Expression Of SF-1 Research Articles

8342 A Rare Case of a Corticotropic and Gonadotropic Pituitary Double Adenoma

Abstract Disclosure: S. Gruber: None. D. Kirschenbaum: None. T. Hortobagyi: None. C. Serra: None. A. Bink: None. F. Beuschlein: None. Z. Erlic: None. Background: Pituitary adenomas account for about 15% of all intracranial neoplasms and typically arise from monoclonal proliferation of anterior pituitary cells. While lactotropic and clinically hormone-inactive adenomas make up by far the largest proportion, somatotropic and corticotropic adenomas have a significantly lower prevalence. This case describes the very rare finding of a corticotropic and gonadotropic double adenoma. Clinical Case: A 63-year-old Caucasian man (BMI 36.7 kg/m2) presented in the obesity clinic. Hypercortisolism was suspected by repeated lack of cortisol suppression in low-dose dexamethasone suppression tests [minimal 87 nmol/l] and disrupted circadian rhythm of cortisol secretion according to salivary cortisol profile. Cortisol levels in a 24-hour sample of urine were normal. ACTH levels were elevated [73 ng/l; n<61 ng/l] leading to suspicion of Morbus Cushing. Concomitant diseases include sleep apnea, arterial hypertension and newly diagnosed type 2 diabetes mellitus. Medical history was notable for deep vein thrombosis and pulmonary embolism. Other stigmata of hypercortisolism were not evident. The rest of endocrine work up confirmed partial secondary hypothyroidism and hypogondotropic hypogonadism. An MRI of the pituitary gland revealed a pituitary macroadenoma with a maximum dimension of 20 mm. The patient underwent an endoscopic transnasal adenomectomy with post-operative corticotropic insufficiency. Contrary to our expectations, the histological examination of the surgical resection revealed a gonadotrophic pituitary adenoma with SF1 and focal FSH expression. The discrepancy between clinical, biochemical and histological findings led to subsequent examinations of the specimen with detection of an small fragment accounting for less than 5% of the total sample with expression of T-Pit and ACTH but neither SF1 nor Pit-1, corresponding to an additional corticotropic adenoma with densely granulated pattern. The tumor cells did not show overlapping expression patterns of transcription factors or hormones, which indicates the coincidence of two distinct adenomas. Three months postoperatively, the patient is biochemically cured. Partial anterior pituitary insufficiency remained. Conclusion: Pituitary double adenomas are found in 0.5-1.5% of all surgical specimens. They are characterized by their origin from different cell lines, which distinguishes them from plurihormonal adenomas, which originate from one single cell line. Here we demonstrate a SF1-T-Pit lineage combination resulting in a neighboring ACTH and FSH-adenoma. In this particular case, the clinically relevant corticotropic adenoma was comparatively tiny, making it easy to miss. This illustrates well the importance of careful correlation of clinical, laboratory and histologic findings as correct diagnosis is crucial for postoperative patient management. Presentation: 6/2/2024

Supplementing ageing male laying breeders with lycopene alleviates oxidative stress in testis and improves testosterone secretion

BackgroundReproductive performance is a crucial aspect of poultry production and is carefully controlled by endocrine, paracrine, and autocrine factors. This study aimed to investigate the effect of lycopene on testosterone synthesis in Leydig cells of laying breeder roosters, clarify the mechanism of lycopene improving Leydig cells function and promoting testosterone production, and explore the role of related signal transduction pathways in testosterone synthesis. ResultsA total of 96 healthy 55-week-old breeding roosters were randomly assigned to one of five dietary treatments. They were provided with a corn-soybean meal-based diet containing different levels of lycopene: 0 mg/kg (control), 50 mg/kg, 100 mg/kg, or 200 mg/kg. The experiment lasted for 6 weeks. With the increase in lycopene levels, the testosterone content in the plasma was significantly higher than in the control group. Testicular Leydig cells were isolated and cultured from fresh testicular tissue of 45-wk-old to 60-wk-old breeding roosters. Various doses of lycopene were administered to Leydig cells, and subsequently, cells were collected for the detection of cell viability and testosterone content. The optimal concentration of lycopene to be added was determined, and changes in mRNA expression and protein levels of key proteins involved in testosterone synthesis were investigated. The results showed that lycopene treatment significantly increased testosterone secretion, mRNA expression, and protein levels of steroid-producing enzymes. Cells were collected to measure the activity of antioxidant enzymes, the mRNA transcription level of apoptotic factors, and the protein expression of apoptotic factors after treatment with lycopene. The results showed that lycopene significantly increased the activities of antioxidant enzymes, and the ability to inhibit oxygen radicals, and decreased the content of malondialdehyde. Apoptosis was inhibited by regulating the expression of apoptosis-inducing and anti-apoptosis factors. After that, the MAPK signaling pathway and downstream SF-1, Nrf2 gene, and protein expression levels were detected. The results showed that lycopene treatment significantly increased the gene and protein expression of JNK, SF-1, and Nrf2, and significantly decreased the gene and protein expression of p38. ConclusionsLycopene treatment could promote testosterone synthesis of testicular Leydig cells by activating MAPK-SF-1 (increasing steroid-producing enzyme level) and MAPK-Nrf2 pathways (resisting oxidative damage).

