Main Steroid Research Articles

Effect of Using Fern Leaf Extract (Diplazium esculentum) in Raw Fish Feed on Growth and Feed Conversion Ratio of Mud Crab (Scylla serrata)

Mangrove crab or known as Scylla sp. is one of the biota whose habitat is in coastal waters, especially in mangrove forest areas. The need for mangrove crabs still largely relies on natural catches that are fluctuating. For this reason, it is necessary to cultivate mangrove crabs in a controlled manner that can support an effective, efficient and economically profitable mangrove crab farming business. Cultivation can increase mangrove crab production to meet domestic and foreign market demand. An alternative that can be chosen for mangrove crab cultivation is an apartment system that is arranged vertically in the form of a storage rack. The water medium used in this apartment system cultivation uses a recirculation system. One of the efforts to accelerate the growth of crabs is by using steroid hormones. Ecdysteroid is the main steroid hormone that has the main function as a skin replacement hormone. In addition, this hormone also regulates physiological functions, such as growth, metamorphosis, and reproduction. Therefore, this study was conducted to determine the effect of using fern leaf extract (Diplazium esculentum) in raw fish feed with different doses on growth and feed conversion ratio in mud crabs. This research method uses a field experimental method with a completely randomized design (CRD) consisting of 5 treatments and 3 replicates with different percentage doses, namely: PA: Percentage of 0 ml/Kg, PB: Percentage of 100 ml/Kg, PC: Percentage of 125 ml/Kg, PD: Percentage of 150 ml/Kg, PE: 175 ml/Kg percentage. The results showed that the use of fern leaf extract in mangrove crab (Scylla serrata) feed had a significant effect on the growth of absolute weight, Absolute Length, SGR (Specific Growth Rate) and FCR (Feed Conversion Ratio) where the most influential treatment was treatment E which was the highest dose of 175 ml/Kg. This indicates that treatment E is the best dose treatment for growth and feed conversion ratio of mud crab (Scylla serrata).

The Action of Testosterone in Anti-aging, Rejuvenation, and Longevity

Testosterone is an anabolic androgenic steroid hormone, that is, it is a lipid synthesized from cholesterol-steroid, produced by glands and with regulatory action on cellular physiology hormone, whose main actions are to promote masculinization -androgenic effect- and the synthesis of complex molecules from simpler molecules anabolic effect. Our body's three main steroid substances are androgens, estrogens, and corticosteroids. This literature review aims to verify testosterone's action on rejuvenation, anti-aging, and longevity. The integrative literature review is a method that aims to synthesize results obtained in research on a topic or issue, in a systematic, ordered, and comprehensive way. It is called integrative because it provides broader information on a subject/problem, thus constituting a body of knowledge. To carry out the study, a search for scientific articles was carried out through the Virtual Health Library (SCIELO), Latin American and Caribbean Literature in Health Sciences (LILACS), US National Library of Medicine National Center for Biotechnology Information (PUBMED), Social Network for Scientists (ResearchGate) and Virtual Social Network for Academics (Academia.edu) using the terminologies registered in the health sciences descriptors: Anti-aging, hormones, longevity, rejuvenation, and testosterone.

Nuclear receptor E75/NR1D2 promotes tumor malignant transformation by integrating Hippo and Notch pathways.

Hormone therapy resistance and the ensuing aggressive tumor progression present a significant clinical challenge. However, the mechanisms underlying the induction of tumor malignancy upon inhibition of steroid hormone signaling remain poorly understood. Here, we demonstrate that Drosophila malignant epithelial tumors show a similar reduction in ecdysone signaling, the main steroid hormone pathway. Our analysis of ecdysone-induced downstream targets reveals that overexpression of the nuclear receptor E75, particularly facilitates the malignant transformation of benign tumors. Genome-wide DNA binding profiles and biochemistry data reveal that E75 not only binds to the transcription factors of both Hippo and Notch pathways, but also exhibits widespread co-binding to their target genes, thus contributing to tumor malignancy. We further validated these findings by demonstrating that depletion of NR1D2, the mammalian homolog of E75, inhibits the activation of Hippo and Notch target genes, impeding glioblastoma progression. Together, our study unveils a novel mechanism by which hormone inhibition promotes tumor malignancy, and describes an evolutionarily conserved role of the oncogene E75/NR1D2 in integration of Hippo and Notch pathway activity during tumor progression.

The Effects of Different Types of Diets on Steroid Hormone Concentrations.

