptors in the human corpus luteum

The molecular mechanisms underlying the control of corpus luteum lifespan in women are not fully understood. Oestradiol has various luteolytic, or luteotrophic, functions in some species, and as it is synthesised within the human corpus luteum, it is an excellent candidate molecule to be a paracrine regulator of luteal function. This study aimed to comprehensively investigate the expression, regulation and effects of oestrogen receptors (ER) in human luteal cells. Genomic oestrogen receptors ERalpha, ERbeta1 and ERbeta2 were immunolocalised in human corpora lutea from throughout the luteal phase. mRNA expression was investigated throughout the luteal phase and after luteal rescue with exogenous human chorionic gonadotrophin (hCG). The regulation of ER expression and oestradiol action was investigated in cultures of luteinised granulosa cells. ER subtypes ERbeta1 and ERbeta2 were localised throughout the luteal phase to steroidogenic cells in the human corpus luteum and cells of the surrounding stroma. Unlike follicular granulosa cells, steroidogenic cells in the corpus luteum showed minimal ERalpha immunostaining. The presence of endothelial cells in the granulosa cell layer with ERbeta1 and ERbeta2 positive nuclei was noted. ERbeta1 and ERbeta2 were differentially regulated across the luteal phase with ERbeta1 maximally expressed in the mid-luteal phase, while ERbeta2 expression was maximal in the early luteal phase. In vivo and in vitro, hCG had no long-term effect on ER expression, although in vitro hCG and oestradiol acutely down-regulated ERs. Treatment with oestradiol in vitro down-regulated 11beta-hydroxysteroid dehydrogenase type 1 and inhibin betaA subunit confirming a functional oestradiol response. These data highlight functional and differentially regulated oestradiol reception in human luteal cells.     

The expression, regulation and function of secreted protein, acidic, cysteine-rich in the follicle-luteal transition

The role of the tissue remodelling protein, secreted protein, acidic, cysteine-rich (SPARC), in key processes (e.g. cell reorganisation and angiogenesis) that occur during the follicle-luteal transition is unknown. Hence, we investigated the regulation of SPARC in luteinsing follicular cells and potential roles of SPARC peptide 2.3 in a physiologically relevant luteal angiogenesis culture system. SPARC protein was detected mainly in the theca layer of bovine pre-ovulatory follicles, but its expression was considerably greater in the corpus haemorrhagicum. Similarly, SPARC protein (western blotting) was up-regulated in luteinising granulosa but not in theca cells during a 6-day culture period. Potential regulatory candidates were investigated in luteinising granulosa cells: LH did not affect SPARC (P>0.05); transforming growth factor (TGF) B1 (P<0.001) dose dependently induced the precocious expression of SPARC and increased final levels: this effect was blocked (P<0.001) by SB505124 (TGFB receptor 1 inhibitor). Additionally, fibronectin, which is deposited during luteal development, increased SPARC (P<0.01). In luteal cells, fibroblast growth factor 2 decreased SPARC (P<0.001) during the first 5 days of culture, while vascular endothelial growth factor A increased its expression (P<0.001). Functionally, KGHK peptide, a SPARC proteolytic fragment, stimulated the formation of endothelial cell networks in a luteal cell culture system (P<0.05) and increased progesterone production (P<0.05). Collectively, these findings indicate that SPARC is intricately regulated by pro-angiogenic and other growth factors together with components of the extracellular matrix during the follicle-luteal transition. Thus, it is possible that SPARC plays an important modulatory role in regulating angiogenesis and progesterone production during luteal development.     