Gene expression of aromatase, SF-1, and HSD17B2 in menstrual blood as noninvasive diagnostic biomarkers for endometriosis

ObjectiveEndometriosis diagnostic delays are still encountered due to the lack of a reliable, noninvasive diagnostic test. Besides, menstrual blood is a relatively untapped field for diagnostics, yet it provides a readily accessible source for investigating common gynecological conditions. In the present study, we aim to evaluate the expression levels of menstrual blood aromatase, SF-1, and HSD17B2 from women with and without endometriosis and their diagnostic performance. MethodsA total of 40 subjects participated in this study, including 20 patients from each endometriosis and non-endometriosis group. The endometriosis group comprised patients with proven endometriosis confirmed by pathological diagnosis and pelvic ultrasound examination, then requiring endometrial biopsy determined by the clinicians. The non-endometriosis group consisted of women who had primary and secondary infertility and underwent endometrial examination without any visible endometriosis lesion. The menstrual blood and eutopic endometrial tissue of enrolled subjects were collected, and the relative expression of the genes was performed by quantitative real-time polymerase chain reaction (qPCR). ROC curve was used to evaluate the diagnostic efficacy of aromatase, SF-1, and HSD17B2. ResultsWe found significantly higher expressions of aromatase, SF-1, and HSD17B2 in the menstrual blood of the endometriosis group compared to non-endometriosis (P < 0.05). In contrast, examination of eutopic endometrial tissue of both groups only found significant in HSD17B2 (P < 0.05), while aromatase and SF-1 showed no statistically significant variance. The Area Under Curve (AUC) of aromatase, SF-1, and HSD17B2 in the menstrual blood was 0.977, 0.862, and 0.807, respectively. The optimal cutoff value was determined to be >1.63 (sensitivity = 95 % and specificity = 90 %) for aromatase, >1.71 (sensitivity = 90 % and specificity = 80 %) for SF-1, and >1.83 (sensitivity = 80 % and specificity = 75 %) for HSD17B2. ConclusionOur study showed that aromatase, SF-1, and HSD17B2 in the menstrual blood solidly discriminate between endometriosis and non-endometriosis patients with high diagnostic accuracy. However, further confirmation in larger cohorts is required to validate the reliability of these biomarkers to endometriosis.

Chemotherapy drug combinations induced maternal ovarian damage and long-term effect on fetal reproductive system in mice

With the postponement of female reproductive age and the higher incidence of cancer in young people, fertility preservation has become increasingly important in childbearing age. Chemotherapy during pregnancy is crucial for maternal cancer treatments and fetal outcomes. It is a need to further study ovarian damage caused by chemotherapy drug combinations and long-term effects on offspring development, and a detailed understanding of side effects of chemotherapy drugs.In this study, chemotherapy drug combinations significantly impacted on ovarian function, especially epirubicin/cyclophosphamide (EC) combination led to an unbalance in the development of the left and right ovary. Exposure to EC and cisplatin/paclitaxel (TP) increased the number of progenitor follicles while decreased the count of antral follicles and corpora luteum. As to the estrus cycle, EC exposure resulted in a longer estrus period and diestrus period, while TP exposure only extended the diestrus period. EC and TP affected steroid biosynthesis by reducing the expression of SF1 and P450arom.γ-H2AX was detected in both EC and TP exposure groups.As to the impact on the offspring from 4T1 tumor-bearing pregnant mice injected with EC, no significant difference was observed in the physical and neurological development compared to the control, but the ovarian weights, estrus cycles of the offspring were significantly different. Chemotherapy drug combinations exhibit ovarian toxicity, not only causing direct damage on the follicle cells but also disrupting steroid biosynthesis. The reproductive system of offspring from maternal tumor-bearing mice exposed to chemotherapy drugs was observed disorder, but the concrete mechanism still needs further exploration.

An extensive immunohistochemical analysis of 290 ovarian adult granulosa cell tumors with 29 markers.

