PSI-B-20 A scan for signatures of selection in Russian Romanov sheep breed based on ROH hot spots analysis

The world-famous Romanov breed of sheep fits easily into different production systems including private households, smallholders, and large farms. Such a resilience and extraordinary reproduction traits combining with good meat qualities have made the Romanov breed as a choice to produce mutton in the Central Russia regions. However, little is known on genetic mechanisms underlying the complex of specific traits of this breed. In this regard, we performed a scan of signatures of selection by identification of runs of homozygosity (ROH) islands in the Romanov sheep population. Forty-eight samples of the Romanov sheep from the Yaroslavl and Kaluga regions of Russia were genotyped using Ovine Infinium® HD SNP BeadChip (Illumina, CA, USA). ROHs were estimated in the R package “detectRUNS” using window-free method for consecutive. The ROH shared in more than 50% of sheep were considered as ROH islands. The genes overlapped with the ROH islands were annotated using Ensembl genome browser 103. Enrichment for functional categories of the identified genes was performed using DAVID Functional Annotation Bioinformatics Microarray Analysis tool. More than 88 SNPs were found in ROH islands located on OAR1, OAR10, OAR11, OAR13, OAR14, OAR15, OAR16, OAR17 and OAR18. We detected a major group of genes related to reproduction traits. The identified candidates included UBE2Q1 (embryo implantation, fertilization, prolactin secretion), RNF17 (spermiogenesis), HSD11B2 (pregnancy, follicle function), FANCA (gonad development), and CUBN (embryonic development). In addition, several identified genes were involved in fatty acid metabolism (ACOX1, HAO2, and EIF6), lipid transport (OSBPL11), and biosynthesis of lipid-binding serum glycoproteins (BPIFB4) as well as in osteoblast differentiation (ADAR). The studies will be continued on a larger sample with further validation of the most promising candidate genes. Genotyping of twenty-four samples was performed within RFBR project No. 20-516-56002. Other twenty-four SNP-genotypes were generated within the theme No. 0445-2019-0024.

Prolactin mediates behavioural rejection responses to avian brood parasitism

Adaptations resulting from co-evolutionary interactions between avian brood parasites and their hosts have been extensively studied, yet the physiological mechanisms underlying antiparasitic host defences remain little known. Prolactin, one of the main hormones involved in the regulation of avian parental behaviour, might play a key role in the orchestration of the host responses to avian brood parasitism. Given the positive association between prolactin and parental behaviour during incubation, decreasing prolactin levels are expected to facilitate egg-rejection decisions. We tested this prediction by implanting Eurasian blackbird (Turdus merula) females with an inhibitor of prolactin secretion, bromocriptine mesylate, to experimentally decrease their plasma prolactin levels. Bromocriptine-implanted individuals ejected mimetic model eggs at higher rates, and showed shorter latency to egg ejection, than placebo-treated birds. To our knowledge, this is the first experimental evidence that behavioural host defences against avian brood parasitism are mediated by prolactin.

P–322 Addressing progesterone and cAMP signalling pathways for decidualization induction of endometrial stromal cells of patients with endometriosis

