bluuebunny/crossref_metadata_2025 huggingface dataset:
FRI592 Pediatric Diabetes Mellitus And The COVID-19 Pandemic
Authors: Cooper Felicia Ilyssa , Thokalath Adrien , Chalise Sweta , Vyas Neha , Gurnurkar Shilpa
Published: October 2023

Disclosure: F.I. Cooper: None. A. Thokalath: None. S. Chalise: None. N. Vyas: None. S. Gurnurkar: None.

Introduction: Evidence suggests that during the COVID-19 pandemic, adults had delayed detection of diabetes mellitus (DM) and its complications as well as an increase in the severity of DM symptoms. Our retrospective study aimed to determine the difference in prevalence of new onset DM and to compare the severity of diabetic ketoacidosis (DKA) in these children prior to and during the COVID-19 pandemic at Nemours Children’s Health in Orlando, FL. Methods: Chart review was performed on children with new onset DM (type 1 and 2) pre-pandemic and during the pandemic (9/1/2017-9/1/2022). 509 participants met inclusion criteria. Demographic information and laboratory data were collected. One sample proportions test was used to compare the proportion of new onset DM diagnoses and severity of presentation. Chi square test was used to test association of demographics with DM onset and DKA status. Statistical significance was set at p<0.05 and all analyses were run on IBM SPSS 28.0. Results: A greater number of pediatric DM diagnoses were made during the COVID-19 pandemic when compared to pre-pandemic [295 vs 214 (p<0.001)]. More patients presented in DKA at initial diagnosis during the pandemic compared to pre-pandemic (52% vs 48%, p=0.046). There was no significant difference in DKA severity between the pre-pandemic and pandemic groups (p=0.179). There was a significant difference in the average Hemoglobin A1C at diagnosis between COVID-19 unknown and COVID-19 negative group (12% vs. 10.6%, p<0.05). Conclusion: Our study found that more children were diagnosed with DM during the COVID-19 pandemic compared to an equal length of time pre-pandemic. We also found that the prevalence of DKA among those with new-onset DM was higher during the pandemic period. It is speculated that SARS-CoV-2 can disturb self-tolerance and trigger autoimmune responses through cross-reactivity with host cells. Additional studies are needed to investigate this mechanism specifically among children with new onset DM.

Presentation: Friday, June 16, 2023
OR04-02 KANK1 Deficiencies Underlie A Subset of Male Congenital Genitourinary Anomalies
Authors: Lamb Dolores J , O'Neill Marisol Ann
Published: October 2023

Disclosure: D.J. Lamb: Advisory Board Member; Self; Roman Health, American Board of Bioanalysts, American Association of Bioanalysts, Northeastern Region. Consulting Fee; Self; Roman Health. Stock Owner; Self; Fellow Health. M.A. O'Neill: None.

Development of the genitourinary (GU) system involves several complex processes directed both by genetic and hormonal signals essential for proper sexual differentiation. Studying genetic and genomic variants can provide insight into genes which drive GU development. Using array comparative genomic hybridization (aCGH), 9p24 was identified as a putative hotspot for copy number variants (CNVs) in patients with non-syndromic GU anomalies. Analysis of genes in the minimal overlap region of these CNVs revealed KANK1 as a candidate for GU development. KANK1, an anchor protein, which has been studied in kidney cancer and cerebral palsy, is an important regulator of cytoskeletal formation through the polymerization of f-actin. Using DECIPHER, ICSA, ClinVar, and published datasets a total of 43 CNVs (30 duplications, 13 deletions) encompassing KANK1, variants were identified in patients with GU anomalies including both upper tract and lower tract anomalies. The incidence of KANK1 CNVs was further established by screening non-syndromic patients with GU anomalies from Texas Children’s Hospital by Taqman CNV qPCR and 2.2% of patients screened have KANK1 CNVs. Since the majority of KANK1 CNVs identified encompassed of more than one gene, KANK1 was further analyzed as a causative gene for GU anomalies through an analysis of exome sequencing variants in the University of Washington Center for Mendelian Genomics (Geno2MP). Through Geno2MP, 46 KANK1 missense variants were identified in patients with GU anomalies including hypospadias, cryptorchidism, and nephrotic syndrome. Of these variants, 31 were predicted to be deleterious or probably deleterious by Polyphen2. A mouse model of Kank1 loss was generated by CRISPR/Cas9 deletion to further analyze Kank1’s role in GU development and function. Kank1 deficient males were found to have decreased penile length, abnormal development of the urethral opening, and renal dysfunction. These phenotypes are consistent with those found in patients with Kank1 CNVs and SNPS supporting the hypothesis that KANK1 regulates GU development and function.

Presentation: Thursday, June 15, 2023
FRI129 Adrenal Insufficiency In Olmsted County, MN, USA: A Population-Based Study
Authors: Moazzami Mitra , Achenbach Sara , (Beth) Atkinson Elizabeth J , Bancos Irina
Published: October 2023

Disclosure: M. Moazzami: Grant Recipient; Self; Rochester Epidemiology Project. S. Achenbach: None. E. Atkinson: None. I. Bancos: None.