The current knowledge about the immunohistochemical features of adult granulosa cell tumor (AGCT) is mostly limited to the "traditional" immunohistochemical markers of sex cord differentiation, such as inhibin, calretinin, FOXL2, SF1, and CD99. Knowledge about the immunohistochemical markers possibly used for predictive purpose is limited. In our study, we focused on the immunohistochemical examination of 290 cases of AGCT classified based on strict diagnostic criteria, including molecular testing. The antibodies used included 12 of the "diagnostic" antibodies already examined in previous studies, 10 antibodies whose expression has not yet been examined in AGCT, and 7 antibodies with possible predictive significance, including the expression of HER2, PD-L1, CTLA4, and 4 mismatch repair (MMR) proteins. The results of our study showed expression of FOXL2, SF1, CD99, inhibin A, calretinin, ER, PR, AR, CKAE1/3, and CAIX in 98%, 100%, 90%, 78%, 45%, 41%, 94%, 82%, 26%, and 9% of AGCT, respectively. GATA3, SATB2, napsin A, MUC4, TTF1, and CD44 were all negative. PTEN showed a loss of expression in 71% of cases and DPC4 in 4% of cases. The aberrant staining pattern (overexpression) of p53 was found in 1% (3/268) of cases, 2 primary tumors, and 1 recurrent case. Concerning the predictive markers, the results of our study showed that AGCT is microsatellite stable, do not express PD-L1, and are HER2 negative. The CTLA4 expression was found in almost 70% of AGCT tumor cells.

Effects of thyroxin and luteinizing hormone releasing hormone on reproductive physiology of Rohu (Labeo rohita): Insights into spawning performance, oocyte maturation, steroidogenesis, and follicular development genes

As the global aquaculture industry grows, attention is increasingly turning towards assisted reproductive technologies. In this study, we examined the impact of D-Ala6, Pro9-Net-mGnRH (LHRHa: 0.4 mL/kg) and two doses (1 and 10 μg/kg fish) of thyroxin (T4) administered through a single injection on oocyte maturation, spawning performance, sex steroid hormone levels, as well as the expression of genes related to steroidogenesis and follicle development (ZP2, Cyp19a1a and SF-1) in Rohu (Labeo rohita). The study found that untreated female Rohu did not spawn, while those treated with LHRHa and thyroxin ovulated and spawned across a hormonal gradient. The highest spawning success was observed with a thyroxin dosage of 10 µg/kg (no significant change with a dose of 1 μg/kg), and female latency period decreased with increasing dosage. Additionally, females treated with thyroxin exhibited significantly higher fecundity than other experimental groups. Treatment with LHRHa and two doses of thyroxin significantly increased the gonadal somatic index compared to the control and sham groups. Hormonal treatment also led to increased fertilization success, hatching rate, and larval survival. At 12 h post-injection, females treated with thyroxin exhibited a significant decline in estradiol levels and expression of Zp2, Cyp19a1a, and SF-1 compared to other experimental groups. Levels of DHP significantly increased across the hormonal gradient. Histological analyses supported a steroidogenic shift, where oocyte maturation was accelerated by hormone administration, particularly with both doses of thyroxin. In conclusion, the findings suggest that thyroxin is a recommended treatment for assisted reproduction of Rohu due to its ability to induce spawning, increase fecundity and improve larval survival.

Atrazine disorganises laminin formation and reduces cell numbers in the tammar testis during early differentiation.

Atrazine, like oestrogen, disorganises laminin formation and reduces the number of germ cells and Sertoli cells in the developing testes of the tammar wallaby. This study suggests that interfering with the balance of androgen and oestrogen affects the integrity of laminin structure and testis differentiation. The herbicide atrazine was banned in Europe in 2003 due to its endocrine disrupting activity but remains widely used. The integrity of the laminin structure in fetal testis cords requires oestrogen signalling but overexposure to xenoestrogens in the adult can cause testicular dysgenesis. However, whether xenoestrogens affect laminin formation in developing testes has not been investigated. Here we examined the effects of atrazine in the marsupial tammar wallaby during early development and compare it with the effects of the anti-androgen flutamide, oestrogen, and the oestrogen degrader fulvestrant. The tammar, like all marsupials, gives birth to altricial young, allowing direct treatment of the developing young during the male programming window (day 20-40 post partum (pp)). Male pouch young were treated orally with atrazine (5 mg/kg), flutamide (10 mg/kg), 17β-oestradiol (2.5 mg/kg) and fulvestrant (1 mg/kg) daily from day 20 to 40 pp. Distribution of laminin, vimentin, SOX9 and DDX4, cell proliferation and mRNA expression of SRY, SOX9, AMH, and SF1 were examined in testes at day 50 post partum after the treatment. Direct exposure to atrazine, flutamide, 17β-oestradiol, and fulvestrant all disorganised laminin but had no effect on vimentin distribution in testes. Atrazine reduced the number of germ cells and Sertoli cells when examined at day 40-50 pp and day 20 to 40 pp, respectively. Both flutamide and fulvestrant reduced the number of germ cells and Sertoli cells. Atrazine also downregulated SRY expression and impaired SOX9 nuclear translocation. Our results demonstrate that atrazine can compromise normal testicular differentiation during the critical male programming window.

