In mares resistant to endometrial infection, periovulatory treatment with ecbolic drugs does not influence uterine clearance or luteal development.

We aimed to determine associations between experimentally impaired uterine clearance or treatment with ecbolic drugs on luteal development in estrous mares after insemination. In a crossover design, eight mares were treated with saline (CON), clenbuterol (CLEN), oxytocin (OXY) and carbetocin (CARB) from the day of first insemination until 2 days after ovulation. Between treatments, the mares rested for one cycle. Estrous mares were examined for the presence of free intrauterine fluid by transrectal ultrasound. Endometrial swabs for cytology and bacteriology were collected on days 1 and 14. Blood samples were collected daily before AI until day 14 after ovulation for determination of progesterone and PGF2α metabolites (PGFM). Differences between treatments were compared by a general linear model for repeated measures (SPSS 29). One mare was excluded because of a uterine infection in the control cycle. In all other mares, only minor amounts of free intrauterine fluid were present after insemination and decreased over time (P<0.05) with no treatment x time interaction. There was no effect of treatment on polymorphonucleated cells (PMN) in endometrial cytology after ovulation or PGFM secretion. Progesterone release from day 1-14 as well as pregnancy rate and conceptus size on day 14 was not influenced by treatment. In conclusion, treatment with clenbuterol does not impair uterine clearance in estrous mares resistant to endometritis. Repeated injection of the oxytocin analogue carbetocin during the early postovulatory period is not detrimental to corpus luteum function and can be recommended to enhance uterine clearance.

Uterine tone: A neglected criterion for the selection of bovine embryo transfer recipients.

This study evaluated the relationship between CL features assessed by ultrasound (luteal tissue area and blood flow, BF) or rectal palpation (size), uterine tone (UT), plasma progesterone (P4) concentration on Day 7 (D7) and subsequent pregnancy outcomes in bovine embryo recipients. A total of 163 cows and heifers were included in this study. The expected day of ovulation after the synchronization protocol was designated as D0. On D7, ovaries and uterus were examined by ultrasonography and rectal palpation, and subjective scores (1-3 scale) were assigned for CL size, area and BF, and for UT. Blood samples were collected for further P4 analysis. Each embryo recipient then received a grade I frozen-thawed in vivo-produced blastocyst, which was transferred to the uterine horn ipsilateral to the CL. Pregnancy diagnosis was performed on D35, and the results were retrospectively compared with the assigned scores for CL and UT. We observed a significant (p < .02) interaction between CL size and UT, with a progressive increase in the likelihood of pregnancy for recipients bearing a large CL among those with turgid UT. Ultrasound scoring of the CL using B-mode and Doppler-mode did not significantly predict pregnancy rates on D35 (p < .6 and p < .5, respectively). However, logistic regression analysis revealed a trend towards a quadratic effect (p < .08 and p < .06) indicating that the probability of pregnancy varied according to the area of luteal tissue and P4 concentrations, respectively. No significant (p > .05) association was found between the probability of pregnancy and the BF area of the CL. In summary, UT before embryo transfer may reflect successful recipient synchronization. Elevated P4 levels, assessed by CL size, may offset uterine contractility, mitigating adverse effects. Additionally, the CL area may be more important than its vascularization area when evaluating recipients D7 after ovulation.

The effect of exogenous melatonin administration before superovulation on embryo yield in Assaf ewes.