Background: Epidemiology of adrenal insufficiency (AI) is incompletely characterized with most data originating from referral centers and registries. We aimed to determine the incidence, morbidity index, risk of hospitalizations, and mortality in patients with AI living in Olmsted County, Minnesota. Methods: We assessed the standardized incidence rate, morbidity index, risk of hospitalizations and mortality of AI in residents (aged 18+) in Olmsted County, MN, USA, from Jan 1, 2005, to December 31, 2021. The data was collected using the Rochester Epidemiology Project (REP), a database of medical records for all Olmsted County residents since 1966. Incidence rates were standardized for age and sex according to the 2020 US white population. Sex- and age- matched referent subjects were selected from the REP. AI was defined based on presence of oral chronic glucocorticoid therapy (hydrocortisone, prednisone, prednisolone, or dexamethasone) defined as (1) any hydrocortisone use or (2) 6 months of consecutive use of prednisone, prednisolone, or dexamethasone or (3) two AI diagnosis codes 6+ months apart plus at least one glucocorticoid prescription within 1 year of the second diagnosis. Elixhauser morbidity index was used. Findings: Between 2005 and 2021, 5419 patients were diagnosed with AI (median age of 62 years, range 18-103), 56% women). Overall standardized incidence rates for AI increased from 206 (95% CI 174-238) in 2005 to 252 (223-280) per 100 000 person-years in 2021. At baseline, patients with AI had a higher morbidity index (median 3 vs median 1, p <0.001) and a higher history of hospitalizations within 5 years prior to baseline (57% vs 30%; p<0.001) when compared to referent subjects. When comparing patients with AI to referent subjects, and after adjusting for age, sex, morbidity index and prior history of hospitalizations, patients with AI were more likely to be hospitalized during follow-up (aHR 1.4, 95% CI 1.3-1.5). When comparing patients with AI to referent subjects, and after adjusting age, sex, and morbidity index, patients with AI had higher mortality (aHR 1.7, 95% CI 1.6-1.9). Patients with AI demonstrated a higher risk of dying from musculoskeletal disorders (aHR 4.9, 95% CI 2.2-10.8), respiratory disorders (aHR 4.7, 95% CI 3.3-6.6), neoplasms (aHR 2.4, 95% CI 2.0-2.9), and infectious disorders (aHR 2.3, 95% CI 1.2-4.3) than referent subjects. Interpretation: We demonstrate an increase in AI incidence between 2005 and 2021. Surprisingly, even after controlling for morbidity and prior hospitalizations, patients with AI were more likely to be hospitalized. In comparison to referent subjects, mortality was higher in patients with AI, even after adjustment for morbidity index.

Presentation: Friday, June 16, 2023
FRI419 Progesterone Regulation Of The GnRH Pulse Generator
Authors: Wall Ellen Gabrielle
Published: October 2023

Disclosure: E.G. Wall: None.

A group of kisspeptin neurons located in the arcuate nucleus (ARN) of the hypothalamus represent the GnRH pulse generator and their activity is thought to be controlled primarily by the feedback actions of estradiol and progesterone (P4). In a healthy menstruating woman, following the mid-cycle ovulation, P4 levels dramatically increase and exert a negative feedback action onto the hypothalamus to slow pulsatile LH secretion. However, the exact mechanism by which P4 negatively feeds back to the hypothalamus remains unknown. The ARN kisspeptin neurons display synchronized episodes (SEs) of activity approximately every hour for most of the cycle but these dramatically reduce in number following the LH surge. As these ARN kisspeptin neurons express the progesterone receptor (PR) it is hypothesized that they may be the site at which P4 negative feedback occurs. Given the lack of detailed information in the mouse, we first assessed the profile of circulating P4 levels in mice at 8-hr intervals throughout the estrous cycle. Terminal blood samples were taken from individual mice along the 4 stages of the estrous cycle at 3-time points 8 hrs apart and the gonadal hormones were analyzed using LC-MS. The concentration of P4 was found to peak around the time of the LH surge at 6 pm on proestrus (21.94 ± 3.78 ng/ml) and was at its lowest at diestrus 10 am (0.42 ± 0.11 ng/ml). We next assessed the impact of circulating P4 on the activity of the ARN kisspeptin pulse generator using GCaMP fibre photometry in freely behaving mice. A prior study found that IP administration of 8 mg/kg P4 resulted in circulating P4 concentrations reaching 34 ng/ml (Wong et al., J Pharm Pharmacol 64, 2012). Hence, in this study, diestrus mice equipped for monitoring the activity of the kisspeptin pulse generator were injected with either 8 or 4 mg/kg P4 at 10 am and SEs monitored for 24 hrs. Both doses resulted in a significantly decreased frequency of SEs compared to the vehicle (p<0.05), which persisted for 6 hrs. To next examine whether P4 may be acting directly on the ARN kisspeptin neurons to slow pulse generation, P4 (25 ng/ml) was infused directly into the ARN while the activity of the pulse generator was monitored. Again, this resulted in a decreased frequency of SEs compared to the vehicle with the effects lasting approximately 5 hrs (p<0.05).These results indicate that P4 exerts powerful inhibitory control over the activity of the kisspeptin pulse generator and that these effects may be direct. Elucidating the mechanisms of P4 negative feedback may provide a platform for the development of new therapeutic strategies in the infertility clinic.

Presentation: Friday, June 16, 2023
FRI421 Immunomodulatory Therapy Can Improve Fertility In Women With Autoimmune Premature Ovarian Insufficiency - A Proof Of Concept Study
Authors: Hirschberg Angelica Linden , Björnsdottir Sigridur , Gunnarsson Iva , Sergouniotis Fotios , Vogt Elinor , Øksnes Marianne , Husebye Eystein Sverre , Bensing Sophie , Kämpe Olle
Published: October 2023

Disclosure: A.L. Hirschberg: None. S. Björnsdottir: None. I. Gunnarsson: None. F. Sergouniotis: None. E. Vogt: None. M. Øksnes: None. E.S. Husebye: None. S. Bensing: None. O. Kämpe: None.

Introduction: Autoimmune disorders are associated with increased risk of premature ovarian insufficiency (POI), leading to early menopause and infertility. The etiology of autoimmune POI remains unknown, but measurable levels of anti-Mullerian hormone indicate that some follicles may be intact. We asked if rituximab, a B cell specific anti-CD20 monoclonal drug could improve the ovarian response to gonadotropin stimulation and restore fertility.