Differentiation of bone marrow mesenchymal stem cells into Leydig-like cells with testicular extract liquid in vitro.

Differentiation of Leydig cells plays a key role in male reproductive function. Bone marrow mesenchymal stem cells (BMSCs) have emerged as a potential cell source for generating Leydig-like cells due to their multipotent differentiation capacity and accessibility. This study aimed to investigate the morphological and genetic expression changes of BMSCs during differentiation into Leydig-like cells. Testicular extract liquid, which simulates the microenvironment in vivo, induced the third passage BMSCs differentiated into Leydig-like cells. Changes in cell morphology were observed by microscopy, the formation of lipid droplets of androgen precursor was identified by Oil Red Staining, and the expression of testicular specific genes 3β-HSD and SF-1 in testicular stromal cells was detected by RT-qPCR. BMSCs isolated from the bone marrow of Sprague-Dawley (SD) rats were cultured for 3 generations and identified as qualified BMSCs in terms of morphology and cell surface markers. After 14 days of induction with testicular tissue lysate, lipid droplets appeared in the cytoplasm of P3 BMSCs by Oil Red O staining. RT-qPCR detection was performed on BMSCs on the 3rd, 7th, 14th, and 21st day after induction. Relative expression levels of 3β-HSD mRNA significantly increased after 14 days of induction, while the relative expression of SF-1 mRNA increased after 14 days of induction but was not significant. BMSCs can differentiate into testicular interstitial cells with reserve androgen precursor lipid droplets after induction by testicular tissue lysate. The differentiation ability of BMSCs provides the potential to reconstruct the testicular microenvironment and is expected to fundamentally improve testicular function and provide new treatment options for abnormal spermatogenesis diseases.

Dorsomedial Ventromedial Hypothalamic Nucleus Growth Hormone-Releasing Hormone Neuron Steroidogenic Factor-1 Gene Targets in Female Rat.

The prospect that the ventromedial hypothalamic nucleus (VMN) transcription factor steroidogenic factor-1/NR5A1 (SF-1) may exert sex-dimorphic control of glucose counterregulation is unresolved. Recent studies in male rats show that SF-1 regulates transcription of co-expressed hypoglycemia-sensitive neurochemicals in dorsomedial VMN growth hormone-releasing hormone (Ghrh) neurons. Gene knockdown and laser-catapult-microdissection/single-cell multiplex qPCR techniques were used here in a female rat model to determine if SF-1 control of Ghrh neuron transmitter marker, energy sensor, and estrogen receptor (ER) variant mRNAs varies according to sex. Data show that in females, hypoglycemia elicits a gain of SF-1 inhibitory control of VMNdm Ghrh neuron Ghrh and Ghrh-receptor gene profiles and loss of augmentation of glutaminase transcription; SF-1 gene silencing diminished eu- and hypoglycemic patterns of neuronal nitric oxide gene transcription. SF-1 imposes divergent control of baseline and hypoglycemic glutamate decarboxylase65 (GAD)-1 (stimulatory) versus GAD2 (inhibitory) mRNAs in that sex. SF-1 stimulates baseline VMNdm Ghrh neuron PRKAA1/AMPKα1 and PRKAA2/AMPKα2 gene expression, yet causes opposite changes in these gene profiles during hypoglycemia. SF-1 exerts glucose-dependent control of ER-alpha and G-protein-coupled ER-1 transcription, but blunts ER-beta gene profiles during eu- and hypoglycemia. In females, SF-1 knockdown did not affect hypercorticosteronemia or hyperglucagonemia, but blunted hypoglycemic suppression of growth hormone secretion. Results show that SF-1 expression is critical for female rat VMNdm Ghrh neuron counterregulatory neurochemical, AMPK catalytic subunit, and ER gene transcription responses to hypoglycemia. Sex differences in direction of SF-1 control of distinctive gene profiles may result in observed disparities in SF-1 regulation of counterregulatory hormone secretion between sexes.