Prostaglandin F2alpha increases endothelial nitric oxide synthase in the periphery of the bovine corpus luteum: the possible regulation of blood flow at an early stage of luteolysis

Prostaglandin F(2)(alpha) (PGF(2)(alpha)) released from the uterus causes alterations in luteal blood flow, reduces progesterone secretion, and induces luteolysis in the bovine corpus luteum (CL). We have recently discovered that luteal blood flow in the periphery of the mature CL acutely increases coincidently with pulsatile increases in a metabolite of PGF(2)(alpha) (PGFM). In this study, we characterized changes in regional luteal blood flow together with regional alterations in endothelial nitric oxide synthase (eNOS) expression during spontaneous luteolysis and in response to PGF(2)(alpha). Smooth muscle actin-positive blood vessels larger than 20 microm were observed mainly in the periphery of mature CL. PGF(2)(alpha) receptor was localized to luteal cells and large blood vessels in the periphery of mid-CL. PGF(2)(alpha) acutely stimulated eNOS expression in the periphery but not in the center of mature CL. Injection of the NO donor S-nitroso-N-acetylpenicillamine into CL induced an acute increase in luteal blood flow and shortened the estrous cycle. In contrast, injection of the NOS inhibitor l-NAME into CL completely suppressed the acute increase in luteal blood flow induced by PGF(2)(alpha) and delayed the onset of luteolysis. In conclusion, PGF(2)(alpha) has a site-restricted action depending on not only luteal phase but also the region in the CL. PGF(2)(alpha) stimulates eNOS expression, vasodilation of blood vessels, and increased luteal blood flow in periphery of mature CL. Furthermore, the increased blood flow is mediated by NO, suggesting that the acute increase in peripheral blood flow to CL is one of the first physiological indicators of NO action in response to PGF(2)(alpha).     

A novel physiological culture system that mimics luteal angiogenesis

Luteal inadequacy is a major cause of poor embryo development and infertility. Angiogenesis, the formation of new blood vessels, is an essential process underpinning corpus luteum (CL) development and progesterone production. Thus, understanding the factors that regulate angiogenesis during this critical time is essential for the development of novel strategies to alleviate luteal inadequacy and infertility. This study demonstrates the development of a physiologically relevant primary culture system that mimics luteal angiogenesis. This system incorporates all luteal cell types (e.g. endothelial, steroidogenic cells, fibroblasts and pericytes). Using this approach, endothelial cells, identified by the specific marker von Willebrand factor (VWF), start to form clusters on day 2, which then proliferate and develop thread-like structures. After 9 days in culture, these tubule-like structures lengthen, thicken and form highly organized intricate networks resembling a capillary bed. Development of the vasculature was promoted by coating wells with fibronectin, as determined by image analysis (P<0.001). Progesterone production increased with time and was stimulated by LH re-enforcing the physiological relevance of the model in mimicking in vivo luteal function. LH also increased the area stained positively for VWF by twofold (P<0.05). Development of this endothelial cell network was stimulated by fibroblast growth factor 2 and vascular endothelial growth factor A, which increased total area of VWF positive staining on day 9, both independently (three- to fourfold; P<0.01) and in combination (tenfold; P<0.001). In conclusion, the successful development of endothelial cell networks in vitro provides a new opportunity to elucidate the physiological control of the angiogenic process in the developing CL.     

Molecular characterization of bovine placental and ovarian 20α-hydroxysteroid dehydrogenase

The enzyme 20α-hydroxysteroid dehydrogenase (20α-HSD) catalyzes the conversion of progesterone to its inactive form, 20α-hydroxyprogesterone. This enzyme plays a critical role in the regulation of luteal function in female mammals. In this study, we conducted the characterization and functional analyses of bovine 20α-HSD from placental and ovarian tissues. The nucleotide sequence of bovine 20α-HSD showed significant homology to that of goats (96%), humans (84%), rabbits (83%), and mice (81%). The mRNA levels increased gradually throughout the estrous cycle, the highest being in the corpus luteum (CL) 1 stage. Northern blot analysis revealed a 1.2 kb mRNA in the bovine placental and ovarian tissues. An antibody specific to bovine 20α-HSD was generated in a rabbit immunized with the purified, recombinant protein. Recombinant 20α-HSD protein produced in mammalian cells had a molecular weight of ～37 kDa. Bacterially expressed bovine 20α-HSD protein showed enzymatic activity. The expression pattern of the 20α-HSD protein in the pre-parturition placenta and the CL1 stage of the estrous cycle was similar to the level of 20α-HSD mRNA expression. Immunohistochemical analysis also revealed that bovine 20α-HSD protein was intensively localized in the large luteal cells during the late estrous cycle.     