Methodology: In this open label pilot study, nine women aged 18-35 years, BMI 19-30 kg/m2 with autoimmune POI (S- FSH > 40 IE/L), of which six had Addison’s disease, were included. The procedure comprised controlled ovarian hyperstimulation according to a standard high dose protocol with daily subcutaneous injections of follitropin alpha, prior to, and 4 months after, two infusions of 1-gram rituximab 2 weeks apart, and a follow-up period of 12 months. If positive response to ovarian stimulation was obtained, GnRH antagonist was given to prevent premature ovulation and choriogonadotropin alpha to induce ovulation. When possible, oocytes were retrieved for cryopreservation and later fertilization.

Results: None of the nine patients responded to ovarian stimulation prior to rituximab treatment (mean stimulation days 15.7 ± 1.6 days). After rituximab therapy, five of seven responded with follicular development, and in four women mature eggs were harvested (oocyte yield 3-13 oocytes) for preservation. Four of these five women experienced spontaneous menstrual bleedings and decreased vasomotor symptoms. All responders had autoimmune Addison’s disease, whereas two women with no response had Myasthenia gravis and Hashimoto thyroiditis, respectively. We observed no serious drug reactions or other complications.

Conclusions: We have shown for the first time that rituximab can restore menstrual bleedings and enable egg retrieval with the potential to restore fertility in patients with autoimmune POI. This could become an alternative fertility treatment to egg donation, which is the only option today.

Presentation: Friday, June 16, 2023
FRI429 Association Between Nonalcoholic Fatty Liver Disease And Fib-4 In Patients With Hypogonadism
Authors: Godar Cassandra M , Brown Kevin F , Spiro Andrew J , Vietor Nicole O , Hoang Thanh D , Shakir Mohamed K M
Published: October 2023

Disclosure: C.M. Godar: None. K.F. Brown: None. A.J. Spiro: None. N.O. Vietor: None. T.D. Hoang: None. M.K. Shakir: None.

Introduction: Recent studies have reported a strong bidirectional association between non-alcoholic fatty liver disease (NAFLD) and hypogonadism in both genders. Men with hypogonadism may have higher noninvasive indices of NAFLD via hepatic steatosis index (HSI, reference range <36) and hepatic fibrosis via FIB-4 (reference range <1.3). However, there is limited clinical data on the association between serum testosterone concentrations and NAFLD in men. We report here HSI and FIB-4 in 3 men with hypogonadism.

Case #1: 56-year-old male with a 20-year history of idiopathic hypogonadism, hypertension on telmisartan and amlodipine, and obstructive sleep apnea with regular CPAP use. Pituitary MRI is normal. He uses 3 pumps of topical testosterone gel daily. Vital signs: BP 139/83mmHg, HR 59bpm and BMI 36.2. Examination of heart and lungs is normal, testes are 15cc bilaterally. Serum testosterone is 400-500 ng/dL, alanine aminotransferase (ALT) 88 U/L (0-50), aspartate aminotransferase (AST) 71 U/L (17-59), alkaline phosphatase (ALP) 78 U/L (38-126), iron saturation 43.1% (20-55), ferritin 452 ng/mL (30-400). Calculated FIB-4: 2.3, HSI: 46.

Case #2: 71-year-old male with 46XX phenotype with hypogonadism and gynecomastia on 2 pumps of transdermal testosterone gel daily. Vital signs: BP 150/79 mmHg, HR 67/min, BMI 36.8. Examination of heart and lungs is normal, and testes are 5cc bilaterally. Serum testosterone is 416 ng/dL, ferritin 509 ng/mL, iron saturation 41.3%, ALT 19 U/L, AST 22 U/L, ALP 136 U/L. Liver ultrasound demonstrated diffuse hepatic steatosis with borderline hepatomegaly. Calculated FIB-4: 1.38, HSI: 48.4.

Case #3: 48-year-old male with central hypogonadism currently taking testosterone 100 mg IM every week. Evaluation for central hypogonadism was unrevealing for pituitary lesion. Vital signs of BP 130/91, HR 76/min, BMI 27.2. Examination of heart and lungs is normal, and testes are 20cc bilaterally. ALT 201U/L, AST 80U/L, ALP 136U/L, serum testosterone 241 ng/dL. Liver ultrasound is consistent with hepatic steatosis. Calculated FIB-4: 1.22, HSI 59.2.

Conclusion: Patients with a combination of hypogonadism, high BMI, mild or moderate elevations of AST/ALT/ALP, elevated serum ferritin or transferrin saturation should undergo screening for NAFLD. In this series, all 3 patients had high HSI, an early indicator of fibrosis although only 1 patient had a high FIB-4 level. As fibrosis is the strongest predictor for long-term clinical outcomes among patients with NAFLD there is growing interest in employing non-invasive methods such as HSI, FIB-4, NFS and APRI for risk stratification. In conclusion, it is recommended that high risk patients with hypogonadism need to be screened for NAFLD and hepatic fibrosis with these simple tools.

Presentation: Friday, June 16, 2023
FRI453 Genetic Heterogeneity of Hypothalamic Hypogonadism DIsorders
Authors: Bheeman Chandni , Astapova Olga
Published: October 2023

Disclosure: C. Bheeman: None. O. Astapova: None.

