[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to document the expression and localization of VEGF system comprising of VEGF isoforms (VEGF 120, VEGF 164 and VEGF 188) and their receptors (VEGFR1 and VEGFR2) in buffalo corpus luteum (CL) obtained from different stages of the oestrous cycle. Real-time RT-PCR (qPCR), Western blot and immunohistochemistry were applied to investigate mRNA expression, protein expression and localization of examined factors. In general, all the components of VEGF system (the VEGF isoforms and their receptors) were found in the water buffalo CL during the oestrous cycle. The mRNA as well as protein expression of VEGF system was highest during the early and mid-luteal phase, which later steadily decreased (p < 0.05) after day 10 to reach the lowest level in regressed CL. As demonstrated by immunohistochemistry, VEGF protein was localized predominantly in luteal cells; however, VEGFR1 and VEGFR2 were localized in luteal cells as well as in endothelial cells. In conclusion, the dynamics of expression and localization of VEGF system in buffalo corpora lutea during the luteal phase were demonstrated in this study, indicating the possible role of VEGF system in the regulation of luteal angiogenesis and proliferation of luteal as well as endothelial cells through their non-angiogenic function.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to evaluate mRNA expression, protein concentration and localization of the assumedly important lymphangiogenic factors VEGFC and VEGFD and the receptor FLT4 in bovine corpora lutea (CL) during different physiological stages. In experiment 1, CL were collected in a slaughterhouse and stages (days 1-2, 3-4, 5-7, 8-12, 13-16, >18) of oestrous cycle and month <3, 3-5, 6-7 and >8 of pregnancy. In experiment 2, prostaglandin F2α (PGF)-induced luteolysis was performed in 30 cows, which were injected with PGF analogue on day 8-12 (mid-luteal phase), and CL were collected before and 0.5, 2, 4, 12, 24, 48 and 64 h after PGF injection. The mRNA expression was characterized by RT-qPCR. All three factors were clearly expressed and showed significant changes during different groups and periods examined in both experiments. Protein concentrations of VEGFD and FLT4 measured by ELISA were not detectable in early cyclic CL but increased to higher plateau levels during pregnancy. After PGF-induced luteolysis FLT4 protein showed an increase within 2-24 h after the injection. FLT4 localization by immunohistochemistry in the cytoplasm of luteal cells was relatively weak in early CL. It increased in late CL and especially in CL during pregnancy. During pregnancy, a positive FLT4 staining in both the nucleus and cytoplasm of lymphatic endothelial cells in peripheral tissue was observed. In conclusion, our results lead to the assumption that lymphangiogenic factors are produced and regulated in CL and may be involved in mechanisms regulating CL function, especially during pregnancy.
[Show abstract][Hide abstract] ABSTRACT: Eosinophilic cells accumulate in the capillaries of the bovine Graafian follicle shortly before ovulation and in the early developing corpus luteum (CL). Suppressing the migration of these eosinophilic cells by dexamethasone allowed us to evaluate their possible function in the CL development. Brown Swiss cows (n = 10) were randomly subdivided into two groups (n = 5). Every group was used once as control group and once as experimental group with two oestrous cycles between each treatment. Eighteen hours (h) after oestrus synchronization, dexamethasone or saline was given. Ovulation was induced 24 h later with gonadotropin-releasing hormone. Another injection of dexamethasone or saline was given 12 h later. Eosinophilic cells in the blood were counted daily until day 7 after the first dexamethasone injection. The collection of ovaries took place at days 1, 2 and 5. Gene expression, protein concentration and location of angiogenic factors, chemokines, insulin-like growth factor 1 (IGF1) and eosinophilic cells were studied. No eosinophilic cells were found in the CL of the treatment group. Blood progesterone decreased significantly in the dexamethasone group from day 8 to 17. The protein concentration of FGF2 increased significantly in CL tissue at day 2 and VEGFA decreased. Local IGF1 gene expression in the CL was not regulated. We assume from our data that the migration of eosinophilic cells into the early CL is not an essential, but an important stimulus for angiogenesis during early CL development in cattle.