Clinical and Molecular Characteristics of Gonadotroph Pituitary Tumors According to the WHO Classification

Since 2017, hormone-negative pituitary neuroendocrine tumors expressing the steroidogenic factor SF1 have been recognized as gonadotroph tumors (GnPT) but have been poorly studied. To further characterize their bio-clinical spectrum, 54 GnPT defined by immunostaining for FSH and/or LH (group 1, n = 41) or SF1 only (group 2, n = 13) were compared and studied for SF1, βFSH, βLH, CCNA2, CCNB1, CCND1, caspase 3, D2R, and AIP gene expression by qRT-PCR. Immunohistochemistry for AIP and/or D2R was performed in representative cases. Overall, patients were significantly younger in group 1 (P = 0.040 vs group 2), with a similar trend excluding recurrent cases (P = 0.078), and no significant difference in gender, tumor size, invasion or Ki67. SF1 expression was similar in both groups but negatively correlated with the patient’s age (P = 0.013) and positively correlated with βLH (P < 0.001) expression. Beta-FSH and AIP were significantly higher in group 1 (P = 0.042 and P = 0.024, respectively). Ki67 was unrelated to gonadotroph markers but positively correlated with CCNB1 (P = 0.001) and negatively correlated with CCND1 (P = 0.008). D2R and AIP were strongly correlated with each other (P < 0.001), and both positively correlated with SF1, βFSH, βLH, and CCND1. AIP immunopositivity was frequently observed in both groups, with a similar median score, and unrelated to Ki67. D2R immunostaining was best detected with a polyclonal antibody and mostly cytoplasmic. This study indicates that hormone-negative GnPT tend to occur in older patients but do not significantly differ from other GnPT in terms of invasion or proliferation. It also points out the current limits of D2R immunostaining in such tumors.

SF-1 Induces Nuclear PIP2.

Metazoan cell nuclei contain non-membrane pools of the phosphoinositide lipid PI(4,5)P2 (PIP2), but how this hydrophobic lipid exists within the aqueous nucleoplasm remains unclear. Steroidogenic Factor-1 (NR5A1, SF-1) is a nuclear receptor that binds PIP2 in vitro, and a co-crystal structure of the complex suggests the acyl chains of PIP2 are hidden in the hydrophobic core of the SF-1 protein while the PIP2 headgroup is solvent-exposed. This binding mode explains how SF-1 can solubilize nuclear PIP2; however, cellular evidence that SF-1 expression associates with nuclear PIP2 has been lacking. Here, we examined if tetracycline induction of SF-1 expression would associate with nuclear accumulation of PIP2, using antibodies directed against the PIP2 headgroup. Indeed, tetracycline induction of wild-type SF-1 induced a signal in the nucleus of HEK cells that cross-reacts with PIP2 antibodies, but did not cross-react with antibodies against the lower abundance phosphoinositide PI(3,4,5)P3 (PIP3). The nuclear PIP2 signal co-localized with FLAG-tagged SF-1 in the nuclear compartment. To determine if the nuclear PIP2 signal was dependent on the ability of SF-1 to bind PIP2, we examined a "pocket mutant" of SF-1 (A270W, L345F) shown to be deficient in phospholipid binding by mass spectrometry. Tetracycline induction of this pocket mutant SF-1 in HEK cells failed to induce a detectable PIP2 antibody cross-reactive signal, despite similar Tet-induced expression levels of the wild-type and pocket mutant SF-1 proteins in these cells. Together, these data are the first to suggest that expression of SF-1 induces a PIP2 antibody cross-reactive signal in the nucleus, consistent with X-ray crystallographic and biochemical evidence suggesting SF-1 binds PIP2 in human cells.

Overview of the New World Health Organization Classification of Pituitary Neuroendocrine Tumors

The 5th edition of the World Health Organization(WHO)classification of endocrine and neuroendocrine tumors was revised in 2022. A major nomenclature change was from "adenoma" to "pituitary neuroendocrine tumor"(PitNET). Tumors are classified based on the cell lineage as determined by expression of transcription factors, such as, PIT1, TPIT, and SF1. Therapeutic modalities available for management of functioning PitNETs include surgery, medical treatment, and irradiation. Transsphenoidal surgery is the gold-standard surgical treatment. The aim of the present study was to summarize changes from the previous edition of the WHO classification and discuss topics regrading PIT1, TPIT, and SF1-lineage PitNETs with reference to clinicopathological findings.