The great popularity of various diets in recent years has led us to reflect on their suitability for our health. The aim of this communication is to review current knowledge on the influence of the most well-known diets on the concentrations of the main steroids and to consider possible mechanisms. The influence of diet on hormone concentrations is expected, but the literature data on this topic are inconsistent and yield conflicting results. The main problem in evaluating these influences is the change in weight that a change in diet induces. This effect needs to be filtered out in order to discover interesting associations between diet and steroid hormones. This is illustrated by the example of the effects of ketogenic diets on testosterone levels in men, where the direct effect of the diet is to reduce testosterone levels, but a number of papers have described increases that are due to diet-related weight loss and the modification of obesity-induced changes. A second major driver is the change in circadian rhythm, and it is necessary to assess hormonal changes induced by changing the time of day of the diet. Such shifts within the circadian rhythm rather than due to a particular type of diet itself are documented by changes in the circadian rhythm of cortisol.

Incidence and treatment of checkpoint inhibitor induced immune related adverse events in the emergency department.

438 Background: Immune checkpoint inhibitors (ICPis) are currently indicated for the treatment of various types of cancers. Through inhibition of the cancer cell’s checkpoint mediated T-cell evasion, ICPis allow for sustained T-cell activation and proliferation, resulting in increased cancer cell death. However, due to increased activation and proliferation of cytotoxic T-cells, ICPis have been associated with a wide spectrum of adverse effects, known as immune related adverse effects (irAEs). Although recently updated, the 2018 NCCN and ASCO guidelines state that treatment is based on the location and severity of irAEs. In general, grade 3 toxicities warrant suspension of ICPis and initiation of high-dose corticosteroids. Corticosteroids should then be tapered over the course of at least 4-6 weeks. The diagnosis of an irAE is challenging, toxicities may be misdiagnosed as there is a diverse familiarity with ICPis and their associated irAEs. Subsequently, care may be delayed, and patients may not receive appropriate guideline directed care. Methods: This multi-center chart review included 7 hospitals within a large health system. Our electronic health record was utilized to compile a report of patients who had received ICPIs within 12 months of presentation to an ED within the health system between 9/1/2019 and 9/1/2022. Assessed outcomes included incidence and type of irAE, time between last ICPi dose and presentation to the ED, steroid used for treatment, time to steroid, treatment driver, and the action that deviated from the 2018 ASCO/NCCN guideline directed treatment if applicable. Results: Of the 241 patients receiving an ICPi, 26 (10.8%) experienced an irAE that prompted a visit to an ED within our health system. Guideline directed treatment of irAE(s) was followed 76.9% of the time, with the steroid dose being too low (15.4%) and the steroid taper occurring too quickly (7.7%) being the two reasons for discordant treatment. Treatment started a median of 1.1 after presentation to an ED, with colitis as the most prevalent irAE in 11 patients (42%) presenting. For all patients, there was an average of 20 days between the last ICPi dose and presentation to ED. The main steroid used was methylprednisolone (42%). Conclusions: The 10.8% of patients who experienced an irAE that prompted a visit to an ED within our health system is a lower rate than what has been described in the literature previously, where rates of presentation to an ED due to an irAE ranged from 25-37%. This could be due to previous data coming from before or shortly after the publication of ASCO and NCCN guidelines in 2018, or because our outpatient medical staff has become more aware of irAEs and their appropriate treatment. Limitations include this review was retrospective, no patients had an irAE that was refractory to steroids, and the study period being soon after national guidelines were published.

P-571 Endometrial cortisol and estradiol levels as indicators of pregnancy prognosis and their association with State-Trait Anxiety Inventory (STAI) psychological questionnaire punctuations in IVF patients