Study question Which compounds/compound combinations are most effective in decidualization induction of endometrial stromal cells (ESCs) of patients with and without endometriosis? Summary answer Combination of compounds addressing different steps in the signalling cascade of decidualization induce decidualization more effectively than application of the individual compounds alone. What is known already Decidualization is the monthly recurring differentiation process of the ESCs in preparation for embryo implantation in human. Undifferentiated ESCs reveal an increased potential to proliferate and invade after retrograde menstruation. This may lead to the formation of ectopic lesions and the manifestation of the chronic gynaecological disease of endometriosis due to an impairment of the decidualization process. Study design, size, duration Compounds and compound combinations addressing the progesterone receptor- or the cAMP-mediated pathway were evaluated with regard to their own and their synergistic potential to induce decidualization of ESCs from women with (n = 10) and without (n = 10) endometriosis during a 6-day treatment. Participants/materials, setting, methods Human primary ESCs were isolated via enzymatic-mechanic digestion from eutopic endometrium from women with and without endometriosis and treated for 6 days in vitro with different progestins (progesterone, medoxyprogesterone acetate (MPA)), 8-Br-cAMP, forskolin, or phosphodiesterase (PDE)-inhibitor (Rolipram) alone or in combination. The degree of decidualization induction was quantified by morphological, biochemical (prolactin) and molecular (HAND2, FOXO1) parameters by means of ELISA, flow cytometric analysis, Realtime PCR and Western blot analysis. Main results and the role of chance After 6 days of treatment, decidualization was induced by forskolin as well as by 8-Br-cAMP whereas progestins or PDE alone hardly induced prolactin secretion by ESCs as a marker of decidualization. A change of morphology from undifferentiated fibroblast-like cells to rounded cells could be observed in parallel with the secretion of prolactin. Forskolin and 8-Br-cAMP-induced decidualization was significantly enhanced by MPA but not by progesterone. These effects were similar in ESCs from women with and without endometriosis. Moreover, forskolin-induced decidualization was significantly enhanced by simultaneous application of PDE. Interestingly, this effect was higher in cells of patients with endometriosis. An induction of decidualization in ESCs was associated with a parallel increase of the process-associated transcription factors HAND2 and FOXO1. This rise of transcription was markedly increased in combination with MPA but not with progesterone. Limitations, reasons for caution Endometrial tissue was obtained from women undergoing infertility treatment and thus may differ from the endometrium of fertile women. Results obtained from primary cells in vitro may not cover the in vivo situation in all respects. Wider implications of the findings: The results of this study provide baseline data for the development of a possible therapeutical approach to induce decidualization as a treatment option for endometriosis. Further research is required to determine the effectiveness of the in vitro tested compound combinations in an in vivo model. Trial registration number Not applicable

P-322 Addressing progesterone and cAMP signalling pathways for decidualization induction of endometrial stromal cells of patients with endometriosis

Study question Which compounds/compound combinations are most effective in decidualization induction of endometrial stromal cells (ESCs) of patients with and without endometriosis? Summary answer Combination of compounds addressing different steps in the signalling cascade of decidualization induce decidualization more effectively than application of the individual compounds alone. What is known already Decidualization is the monthly recurring differentiation process of the ESCs in preparation for embryo implantation in human. Undifferentiated ESCs reveal an increased potential to proliferate and invade after retrograde menstruation. This may lead to the formation of ectopic lesions and the manifestation of the chronic gynaecological disease of endometriosis due to an impairment of the decidualization process. Study design, size, duration Compounds and compound combinations addressing the progesterone receptor- or the cAMP-mediated pathway were evaluated with regard to their own and their synergistic potential to induce decidualization of ESCs from women with (n = 10) and without (n = 10) endometriosis during a 6-day treatment. Participants/materials, setting, methods Human primary ESCs were isolated via enzymatic-mechanic digestion from eutopic endometrium from women with and without endometriosis and treated for 6 days in vitro with different progestins (progesterone, medoxyprogesterone acetate (MPA)), 8-Br-cAMP, forskolin, or phosphodiesterase (PDE)-inhibitor (Rolipram) alone or in combination. The degree of decidualization induction was quantified by morphological, biochemical (prolactin) and molecular (HAND2, FOXO1) parameters by means of ELISA, flow cytometric analysis, Realtime PCR and Western blot analysis. Main results and the role of chance After 6 days of treatment, decidualization was induced by forskolin as well as by 8-Br-cAMP whereas progestins or PDE alone hardly induced prolactin secretion by ESCs as a marker of decidualization. A change of morphology from undifferentiated fibroblast-like cells to rounded cells could be observed in parallel with the secretion of prolactin. Forskolin and 8-Br-cAMP-induced decidualization was significantly enhanced by MPA but not by progesterone. These effects were similar in ESCs from women with and without endometriosis. Moreover, forskolin-induced decidualization was significantly enhanced by simultaneous application of PDE. Interestingly, this effect was higher in cells of patients with endometriosis. An induction of decidualization in ESCs was associated with a parallel increase of the process-associated transcription factors HAND2 and FOXO1. This rise of transcription was markedly increased in combination with MPA but not with progesterone. Limitations, reasons for caution Endometrial tissue was obtained from women undergoing infertility treatment and thus may differ from the endometrium of fertile women. Results obtained from primary cells in vitro may not cover the in vivo situation in all respects. Wider implications of the findings The results of this study provide baseline data for the development of a possible therapeutical approach to induce decidualization as a treatment option for endometriosis. Further research is required to determine the effectiveness of the in vitro tested compound combinations in an in vivo model. Trial registration number not applicable