Mechanisms of DHEA-induced AMH increase in the ovarian tissues of PCOS rat

The present study aimed to build a DHEA-induced polycystic ovary syndrome (PCOS) rat model to evaluate the potential mechanism of DHEA-induced AMH rise in these rat ovarian tissues. A total of 36 female 3-week-old rats were allocated into two groups at random. The control group received merely the same amount of sesame oil for 20 days while the experimental group received 0.2 mL of sesame oil Plus DHEA 6 mg/100 g daily. Both groups' vaginal opening times were noted, and vaginal smears were taken. By using RT-qPCR and Western blot, the mRNA and protein expression of AMH, GATA4, SF1, and SOX9 in the ovarian tissues of the two groups was investigated.The rats in the experimental group appeared to have obvious disorders of the estrus cycle, as evidenced by the ratio of estrus being significantly higher than that in the control group (P < 0.05); HE staining revealed that the ovarian volume, follicular vacuoles, and follicular lumen of the rats in the experimental group increased significantly.The ELISA results revealed that T and AMH in the experimental group were higher than those in the control group at day 15 and 20. AMH、GATA4 and SF1 mRNA and protein expression were higher in the experimental group than in the control group on day 15 and 20 (P < 0.05). On day 20, the experimental group outperformed the control group (P < 0.05). In the DHEA-induced PCOS rat model, androgen may have enhanced AMH expression via increasing the expression of genes associated to the AMH promoter binding site (GATA4, SF1, SOX9).

Effect ofhigh temperatures onsex ratio anddifferential expression analysis (RNA-seq) ofsex-determining genes inOreochromis niloticus from different river basins inBenin.

The high temperature sex reversal process leading to functional phenotypic masculinization during development has been widely described in Nile tilapia (Oreochromis n iloticus) under laboratory or aquaculture conditions and in the wild. In this study, we selected five wild populations of O. niloticus from different river basins in Benin and produced twenty full-sib families of mixed-sex (XY and XX) by natural reproduction. Progenies were exposed to room temperature or high (36.5°C) temperatures between 10 and 30 days post-fertilization (dpf). In control groups, we observed sex ratios from 40% to 60% males as expected, except for 3 families from the Gobé region which showed a bias towards males. High temperature treatment significantly increased male rates in each family up to 88%. Transcriptome analysis was performed by RNA-sequencing (RNA-seq) on brains and gonads from control and treated batches of six families at 15 dpf and 40 dpf. Analysis of differentially expressed genes, differentially spliced genes, and correlations with sex reversal was performed. In 40 dpf gonads, genes involved in sex determination such as dmrt1, cyp11c1, amh, cyp19a1b, ara, and dax1 were upregulated. In 15 dpf brains, a negative correlation was found between the expression of cyp19a1b and the reversal rate, while at 40 dpf a negative correlation was found between the expression of foxl2, cyp11c1, and sf1 and positive correlation was found between dmrt1 expression and reversal rate. Ontology analysis of the genes affected by high temperatures revealed that male sex differentiation processes, primary male sexual characteristics, autophagy, and cilium organization were affected. Based on these results, we conclude that sex reversal by high temperature treatment leads to similar modifications of the transcriptomes in the gonads and brains in offspring of different natural populations of Nile tilapia, which thus may activate a common cascade of reactions inducing sex reversal in progenies.

Role of miR-300-3p in Leydig cell function and differentiation: A therapeutic target for obesity-related testosterone deficiency

MicroRNAs (miRNAs) regulate various cellular functions, but their specific roles in the regulation of Leydig cells (LCs) have yet to be fully understood. Here, we found that the expression of miR-300-3p varied significantly during the differentiation from progenitor LCs (PLCs) to adult LCs (ALCs). High expression of miR-300-3p in PLCs inhibited testosterone production and promoted PLC proliferation by targeting the steroidogenic factor-1 (Sf-1) and transcription factor forkhead box O1 (FoxO1) genes, respectively. As PLCs differentiated into ALCs, the miR-300-3p expression level significantly decreased, which promoted testosterone biosynthesis and suppressed proliferation of ALCs by upregulating SF-1 and FoxO1 expression. The LH/METTL3/SMURF2/SMAD2 cascade pathway controlled miR-300-3p expression, in which luteinizing hormone (LH) upregulated SMAD-specific E3 ubiquitin protein ligase 2 (SMURF2) expression through methyltransferase like 3 (METTL3)-mediated Smurf2 N6-methyladenosine modification. The Smurf2 then suppressed miR-300 transcription by inhibiting SMAD family member 2 (SMAD2) binding to the promoter of miR-300. Notably, miR-300-3p was associated with an obesity-related testosterone deficiency in men and the inhibition of miR-300-3p effectively rescued testosterone deficiency in obese mice. These findings suggested that miR-300-3p plays a pivotal role in LC differentiation and function, and could be a promising diagnostic or therapeutic target for obesity-related testosterone deficiency.