Abstract Study question How anxiety and stress questionnaires punctuations and endometrial steroid biosynthesis metabolites levels are associated with embryo implantation and pregnancy outcomes in IVF patients? Summary answer Those patients with high cortisol and low estradiol endometrial levels, were stressed according to STAI State punctuations and showed lower rates of implantation and pregnancy. What is known already Although IVF treatments are known to be a source of stress, its relevance in reproductive success remains controversial, mainly due to the lack of accurate biomarkers. Steroid hormones, such as progesterone and estrogens, are essential for reproductive physiology, and directly related to common stress biomarkers such as cortisol. Nowadays, clinical stress evaluation is exclusively performed via highly subjective psychological questionnaires, not being standardized in the clinical setting. In this study, we measured endometrial levels of main steroid hormones -including cortisol-, stress psychological questionnaires, and reproductive outcomes for determining the relationship among potential stress biomarkers and their influence on fertility. Study design, size, duration In this prospective cohort study, a total of 55 IVF patients were included (<45 years old, no uterine or systemic pathologies and good quality embryos). All endometrial biopsies were collected in mid-secretory phase for metabolites measurement between 2019 and 2021. Also, these patients were psychologically evaluated by stress related questionnaires the same day of biopsy collection or days later in the same cycle. Reproductive outcomes were compared between groups established according to stress biomarkers thresholds. Participants/materials, setting, methods Concentration of eleven selected steroid metabolites was measured by ultra-performance liquid chromatography-tandem mass spectrometry. A psychological questionnaire was also assessed to measure patients’ stress and anxiety (STAI). According to metabolites’ concentration distribution, Barnard’s test was applied to compare the proportion of patients with successful and unsuccessful treatments at different thresholds. A Wilcoxon test was performed for mean metabolites’ levels comparison between stressed and unstressed patients. A p-value of 0.05 was stablished as significant. Main results and the role of chance All patients with high endometrial cortisol levels (>12.5 ng/g; n = 3) failed to implant (100% vs 45.8% non-implanted in > 12.5 ng/g and <12.5 ng/g patients respectively, p-value=0.047), while the totality of patients with high endometrial estradiol levels (>0.9 ng/g; n = 5) achieved embryo implantation and on-going pregnancy (100% vs 50% implanted in > 0.9 ng/g and <0.9 ng/g patients respectively, p-value=0.038). Those patients with high cortisol levels, all not pregnant, had also low estradiol levels and high stress state punctuation (STAI State punctuations ≥ 60). Conversely, patients with high estradiol levels, all pregnant, had also low cortisol levels and were not stressed (STAI State punctuations < 60). Stratification of patients into stressed (STAI State>60) and unstressed (STAI State<60) independently of their implantation results, revealed that 27% of stressed women presented high cortisol levels, while 100% of unstressed patients had low cortisol levels. In the same way, 42% of unstressed patients had high estradiol levels, while 100% of stressed patients presented low estradiol values. Hence, mean values differences of cortisol (10.54 vs 3.05 ng/g) and estradiol (0.55 vs 4.10 ng/g) between stressed and unstressed patients were not significant (p-value=0.08862 and p-value=0.2327, respectively). No relevant associations were observed with stress as a trait. Limitations, reasons for caution Due to the heterogeneous and non-normal metabolites’ concentration distribution among patients, this study would need a largest sample size for having a representative population in the different subgroups. Although we have used molecular and psychological validated stress biomarkers, a causal relationship between stress and reproductive outcomes should be further explored. Wider implications of the findings This study highlighted cortisol and estradiol as potential endometrial biomarkers that better reflect pregnancy prognosis. Further investigation is needed to find if other psychological questionnaires molecularly recognize stressed patients as a noninvasive diagnosis associated to reproductive outcomes. These patients would benefit from psychological counseling to improve their reproductive success. Trial registration number Not applicable

Knowledge and Attitude of Using Anabolic Androgenic Steroids Among Male Bodybuilders in Al-Russafa Baghdad Province

Anabolic androgenic steroids are being more popular between bodybuilders and people who are using gym to enhance their physiques. However, according to the increase of the laws prohibiting sales of these substances without physician prescription, the route of getting and administration practices have become more and more dangerous. Anabolic androgenic steroids include many synthetic derivatives of testosterone its play a significant role in medical treatment. These groups of medications are extensively abused at young ages to rising lean body mass and advance the athletic quality of performance. It has been described that anabolic steroids use can cause many long and short-term side effects, this study intended to assess the level of knowledge about AAS and its effects among gym users. 50 bodybuilders were using gym routinely and were using AASs for at least 2 months and another who were using gym routinely but never use AAS before were interviewed to show their knowledge about short and long term AASs using side effects. Participants used androgenic hormones for three main reasons health, social and personal, vast majority of the participants obtained the required results by using AAS, The majority of participants were going into the gym five to six times weekly for an hour daily. Enanthate was the main steroid abused by the participants and the gym coaches were the major source of selling steroids to the participants. The anabolic androgenic steroids had a harmful effect and abused by large number of young ages.

Comparison of viscous budesonide and fluticasone in the treatment of patients with eosinophilic esophagitis: a systematic review and meta-analysis.