Potential role for peroxisome proliferator-activated receptor gamma in regulating luteal lifespan in the rat

Peroxisome proliferator-activated receptor gamma (PPARgamma) has been shown to stimulate progesterone production by bovine luteal cells. We previously reported higher expression of PPARgamma in old compared with new luteal tissue in the rat. The following studies were conducted to determine the role of PPARgamma in rat corpora lutea (CL) and test the hypothesis that PPARgamma plays a role in the metabolism of progesterone and/or luteal lifespan. Ovaries were removed from naturally cycling rats throughout the estrous cycle, and pseudopregnant rats. mRNA for PPARgamma and P450 side-chain cleavage (SCC) was localized in luteal tissue by in situ hybridization, and protein corresponding to PPARgamma and macrophages identified by immunohistochemistry. Luteal tissue was cultured with agonists (ciglitazone, prostaglandin J2) or an antagonist (GW-9662) of PPARgamma. Progesterone was measured in media by RIA and levels of mRNA for 20alpha-hydroxysteriod dehydrogenase (HSD) and bcl-2 were measured in luteal tissue after culture by RT-PCR. An inverse relationship existed between the expression of mRNA for SCC and PPARgamma. There was no effect of PPARgamma agonists or the antagonist on luteal progesterone production in vitro, or levels of mRNA for 20alpha-HSD. PPARgamma protein was localized to the nuclei of luteal cells and did not correspond with the presence of macrophages. In new CL, ciglitazone decreased mRNA for bcl-2 on proestrus, estrus, and metestrus. Interestingly, GW-9662 also decreased mRNA for bcl-2 on proestrus and diestrus in old and new CL, and on metestrus in new CL. These data indicate that PPARgamma is not a major player in luteal progesterone production or metabolism but may be involved in regulating luteal lifespan.     

Characterization of endothelin-1 and nitric oxide generating systems in corpus luteum-derived endothelial cells

Endothelium-derived endothelin-1 (ET-1) and nitric oxide (NO) are pivotal regulators of corpus luteum (CL) function. To have a better insight into their synthesis and action, members of the ET system (ET-1, ET converting enzyme (ECE-1) isoforms a-d, ETA and ETB receptors) along with NO synthase (NOS) isoforms--endothelial (e)NOS and inducible (i)NOS--were quantified in CL-derived endothelial cells (CLEC). The expression of these genes in microvascular CLEC, obtained by lectin-coated magnetic beads, was compared with cells removed from the luteal microenvironment and maintained in culture for different durations, and with endothelial cells (EC) derived from a large blood vessel (i.e. bovine aortic endothelial cells, BAEC). The profile of gene expression in the different EC types was determined by quantitative real-time PCR. Freshly isolated EC from mid-cycle CL exhibited high ET-1 receptor expression (both ETA and ETB), low ET-1 synthesizing ability (both prepro (pp) ET-1 and ECE-1), but elevated iNOS - the high throughput NOS isoform. The distinct phenotype of CLEC was lost soon after an overnight culture. ETA and ETB receptor levels declined, ppET-1 levels increased while iNOS was reduced. These changes were extenuated during long-term culture of CLEC. The general pattern of gene expression in BAEC and long-term cultured CLEC was similar yet some differences, reminiscent of freshly isolated CLEC, remained: ECE-1c, ETB receptor and NOS isoforms were expressed differently in BAEC as compared with lines of CLEC. This study suggests that the luteal microenvironment is necessary to sustain the selective phenotype of its resident endothelial cells. The inverse relationship between ppET-1 and iNOS observed in freshly isolated CLEC and in cultured cells is physiologically significant and suggests that ET-1 and NO may modulate the production of each other.     