The aim of this study was to determine the effect of exogenous melatonin administration on transferable embryos by increasing total antioxidant status before superovulation in Assaf ewes. Selected ewes were randomly divided into two equal groups: melatonin (n = 9) and control (n = 9). In the melatonin group, a melatonin implant (18 mg melatonin, Regulin®, Ceva, Turkey) was placed under the skin of the ear 7 days prior to insertion of the progesterone-containing sponge. In the control group, a physiological saline solution was injected under the skin of the ear on the same day. The same superovulation protocol was used in both groups. In addition, blood samples for determination of Glutathione peroxidase, superoxide dismutase, total antioxidant status and total oxidant status concentrations were collected on five different days, including the day of melatonin implant placement (Day-7), vaginal sponge insertion (Day 0), vaginal sponge removal (Day 11), mating (Day 12-13) and uterine flushing (Day 19). Embryos were collected by laparotomy on the 7th day after mating. Uterine flushing taken into petri dishes were scanned under a stereomicroscope, and the quality and developmental stages of the embryos were recorded. In the study, total corpus luteum count and total cell count were found to be higher in the control group than in the melatonin group (p < .05). When the results were evaluated in terms of oxidative stress index, a negative correlation was found between the total number of corpus luteum, number of cells obtained, count of transferable embryos and number of Grade 1 embryos on Day 0. There was also a positive correlation oxidative stress index and the number of unfertilized oocytes on Day-7. As a result, exogenous melatonin administration prior to superovulation during the breeding season is thought to have a negative effect on embryo yield and quality. Therefore, the use of exogenous melatonin in MOET studies during the breeding season is recommended to be investigated in new studies.

In vitro effect of visfatin on endocrine functions of the porcine corpus luteum.

Previously, we demonstrated the expression of visfatin in porcine reproductive tissues and its effect on pituitary endocrinology. The objective of this study was to examine the visfatin effect on the secretion of steroid (P4, E2) and prostaglandin (PGE2, PGF2α), the mRNA and protein abundance of steroidogenic markers (STAR, CYP11A1, HSD3B, CYP19A1), prostaglandin receptors (PTGER2, PTGFR), insulin receptor (INSR), and activity of kinases (MAPK/ERK1/2, AKT, AMPK) in the porcine corpus luteum. We noted that the visfatin effect strongly depends on the phase of the estrous cycle: on days 2-3 and 14-16 it reduced P4, while on days 10-12 it stimulated P4. Visfatin increased secretion of E2 on days 2-3, PGE2 on days 2-3 and 10-12, reduced PGF2α release on days 14-16, as well as stimulated the expression of steroidogenic markers on days 10-12 of the estrous cycle. Moreover, visfatin elevated PTGER mRNA expression and decreased its protein level, while we noted the opposite changes for PTGFR. Additionally, visfatin activated ERK1/2, AKT, and AMPK, while reduced INSR phosphorylation. Interestingly, after inhibition of INSR and signalling pathways visfatin action was abolished. These findings suggest a regulatory role of visfatin in the porcine corpus luteum.

The regulation of the expression of prokineticin 1 and its receptors and its mechanism of action in the porcine corpus luteum.

Prokineticin 1 (PROK1) is an important factor in pregnancy establishment in pigs, acting at the embryo-maternal interface and the corpus luteum (CL). Estradiol-17ß (E2) is the primary pregnancy recognition signal in pigs, and its effects are augmented by luteotropic prostaglandin E2 (PGE2). On the contrary, prostaglandin F2α (PGF2α) exerts mainly a luteolytic effect. The present study aimed to elucidate whether E2, PGE2, and PGF2α regulate the expression of PROK1 and its receptors in the porcine CL and to determine the PROK1 effect on luteal endothelial cells and pathways that may be involved in this regulation. The effects of E2, PGE2, and PGF2α on the expressions of PROK1 and its receptors in the CL were studied using an in vitro model of ultrathin luteal tissue explants model. Additionally, the effects of E2 and PGE2 on the PROK1 system were determined using an in vivo approach, in which the hormones were administered into the uterine lumen to imitate their secretion by embryos. Endothelial cell proliferation was measured using the colorimetric method. E2 acting via estrogen receptors simulated the mRNA and protein expressions of PROK1 and PROKR1 in CL explants in vitro (p < 0.05). The simultaneous action of E2 with PGE2 enhanced the expression of luteal PROK1 mRNA in vitro (p < 0.05). Estradiol-17ß acting alone significantly increased PROK1 mRNA levels in vivo, whereas E2 simultaneously administered with PGE2 significantly elevated the PROK1 mRNA expression and PROKR1 mRNA and protein contents in CLs adjacent to uterine horns receiving hormonal infusion compared with CLs adjacent to placebo-treated uterine horns (p < 0.01). The PROK1 protein expression was significantly higher in the CLs of pigs treated with E2, PGE2, and E2 together with PGE2 than in the control group. PGF2α increased the PROK1 mRNA content in CLs on days 12 and 14 of the estrous cycle (p < 0.05). The expression of PROKR2 at the mRNA and protein levels remained unchanged in response to in vitro and in vivo treatments. PROK1 stimulated the proliferation of luteal endothelial cells by activating the MAPK, AKT, and mTOR pathways (p < 0.05). In summary, the luteal expressions of PROK1 and PROKR1 in early pregnancy are regulated by E2 and PGE2. PROK1 stimulates luteal angiogenesis by activating the MAPK, AKT, and mTOR pathways. The regulation of luteal PROK1 expression by PGF2α indicates PROK1's putative role during luteolysis. We conclude that PROK1-PROKR1 signaling supports luteal function during CL rescue in pregnancy in pigs.