Title: Genetic Heterogeneity of Hypothalamic Hypogonadism disorders. Introduction: Hypothalamic hypogonadism (HH) can be defined as having inappropriately low serum concentrations of gonadotropins in the presence of low circulating concentrations of sex steroids. It represents a group of genetically and clinically heterogeneous disorders. Case A 32-year-old female who presented to the endocrine clinic for evaluation of infertility. Patient had menarche at age 12 but became amenorrhoeic at age 14. Her physical exam was notable for normal female secondary sexual characteristics and intact sense of smell. She had no history of bone fracture or menopausal symptoms. The patient had failed the progesterone withdrawal test twice, but had withdrawal bleeding with the birth control pill or with hormonal replacement therapy. She had failed ovulation induction with clomiphene citrate, but was able to conceive 3 pregnancies through in vitro fertilization. Patient’s labs off of hormonal replacement and birth control were: LH <0.3. FSH 0.6, Estradiol <5, Progesterone 0.4. Her prolactin level was within normal limits at 6.6 and her TSH was 1.08. MRI of the brain showed no abnormalities in the pituitary gland. Genetic testing showed that the patient was heterozygous for a duplication mutation in the ANOS1 gene. Conclusion The genotypic and phenotypic heterogeneity that can be seen in HH disorders is due to the numerous genes involved in hypothalamic hypogonadism and their intricate interactions. Mutations contributing to HH can be involved in either the neurodevelopmental or neuroendocrine pathways of the hypothalamic pituitary gonadal axis or in both. There are mutations that can be inactivating variants while some can play the role of the modifier genes. The later may be able to explain the incomplete penetrance and variable expressivity sometimes seen in HH. It is important to note that though > 60 genes have been implicated in HH 50% of the cases remain genetically undiagnosed. This case is particularly interesting as this patient is a female with a heterozygous mutation on the ANOS1 gene, the primary gene associated with Kallmann syndrome. Kallmann syndrome is a X-linked recessive disorder and therefore the patient having phenotypical expression of the mutation is unexpected. Through the presentation of this case we aim to explore different modes of genetic expression and the different mechanisms that could have led to this patient hypothalamic hypogonadism.

Presentation Date: Friday, June 16, 2023
FRI170 Ultrasound-Based Evaluation Of Muscles And Tendons In Patients With Cushing Syndrome
Authors: Bolayir Basak , Gogus Feride , Demir Beril , Yalcin Mehmet Muhittin , Meray Jale , Altinova Alev , Toruner Fusun Saadet , Akturk Mujde Yasim
Published: October 2023

Disclosure: B. Bolayir: None. F. Gogus: None. B. Demir: None. M.M. Yalcin: None. J. Meray: None. A. Altinova: None. F.S. Toruner: None. M.Y. Akturk: None.

Background: Myopathy, being induced by antianabolic and catabolic effects of excess corticosteroids is one of the characteristic features of Cushing Syndrome (CS). Muscle thickness which has been shown to be correlated with muscle cross sectional area can be used as an indicator of muscle mass. Although many studies have demonstrated effects of corticosteroids on collagen metabolism and case reports documented rupture of tendons in patients with chronic exposure to corticosteroids, tendon changes of patients with endogenous hypercortisolism have not been studied. Ultrasound, although not studied well in this group of patients, can be an alternative imaging method that provides information about muscle and tendon thickness. The aim of this study was to investigate muscle, tendon thicknesses and serum levels of myostatin in patients with CS. Methods: Twenty-two patients with CS and nineteen healthy control subjects similar for age, sex, height and weight were enrolled. Lower extremity tendon and muscle groups’ thickness were evaluated by ultrasound. Serum myostatin levels were measured by ELISA. Results: Age, height, weight and body mass index were similar between groups (p>0.05). The thicknesses of rectus femoris, vastus medialis, vastus lateralis, gastrocnemius medialis and gastrocnemius lateralis were lower in the CS group than control group (p≤0.001). However, thickness of soleus muscle was not significantly different between the groups (p > 0.05). No significant difference was revealed between groups in terms of Achilles and patellar tendon thickness (p>0.05). Median serum myostatin levels were found to be lower in the CS group than the control group (p<0.001).Conclusions: Ultrasound based measurement of muscle thickness can be used to assess and monitor steroid-induced changes of muscle mass. The fact that the thickness of the proximal muscles of lower extremity was found to be lower by ultrasound in patients with CS in our study supports the proximal myopathy feature of CS. Besides that, it was shown that the gastrocnemius lateralis and medialis thicknesses, which are the distal muscles of the lower extremity, were also shown to be lower in the Cushing group. On the other hand, CS might not affect the thickness of the lower extremity tendons. Moreover, the mechanism of the decrease in the level of myostatin, the negative regulator of muscle growth, in patients with needs to be elucidated with CS further studies.

Presentation: Friday, June 16, 2023
SAT353 Relevance of Ovarian Morphology Among Adolescents With Polycystic Ovary Syndrome (PCOS)
Authors: Christ Jacob P , Shinkai Kanade , Corley Jamie , Pasch Lauri , Cedars Marcelle Ivonne , Huddleston Heather
Published: October 2023

Disclosure: J.P. Christ: None. K. Shinkai: None. J. Corley: None. L. Pasch: None. M.I. Cedars: None. H. Huddleston: None.