Steroids are an important pharmacologic treatment in patients with eosinophilic esophagitis (EoE). Fluticasone and budesonide are the 2 main steroid medications used in EOE treatment, but current United States (US) guidelines do not recommend one agent over the other. In this study, we conducted a meta-analysis to compare important patient outcomes when both agents are used. A comprehensive search of MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, Cochrane Database of Systematic Reviews, and Scopus was performed from each database's inception to March 29th, 2023. Two independent reviewers systematically identified trials that compared the effect of budesonide vs. fluticasone in the management of EoE. A meta-analysis was performed using a fixed-effects model. The primary outcome was the histologic response (defined as an eosinophil count <15 per high-power field) which reflects the response to treatment. Three studies met our inclusion criteria and were included in the analysis, with a total of 272 patients. All studies were carried out in the US and 1 was a randomized controlled trial. Our meta-analysis showed no statistically significant difference with the use of budesonide compared to fluticasone in achieving a histologic response (odds ratio 1.29, 95% confidence interval 0.77-2.14; P=0.34; I2=0%). Our systematic review and meta-analysis indicated no difference between budesonide and fluticasone in achieving a histologic response in patients with EoE.

An ester derivative of tenacigenin B from Marsdenia tenacissima (Roxb.) Wight et Arn reversed paclitaxel-induced MDR in vitro and in vivo by inhibiting both P-gp and MRP2

Ethnopharmacological relevanceMarsdenia tenacissima is a medicinal plant, used as a raw material for cancer treatment in China. In our previous studies, 11α-O-2-methylbutanoyl-12β-O-tigloyl-tenacigenin B (MT2), the main steroid aglycone isolated from M. tenacissima, was found to significantly enhance the antitumor activity of paclitaxel (PTX) in vivo. However, it is unclear whether MT2 reverses multidrug resistance (MDR) in tumors. Aim of the studyTo determine the role and mechanism of MT2 in reversing tumor MDR. Materials and methodsMDR cell line HeLa/Tax was established from the human cervical carcinoma cell line HeLa by long-term exposure to subtoxic concentrations of PTX and was used to evaluate the ability of MT2 to restore chemosensitivity of cells both in vitro and in a nude mouse model. The expression of P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2) was determined using western blotting and immunohistochemistry. The substrate transport function was assessed using an MDR function assay kit. The binding modes of MT2 and P-gp were determined using the conformation-sensitive anti-P-gp antibodies. The permeability and transport properties of MT2 were analyzed in Caco-2 cell monolayers. ResultsCompared to parental cells, HeLa/Tax cells overexpress P-gp and MRP2 and are approximately 100–360 fold more resistant to the anticancer drugs PTX, docetaxel, and vinblastine. MT2 at 5 or 10 μmol/L significantly increased the sensitivity of HeLa/Tax to these three anticancer drugs (18–56-fold decrease in IC50 value) and suppressed the expression of P-gp and MRP2. Knockdown of P-gp with small interfering RNA partially reversed MT2-induced sensitivity to PTX in HeLa/Tax cells. Moreover, MT2 directly inhibited P-gp-mediated substrate transport while interacting with membrane P-gp in non-substrate ways. MT2 was highly permeable and could not be transported in the Caco-2 cell monolayers. In nude mice bearing HeLa/Tax xenografts, the combination treatment with MT2 and PTX exerted a synergistic inhibitory effect on the growth of tumors and the expression of P-gp and MRP2 without increasing toxicity. ConclusionMT2 is a potential agent for reversing MDR. It impedes membrane drug efflux pumps by suppressing P-gp and MRP2 expression, and directly inhibiting the transport function of P-gp.

A mitochondrial ADXR–ADX–P450 electron transport chain is essential for maternal gametophytic control of embryogenesis in Arabidopsis

Mitochondrial adrenodoxins (ADXs) are small iron-sulfur proteins with electron transfer properties. In animals, ADXs transfer electrons between an adrenodoxin reductase (ADXR) and mitochondrial P450s, which is crucial for steroidogenesis. Here we show that a plant mitochondrial steroidogenic pathway, dependent on an ADXR-ADX-P450 shuttle, is essential for female gametogenesis and early embryogenesis through a maternal effect. The steroid profile of maternal and gametophytic tissues of wild-type (WT) and adxr ovules revealed that homocastasterone is the main steroid present in WT gametophytes and that its levels are reduced in the mutant ovules. The application of exogenous homocastasterone partially rescued adxr and P450 mutant phenotypes, indicating that gametophytic homocastasterone biosynthesis is affected in the mutants and that a deficiency of this hormone causes the phenotypic alterations observed. These findings also suggest not only a remarkable similarity between steroid biosynthetic pathways in plants and animals but also a common function during sexual reproduction.