Ovulation Induction in Patients with Polycystic Ovarian Syndrome (PCOS) & Hyperprolactinemia (HPRL): Efficacy of Letrozole (LE) Combined with Cabergoline (CE) in Comparison to (LE) Alone

Objectives: (PCOS) is the most common cause of anovulatory infertility, with majority of patients having mild (HPRL). (CE), a dopamine receptor agonist, inhibits prolactin secretion, leading to better ovulatory response. (LE), an aromatase inhibitor, without adverse effects on endometrium & induces fewer mature follicles with less risk of OHSS. Our aim was to investigate effects of combined (LE) & (CE) in comparison to (LE) alone on ovulation & clinical pregnancy rates in (PCOS) patients with (HPRL). Patients & Methods: 180 women with (PCOS) and of 22-38 years old, were enrolled in a hospital based clinical trial. Patients randomly allocated into 2 groups, (A&B). All with a serum prolactin > 32 ng/ml. Patients in (A): (92) were given (LE), 5mg for 5days: (3 – 7 of the cycle)/3 cycles and (CE), 0.5mg weekly for 12 weeks. Those in (B): (88) received only (LE), same dose & duration as in (A). All patients were matched for their age and BMI. Exclusion criteria: other causes of (HPRL). Outcome measure: ovulation rate & detection of both chemical & clinical pregnancies by βhCG and ultrasound of fetal cardiac activity, 2-4 weeks after missed period. Follow-up for 6 months. Data analysis by using SPSS version for windows, P-value significant if (< 0.05). Results: 3 patients from (A) & 5 from (B) had drug side effects and were excluded. None of patients were lost during the follow-up period. In (A), difference between mean serum prolactin level before & after treatment was statistically significant (P<0.001): 48.3±4.2ng/ml and 8.1±5.2ng/ml, respectively. No significant decrease of prolactin level in (B) (P >0.05). After treatment, BMI in (A) 24.1± 3.2, & 24.2 ± 3.6 in (B) (P=0.567). (56.2%) of women in (A) became regularly menstruating but only (30.1%) in (B) (P< 0.05). Ovulation rate was higher in (A) (50.6%) in comparison to (B) (26.5%), (P<0.05). Clinical pregnancy rate in (A) (41.6%) and (21.6%) in (B) (P<0.05). Neither twin pregnancy, nor OHSS were recorded in both groups. Conclusions: The combination of (LE) & (CE) is superior to (LE) alone in management of anovulatory patients with (PCOS) and should be used as the first-line treatment for them

Ovulation Induction in Patients with Polycystic Ovarian Syndrome (PCOS) & Hyperprolactinemia (HPRL): Efficacy of Letrozole (LE) Combined with Cabergoline (CE) in Comparison to (LE) Alone