[Show abstract][Hide abstract] ABSTRACT: Blood plasma levels of the hormones bovine growth hormone (bGH), insulin and insulin-like growth factor 1 (IGF-1) and the metabolites glucose, free fatty acids (FFA), urea, creatine and creatinine in 70 bulls (Fleckvieh, Braunvieh, Grauvieh) kept at four AI stations were measured. Plasma determinations were performed on day 1 with normal feeding (N), day 4 after fast (F), and day 5 (R) after realimentation. Fleckvieh bulls had lower bGH but higher IGF-1 levels than Braunvieh bulls; young bulls had lower IGF-1 and insulin but higher glucose levels than mature bulls. The effect of fasting led to higher plasma levels of FFA, urea and creatinine and to lower levels of bGH and IGF-1. Correlations between plasma levels and breeding values for yield of milk, fat and protein were estimated for mature bulls. All three breeding values correlated positively with bGH and negatively with insulin, while correlations with other indicators depended on the feeding status of bulls. Prediction equations for breeding values which were limited to three indicators, included FFA, urea, and creatinine, and the r2 values ranged up to 0.32. Validation of these equations, derived from Fleckvieh bulls, on Braunvieh and Grauvieh bulls resulted in correlations of about 0.2. The use of indicators, in addition to information from relatives (dam, full and half-sisters), improves the accuracy of selection of young bulls.ZusammenfassungBeziehungen zwischen Plasmahormon und -metabolitenspiegel in MilchzuchtwertenVon 70 Bullen der Rassen Fleckvieh, Braunvieh und Tiroler Grauvieh aus vier Besamungsstationen wurden die Blutkonzentrationen der Hormone bGH, IGF-1 und Insulin sowie die der Metaboliten Glucose, Freie Fettsäuren (FFA), Harnstoff, Kreatin und Kreatinin während normaler Fütterung (N), nach einer Fastenperiode (F) und nach Wiederfütterung (R) bestimmt. Fleckviehbullen hatten signifikant mehr bGH und IGF-1 als Braunviehbullen, Jungbullen signifikant weniger IGF-1 und Insulin, aber mehr Glucose als ausgewachsene Bullen. Fasten der Tiere führte zu signifikant höherem Blutspiegel an Freien Fettsäuren, Harnstoff und Kreatin und zu niedrigerem Blutspiegel an bGH und IGF-1. Korrelationen zwischen Blutserumspiegel und Zuchtwert für Milchmenge, Fettmenge und Proteinmenge wurden geschätzt. bGH hatte eindeutig einen positiven, Insulin einen negativen Zusammenhang zu alien Zuchtwerten, während er bei den anderen Blutparametern je nach Fütterungsstatus unterschiedlich war. Schätzungsgleichungen für Zuchtwerte aus drei Blutwerten wurden erstellt, die Freie Fettsäuren, Harnstoff und Kreatinin berücksichtigten. Validierungen der Schätzungsgleichungen für Fleckvieh an Braunvieh und Grauvieh ergaben Korrelationen von etwa 0,20. Die Wirksamkeit der Verwendung der Indikatoren zur Zuchtwertschätzung eines Jungbullen-und MOET-Nukleus-Programm wurde untersucht und dabei jeweils die Genauigkeit der Schätzung aus Verwandten (Mutter, Voll-bzw. Halbschwestern) deutlich erhöht.