Cbx2 is a functional target of the let-7 family in the gonad of Japanese flounder (Paralichthys olivaceus).

As a key member of the miRNA family, the role and target gene of the let-7 family in the gonad of Japanese flounder (Paralichthys olivaceus) is unclear. Chromobox homolog 2 (CBX2) is one of the core components of the polycomb group complex (PcG) and significantly influences gonadal development. The deletion of CBX2 can lead to sex reversal in mammals. Therefore, exploring the relationship between the let-7 family and cbx2 is crucial to clarify the role played by the let-7 family in the gonad of Japanese flounder. We predicted and verified the target interaction between the let-7 family and cbx2. The results showed that cbx2 was a direct target of let-7d, let-7e, let-7g, let-7j, and let-7b. Among them, let-7d, let-7e, let-7g, and let-7j exhibited an extremely significant targeting relationship with cbx2 (p<0.001). Taking let-7g as an example, we further investigated the regulatory role between let-7g and cbx2 in the gonad by miRNA overexpression and inhibition experiments in primary testis cells. The results revealed that let-7g could negatively regulate cbx2 at the level of primary testis cells. And the expression of sf1 (steroidogenic factor 1) was also significantly decreased after the interference of cbx2 siRNA. This suggests that the let-7 family may be involved in the Japanese flounder gonadal development via targeting cbx2.

Transcriptomic Classification of Pituitary Neuroendocrine Tumors Causing Acromegaly

Acromegaly results from growth hormone hypersecretion, predominantly caused by a somatotroph pituitary neuroendocrine tumor (PitNET). Acromegaly-causing tumors are histologically diverse. Our aim was to determine transcriptomic profiles of various somatotroph PitNETs and to evaluate clinical implication of differential gene expression. A total of 48 tumors were subjected to RNA sequencing, while expression of selected genes was assessed in 134 tumors with qRT-PCR. Whole-transcriptome analysis revealed three transcriptomic groups of somatotroph PitNETs. They differ in expression of numerous genes including those involved in growth hormone secretion and known prognostic genes. Transcriptomic subgroups can be distinguished by determining the expression of marker genes. Analysis of the entire cohort of patients confirmed differences between molecular subtypes of tumors. Transcriptomic group 1 includes ~20% of acromegaly patients with GNAS mutations-negative, mainly densely granulated tumors that co-express GIPR and NR5A1 (SF-1). SF-1 expression was verified with immunohistochemistry. Transcriptomic group 2 tumors are the most common (46%) and include mainly GNAS-mutated, densely granulated somatotroph and mixed PitNETs. They have a smaller size and express favorable prognosis-related genes. Transcriptomic group 3 includes predominantly sparsely granulated somatotroph PitNETs with low GNAS mutations frequency causing ~35% of acromegaly. Ghrelin signaling is implicated in their pathogenesis. They have an unfavorable gene expression profile and higher invasive growth rate.

Sarcomatoid Adrenal Cortical Carcinoma: Report of a Rare Case

Abstract Introduction/Objective Adrenal cortical carcinoma (ACC) is a rare neoplasm with oncocytic, myxoid, and sarcomatoid variants. Sarcomatoid is the least common variant. To our knowledge, there are only 23 cases of sarcomatoid ACC reported in the English literature. Typically, ACC’s express SF1, Melan A, inhibin, and synaptophysin. It has been reported that the sarcomatoid component does not express the aforementioned immunomarkers; however, it can display mutational expression of p53 and β-catenin. Methods/Case Report A 77-year-old man was incidentally found to have a left adrenal gland mass. Hormone levels were unremarkable, and adrenalectomy was performed. Gross examination of the specimen revealed a 4.5 cm well-circumscribed tan and focally hemorrhagic mass confined to the adrenal gland. Histologically, the mass was composed of pleomorphic round to polygonal cells with clear to eosinophilic cytoplasm and spindle cells. The mitotic count exceeded 20 in 50 high power fields. Lymphovascular invasion and extensive necrosis were present. Immunohistochemical studies revealed that the sarcomatoid component expressed SF1 and synaptophysin focally. The non-sarcomatoid component exhibited focal expression of synaptophysin and diffuse expression of SF1 and Melan A. Both components displayed null expression of p53, and neither expressed β-catenin. Ki67 proliferation index was &amp;gt; 95% in the sarcomatoid component. Results (if a Case Study enter NA) NA Conclusion In sarcomatoid ACC, both sarcomatoid and non-sarcomatoid components have been shown to harbor TP53 and CTNNB1 mutations, with the latter leading to activation of Wnt pathway and causing diffuse and strong nuclear expression of β-catenin. In the present case, tumor cells lacked expression of p53, which might have played a role in the evolution of the tumor.