Expression and localization of anti-Müllerian hormone and its receptors in bovine corpus luteum.

Although anti-Müllerian hormone (AMH) is involved in the regulation of granulosa cell function in female animals, its role in tissues other than ovarian follicles remains poorly understood. It has also been suggested that cows with high circulating AMH concentrations have increased fertility; however, the mechanism has not been elucidated. This study was conducted to identify the presence of the AMH-signaling system and its target cells in the bovine corpus luteum formed from an ovulated follicle. Immunoblotting revealed that the proteolytically cleaved C-terminal region in AMH (AMHC), a biologically active peptide, was present in trace amounts in the early corpus luteum and significantly increased during the mid to regressed stages. AMHC and cleaved N-terminal region (AMHN) in AMH generate a noncovalent isoform that improves the activity of AMH signaling. An immunohistochemical analysis revealed that AMHC, AMHN, and type II AMH receptor (AMHR2) were localized to luteal cells during the entire estrous cycle. AMH in the corpus luteum seemed to be newly synthesized since AMH expression was detected. These findings suggest that AMH signaling is involved in the regulation of luteal cell function through an autocrine and post-translational processing mechanism. The level of AMHR2 and mRNA expression of AMHR2 and type I AMH receptors (activin-like kinase 2, 3, and 6) were highest in the mid stage. Thus, AMH signaling in the corpus luteum may also be regulated by changes in the receptor levels. Since the transforming growth factor-beta superfamily, to which AMH belongs, is a multifunctional polypeptide growth factor, further studies are needed to evaluate whether AMH signaling has a role in facilitating or inhibiting luteal cell functions.

Inter-software reliability and agreement for follicular and luteal morphometric and echotextural ultrasonographic parameters in beef cattle.

This study aimed to compare the inter-software and inter-observer reliability and agreement for the assessment of follicular and luteal morphometry and echotexture parameters in beef crossbreed females (3/8 Bos taurus indicus and 5/8 Bos taurus taurus). B-mode and color Doppler ultrasonographic ovarian images were obtained at specific time points of estradiol-progesterone-based protocols for timed artificial insemination (TAI). Sonograms were analyzed by two observers using a licensed (IASP1) and an open access (IASP2) software package. A total of 292 snap-shot sonograms were analyzed for morphometric parameters and 504 for echotexture parameters. inter-software reliability was judged moderate to excellent (ICC or CCC=0.73-0.98), whereas inter-observer reliability for morphometric parameters was deemed good to excellent (ICC or CCC=0.82-0.98). A small percentage (up to 10.95 %) of measured parameters fell outside the limits of inter-software and inter-observer agreement. For echotexture parameters, inter-software reliability varied widely (ICC or CCC=0.16-0.95) based on the size of regions of interest (ROI), while inter-observer reliability ranged from moderate to excellent (ICC or CCC= 0.71-0.97). The highest inter-software reliability for pixel value and heterogeneity value was observed for the corpus luteum (ICCs=0.81-0.95; P>0.05), followed by the peripheral follicular antrum (ICCs=0.75-0.78; P<0.05). However, lower reliability was determined for the follicular wall (ICCs=0.08-0.33; P<0.0001) and perifollicular stroma (ICCs=0.16-0.46; P<0.05). In conclusion, both software packages showed high reproducibility for morphometric measurements, while echotexture measurements were more challenging to replicate based on ROI sizes. Caution is advised when selecting ROI sizes for echotexture measurements in bovine ovaries.