Journal of Animal Breeding and Genetics 03/2011; 112(1‐6):313 - 326. DOI:10.1111/j.1439-0388.1995.tb00573.x · 1.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Despite their economic and cultural importance, dromedary camel is considered as a slow breeding animal, because of the higher incidence of early embryonic death. The present study was designed to investigate: 1) Expression and cloning of progesterone receptors (PR) and oestradiol receptor α (ERα) in CL and endometrium of pregnant camel; 2) Detection of interferon stimulated gene 15 (ISG15) in corpus luteum (CL) and endometrium of pregnant dromedary camels. For PR and ERα, RNA was extracted from CL and endometrium of dromedary camels during early (1 to 3 months), mid (4 to 9 months), and late stage (10 to 13 months) of pregnancy. Messenger RNA expression of PR and ERα was performed using RT-qPCR. Detection of ISG15 was performed using immunohistochemistry and Western blot analysis. In CL, both PR and ERα±showed the same pattern with significantly high (P<0.01) expression during early stage compared to mid or late stages of pregnancy. The lowest (P<0.01) expression was detected during the late stage of pregnancy compared with the mid stage. There was no difference inmRNA expression for PR and ERα in endometrium of during the different stages of pregnancy in dromedary camels. ISG15 conjugated protein showed no expression in CL or endometrium of pregnant dromedary camels either by immunohistochemistry or Western blot. In conclusion, PR and ERα potentially play a role in regulating luteal function in CL during pregnancy in dromedary camels, further work is necessary to study the mechanism of pregnancy recognition in dromedary camels.
Reproduction Fertility and Development 01/2011; 23(1):160. DOI:10.1071/RDv23n1Ab110 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The objective of this study was to assess the effect of a change in the social composition in a group of red deer males on the relationship between their rank and testosterone. A group of twelve adult red deer males (Cervus elaphus) was tested in two social settings. From April 15 to June 9 (Period 1) this group was kept separately in an enclosure. On June 10, nine 3-year-old males were added to that group of adult males. They were kept together until August 31. We performed 10 observations of the group when the agonistic interactions of the males were recorded and we took 9 blood samples per male in Period 1; 11 observations were made and 10 samples were taken in Period 2. Concentrations of testosterone and cortisol were later determined in plasma. Adding much younger and smaller sparring partners into the experimental group of adult males in Period 2 altered the agonistic behaviour of the adults even though this did not trigger any change in rank position of the experimental males except one. Adult males targeted preferentially their attacks on individuals much lower in the hierarchy. Experimental male deer with higher social rank had lower levels of testosterone in Period 1; in Period 2 it was just the opposite. In Period 1 the animals had higher cortisol levels than in Period 2. As controls we used four adult (5years old) males sharing the enclosure with four 3-year-old males. No changes in hormone concentrations were observed in the control group. Thus, changing the social environment of adult red deer males resulted in change of the relationship between rank and testosterone and cortisol concentrations.
[Show abstract][Hide abstract] ABSTRACT: Development of the corpus luteum (CL) in ruminants occurs in a rapid and time-dependent manner within 1 week after ovulation, with morphologic and biochemical changes in the cells of the theca interna and granulosa cells of the preovulatory follicle. These changes involve luteinisation of steroidogenic cells and angiogenesis to establish normal luteal function (progesterone secretion). The CL is composed of a large number of vascular endothelial cells, large and small steroidogenic luteal cells, smooth muscle cells, pericytes, fibrocytes and immune cells, indicating that the CL is a heterogeneous tissue. Moreover, the CL produces and secretes growth factors (fibroblast growth factor, vascular endothelial growth factor and insulin-like growth factor), vasoactive factors (nitric oxide, angiotensin II and endothelin-1), steroids (progesterone is important for its own production), oxytocin and prostaglandins (PGF2alpha and PGE2) to regulate luteal formation and development. Clearly, the main function of the CL is to produce progesterone, which is a prerequisite for survival of the embryo, implantation and maintenance of pregnancy. Inadequate luteinisation and angiogenesis during the early luteal phase results in poor progesterone secretion and causes compromised embryo development and reduced fertility. Secretion of adequate amounts of progesterone during luteal development requires "precise luteinisation" of theca and granulosa cells to form luteal cells, neovascularization, and the establishment of a blood supply (angiogenesis). PGF2alpha in the developing CL acts as a local regulator to enhance progesterone secretion directly and indirectly by stimulating angiogenic factors, VEGF and FGF2. The preceding role of PGF2alpha may explain why the developing CL does not acquire luteolytic capacity until several days following ovulation. The balance between luteotrophic and luteolytic factors as well as stimulation and inhibition of angiogenic factors during luteal formation, development and maintenance can have a profound effect on the fate of the CL.