Vascular composition, apoptosis, and expression of angiogenic factors in the corpus luteum during prostaglandin F2alpha-induced regression in sheep

Corpora lutea and blood samples were collected from superovulated ewes 0, 4, 8, 12 and 24 h after prostaglandin F(2alpha) (PGF) analog injection on day 10 of the estrous cycle. Changes in vascular cell and fibroblast composition, apoptosis and mRNA expression for several angiogenic factors in the corpus luteum (CL) were determined. While peripheral progesterone concentration decreased at 24 h after PGF injection, CL weight did not change. The area of positive BS-1 lectin staining (endothelial cell marker), smooth muscle cell actin (SMCA; pericyte and SMC marker), collagen type 1 (fibroblast marker), and the rate of cell death changed in luteal tissues after PGF treatment. In association with these cellular changes, mRNA for several angiogenic factors including vascular endothelial growth factor (VEGF) and receptors (Flt and KDR), basic fibroblast growth factor (FGF2) and receptor, angiopoietin (ANGPT) 1 and receptor Tie-2, endothelial nitric oxide synthase (NOS3), and angiotensin II receptor 1 (AT1) were altered. Changes in endothelial cell marker expression were positively correlated with changes in VEGF and NO systems. In addition, changes in mRNA expression for VEGF, Flt and KDR were positively correlated with changes in ANGPT2, Tie-2, and NOS3, indicating a functional relationship. This data demonstrates that after an initial increase, the endothelial component of the vascular bed decreases during PGF-induced luteal regression. However, SMCA expression remained high during luteal regression, potentially indicating a role of pericytes and vascular SMC in luteolysis, likely to regulate tissue remodeling and to maintain the integrity of larger blood vessels. Further, it appears that early regression may increase collagen type 1 production and/or expression by fibroblasts. Expression of angiogenic factors is influenced by PGF-induced luteolysis and may serve to maintain vascular structure in order to aid luteal regression.     

KIT receptor-positive cells in the bovine corpus luteum are primarily theca-derived small luteal cells

The tyrosine kinase KIT receptor, the protooncogene CD117, plays a key role in growth and maturation of oocytes and follicles. Relevant data are sparse for the corpus luteum (CL). We first confirmed the presence of KIT mRNA and KIT protein in bovine CL homogenates. We then localized KIT-positive (KIT+) cells in CL sections by immunohistochemistry. At the CL stage of early development, the former theca transforming into capsule/septa showed a strong band-like KIT+ immunoresponse. In addition, CD45+ leukocytes in septa included subpopulations of CD45+/KIT+ and CD14+/KIT+ leukocytes as validated by double immunofluorescence localization. At the early secretory stage, KIT+ cells appeared within the septa/capsule region and in the periphery of the CL parenchyma, there forming a complex network. This was separate from the capillary bed as determined by double staining for CD117 and FVIII-related endothelial cell antigen (FVIIIr). The KIT+ network coincided with cells positive for cytochrome P450 17alpha-hydroxylase, a thecal cell-specific enzyme. The late secretory stage was defined by an advanced manifestation of the KIT+ network in the CL periphery. At the stage of regression, the KIT+ network was absent. The CL of pregnancy expressed high levels of KIT mRNA and KIT protein uniformly throughout pregnancy. The KIT+ immunolocalization revealed small fibroblast-like cells, luteal cells with granules, and clusters of large luteal cells with staining of the cell membrane. We conclude that a majority of KIT+ cells in the bovine CL are primarily theca-derived small luteal cells, and that a minority represent KIT+ leukocytes, in some cases KIT+ monocytes.     

Changes in vascular leakage and expression of angiopoietins in the corpus luteum during pregnancy in rats

The present study investigates changes in blood vessel stability and its regulation in the corpus luteum (CL) during pregnancy in the rat. First, blood vessel stability in the CL was evaluated during pregnancy based on vascular leakage, which was quantified by the Evans blue assay. Vascular leakage was highest on day 3, thereafter decreased until day 15 and increased again on day 21. Secondly, to study the regulation of vascular leakage, the expression of angiopoietins was examined in the CL during pregnancy. Angiopoietin-1 (Ang-1) effects maturation and stabilization of newly formed blood vessels, while Ang-2 produces the opposite effect by allowing vascular remodeling. An immunohistochemical study showed both Ang-1 and Ang-2 expression in luteal cells. mRNA and protein levels of Ang-1 were significantly higher on days 12 and 15 than those on days 3 and 21, whereas there was no significant change in Ang-2 expression. Since estradiol contributes to CL development during mid-pregnancy, we finally studied whether estradiol regulates vascular leakage and angiopoietin expression. Rats undergoing hypophysectomy and hysterectomy (hypox-hect) on day 12 were treated with estradiol until day 15. Vascular leakage was increased and Ang-1 expression was decreased by hypox-hect, and these effects were completely reversed by estradiol treatment. In conclusion, blood vessel stability in the CL is likely to be associated with CL development and CL regression, and may be regulated by angiopoietins. Estradiol contributes to blood vessel stabilization in the CL during mid-pregnancy, which is associated with an increase in Ang-1 expression.     