Unraveling the role of lipid droplets and perilipin 2 in bovine luteal cells.

Steroidogenic tissues contain cytosolic lipid droplets that are important for steroidogenesis. Perilipin 2 (PLIN2), a structural coat protein located on the surface of lipid droplets in mammalian cells, plays a crucial role in regulating lipid droplet formation and contributing to various cellular processes such as lipid storage and energy homeostasis. Herein, we examine the role that PLIN2 plays in regulating progesterone synthesis in the bovine corpus luteum. Utilizing gene array databases and Western blotting, we have delineated the expression pattern of PLIN2 throughout the follicular to luteal transition. Our findings reveal the presence of PLIN2 in both ovarian follicular and steroidogenic luteal cells, demonstrating an increase in its levels as follicular cells transition into the luteal phase. Moreover, the depletion of PLIN2 via siRNA enhanced progesterone production in small luteal cells, whereas adenovirus-mediated overexpression of both PLIN2 and Perilipin 3 (PLIN3) induced an increase in cytosolic lipid droplet accumulation and decreased hormone-induced progesterone synthesis in these cells. Lastly, in vivo administration of the luteolytic hormone prostaglandin F2α resulted in an upregulation of PLIN2 mRNA and protein expression, accompanied by a decline in serum progesterone. Our findings highlight the pivotal role of PLIN2 in regulating progesterone synthesis in the bovine corpus luteum, as supported by its dynamic expression pattern during the follicular to luteal transition and its responsiveness to luteotropic and luteolytic hormones. We suggest PLIN2 as a potential therapeutic target for modulating luteal function.

Effect of prostaglandin treatment on the estrus behaviour, follicular and luteal morphometry and serum hormone profile in sub-estrus buffaloes during non-breeding season.

Sub-estrus buffaloes do not exhibit estrus signs despite being cyclic contributing to extended service periods and inter-calving intervals causing significant economic loss. The present study described the effect of synthetic prostaglandin (PGF2α) on estrus behaviour, follicular and luteal morphometry, and serum estradiol (E2) and progesterone (P4) profile in sub-estrus buffaloes during the non-breeding season. The incidence of sub-estrus was 38.4% during the non-breeding season. The sub-estrus buffaloes (n = 33) were divided into two groups, viz., Control (n = 16) and PGF2α treatment (Inj. Cloprostenol 500 µg, i.m., n = 17). Estrus induction response was significantly greater in the treatment (100 vs. 18.75%, p < .001), and a relatively greater proportion of animals conceived in the treatment group (29.41 vs. 6.25%, p = .08). The time elapsed to induction of estrus and insemination following treatment was significantly lower in the treatment group than control. A significant increment in the follicle diameter (9.72 ± 0.45 vs. 13.00 ± 0.45 mm, P < .0001) and serum estradiol (E2) concentration (66.01 ± 11.92 vs. 104.9 ± 13.21 pg/mL, p = .003) observed at the post-treatment period in the PGF2α treatment group. At the same time, CL diameter was reduced significantly at a higher regression rate in the PGF2α treated buffaloes than those of control. Of the responded buffaloes, only 30% showed high-intensity estrus attributed to the expulsion of cervico-vaginal mucus (CVM), uterine tonicity, micturition, and mounting response by a teaser bull. From this study, it can be concluded that the administration of PGF2α could induce estrus in the sub-estrus buffaloes during the non-breeding season. Behavioural changes, along with sonographic observation of POF, regressing CL, and serum E2 and P4 concentration would be useful to determine the right time of insemination in sub-estrus buffaloes during non-breeding season.

Expression dynamics of adipokines during induced ovulation in the bovine follicles and early corpus luteum.