Objectives: (PCOS) is the most common cause of anovulatory infertility, with majority of patients having mild (HPRL). (CE), a dopamine receptor agonist, inhibits prolactin secretion, leading to better ovulatory response. (LE), an aromatase inhibitor, without adverse effects on endometrium & induces fewer mature follicles with less risk of OHSS. Our aim was to investigate effects of combined (LE) & (CE) in comparison to (LE) alone on ovulation & clinical pregnancy rates in (PCOS) patients with (HPRL). Patients & Methods: 180 women with (PCOS) and of 22-38 years old, were enrolled in a hospital based clinical trial. Patients randomly allocated into 2 groups, (A&B). All with a serum prolactin > 32 ng/ml. Patients in (A): (92) were given (LE), 5mg for 5days: (3 – 7 of the cycle)/3 cycles and (CE), 0.5mg weekly for 12 weeks. Those in (B): (88) received only (LE), same dose & duration as in (A). All patients were matched for their age and BMI. Exclusion criteria: other causes of (HPRL). Outcome measure: ovulation rate & detection of both chemical & clinical pregnancies by βhCG and ultrasound of fetal cardiac activity, 2-4 weeks after missed period. Follow-up for 6 months. Data analysis by using SPSS version for windows, P-value significant if (< 0.05). Results: 3 patients from (A) & 5 from (B) had drug side effects and were excluded. None of patients were lost during the follow-up period. In (A), difference between mean serum prolactin level before & after treatment was statistically significant (P<0.001): 48.3±4.2ng/ml and 8.1±5.2ng/ml, respectively. No significant decrease of prolactin level in (B) (P >0.05). After treatment, BMI in (A) 24.1± 3.2, & 24.2 ± 3.6 in (B) (P=0.567). (56.2%) of women in (A) became regularly menstruating but only (30.1%) in (B) (P< 0.05). Ovulation rate was higher in (A) (50.6%) in comparison to (B) (26.5%), (P<0.05). Clinical pregnancy rate in (A) (41.6%) and (21.6%) in (B) (P<0.05). Neither twin pregnancy, nor OHSS were recorded in both groups. Conclusions: The combination of (LE) & (CE) is superior to (LE) alone in management of anovulatory patients with (PCOS) and should be used as the first-line treatment for them.

Deciphering the role of PGRMC1 during human decidualization using an in vitro approach

Context Non-classical membrane progesterone receptor (mPRs) and PGRMC1 expression have been detected in endometrium, but their role in decidualization was not yet investigated. We previously demonstrated PGRMC1 downregulation in receptive endometrium and that its overexpression inhibits decidualization. Furthermore, during decidualization, PGRMC1 mainly interacts with proteins involved in biosynthesis, intracellular transport and mitochondrial activity. Objective To determine PGRMC1 and mPRs signaling role during decidualization. Design and Interventions Isolated primary endometrial stromal cells (EnSC) were in vitro decidualized in presence of classic stimuli (E2+P4), PGRMC1 inhibitor (AG205), or membrane-impermeable P4 (P4-BSA). Setting and Participants Endometrial biopsies from 19 fertile oocyte donors attending IVI-Valencia IVF clinic. Main Outcome Measure(s) EnSC decidualization was evaluated by prolactin ELISA and F-actin immunostaining. Progesterone receptor localization was evaluated by immunofluorescence. EnSC transcriptomic profiles were analyzed by microarray technology. Result(s) PGRMC1 inhibition during EnSC decidualization (AG205dEnSC) does not interfere with EnSC cytoskeletal rearrangements and prolactin secretion. However, global transcriptional profiling revealed more differentially expressed genes in AG205dEnSC than in dEnSC, compared with non-decidualized EnSC (ndEnSC). In silico analysis showed that PGRMC1 inhibition upregulated more genes related to metabolism, molecular transport, and hormonal biosynthesis compared to control dEnSC. EnSC decidualized in the presence of P4-BSA showed a similar behavior as ndEnSC in terms of morphological features, absence of prolactin secretion, and transcriptomic pattern. Conclusion(s) Our findings associate PGRMC1 to hormonal biosynthesis, metabolism, and vesicular transport—important cellular functions for dEnSC supporting pregnancy. Activation of membrane P4 receptor signaling alone was unable to induce downstream effects needed for proper decidualization.