[Show abstract][Hide abstract] ABSTRACT: Thrombopoietin (TPO) is known to be involved in megakaryocytopoiesis, but its role in the control of ovarian function is unknown in cattle. The aims of this study were to demonstrate the expression of TPO and its receptor (c-MPL) in detail in bovine corpus luteum (CL) obtained from different stages of the oestrous cycle and during pregnancy--and to demonstrate that TPO/c-MPL system is expressed clearly in bovine follicles. Real-time RT-PCR (qPCR) and ELISA were applied to investigate mRNA expression of examined factors and TPO protein, respectively. In this investigation, increases in the concentrations of TPO protein and the mRNA expression of TPO and c-MPL were noticed during both early luteal stage and late luteal stage of the oestrous cycle. Furthermore, the expression of TPO/c-MPL system does not show any significant regulation in the CL throughout pregnancy. Highest co-expression of TPO/c-MPL system in both theca interna (TI) and granulosa cells (GC) in small follicles (<10 mm in diameter) was observed in this study that may suggest the possible role of TPO/c-MPL system in proliferation of TI and GC cells. To conclude, the results demonstrate the possible involvement of locally produced TPO/c-MPL system as a 'physiological filter' in bovine ovary where they may promote cell selection by inducing proliferation of viable cells and scavenging non-viable cells and thereby may play an important role in modulation of ovarian function.
[Show abstract][Hide abstract] ABSTRACT: Seminal fluids introduced to the female reproductive tract at mating can affect subsequent events, such as ovulation, fertilization, conception, and pregnancy. Bioactive molecules present in seminal plasma can modify the cellular composition, structure, and function of local tissues and of tissues distal to the tract. The oviduct plays a decisive role in reproduction providing a beneficial milieu for gamete maturation, fertilization, and early embryonic development. Therefore we have investigated whether intrauterine infusion of seminal plasma can modulate prostaglandin (PG) synthesis in the porcine oviduct through regulation of gene and protein expression of enzymes of prostaglandin synthesis pathway. Among several enzymes involved in the prostaglandin synthesis pathway tested in the present study PGF(2α) synthase (PTGFS) and prostaglandin 9-ketoreductase (CBR1), which convert PGE(2) to PGF(2α), expression were significantly down-regulated in the oviducts on Day 1 after seminal plasma infusion into the uterine horns. The effects of the treatment were transient and by Day 5 levels of PTGFS and CBR1 were comparable in seminal plasma-treated and control animals. Additionally, increased PGE(2) to PGF(2α) and PGFM to PGF(2α) ratios in the oviductal tissues were indicated. Our results clearly demonstrate that seminal plasma affects prostaglandin synthesis in the porcine oviduct. Altered PTGFS and CBR1 expression in consequence changed PGE(2) to PGF(2α) and PGFM to PGF(2α) ratios in the porcine oviduct.
[Show abstract][Hide abstract] ABSTRACT: To cope with rising demands for increased blood supply during pregnancy, the vasculature of the uterus undergoes several adaptive changes, including increased permeability, angiogenesis and vasodilatation. Although it is clear that vascular endothelial growth factor (VEGF) plays a paramount role in achieving these adaptations, little is known about regulation of VEGF expression in endometrium during pregnancy. Thus, we have investigated whether luteinizing hormone (LH) and tumour necrosis factor-alpha (TNFalpha) may affect VEGF secretion by stromal cells during early pregnancy in pigs. Real-time reverse transcription/polymerase chain reaction (RT/PCR) of VEGF120 and VEGF164 gene expression revealed significantly higher levels of VEGF164 mRNA in cultured stromal cells (p < 0.0001). The LH-stimulated secretion of VEGF was detected after 24 and 48 h of treatment when doses 50 and 100 ng/ml were used (p < 0.05 and p < 0.01, respectively). The TNFalpha-induced secretion of VEGF by stromal cells was detected only after 24-h treatment with the highest dose used in the experiment (50 ng/ml; p < 0.05). Although the influence of LH on VEGF secretion was more visible compared with TNFalpha, both factors may be considered as potential modulators of adaptive changes in uterine vasculature occurring during pregnancy in the pig.