The study aimed to assess the local gene expression of adipokine members, namely vaspin, adiponectin, visfatin, resistin and their associated receptors - heat shock 70 protein 5 (HSPA5), adiponectin receptor 1 (AdipoR1) and adiponectin receptor 2 (AdipoR2) - in bovine follicles during the preovulatory period and early corpus luteum development. Follicles were collected before gonadotropin-releasing hormone (GnRH) treatment (0 h) and at 4, 10, 20, 25 and 60 h after GnRH application through transvaginal ovariectomy (n = 5 samples/group). Relative mRNA expression levels were quantified using real-time reverse transcription polymerase chain reaction (RT-qPCR). Vaspin exhibited high mRNA levels immediately 4 h after GnRH application, followed by a significant decrease. Adiponectin mRNA levels were elevated at 25 h after GnRH treatment. AdipoR2 exhibited late-stage upregulation, displaying increased expression at 20, 25 and 60 h following GnRH application. Visfatin showed upregulation at 20 h post-GnRH application. In conclusion, the observed changes in adipokine family members within preovulatory follicles, following experimentally induced ovulation, may constitute crucial components of the local mechanisms regulating final follicle growth and development.

Autophagy favors survival of corpora lutea during the long-lasting pregnancy of the South American plains vizcacha, Lagostomus maximus (Rodentia, Caviomorpha).

The corpus luteum (CL) is a transient endocrine gland that plays a crucial role in establishing and maintaining pregnancy. Although autophagy and apoptosis have been suggested as cooperative mechanisms, their interaction within the CL of pregnant mammals has not been thoroughly investigated. To understand the collaborative function of autophagy and apoptosis in the CL, we analyzed both mechanisms during pregnancy in the South American plains vizcacha, Lagostomus maximus. This rodent undergoes a decline in progesterone levels during mid-gestation, a reactivation of the hypothalamus-hypophysis-gonadal axis, and the incorporation of new functional secondary CL. Our analysis of autophagy markers BECLIN 1 (BECN1), SEQUESTOSOME1 (SQSTM1), Microtubule-associated protein light chain 3 (LC3B), and lysosomal-associated membrane protein 1 (LAMP1) and anti- and pro-apoptotic markers BCL2 and ACTIVE CASPASE 3 (A-C3) revealed interactive behaviors between both processes. Healthy primary and secondary CL exhibited positive expression of BECN1, SQSTM1, LC3B, and LAMP1, while regressed CL displayed enhanced expression of these autophagy markers along with nuclear A-C3. Transmission electron microscopy revealed a significant formation of autophagic vesicles in regressed CL during full-term pregnancy, whereas healthy CL exhibited a low number of autophagy vesicles. The co-localization between LC3B and SQSTM1 and LC3B with LAMP1 was observed in both healthy and regressed CL during pregnancy, while co-localization of BECN1 and BCL2 was only detected in healthy CL. LC3B and ACTIVE CASPASE 3 co-localization were detected in a subset of luteal cells within the regressing CL. We propose that autophagy could act as a survival mechanism in the CL, allowing the pregnancy to progress until full-term, while also serving as a mechanism to eliminate remnants of regressed CL, thereby providing the necessary space for subsequent follicular maturation.

Effect of prostaglandin alone and in combination with trace minerals on the follicular and luteal dynamics, estrus response and pregnancy in sub-estrus buffaloes (Bubalus bubalis).

Sub-estrus is a condition when buffaloes do not display behavioural estrus signs, despite being in estrus and causes a delay in conception and increases the service period. The present study describes the effect of synthetic prostaglandin (PGF2α) alone and in combination with trace minerals on the follicular and corpus luteum (CL) dynamics, serum estradiol (E2) and progesterone (P4) concentration correlating estrus response and pregnancy outcome in sub-estrus buffaloes during the breeding season. A total of 50 sub-estrus buffaloes, identified through ultrasonography (USG) examination, were randomly allocated into three groups, viz. T1 (Synthetic PGF2α, Inj. Cloprostenol 500 µg, i.m, n = 17), T2 (Synthetic PGF2α + Trace mineral supplementation, Inj. Stimvet 1 mL/100 kg body weight, i.m., n = 17) and control (untreated; n = 16). Following treatment, 100% of sub-estrus buffaloes were induced estrus in the T1 and T2 groups, while only 18.75% were induced in the control. The CL diameter and serum P4 concentration were significantly lower at post-treatment, whereas the pre-ovulatory follicle (POF) size and serum E2 concentration were significantly higher in the T1 and T2 groups as compared to the control (p < .05). The buffaloes of the T2 group had a greater proportion of moderate intensities estrus than those of T1. Moreover, the proportion of buffaloes conceived in the T1 and T2 were 41.2% and 52.95%, respectively. The larger POF diameter and higher serum E2 concentration were associated with intense intensity estrus and higher conception rate (66.7%) in sub-estrus buffaloes. Similarly, CL regression rate, POF size and serum E2 concentration were relatively higher in the buffaloes conceived as compared to those not conceived. It is concluded that synthetic PGF2α in combination with trace minerals induces moderate to intense intensities estrus in a greater proportion of sub-estrus buffaloes and increases the conception rate during the breeding season. Moreover, behavioural estrus attributes correlating follicle and luteal morphometry, serum E2 and P4 concentration could be used to optimise the breeding time for augmenting the conception rate in sub-estrus buffaloes.