Prolactinoma in a Male to Female Transgender Woman on Gender Affirming Hormone Therapy

Background: Estrogen promotes prolactin secretion and its role in prolactin secreting adenomas is still under investigation. The genesis and resolution of prolactinomas may be affected by gender affirming hormone therapy. Case Description: A 50 year old transgender female presented with new onset tunnel vision and extremity parathesias for one day. She had been taking oral estrogen for six years. Physical exam showed bilateral gynecomastia as well as galactorrhea. Her visual fields were intact by confrontation. Her brain MRI revealed a pituitary macroadenoma measuring 1.4cm x 1.3cm x 1.3cm. Laboratory studies showed prolactin 538 ng/mL (&lt;20 ng/mL), IGF-1 89.0 (66-303 ng/ml). TSH 1.7 (0.4-4.7 mcIU/mL), Free T4 0.997 (0.58-1.76 ng/dL) and estradiol 103 pg/ml (N/A). She was treated with bromocriptine 5mg daily. Three months later galactorrhea had resolved, but gynecomastia was unchanged. The patient had a prolactin level of 3 ng/mL. Follow up brain MRI at one and three years showed no significant decrease in size of macroadenoma. Discussion: Despite this patient’s biochemical and symptomatic response to dopamine agonist therapy, her pituitary adenoma did not decrease in size over 3 years, which is a common occurrence with macroprolactinomas. However, her prolactin levels declined and her galactorrhea resolved with therapy. Research into the long term effects of gender transition therapy remains mainly in the realm of case reports and retrospective studies. Our review of the medical literature suggests that some transgender patients on estrogen therapy may have an increased risk of developing pituitary adenomas. However, guidelines have not yet been published and currently there are no recommendations for routine imaging or biochemical assessment during follow-up of these patients. While there are established reference ranges for prolactin levels for men, non-pregnant and pregnant women, no established reference ranges for prolactin levels are available for transgender women on gender affirming hormone therapy. Studies indicate that 76-86% of patients have a reduction in the size of their prolactinomas after dopamine agonist therapy. However, the same may not be true for transgender women on transfeminine hormone therapy with estrogen. Conclusion: Prolactinomas may commonly occur and be masked in transgender women on gender affirming hormone therapy due to the expected symptoms of gynecomastia, erectile dysfunction, and diminished libido from reduced testosterone levels. Estrogen promotes prolactin secretion by the pituitary gland and may have a role in development and expansion of prolactinomas. More research is needed in regards to the assessment and monitoring of the pituitary gland in transgender women on gender affirming hormone therapy with estrogen.

Status epilepticus and diabetes ketoacidosis: uncommon clinical presentations of acromegaly

Summary Acromegaly is a rare disease caused by hypersecretion of the growth hormone (GH). Most cases are caused by either pituitary microadenoma or macroadenoma. The GH producing tumors present with clinical manifestations of acromegaly due to excessive GH secretion or symptoms resulting from mass effects of the enlarging tumor. The physical changes are usually slow and, therefore, recognition of the disease is delayed. These adenomas are never malignant but can have significant morbidity and mortality. A subgroup of patients with acromegaly present with severe hyperglycemia resulting in diabetic ketoacidosis (DKA) which requires insulin. Rarely are pituitary tumors responsible for generalized convulsions except when they are too large. We hereby present two cases, the first is that of a 26-year-old female who presented with new onset status epilepticus, DKA with a 1-year history of diabetes mellitus (DM). On examination, she had clinical features of acromegaly. The second case is that of a 34-year-old female who presented with new onset status epilepticus, hyperglycemia with a history of recently diagnosed DM, and features of gigantism. In both cases, their diagnosis was confirmed by elevated serum GH and later by elevated insulin-like growth factor type 1 levels, and CT of the head demonstrating large pituitary macroadenoma. The importance of clinical history and examination, as well as investigations is vital in the recognition of acromegaly. The prognosis of acromegalic patients depends on early clinical recognition and tumor size reduction by either medical or surgical therapy. Learning points Conditions such as status epilepticus and DKA may be clinical presentations in patients presenting with acromegaly. Seizures are rare in people with pituitary adenoma and typically occur when the tumor invades the suprasellar area due to mass effect on the brain. This article shows how best we were able to manage the acromegaly complications in a low resource setting. Hyperprolactinemia in acromegaly may be due to disruption of the normal dopaminergic inhibition of prolactin secretion due to mass effect of the macroadenoma, and around 25% of GH-secreting adenomas co-secrete prolactin.