[Show abstract][Hide abstract] ABSTRACT: The essential role of endometrial prostaglandin F2 alpha (PTGF) for induction of the corpus luteum (CL) regression is well documented in the cow. However, the acute effects of PTGF on known local luteotropic factors (oxytocin [OXT] and its receptor, insulin-like growth factor [IGF] 1, and progesterone and its receptor), the principal angiogenic factor vascular endothelial growth factor (VEGF) A and the capillary destabilization factor angiopoietin (ANGPT) 2 were not thoroughly studied in detail. The aim of this study was therefore to evaluate the tissue concentration of these factors during PTGF induced luteolysis. In addition the mRNA expression of progesterone receptor (PGR), OXT receptor (OXTR), IGF1, IGFBP1, ANGPT1, and ANGPT2 was determined at different times after PTGF treatment. Cows (n = 5 per group) in the mid-luteal phase (Days 8-12, control group) were injected with the PTGF analog (cloprostenol), and CL were collected by transvaginal ovariectomy at 0.5, 2, 4, 12, 24, 48, and 64 h after injection. The mRNA expression was analyzed by quantitative real-time PCR, and the protein concentration was evaluated by enzyme immunoassay or radioimmunoassay. Progesterone concentrations, as well as mRNA expression of PGR, in CL tissue were significantly down-regulated by 12 h after PTGF. Tissue OXT peptide and OXTR mRNA decreased significantly after 2 h, followed by a continuous decrease of OXT mRNA. IGF1 and VEGFA protein already decreased after 0.5 h. By contrast, the IGFBP1 mRNA was up-regulated significantly after 2 h to a high plateau. ANGPT2 protein and mRNA significantly increased during the first 2 h, followed by a steep decrease after 4 h. The acute decrease of local luteotropic activity and acute changes of ANGPT2 and VEGFA suggest that modulation of vascular stability may be a key component in the cascade of events leading to functional luteolysis.
Biology of Reproduction 05/2010; 82(5):940-7. DOI:10.1095/biolreprod.109.076752 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The pattern and regulation of endothlin-2 (EDN2) expression and its putative roles in bovine ovaries were investigated. EDN2 mRNA was determined in corpus luteum (CL) and during folliculoluteal transition induced by GnRH in vivo. EDN2 was elevated only in the early CL and was not present in older CL. In the young CL, EDN2 mRNA was identified mainly in luteal cells but not endothelial cells that expressed the EDN1 gene. Similarly, in preovulatory follicles, EDN2 was expressed in the granulosa cells (GCs) and not in the vascular theca interna. LH and hypoxia are two major stimulants of CL formation. Therefore, GCs were cultured with bovine LH, under hypoxic conditions. GCs incubated with bovine LH resulted in increased EDN2 mRNA 42 h later. CoCl2, a hypoxia-mimicking agent, elevated EDN2 in GCs in a dose-dependent manner. Incubation of the human GC line (Simian virus 40 large T antigen) under low oxygen tension (1%) augmented EDN2 6 and 24 h later. In these two cell types, along with EDN2, hypoxia augmented VEGF. EDN2 induced in GCs changes that characterize the developing CL: cell proliferation as well as up-regulation of vascular endothelial growth factor and cyclooxygenase-2 (mRNA and protein levels). Human chorionic gonadotropin also up-regulated these two genes. Small interfering RNA targeting EDN-converting enzyme-1 effectively reduced its mRNA levels. This treatment, expected to lower the mature EDN2 peptide production, inhibited VEGF mRNA levels and GC numbers. Together these data suggest that elevated EDN2 in the early bovine CL, triggered by LH surge and hypoxia, may facilitate CL formation by promoting angiogenesis, cell proliferation, and differentiation.