Changes on corpus luteum structure and progesterone synthesis pathway after hCG or GnRH treatment during the early luteal phase in sheep.

This study investigated the effect of hCG or GnRH on structural changes of the corpora lutea (CL) and the regulation of the expression of steroidogenic enzymes involved in P4 secretion in post-ovulatory (po-CL) and accessory CL (acc-CL). Sixty-four ewes were assigned to three groups receiving: 300 IU of hCG (hCG) or 4 µg Buserelin (GnRH) or 1 mL of saline solution (Control) on Day (d) 4 post artificial insemination (FTAI). Laparoscopic ovarian were performed on d 4, 14 and, 21 post-FTAI to determine the numbers of CL. Blood samples were collected for serum LH and P4 analysis. On d 14 post-FTAI, both CL were removed from the ovary to determine large luteal cell (LLC) number and to evaluate the expression of steroidogenic enzymes (HSD3B1, STAR, CYP11A1). Only hCG and GnRH treated ewes generated acc-CL. The LLC in both po- and acc-CL were significantly greater in the hCG group compared to GnRH and Control groups (P<0.05). Overall, hCG group showed the greatest immunodetection of HSD3B1and STAR in both po- and acc-CL (P<0.05). rnRNA expression of HSD3B1, STAR and CYP11A1 in the acc-CL tended to be greater in hCG group than in GnRH group (P<0.1). The LH concentration was increased in GnRH group (P<0.05) and P4 concentration was greater in hCG group compared to the other groups (P<0.05). In conclusion, administration of hCG has a notably impact on acc-CL development and the expression of steroidogenic enzymes compared to GnRH treatment in ewes. This leads to elevated P4 concentration and improved luteal function.

Adropin may regulate ovarian functions by improving antioxidant potential in adult mouse.

The corpus luteum (CL) is a temporary endocrine gland that synthesizes progesterone. The luteal progesterone plays a central role in the regulation of the estrous cycle as well as the implantation and maintenance of pregnancy. Our previous study showed the expression of adropin and its receptor, GPR19, in the luteal cells and its significant role in luteinization. The aim of the present study was to investigate the in vitro effect of adropin on hCG-induced ovarian functions in adult mice. We also evaluated the effect of exogenous treatment with adropin on ovarian steroidogenesis and anti-oxidant parameters, with special emphasis on CL function. Our results demonstrated that adropin acts synergistically with hCG to promote ovarian steroidogenesis and survival by increasing the expression of StAR, 3ß-HSD, and aromatase proteins and decreasing the BAX/BCL2 ratio. Exogenous adropin treatment increased progesterone production by increasing the expression of GPR19, StAR and 3ß-HSD enzymes in the mouse ovary. Also, adropin inhibited the luteal oxidative stress by increasing nuclear translocation of NRF-2 in CL, which resulted in increased HO-1 expression and SOD, catalase activity. Decreased oxidative stress might inhibit the translocation of NF-κB into the nucleus of luteal cells, resulting into increased survival and decreased apoptosis, as evident by decreased lipid peroxidation, BAX/BCL2 ratio, caspase 3, active caspase 3 expression, and TUNEL-positive cells in adropin treated mice. Our findings suggest that adropin can be a promising candidate that can enhance the survivability of the CL.

Symptomatic corpus luteum hemorrhage in adolescent females with ITP.