Pharmacokinetics, plasma protein binding, and metabolism of a potential natural chemosensitizer from Marsdenia tenacissima in rats

Ethnopharmacological relevanceMarsdenia tenacissima (Roxb.) Wight et Arn is a medicinal plant mainly distributed in southwest China. It is used in folk medicine for the treatment of tumors and is synergistic with chemotherapies. In our previous study, 11α-O-2-methybutyryl-12β-O-tigloyl-tenacigenin B (MT2), a main steroid aglycone isolated from the total aglycones of M. tenacissima, significantly enhanced the in vivo antitumor effect of paclitaxel in mice bearing human tumor xenografts, showing its potential as a chemosensitizer. However, the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 remain unclear. Aim of the studyTo elucidate the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 in rats. Materials and methodsMT2 in rat plasma and phosphate-buffered saline was quantified using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method, while the MT2 metabolites in rat liver microsomes were analyzed using UPLC-triple time-of-flight MS/MS. ResultsFor intravenously administered MT2, the maximum plasma concentration and the area under the plasma concentration-time curve indicated dose dependency, while the elimination half-life time, the mean residence time, apparent volume of distribution and total apparent clearance values remained relatively unchanged in both the 5 mg/kg and 10 mg/kg groups. For orally administered MT2, the bioavailability was 1.08–1.11%. In rat plasma, MT2 exhibited a protein binding rate of 93.84–94.96%. In rat liver microsomes, MT2 was metabolized by oxidation alone or in combination with demethylation, and five MT2 metabolites were identified. ConclusionMT2 has low oral bioavailability and a high plasma protein binding rate in rats. After administration, MT2 is transformed into oxidative metabolites in the liver. To achieve a high blood concentration of MT2, it should be administered intravenously. These findings would serve as a reference for further MT2-based pharmacological study and drug development.

P298 Budesonide MMX versus Prednisolone for Ulcerative Colitis: Comparative outcomes from a British retrospective multi-centre study

BackgroundBudesonide MMX (Cortiment) is superior to placebo for mild to moderate UC flares and has a favourable side effect profile. However, no head-to-head data with Prednisolone exist. During the COVID-19 pandemic many IBD units chose Cortiment as first line treatment for outpatients’ flares of UC. The aim of this retrospective study was to compare outcomes of Cortiment vs Prednisolone treatment for UC.MethodsHospital based prescriptions from of 3 UK IBD units were extracted from computerized pharmacy records for the time periods between 1/3/2019 – 30/6/2019 and 1/3/2020 – 30/6/2020. All adult outpatients treated with oral steroids for a flare of UC were included.Baseline data included age, sex, phenotype, IBD medications, symptoms and changes to medication at time of steroid prescription. Follow up data included need for hospital admission for acute severe ulcerative colitis, symptoms at 4 weeks and end of treatment, need for rescue Prednisolone (Cortiment group only). Primary outcome was symptomatic improvement at 4 weeks. ResultsThe 2019 (94 patients) and 2020 (127 patients) cohorts did not differ significantly with regards to age, sex, phenotype and baseline characteristics. The proportion of Cortiment prescriptions rose significantly from 24.5% in 2019 to 70.1% in 2020 (p<0.001). At week 4 of treatment there were statistically significant differences in mean bowel frequency (3.49 in 2019 vs 5.85 in 2020, p=0.001), rectal bleeding <50% (89.7% of patients in 2019 vs 73.1% in 2020, p=0.039) and physician global assessment (39.2% of patients in remission in 2019 compared to 19.8% in 2020, p=0.045).There was no significant difference in hospital admissions, rectal bleeding and physician global assessment at end of treatment.Patients prescribed Cortiment in 2019 had similar baseline characteristics to those prescribed Cortiment in 2020. Mean bowel frequency at four weeks was significantly higher in 2020 (6.18) compared to 2019 (3.69, p=0.034), but rectal bleeding (p=0.388) and physician global assessment at week four did not differ between 2019 and 2020 (p=0.422). Rescue Prednisolone was required in 10% of Cortiment patients in 2019 vs 31.3% in 2020 (p=0.058).ConclusionCortiment was used as the main first line steroid for UC during the pandemic to reduce the risk of adverse COVID-19 outcomes. This change in treatment was, however, associated with worse UC outcomes at 4 weeks and 31% needed rescue Prednisolone. As active IBD is associated with worse COVID-19 outcomes clinicians should carefully evaluate the choice of steroid to achieve optimal disease control and COVID-19 risk minimization.

Prospective evaluation of urinary steroids and prostate carcinoma-induced deviation: preliminary results.