Up-regulation of expression of interferon-stimulated gene 15 in the bovine corpus luteum during early pregnancy

Interferon-τ (IFNT), the pregnancy recognition signal in ruminant species, is secreted by conceptus trophectoderm cells and induces expression of IFN-stimulated gene 15 (ISG15) in the uterus and corpus luteum (CL) in ewes. Expression of ISG15 in ovine CL is speculated to be through an endocrine pathway, but it is unclear whether expression of ISG15 in bovine CL is via such a pathway. In this study, CL were obtained from cows on d 16, 25, 60, 120, 180, and 270 of pregnancy, and endometrium, mammary gland, ovarian stroma, and CL were also collected from cows on d 18 of pregnancy and on d 15 and 18 of the estrous cycle. All tissue explants from d 15 of the estrous cycle were cultured in the absence or presence of 100 ng/mL of recombinant bovine IFNT for 24 h. The results indicated that ISG15 and conjugated proteins were expressed in CL of both cyclic and pregnant cows regardless of pregnancy status and were upregulated during early pregnancy. The mammary gland from d 18 of pregnancy did not express ISG15, but explants of the mammary gland from d 15 of the estrous cycle did express ISG15 after being treated with IFNT. However, luteal explants from d 15 of the estrous cycle did not express ISG15 after being cultured for 24 h. In conclusion, ISG15 expression is upregulated in the bovine CL during early pregnancy. Interestingly, cultured CL cells do not respond to IFNT, suggesting that the pregnancy-dependent stimulation of ISG15 expression is controlled by something other than IFNT in the bloodstream.     

Dickkopf-1 secreted by decidual cells promotes trophoblast cell invasion during murine placentation

To investigate the physiological characteristics of the corpus luteum (CL) of pregnancy, we raised a mAb, human corpus luteum (HCL)-4, against human luteal cells obtained from CL of pregnancy. The affinity-purified antigen from human CL of pregnancy or placenta using HCL-4 was a 61 kDa protein. The partial amino acid sequence of the antigenic protein was identical to that of human monoamine oxidase A (MAOA, EC1.4.3.4). MAOA has been shown to catabolize catecholamines that were reported to regulate luteal function in CL and vasoconstriction in various organs. Immunohistochemistry using HCL-4 mAb showed that MAOA was intensely expressed on large luteal cells and moderately expressed on small luteal cells in the CL of pregnancy. In the CL of menstrual cycle, MAOA was weakly detected on large luteal cells but not detected at all on small luteal cells. Western blotting analysis confirmed the high expression of MAOA in CL of pregnancy. Northern blot analysis also showed the expression of MAOA mRNA in human CL, and showed that its expression was higher in CL of pregnancy than in CL of menstrual cycle. The increased expression of MAOA in the CL of pregnancy suggests the contribution of MAOA to the function of the CL of pregnancy.     

Regulation and manipulation of angiogenesis in the primate corpus luteum

Intense physiological angiogenesis occurs during the early stages of luteal development, providing a model in which the complex processes regulating the angiogenic pathway may be studied. Here, a working hypothesis is presented to explain the diverse changes in the vasculature of the corpus luteum that occur over a short period, based around changes in vascular endothelial growth factor, the angiopoietins and matrix metalloproteinases. An illustration is given of how angiogenesis can be monitored in a primate model and how the role of individual angiogenic factors such as vascular endothelial growth factor may be explored in vivo. Because of the marked effect of inhibition of angiogenesis on luteal function, it is predicted that the normal processes of follicular development, ovulation and luteal function could all be profoundly influenced by the manipulation of angiogenesis.