Patients with immune thrombocytopenia (ITP) usually present with minor mucocutaneous bleeding. Corpus luteum hemorrhage (CLH) is generally asymptomatic but may, rarely, lead to severe intraperitoneal bleeding, mostly in patients with coagulation disorders. CLH causing intraperitoneal bleeding has only been described in few individuals with ITP. The objective of this retrospective observational study was to assess the clinical course and incidence of symptomatic CLH in adolescent females with newly diagnosed or chronic ITP. Additionally, a comprehensive literature review was conducted to scrutinize cases of pediatric female patients with ITP, complicated by CLH. We identified three patients with ITP and hemoperitoneum secondary to CLH. They presented with acute abdominal pain, had severe thrombocytopenia (platelet counts below 20 × 109/L), and required blood transfusions as well as ITP-directed therapy. All the patients were hemodynamically stable and did not require emergency surgical intervention.  Conclusion: CLH could potentially pose a significant complication in the context of adolescent females with ITP, requiring a strong index of suspicion to direct expedient therapy. What is Known: • Immune thrombocytopenia is typically associated with minor bleeding tendency. • Corpus luteum hemorrhage is generally asymptomatic; however, in women with bleeding disorders, it has the potential to result in substantial intra-abdominal bleeding. What is New: • Corpus luteum hemorrhage leading to intra-abdominal bleeding is a potential severe complication of immune thrombocytopenia in adolescent females.

Histological, immunohistochemical and serological investigations of the ovary during follicular phase of estrous cycle in Saidi sheep.

BACKGROUND: Saidi sheep are the most abundant ruminant livestock species in Upper Egypt, especially in the Assiut governorate. Sheep are one of the most abundant animals raised for food in Egypt. They can convert low-quality roughages into meat and milk in addition to producing fiber and hides therefore; great opportunity exists to enhance their reproduction. Saidi breed is poorly known in terms of reproduction. So this work was done to give more information on some hormonal, oxidative, and blood metabolites parameters in addition to histological, histochemical and immunohistochemical investigations of the ovary during follicular phase of estrous cycle. The present study was conducted on 25 healthy Saidi ewes for serum analysis and 10 healthy ewes for histological assessment aged 2 to 5 years and weighted (38.5 ± 2.03 kg). RESULTS: The follicular phase of estrous cycle in Saidi sheep was characterized by the presence of ovarian follicles in different stages of development and atresia in addition to regressed corpus luteum. Interestingly, apoptosis and tissue oxidative markers play a crucial role in follicular and corpus luteum regression. The most prominent features of the follicular phase were the presence of mature antral (Graafian) and preovulatory follicles as well as increased level of some blood metabolites and oxidative markers. Here we give a new schematic sequence of ovarian follicles in Saidi sheep and describing the features of different types. We also clarified that these histological pictures of the ovary was influenced by hormonal, oxidative and blood metabolites factors that characterizes the follicular phase of estrous cycle in Saidi sheep. CONCLUSION: This work helps to understanding the reproduction in Saidi sheep which assist in improving the reproductive outcome of this breed of sheep. These findings are increasingly important for implementation of a genetic improvement program and utilizing the advanced reproductive techniques as estrous synchronization, artificial insemination and embryo transfer.

Enhancement of progesterone biosynthesis via kisspeptin stimulation: Upregulation of steroidogenic transcripts and phosphorylated extracellular signal-regulated kinase (p-ERK1/2) expression in the buffalo luteal cells.