Background: The 2018 international guidelines for the assessment and management of PCOS exclude ovarian morphology from the diagnosis of PCOS in adolescents due to limited data regarding normative values in this population. Adolescents with oligo-amenorrhea (OA) or hyperandrogenism (HA) alone are thus excluded from diagnosis and ultrasonography has been removed from clinical use without an assessment of its utility in this population. Objective: To characterize ovarian morphology among adolescents with PCOS and evaluate the relevance of PCOM among those currently excluded from diagnosis. Methods: A cross-sectional analysis of patients that underwent a standardized evaluation for PCOS at a single tertiary center from 2006 to 2022 was completed. Participants were included if they were within 8 years of menarche or were <20 years of age at screening and had data on ovarian morphology. Follicle number per ovary (FNPO) and ovarian volume (OV) were assessed by transvaginal ultrasound, or if not tolerated, transabdominal (in which case FNPO was omitted) or transrectal approach. PCOM was defined as a FNPO of >20 follicles or an OV >10cc in either ovary. Maximum FNPO and OV in either ovary was used for analyses. Age based criteria were used to define irregular menstrual cycles and were characterized among those >1 year post menarche. Patients were grouped into those with OA and HA (PCOS-NIH), with OA or HA as well as PCOM (PCOS-Rotterdam), and with one or no features (non-PCOS). Results: Of the 133 subjects included, 95 met PCOS-NIH criteria, 22 met PCOS-Rotterdam criteria and 16 met non-PCOS criteria. Prevalence of PCOM among PCOS-NIH was 82.1%. Significant differences were seen between Non-PCOS, PCOS-Rotterdam, and PCOS-NIH for FNPO (15.5±8.7 vs 30.9±12.5 vs 31.8±14.6), OV (6.1±4 vs 10.8±4.5 vs 11.4±6.4), waist:hip ratio (WHR) (0.7±0.1 vs 0.8±0.1 vs 0.8±0.1), 2-hour glucose (99.5±17.1 vs 84.8±18.9 vs 105.6±41 uIU/mL), QUICKI (0.4±0 vs 0.4±0.1 vs 0.3±0.1), and HOMA-IR (1.9±1.2 vs 2.4±2.1 vs 4.7±5.7) (p<0.05 for overall comparisons between groups). Among PCOS-Rotterdam and Non-PCOS, in multivariable models controlling for WHR, systolic blood pressure (β=0.5, p=0.09) and free testosterone (β=0.03, p=0.03) associated with FNPO and QUICKI (β=69.1, p=0.003) associated with OV. Conclusion: Adolescents meeting NIH criteria appear to be at greatest metabolic risk. Ovarian morphology, however, may reflect degree of hyperandrogenism and metabolic dysfunction among adolescents currently excluded from diagnosis of PCOS (i.e. PCOS-Rotterdam and non-PCOS groups). The inclusion of ultrasound assessment may thus be able to aid in risk stratification and phenotyping among those without a definitive PCOS diagnosis.

Presentation Date: Saturday, June 17, 2023
SAT362 Low-dose DHT’s Effects On Gluconeogenesis And Energy Metabolism Are Ameliorated By Central Nervous System Knockout Of The Androgen Receptor
Authors: Ubba Vaibhave , Joseph Serene , Akbar Razeen , Dsilva Milan , Wu Sheng
Published: October 2023

Disclosure: V. Ubba: None. S. joseph: None. R. Akbar: None. M. Dsilva: None. S. Wu: None.

Polycystic ovarian syndrome (PCOS) is a leading cause of infertility in women of reproductive age. Mostly, the level of androgens are deviated from their physiological values in females, resulting in hyperandrogenemia. Androgen manifests its function in the cells via the Androgen Receptor (AR), making it a prime factor for studying PCOS pathophysiology. We generated central nervous system (CNS) specific AR knockout mouse (SynARKO), using synapsin promoter driven cre, to examine its role under normal and hyperandrogenemic conditions in female mice. Assessment of reproductive parameters under physiological levels of androgens, revealed no differences in the cyclicity and fertility of SynARKO mice when compared to Control (Con-veh) mice. These parameters were altered under hyperandrogenemia (induced using DHT) in Control mice (Con-DHT) mice and were not rescued in the SynARKO- DHT mice, suggesting CNS AR is dispensable to reproductive features in females. Evaluation of metabolic functions by Glucose tolerance test, we observed alleviated glucose intolerance in SynARKO-DHT (4 months). Energy expenditure and food intake were increased in Con-DHT mice compared to Con-veh, while showing no further differences in SynARKO-DHT mice compared to SynARKO-veh. At the molecular level, DHT treatment showed increased phosphorylated Akt in hypothalamus of control mice. Interestingly, it was restored in synARKO upon DHT treatment. Further, the inflammation marker, phosphorylated NF-kB p-65, was increased in the hypothalamus of Con-DHT mice. Additionally, IBA, a marker of macrophage activation was also increased upon DHT treatment in control mice. Taken together, our results suggest that while the reproductive functions are not mediated by the CNS AR, metabolic effects caused by hyperandrogenemia, is regulated by the central nervous system AR.

Presentation Date: Saturday, June 17, 2023
SAT485 Knockout Of TLR4 Alters Reproductive And Metabolic Outcomes In Mouse Model Of Polycystic Ovary Syndrome
Authors: Dominique Wiggins Kiara , De Robles Gabriela , De La Torre Jamie-Jean , Nicholas Dequina
Published: October 2023

Disclosure: K.D. Wiggins: None. G. De Robles: None. J. De La Torre: None. D. Nicholas: None.

Title: Knockout of TLR4 Alters Reproductive and Metabolic Outcomes in Mouse Model of Polycystic Ovary Syndrome. Polycystic Ovarian Syndrome (PCOS) is a heterogeneous endocrine disorder often characterized by reproductive and metabolic dysfunction. Prior literature has found a correlation between PCOS and chronic low-grade inflammation. However, it remains unknown if inflammation plays a causal role in the pathology of PCOS. In women with PCOS, serum levels of Lipopolysaccharides (LPS), a ligand for Toll-like receptor 4 (TLR4), are elevated. The binding of LPS to TLR4 leads to the production of pro-inflammatory cytokines, which can result in low-grade inflammation and metabolic syndrome, a prominent feature of PCOS. Given the role of TLR4, we hypothesize that the induced inflammatory environment mediates the development and pathogenesis of PCOS. To test this hypothesis, we subcutaneously implanted a nonsteroidal aromatase inhibitor, Letrozole, in TLR4 knockout mice (TLR4KO) and C57BL/6J (wild type) mice. Following five weeks of letrozole treatment, wild type mice failed to cycle, whereas TLR4KO mice were able to enter each estrous cycle phase. Furthermore, in contrast to wild type mice, TLR4KO mice display normalized Luteinizing Hormone (LH) levels with increased levels of Follicle Stimulating Hormone (FSH). We also characterized how inflammation influences the metabolic phenotypes of PCOS. Our results reveal that the deletion of TLR4 reduces weight gain and improves glucose intolerance. Moreover, we show that the development of insulin resistance in letrozole-treated TLR4KO mice was independent of other metabolic outcomes. Our study demonstrates that disruption of this pro-inflammatory pathway mediates the development of reproductive and metabolic phenotypes. Thus, research on the modulation of chronic low-grade inflammation may lead to the development of new therapeutics for PCOS patients.