ODP293 Aberant Expression of SF-1 Transcription Factor in ACTH-secreting Pituitary Tumors: Possible Association With Aggressive Disease Evolution

Abstract Cushing's disease represents a rare endocrine disorder caused by hypercortisolism due to an ACTH-secreting pituitary tumor. Pituitary adenomas (PAs) are mostly benign tumors but may have features of invasiveness and aggressiveness. They are clinically classified by their hormone-secreting characteristics. Expression of pituitary hormones in adenohypophyseal cells is controlled by several transcription factors (PIT1 for GH, PRL and TSH lineage, SF1 for gonadotroph lineage and TPIT for ACTH lineage). Eight cases of Cushing's disease (CD) were included in the current study. The diagnostic was sustained by clinical features, hormonal assessment, and radiological findings (pituitary CT or MRI). All patients underwent surgical removal of pituitary tumor using transsphenoidal resection. All tissue specimens were evaluated using histological routine staining (hematoxylin and eosin) and immunohistochemical staining for vimentin, pituitary hormones (GH, PRL, TSH, FSH, LH, ACTH), and transcription factors (PIT1, TPIT, SF1). The intensity IHC scores ranged from 0 to 3+ and a staining higher than 30% (2+) was considered positive for interpreting the results. Based on the histopathological and immunohistochemical data correlated with the evolution of the disease (remission versus persistent disease), five molecular subgroups were identified. Subgroups 1 to 3 had a positive expression of ACTH and TPIT, with variations regarding SF1 expression: there was no expression in subgroup 1, nuclear positivity in 2, and cytoplasmic reaction in 3. Subgroups 4 and 5 consisted of plurihormonal adenomas with unusual immunohistochemical combination. They had IHC positive score for ACTH, GH, TPIT and were differentiated by the expression of SF1: lack of expression in subgroup 4 and intense cytoplasmic reaction in 5. The expression of SF1 was linked to the evolution of the disease, patients from subgroups which did not have SF1 cytoplasmic reaction (1, 2, 4) had remission of CD, while patients in subgroups 3 and 5 had persistent disease (including one case who evolved to exitus due to cardiovascular complications of the severe hypercortisolism). Transcription factors evaluation is important for classifying pituitary adenomas and finding clinicopathological correlations with the aim of providing a better therapeutical approach. In the current study, although SF1 is a marker of gonadotroph lineage (1), its aberrant cytoplasmic expression in CD may contribute to an aggressive evolution and to other cell lineage differentiation (PIT1, TPIT). Neou M et al, Pangenomic Classification of Pituitary Neuroendocrine Tumors. Cancer Cell. 2020 Jan 13;37(1): 123-134. e5. doi: 10.1016/j. ccell.2019.11. 002. Epub 2019 Dec 26. PMID: 31883967. Presentation: No date and time listed

Integration of RNAi and RNA-seq uncovers the regulation mechanism of DDX20 on vitellogenin expression in Scylla paramamosain

Vitellogenesis in crustaceans is controlled by several steroid hormones. In humans, the expression of SF-1, a gene that regulates gonadal development and the synthesis of steroid hormones, is affected by DDX20. However, how the homologous gene FTZ-F1 is regulated by DDX20 and its association with vitellogenesis remains unknown in the mud crab Scylla paramamosain. In this study, SpDDX20 and SpFTZ-F1 were identified in the transcriptome of mature ovarian tissue from the mud crab. qRT-PCR results revealed that the expression levels of SpFTZ-F1 and SpVTG in the ovaries of crab in the experimental group injected with dsDDX20 (EO) were significantly higher (P < 0.05) than those in the negative control group injected with dsEGFP (NO) and the blank control group injected with SPSS (BO). The differentially expressed genes (DEGs) identified by comparative transcriptome analysis of the EO group and NO group were enriched into five pathways related to ovarian steroidogenesis. The expression of CYP17, CYP3A4, CYP1A1 and 3β-HSD were up-regulated in pathways related to steroid hormone production and biosynthesis. The expression of the INSR, IRS and PI3K genes in the insulin signaling pathway were significantly increased (P < 0.05). The expression level of the TGF-β gene was up-regulated (P < 0.05) in the transforming growth factor pathway, whereas the expression level of the Smad2 gene was down-regulated (P < 0.05). The expression of GnRHR, GS, AC and PKA genes in the gonadotropin-releasing hormone signaling pathway were up-regulated. Our data provide a foundation for investigating the relationship between DDX20 and FTZ-F1 in the regulation of vitellogenin expression in S. paramamosain.