The presence of Kisspeptin (Kp) and its receptors in the corpus luteum (CL) of buffalo has recently been demonstrated. In this study, we investigated the role of Kp in the modulation of progesterone (P4) synthesis in vitro. The primary culture of bubaline luteal cells (LCs) was treated with 10, 50, and 100 nM of Kp and Kp antagonist (KpA) alongside a vehicle control. The combined effect of Kp and KpA was assessed at 100 nM concentration. Intracellular response to Kp treatment in the LCs was assessed by examining transcript profiles (LHR, STAR, CYP11A1, HSD3B1, and ERK1/2) using quantitative polymerase chain reaction (qPCR). In addition, the immunolocalization of ERK1/2 and phosphorylated ERK1/2 (p-ERK1/2) in the LCs was studied using immunocytochemistry. Accumulation of P4 from the culture supernatant was determined using enzyme-linked immunosorbent assay (ELISA). The results indicated that LCs had a greater p-ERK1/2 expression in the Kp treatment groups. A significant increase in the P4 concentration was recorded at 50 nM and 100 nM Kp, while KpA did not affect the basal concentration of P4. However, the addition of KpA to the Kp-treated group at 100 nM concentration suppressed the Kp-induced P4 accumulation into a concentration similar to the control. There was significant upregulation of ERK1/2 and CYP11A1 expressions in the Kp-treated LCs at 100 nM (18.1 and 37fold, respectively, p < 0.01). However, the addition of KpA to Kp-treated LCs modulated ERK1/2, LHR, STAR, CYP11A1, and HSD3B1 at 100 nM concentration. It can be concluded that Kp at 100 nM stimulated P4 production, while the addition of KpA suppressed Kp-induced P4 production in the buffalo LCs culture. Furthermore, an increment in p-ERK1/2 expression in the LCs indicated activation of the Kp signaling pathway was associated with luteal steroidogenesis.

Local effect of allopregnanolone in rat ovarian steroidogenesis, follicular and corpora lutea development.

Allopregnanolone (ALLO) is a known neurosteroid and a progesterone metabolite synthesized in the ovary, CNS, PNS, adrenals and placenta. Its role in the neuroendocrine control of ovarian physiology has been studied, but its in situ ovarian effects are still largely unknown. The aims of this work were to characterize the effects of intrabursal ALLO administration on different ovarian parameters, and the probable mechanism of action. ALLO administration increased serum progesterone concentration and ovarian 3ß-HSD2 while decreasing 20α-HSD mRNA expression. ALLO increased the number of atretic follicles and the number of positive TUNEL granulosa and theca cells, while decreasing positive PCNA immunostaining. On the other hand, there was an increase in corpora lutea diameter and PCNA immunostaining, whereas the count of TUNEL-positive luteal cells decreased. Ovarian angiogenesis and the immunohistochemical expression of GABAA receptor increased after ALLO treatment. To evaluate if the ovarian GABAA receptor was involved in these effects, we conducted a functional experiment with a specific antagonist, bicuculline. The administration of bicuculline restored the number of atretic follicles and the diameter of corpora lutea to normal values. These results show the actions of ALLO on the ovarian physiology of the female rat during the follicular phase, some of them through the GABAA receptor. Intrabursal ALLO administration alters several processes of the ovarian morpho-physiology of the female rat, related to fertility and oocyte quality.

Corpus luteum presence in the bovine ovary increase intrafollicular progesterone concentration: consequences in follicular cells gene expression and follicular fluid small extracellular vesicles miRNA contents.

BACKGROUND: It is well described that circulating progesterone (P4) plays a key role in several reproductive events such as oocyte maturation. However, during diestrus, when circulating P4 is at the highest concentrations, little is known about its local impact on the follicular cells such as intrafollicular P4 concentration due to corpus luteum (CL) presence within the same ovary. Based on that, our hypothesis is that the CL presence in the ovary during diestrus alters intrafollicular P4 concentrations, oocyte competence acquisition, follicular cells gene expression, and small extracellular vesicles (sEVs) miRNAs contents. RESULTS: P4 hormonal analysis revealed that ipsilateral to the CL follicular fluid (iFF) presented higher P4 concentration compared to contralateral follicular fluid (cFF). Furthermore, oocyte maturation and miRNA biogenesis pathways transcripts (ADAMTS-1 and AGO2, respectively) were increased in cumulus and granulosa cells of iFF, respectively. Nevertheless, a RT-PCR screening of 382 miRNAs showed that three miRNAs were upregulated and two exclusively expressed in sEVs from iFF and are predicted to regulate cell communication pathways. Similarly, seven miRNAs were higher and two exclusively expressed from cFF sEVs and are predicted to modulate proliferation signaling pathways. CONCLUSION: In conclusion, intrafollicular P4 concentration is influenced by the presence of the CL and modulates biological processes related to follicular cell development and oocyte competence, which may influence the oocyte quality. Altogether, these results are crucial to improve our knowledge about the follicular microenvironment involved in oocyte competence acquisition.