Keywords: Reproduction, Inflammation, Polycystic Ovary Syndrome,

Presentation Date: Saturday, June 17, 2023
THU578 Concept: Role Of Spiral Steroids (SS) During Pregnancy
Authors: Chasalow Fred I
Published: October 2023

Disclosure: F.I. Chasalow: None.

Background: Earlier, my laboratory reported 3 novel endogenous steroids (SS) that cross react with digoxin-specific antibodies (DLM). The SS are readily distinguished from all other steroids by 3 characteristics: [1] each is a phosphocholine diester, [2] each has more than 21 carbon atoms in the steroid fragment and [3] the extra carbons form an E-ring with a cyclic lactone. The discovery included a biosynthetic scheme with known enzymes and no need for a ‘magic’ step. Function: NaK-ATPase has 3 subunits, designated FXYD, α, and β. Affinity for Na+ and K+ ions is regulated by FXYD proteins. Each SS matches a tissue-specific α-subunit. Each α-subunit has 2 different binding sites for steroids. One site binds a SS; the ligand of the other site is unknown. For example, Ionotropin is specific for the α subunit present in cardiac and renal tissue. Kaleotropin is isolated from gonads, but the α subunit which matches each specific ovarian cell type is unknown. Galotropin (G-trop): One of the SS, G-trop, was detected as a DLM in human breast cyst fluids, specifically cysts with high K+ ion levels. Solubility properties distinguished it from previously known cardiac glycosides and, like other DLM, it inhibited NaK-ATPase. G-trop was also detected in milk from cows, sheep and goats. Ions derived from Ionotropin (m/z=341 Da) or Kaleotropin (m/z=369 Da) were not present in milk extracts. On LC-MS, G-trop fragmented to an ion at m/z= 381 Da. Trial and error analysis suggests C25O4H34. No other formula is consistent with m/z=381 Da. G-trop: G-trop wasn’t present in serum from children. At 22-26 weeks of gestation, whether or not a pregnant woman had pre-eclampsia, serum had low levels of G-trop. Maternal G-trop increases during the 3rd trimester, coincidentally with initiation of milk production; simultaneously, K+ ion levels in milk increased. Pre-eclampsia. In a pilot study, 65% of patients (n=20) with pre-eclampsia had elevated levels of Ionotropin precursor and 30% had elevated levels of Kaleotropin precursor. 25% of patients with pre-eclampsia develop life-threatening hypertension or seizures, typical of the expected response to elevated Ionotropin levels. Therapy includes immediate delivery of the infant at 30-36 weeks of gestation. Maternal G-trop synthesis has not yet begun and mother’s milk has low K+ ion levels. Elevated maternal Kaleotropin is not the cause of maternal hypertension, suggesting a specific gestational maternal α-subunit is its target. Kaleotropin from placenta may be the signal to respond to maternal hypokalemia. Necrotizing enterocolitis (NEC):In utero the fetus does not absorb K+ from the GI tract because fetal need for K+ ion is provided via the placenta. Concept: After premature delivery, without G-trop in milk, the infant can’t absorb K+ from the GI tract; the GI tract fails to grow and tight junctions do not form. As a result, infants are at high risk for developing NEC.

Presentation Date: Thursday, June 15, 2023
FRI159 Syndrome of Apparent Mineralocorticoid Excess (AME) from Licorice Tea
Authors: Clark Lambert Peter
Published: October 2023

Disclosure: P.C. Lambert: None.

We report a patient who had Syndrome of Apparent Mineralocorticoid Excess (AME) from licorice tea. Pt is a 65 y/o man with past medical history of HLD, HTN who presented to clinic for routine follow up. BMP was normal. Six months later patient reported diarrhea, bloating and gas after visiting family. To alleviate symptoms, he tried apple cider vinegar, herbal teas and stopped coffee and caffeinated drinks. His BP was 162/92. BMP showed Na 145, K 3.1, CO 37 and 20 meq of potassium daily was started. Follow up in 5/2020 found persistent elevated BP of 160/102 with elevated BP on home monitor and leg edema. Lisinopril-HCTZ was changed to amlodipine 5 mg. BMP showed Na 146, K 2.2, CO 44. Potassium was increased to 40 meq bid. Two weeks later BP was 160/90. Amlodipine was increased to 10 mg daily. He reported resolution of his edema. BMP showed Na 131, K 3.1, CO 33. At this point, the renin/aldosterone axis was tested with aldosterone <1.0 ng/dL and PRA <0.167 ng/mL/hr. On subsequent discussion it was found pt was taking black licorice tea. After cessation, pt’s BP improved and pt was gradually titrated off potassium supplements and blood pressure meds entirely. Repeat lab testing on 6/2020 found aldosterone 2.4 ng/dL, PRA 0.455 ng/mL/hr, Na 142, K 4.8, CO 31. On return to clinic on 8/2020 BP was 124/72 off medication. His labs were Na 143, K 4.1, CO 30. Syndrome of Apparent Mineralocorticoid Excess (AME) is caused by a deficiency in the 11-beta-hydroxysteroid dehydrogenase enzyme type 2 isoform (11-beta-HSD2). The enzyme converts cortisol to cortisone which cannot bind to the mineralocorticoid receptor. Deficiency of this enzyme will lead to more cortisol binding to the mineralocorticoid receptor which binds just as avidly as aldosterone. This leads to increased mineralocorticoid activity without increased aldosterone. Causes of AME can be genetic or induced by the steroid glycyrrhetinic acid, in licorice, that inhibits 11-beta-HSD2. Licorice candy in the USA generally does not have licorice root. It can be found in many sources including chewing tobacco and flavored chewing gum besides tea. Pt can take as low as 50 g daily of confectionery licorice for two weeks to produce elevated blood pressure. Patients will present with hypokalemia hypertension, metabolic alkalosis, low plasma renin activity and low plasma aldosterone levels. Urinary free cortisone and cortisol levels can assist with diagnosis but as in this case review of history was enough to diagnose and treat the patient.