Background The serum prostate-specific antigen is the most widespread biomarker for prostate disease. Its low specificity for prostatic malignancies is a matter of concern and the reason why new biomarkers for screening purposes are needed. The correlation between altered production of the main steroids and prostate carcinoma (PCa) occurrence is historically known. The purpose of this study is to evaluate the modifications of a comprehensive urinary endogenous steroidal profile (USP) induced by PCa, by multivariate statistical methods. Methods A total of 283 Italian subjects were included in the study, 139 controls and 144 PCa-affected patients. The USP, including 17 steroids and five urinary steroidal ratios, was quantitatively evaluated using gas chromatography coupled with single quadrupole mass spectrometry (GC-MS). The data were interpreted using a chemometric, multivariate approach (intrinsically more sensible to alterations with respect to traditional statistics) and a model for the discrimination of cancer-affected profiles was built. Results Two multivariate classification models were calculated, the former including three steroids with the highest statistical significance (e.g. testosterone, etiocholanolone and 7β-OH-DHEA) and PSA values, the latter considering the three steroids' levels only. Both models yielded high sensitivity and specificity scores near to 70%, resulting significantly higher than PSA alone. Conclusions Three USP steroids resulted significantly altered in our PCa population. These preliminary results, combined with the simplicity and low-cost of the analysis, open to further investigation of the potential role of this restricted USP in PCa diagnosis.

TGF-β1 stimulates aromatase expression and estradiol production through SMAD2 and ERK1/2 signaling pathways in human granulosa-lutein cells.

Estradiol (E2), one of the main steroid hormones secreted by the ovaries, plays an important rolein maintaining normal female reproductive function. Ovarian granulosa cells are the main source of E2 production because these cells express aromatase, which is encoded by the CYP19A1 gene and catalyzes the final step in E2 biosynthesis from androgens. Transforming growth factor-beta 1 (TGF-β1) and its receptors are expressed in human granulosa cells, and TGF-β1 expression can be detected in human follicular fluid. To date, TGF-β1 has been shown to regulate various ovarian functions. However, whether aromatase can be regulated by TGF-β1 in human granulosa cells has not been determined. In the present study, we demonstrate that TGF-β1 stimulates aromatase expression in primary human granulosa-lutein cells and in the human ovarian granulose-like tumor cell line, KGN. We used pharmacological inhibitors and small interfering RNA-mediated knockdown approaches to reveal that the SMAD2 and ERK1/2 signaling pathways are involved in TGF-β1-induced aromatase expression and E2 production. These results not only provide important insights into the molecular mechanisms that mediate TGF-β1-induced aromatase expression and E2 production in human granulosa cells but also increase the understanding of the normal physiological roles of TGF-β1 in the ovary.

Developmental mechanisms of adrenal cortex formation and their links with adult progenitor populations

The adrenal cortex is the main steroid producing organ of the human body. Studies on adrenal tissue renewal have been neglected for many years, but recent intensified research has seen tremendous progress in our understanding of the formation and homeostasis of this organ. However, cell turnover of the adrenal cortex appears to be complex and several cell populations have been identified that can differentiate into steroidogenic cells and contribute to adrenal cortex renewal. The purpose of this review is to provide an overview of how the adrenal cortex develops and how stem cell populations relate to its developmental progenitors. Finally, we will summarize present and future approaches to harvest the potential of progenitor/stem cells for future cell replacement therapies.

MiR-18b regulates the function of rabbit ovary granulosa cells

MicroRNAs (miRNAs) have been determined to participate in the process of oestradiol production. Generally, there are two pathways by which oestradiol levels change, one being the state of cells (i.e. the status of enzymes involved in the synthesis of hormones such as oestradiol) and the other being the number of cells that secrete oestradiol. It is known that oestrogens are the main steroids produced by granulosa cells (GCs) of mature ovarian follicles. In this study we explored the function of miR-18b in rabbit GCs by overexpressing or inhibiting its activity. We found that miR-18b silencing promoted the secretion of oestradiol by significantly affecting the expression of steroidogenesis-related genes. Thus, miR-18b may act as a negative regulator of the production of enzymes related to oestradiol synthesis and affect oestradiol production. Furthermore, the effects of miR-18b on the proliferation, cell cycle and apoptosis of GCs were investigated using a cell counting kit (CCK-8) proliferation assay, detection of annexin V-fluorescein isothiocyanate apoptosis, flow cytometry and quantitative polymerase chain reaction. The results showed that miR-18b upregulated GC apoptosis (miR-18b overexpression decreases cell growth and stimulates apoptosis). These findings suggest that miR-18b and the oestrogen receptor 1 (ESR1) gene may be attractive targets to further explore the molecular regulation of GCs. The miR-18b may also explain, in part, the abnormal folliculogenesis in mammals caused by conditions such as polycystic ovary syndrome, primary ovarian insufficiency, and others.

Effects of a gestational level of estradiol on cellular transition, migration, and inflammation in cervical epithelial and stromal cells.