[Show abstract][Hide abstract] ABSTRACT: ZusammenfassungEinfluß von Geschlecht und Gewicht auf die Plasmakonzentrationen von Wachstumshormon (GH), Insulin-like growth factor I (IGF-I) und Insulin bei Mastrindern der Rasse Deutsches FleckviehBei wachsenden Bullen, Ochsen und Färsen (Deutsches Fleckvieh) wurden die Blutplasma-Konzentrationen von Wachstumshormon (GH), Insulin-like-growth-factor I (IGF-I) und Insulin bestimmt. Der Versuch bestand aus drei Versuchsdurchgängen und begann bei einem durchschnittlichen Alter von 3 Monaten beziehungsweise einem Lebendgewicht von 110 kg. Das Mastendgewicht betrug 510 kg (Färsen) und 660 kg (Bullen, Ochsen). Während der Mast wurden in jedem Versuchsdurchgang jeweils drei Tiere bei Mastgewichten von 200 kg, 360 kg und 425 kg (Färsen) bzw. 200 kg, 360 kg, 510 kg und 585 kg (Bullen, Ochsen) geschlachtet. Die Blutproben wurden durch Punktion in 14-tägigem Abstand gewonnen. Die GH-, IGF-I- und Insulin-Konzentrationen waren durch das Geschlecht signifikant beeinflußt (p<0.05). Die höchsten GH-und IGF-I-Konzentrationen waren bei den Bullen gefolgt von den Ochsen und Färsen festzustellen. Die GH-Konzentrationen zeigten einen gewichtsabhängigen Abfall bei Ochsen und Färsen, während die GH-Konzentrationen bei Bullen wahrscheinlich aufgrund von Streß nur tendenziell mit zunehmendem Gewicht abfielen. Mit zunehmendem Gewicht stiegen die IGF-I-und Insulin-Konzentrationen an. Die Veränderungen des Blutplasmaspiegels der Hormone werden hinsichtlich der Geschlechts- und Wachstumseinflüsse diskutiert. Aufgrund der physiologischen Effekte der Hormone und ihrer Blutplasmakonzentrationen im Hinblick auf die Schlachtkörperzusammensetzung wird gefolgert, daß IGF-I beziehungsweise die Somatotropin-Achse in Beziehung zum Proteinansatz und dem Muskelfleischanteil am Schlachtkörper steht, während der Fettansatz und die Schlachtkörperverfettung eher durch die Insulin-Konzentration wiedergegeben wird.
[Show abstract][Hide abstract] ABSTRACT: ZusammenfassungEinfluß von exogenem Wachstumshormon (GH) auf Mastleistung und Plasma-GH-Konzentration bei MastkälbernEin 9wöchiger Versuch mit 4 × 12 weiblichen Fleckviehkälbern (Alter 6 Wochen) wurde durchgeführt, um den Einfluß von exogenem bovinem Wachstumshormon (bGH; 0, 25, 50 und 100 μg/kg LG täglich s.c. injiziert) auf die Mastleistung und GH-Konzentration im Plasma zu untersuchen. Die tägliche Gewichtszunahme in der Kontrollgruppe betrug bereits 1212 g. Der Einsatz von 25, 50 bzw. 100 μg/kg bGH steigerte die tägliche Zuwachsrate gegenüber den Kontrolltieren signifikant um 7.1, 8.3 und 10.8%. Die Futterverwertung war um 3.8, 5.7 und 7.0% verbessert, ab der Dosisstufe 50 μg/kg signifikant. Die Schlachtkärperbewertung zeigte zwischen den Gruppen nur geringe Unterschiede. Im M. semitendinosus konnte bei gleichem Protein- und Wassergehalt eine leichte Abnahme des Fettgehaltes nach Behandlung mit bGH beobachtet werden. Das applizierte bGH steigerte dosisabhängig die GH-Konzentration im Plasma. In der 5., 7. und 9. Versuchswoche wurden in den behandelten Gruppen höhere Plasma-Spiegel gemessen als in der 2. Woche. Die Ausscheidungsrate des bGH war durch die chronische Behandlung stark verzögert. 51 Stunden nach der letzten Injektion wurden Normalwerte noch nicht erreicht. Dagegen wurde in einem separaten Versuch festgestellt, daß sich die Werte bei nur lmaliger Injektion dosisabhängig bereits nach 8–12 Stunden wieder den Basiswerten angenähert hatten. Die Ergebnisse zeigten, daß exogenes bGH auch bei hohem Leistungsniveau wachstumsfördernd wirkte.