Presentation Date: Friday, June 16, 2023
FRI444 Testosterone Therapy in Men with Classical vs Functional Hypogonadism: Results from a Controlled 9-year, Real-world Registry Study
Authors: Zitzmann Michael
Published: October 2023

Disclosure: M. Zitzmann: None.

Background and Significance: Long-term data on Testosterone therapy (TTh) in hypogonadal men are limited and the clinical value, especially in men with functional hypogonadism (FH) is debated. A long-term, real-world registry study comprising groups of patients with hypogonadism of various etiologies provides a suitable and novel approach to this clinical issue. Methods: A registry spanning 9 years comprising 650 patients with hypogonadism included 188 patients with FH (mean age 42.3±11.3 years) and 462 men with classical hypogonadism (CH). Of these, 266 men had primary hypogonadism (PH, mean age 34.0±11.7 years) and 196 secondary hypogonadism (SH, mean age 31.9±12.0 years). All men uniformly received intramuscular T undecanoate (1000 mg). Effects of TTh on anthropometric parameters, as well as metabolic and safety parameters were compared. Results: The registry contained metabolic and safety parameters in individual duration of TTh spanning 1 to 9 years. Serum T concentrations increased from 6.6±2.4 nmol/L to 19.3±2.9 nmol/L in all patients (duration-of-treatment-dependent averages, mixed linear model for repeated measurements with the fixed variable “time x visit interaction”: p<0.001). In both categories of hypogonadism, TTh was associated with significant weight loss and decrease in waist circumference (WC, both p<0.001). Cox regression and Kaplan-Meier models revealed differences of inter-individual check-points: men with FH were more likely to lose >10% weight and >5% of WC than men with CH (hazard ratio 1.3 [1.1-1.4], p=0.008 and hazard ratio 1.4 [1.3-1.5], p=0.001). There was no difference between groups for the overall marked increase in hematocrit. Changes in PSA levels were more likely to occur in FH (hazard ratio 1.3 [1.1-1.6], p=0.003). During TTh, patients with FH exhibited significantly more pronounced changes of favourable nature in metabolic parameters than patients with CH (total cholesterol, triglycerides, LDL- and HDL-cholesterol and fasting glucose). The same applied to scores of AMS and IIEF-EF questionnaires. Effects on most parameters, especially hematocrit, were significantly modulated by age and baseline values for weight, WC und T. Conclusions: Findings regarding effects and safety of TTh in different groups of hypogonadal men are provided. Effects on factors associated with cardiovascular health are modulated by diagnosis and age. Patients with FH seem to benefit to a larger extent from TTh, most likely attributable to their more pronounced risk factor profile at baseline.

Presentation Date: Friday, June 16, 2023
SAT006 Activation Of The CAR/RXR Heterodimer By Pesticides And Induced Metabolic Effects
Authors: Dauwe Yannick , Lakhal Laila
Published: October 2023

Disclosure: Y. Dauwe: None. L. Lakhal: None.

The nuclear receptor Constitutive Androstane Receptor (CAR or NR1I3) was initially reported as a transcription factor regulating the hepatic genes involved in detoxication function. Thereby CAR protects against xenobiotics but also endogenous molecules such as biliary acid, bilirubin, thyroids and steroids hormones. Different studies showed that CAR activation results in metabolic disorders including Non-Alcoholic Fatty Liver Disease (NAFLD) by promoting lipogenesis in the liver. Environmental contaminants referred to as endocrine disrupting chemicals are suspected to be involved in the increased incidence of metabolic disorders through the activation of nuclear receptors such as CAR and PXR. Previous in vitro studies have shown that the synergistic activation of the Pregnane X Receptor (PXR) by two PXR ligands is potentiated in presence of a Retinoid X Receptor (RXR) environmental ligand, tributyltin (TBT). The objective of our study was to determine whether the synergic activation of PXR/RXR observed in vitro could be transposed in vivo with environmental chemicals on the PXR/RXR and CAR/RXR heterodimers. But also, to asses if this activation could exacerbate the metabolic effects. In that aim, binary mixtures of six, CAR and PXR ligands, pesticides and TBT were first orally administrated to C57BL/6 mice for 3 days, each close to the minimal dose able to activate CAR which was determined in a previous dose response study. A synergistic activation of the CAR/RXR heterodimer by pesticides in combination with TBT was observed on the expression of prototypical CAR target gene Cyp2b10 and on genes involved in lipid metabolism. Combined effects where observed on lipid accumulation in the liver. A synergic elevation of plasma cholesterol was observed and decreased free fatty acid levels were also evidenced for some pesticides in combination with TBT. Two mixtures of five pesticides ligand of CAR and PXR with or without TBT, each at the No Observed Adverse Effect Level (NOAEL), were then administered for 120 days to determine if these threshold safe doses are underestimated in the context of human chronic exposure to a mixture of endocrine disrupting chemicals. The experiment was performed with wild type, CAR null and PXR null mice to check the involvement of nuclear receptors in the measured effects. We observed an activation of CAR by the pesticide cocktail alone or in combination with TBT. The cocktail effect is now tested on the expression of genes involved in lipid metabolism, circulating plasma lipids, lipid accumulation in the liver as well as inflammation and fibrosis. Our first results show an increased transcription of lipogenic genes but no significant effects on lipid plasma levels. Considering these cooperative effects of pesticides may help refine risk assessment, which could lead to a better prevention policy.