Estrogen (E2) is one of the main steroid hormones associated with pregnancy and parturition. High levels of E2 increase uterine contractions, promote fetal membrane weakening, and induce degradation of the cervical extracellular matrix (ECM). Current evidence supports the role of E2 in epithelial-to-mesenchymal transition (EMT) and inflammation in different cell types; however, its effects on the cellular components of the cervix are still unknown. In this study, we assessed the effects of gestational levels of E2 in: (a) the cellular transition of endocervical epithelial cells (EEC) and cervical stromal cells (CSC) in vitro using immunocytochemical staining and Western blot analyses for EMT markers (cytokeratin-18, E-cadherin, N-cadherin, SNAIL, and vimentin); (b) cell migration using in vitro scratch assays; (c) inflammatory cytokine (interleukin 1β and TNF-α) and MMP9 production under untreated and lipopolysaccharide (LPS)-treated conditions using immunoassays. E2 treatment and co-treatment with LPS as a proxy for infection maintained the metastate of EEC (expression of both cytokeratin and vimentin) and the mesenchymal state of CSC. E2 delayed wound healing, which mimics the tissue remodeling process, in EEC and CSC. E2 led to persistently elevated levels of vimentin throughout the EEC wound healing process. E2 did not affect inflammatory cytokine production by EEC and CSC but increased MMP9 production by EEC. Collectively, these results show that third trimester levels of E2 may permit localized inflammation, increase MMP-9 production, and cause an EMT-mediated impairment of the remodeling process in the cervix in vitro. These data suggest a potential contribution of E2 in cervical ripening.

Actaea racemosa L. extract inhibits steroid sulfation in human breast cancer cells: Effects on androgen formation

BackgroundActaea racemosa L., also known as black cohosh, is a popular herb commonly used for the treatment of menopausal symptoms. Because of its purported estrogenic activity, black cohosh root extract (BCE) may trigger breast cancer growth. Study design/methodsThe potential effects of standardized BCE and its main constituent actein on cellular growth rates and steroid hormone metabolism were investigated in estrogen receptor alpha positive (ERα+) MCF-7 and -negative (ERα-) MDA-MB-231 human breast cancer cells. Cell numbers were determined following incubation of both cell lines with the steroid hormone precursors dehydroepiandrosterone (DHEA) and estrone (E1) for 48 h, in the presence and absence of BCE or actein. Using a validated liquid chromatography-high resolution mass spectrometry assay, cell culture supernatants were simultaneously analyzed for the ten main steroids of the estrogen pathway. ResultsInhibition of MCF-7 and MDA-MB-231 cell growth (up to 36.9%) was observed following treatment with BCE (1-25 µg/ml) or actein (1-50 µM). Incubation of MCF-7, but not of MDA-MB-231 cells, with DHEA and BCE caused a 20.9% reduction in DHEA-3-O-sulfate (DHEA-S) formation, leading to a concomitant increase in the androgens 4-androstene-3,17-dione (AD) and testosterone (T). Actein was shown to exert an even stronger inhibitory effect on DHEA-S formation in MCF-7 cells (up to 89.6%) and consequently resulted in 12- to 15-fold higher androgen levels compared with BCE. The formation of 17β-estradiol (E2) and its glucuronidated and sulfated metabolites was not affected by BCE or actein after incubation with the estrogen precursor estrone (E1) in either cell line. ConclusionsThe results of the present study demonstrated that actein and BCE do not promote breast cancer cell growth or influence estrogen levels. However, androgen formation was strongly stimulated by BCE and actein, which may contribute to their ameliorating effects on menopausal symptoms in women. Future studies monitoring the levels of AD and T upon BCE supplementation of patients are warranted to verify an association between BCE and endogenous androgen metabolism.

Abstract P6-04-13: Preclinical and clinical analyses of estrogen receptor alpha mutant metastatic breast cancer