[Show abstract][Hide abstract] ABSTRACT: The present study deals with immunohistochemical localization of S-100 protein in the bovine testis. Immunoreactivity for the protein was seen in Sertoli cells of the seminiferous tubules and as a particularly intense staining in the terminal tubular segment (transitional region, middle portion, and terminal plug), which is mainly composed of modified Sertoli cells. Immunoreactivity was also found in epithelial cells of the straight testicular tubules and rete testis, and in the endothelium of capillaries, veins and lymphatic vessels. Although the functional significance of S-100 protein immunoreactivity in the Sertoli cells remains unclear, the present results suggest that it may be involved in the microtubule assembly-disassembly system. The specificity of the immunolabelling observed should enable the antigen and/or antibody to S-100 to be used as an investigative and diagnostic tool in the study of bovine Sertoli cell function.
[Show abstract][Hide abstract] ABSTRACT: Recent studies implicate that apelin and its receptor APJ may have important role for the modulation of angiogenesis. The aim of this study was to further characterise the regulation of apelin/APJ system in bovine ovary. Experiment 1: corpora lutea (CL) were assigned to the following stages: days 1-2, 3-4, 5-7, 8-12, 13-16, >18 (after regression) of oestrous cycle and of gravidity (month <3, 3-5, 6-7 and >8). Experiment 2: Follicles during maturation were divided into granulosa cells (GC) and theca interna (TI) and were examined separately. Classification of follicles occurred by follicle size and oestradiol-17beta (E2) concentration in the follicular fluid (FF) (<0.5 ng/ml, 0.5-5 ng/ml; 5-40 ng/ml; 40-180 ng/ml; >180 ng/ml). Real-time RT-PCR (qPCR) was applied to investigate mRNA expression of examined factors. In general, the expression level of apelin during the oestrous cycle was significantly higher compared to the one during pregnancy. Apelin mRNA levels were always high during the cycle with a tendency of decrease after CL regression. The APJ mRNA in the CL was significantly up regulated on days 5-7 and 8-12 followed by a decrease on days 13-16, and further on days >18. The expression of APJ does not show any significant regulation in the CL throughout pregnancy. The expression of apelin and APJ was not statistically regulated in GC, but was significantly up regulated in follicles with an E2 concentration of more than 5 ng/ml and showed an increase according to growth and maturation of follicles. In conclusion, our data suggest that apelin/APJ system is involved in the mechanism regulating angiogenesis during follicle maturation as well as during CL formation and function in the bovine ovary.
International journal of biological sciences 05/2009; 5(4):344-50. · 4.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to investigate the possible participation of fibroblast growth factor (FGF) family members (FGF1, FGF2 and FGF7 and their receptors) in porcine follicles (polyovulatory species) under special consideration for FGF2 during final growth. A classification of follicles was done by size and follicular fluid content of oestradiol-17beta, progesterone and prostaglandin F2alpha. The mRNA expression of examined factors was analysed by real-time PCR. The hormone concentration was estimated by enzyme immunoassay, protein characterisation by western blotting and localisation by immunohistochemistry. Follicle tissue separated in theca interna and granulosa cells was extracted and tested for mRNA of FGF1, FGF2, FGF7 and receptors (FGFR1IIIc, FGFRIIIb and FGFR2IIIc). Additionally, the mRNA expression of FSHR, LHR and aromatase cytochrome P450 for further characterisation of follicles was analysed. Significantly, higher FGF2 protein levels were measured in stroma when compared with total follicle or corpus luteum tissue. This result was confirmed by western blot with two strong bands. Immunological localisation of FGF2 only in stroma (fibroblasts) confirms the protein measurements. The results show a clear difference for FGF2 protein expression during final growth of follicles if monovulatory (bovine) and polyovulatory (porcine) species are compared. FGF2 protein in porcine ovary may be (due to localisation and concentration in stroma) important for support of angiogenesis of more follicles (polyovulatory species) and not of a single follicle like in cows.