Abstract Background: Estrogen receptor alpha (ER) positive breast cancer (BC) that recurs as metastatic disease in patients on aromatase inhibitor therapy often harbors mutations in the last exon of the ESR1gene. Such mutations result in amino acid alterations in the ligand binding domain that confer a selective advantage because they render ER constitutively active under estrogen depleted conditions. Hypothesis: We hypothesized that the selective survival advantages gained by metastatic ER mutant (ERmut) tumor cells would become evident in preclinical models simulating the clinical context in which they arise (estrogen deprivation, anchorage independence or metastatic outgrowth) that could then be confirmed in ER+ patient derived xenografts (PDX) and biopsies of metastatic disease. Methods: MCF7 and T47D cells expressing flag-tagged ERmut Y537S or D538G or flag-tagged ER wild type (ERw/t) at the endogenous locus were generated using CRISPR/Cas9. These cells were studied in standard culture conditions as compared to anchorage independent soft agar or poly-HEMA coated plate assays and under long-term estrogen deprivation (LTED) conditions in vitro as well as in vivo in ovariectomized (ovx) mice in late stage metastasis models. ER+ patient derived xenografts (PDX) with naturally occurring ERmutY537S (HCI-013EI) and ERmutL536P (HCI-007) (Dr. Alana Welm) were also studied in vivo. Colony formation assays and tumor growth were evaluated in the context of treatments targeting ER (fulvestrant), androgen receptors (AR) (enzalutamide), steroid hormone synthesis (CYP17 lyase inhibitor seviteronel), or BRD4 Bromo- and Extra-terminal Domain (BET) inhibitor (BETi: JQ1). Drug synergy was determined in vitro using CalcuSyn software or in vivo by tumor caliper measurement and in vivo imaging of tumor luciferase activity. Biopsies of metastatic ER+ BC from clinical trial NCT02953860 were assayed for ESR1 exon 8 mutations using a modified Archer VariantPlex Solid Tumor Assay and immunohistochemistry performed for the four main steroid hormone receptors. Results: ERmutY537S or D538G containing MCF7 and T47D cells exhibited significantly increased anchorage independent survival, a condition in which AR is uniquely upregulated, while ER, progesterone receptors (PR), and glucocorticoid receptors (GR) are not. Furthermore, anti-androgens abrogated the ERmut survival advantage in soft agar. LTED ERmut lines expressed significantly more AR than their non-LTED predecessors or ERw/t counterparts as did ERmut PDX as compared to ERw/t PDX. MCF7 and T47D ERmutY537S and D538G demonstrated a significant increase in AR protein upon anchorage independent survival as compared to ERw/t, with ERmutD538G expressing particularly elevated AR. Biopsies of metastases from consenting patients with ER+ BC confirmed significantly higher AR protein in ERmut metastases as compared to ERw/t by IHC. Treatment of mice with ERmut PDX with combination fulvestrant and anti-androgen decreased tumor growth as compared mice given vehicle control or fulvestrant only. The BET inhibitor JQ1 synergized with anti-androgen to significantly decrease proliferation of ERmut cells. In ovx mice, only ERmut metastases were detectable, while ERw/t did not thrive at metastatic sites. ERmut cells differentially expressed AR-regulated, metastasis-promoting genes and proteins, including chitinase 3-like 1 (CHI3L1), when compared to those with ERw/t. Conclusions: AR is induced by stress encountered during disease progression and treatment, such as anchorage independence and estrogen deprivation. Targeting AR may improve the efficacy of agents such as fulvestrant that inhibit ERmut metastatic disease or delay recurrence, as may agents such as BETi that affect the activity of both ER and AR. Citation Format: Jennifer K Richer, Michelle M Williams, Nicole S Spoelstra, Spencer Arnesen, Jessica L Christenson, Kathleen O’Neill, Jordan M Reese, Zannel D Blanchard, Toru Hanamura, Britta M Jacobsen, Jason Gertz, Anthony Elias. Preclinical and clinical analyses of estrogen receptor alpha mutant metastatic breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-04-13.

Metabolism of Estrogens: Turnover Differs Between Platinum-Sensitive and -Resistant High-Grade Serous Ovarian Cancer Cells.

High-grade serous ovarian cancer (HGSOC) is currently treated with cytoreductive surgery and platinum-based chemotherapy. The majority of patients show a primary response; however, many rapidly develop drug resistance. Antiestrogens have been studied as low toxic treatment options for HGSOC, with higher response rates in platinum-sensitive cases. Mechanisms for this difference in response remain unknown. Therefore, the present study investigated the impact of platinum resistance on steroid metabolism in six established HGSOC cell lines sensitive and resistant against carboplatin using a high-resolution mass spectrometry assay to simultaneously quantify the ten main steroids of the estrogenic metabolic pathway. An up to 60-fold higher formation of steroid hormones and their sulfated or glucuronidated metabolites was observed in carboplatin-sensitive cells, which was reversible by treatment with interleukin-6 (IL-6). Conversely, treatment of carboplatin-resistant cells expressing high levels of endogenous IL-6 with the monoclonal anti-IL-6R antibody tocilizumab changed their status to “platinum-sensitive”, exhibiting a decreased IC50 value for carboplatin, decreased growth, and significantly higher estrogen metabolism. Analysis of these metabolic differences could help to detect platinum resistance in HGSOC patients earlier, thereby allowing more efficient interventions.