Presentation: Saturday, June 17, 2023
SAT007 Excess Bile Acids Can Compensate To Promote Regeneration In Constitutive Androstane Receptor Knockout Mice
Authors: Zhou Weinan , Chembazhi Ullas V , Huang Jiansheng , Bangru Sushant , Dean Angela E , Kalsotra Auinash , Rudnick David A , Anakk Sayeepriyadarshini
Published: October 2023

Disclosure: W. Zhou: None. U.V. Chembazhi: None. J. Huang: None. S. Bangru: None. A.E. Dean: None. A. Kalsotra: None. D.A. Rudnick: None. S. Anakk: None.

The liver plays a key role for the metabolism of nutrients and xenobiotics. Nuclear receptor constitutive androstane receptor (CAR) regulates the transcription of detoxification and proliferation genes. The liver can also regenerate, which can be tested using a model of 2/3rd partial hepatectomy (PHx). We investigated the role for CAR during liver regeneration by mining publicly available single-cell RNA seq data after PHx. Car and its target expression was enriched in hypermetabolic hepatocytes, which was unexpected since based on previous studies we anticipated a proliferative role for CAR. We also examined liver weight, hepatocyte proliferation, and levels of bile acid (BAs) and liver enzymes in Car knockout (CarKO) and wild-type (WT) mice after PHx, and found them comparable between the two genotypes. Intriguingly, we noted a dramatic BA overload and concomitant changes in the expression of genes responsible for BA synthesis and detoxification in CarKO mice after PHx. This led us to speculate if the BA excess observed in CarKO mice after PHx promoted regeneration. When this excess BAs was reduced with a resin treatment in CarKO mice, we found deficits in proliferation and Yes-associated protein signaling demonstrating that elevated BAs act as mitogens to counteract the regenerative defects in CarKO livers.

Presentation: Saturday, June 17, 2023
SAT014 Involvement Of Cytochrome P450 1B1 In Synthesis Of Neurosteroid Pregnenolone In Human Brain Cells
Authors: Christina Lin Yiqi , Cheung Garett , Zhang Zeyu , Papadopoulos Vassilios
Published: October 2023

Disclosure: Y. Lin: None. G. Cheung: None. Z. Zhang: None. V. Papadopoulos: None.

Neurosteroids, steroids synthesized in the nervous system, have important functions in modulating brain function, neuroprotection, and reducing neuroinflammation. The neurosteroid pregnenolone, the precursor to all other neurosteroids, is one of the most abundant neurosteroids in the brain. Still, the enzyme known to make it, CYP11A1, has been difficult to detect in the human brain. We recently reported that unlike in peripheral steroidogenic organs, pregnenolone is not synthesized by CYP11A1 in human brain cells but rather by another mitochondrial cytochrome P450 enzyme (J Biol Chem. 2022 Jul;298(7):102110). Therefore, we aimed to identify the enzyme responsible for producing pregnenolone in the current study. Upon bioinformatic analyses of publicly available datasets for mitochondrial CYP450s expressed in the human brain, we identified three potential candidates: CYP27A1, CYP1A1, and CYP1B1. Given that CYP27A1 is highly expressed in the brain and is involved in cholesterol metabolism, we first tested the effects of CYP27A1 inhibition via drug treatment and siRNA knockdown on pregnenolone synthesis in steroidogenic MGM-1 human glioma cells. Neither treatment with CYP27A1 inhibitors anastrozole or dexmedetomidine nor CYP27A1 siRNA knockdown resulted in a significant reduction in pregnenolone formation by MGM-1 cells, suggesting that CYP27A1 likely is not involved in pregnenolone synthesis in human brain cells. On the other hand, MGM-1 cells treated with the CYP1A1/CYP1B1 inhibitors α-napthoflavone and 2,4,3’,5’-tetramethoxystilbene showed a significant reduction in pregnenolone production in the presence of the pregnenolone precursor 22R-hydroxycholesterol. To investigate whether this effect was due to the inhibition of CYP1A1 or CYP1B1, we knocked down CYP1A1 or CYP1B1 in MGM-1 cells using siRNA. While CYP1A1 knockdown did not result in any significant changes to pregnenolone synthesis, CYP1B1 knockdown significantly decreased secreted pregnenolone formation by MGM-1 cells. Together, these results suggest that CYP1B1 is likely involved in the alternative pathway for pregnenolone synthesis in human brain cells.

Presentation: Saturday, June 17, 2023
SAT018 Investigate If Sex Differences In Microbiota Is Sufficient To Promote Bile Acid-Mediated Hepatocellular Carcinoma In Mice
Authors: Dean Angela E , Gaulke Christopher A , Anakk Sayeepriyadarshini
Published: October 2023

Disclosure: A.E. Dean: None. S. Anakk: None.

Liver cancer is one of the leading causes of cancer-related deaths with men having a higher prevalence than women. It is becoming clearer that changes in bile acid levels are seen in patients diagnosed with liver cancer. Although initially made in the liver, these primary bile acids can also be modified by bacteria in the intestines to form secondary bile acids. The secondary bile acids are more hydrophobic and cytotoxic and when in excess, can promote tumorigenesis. We, therefore, examined if microbial differences between males and females explain sex-dependent HCC development when bile acid metabolism is altered. In this study, we used a mouse model of bile acid excess and HCC, where the bile acid-related nuclear receptors, farnesoid X receptor (Fxr) and small heterodimer partner (Shp), are knocked out. This double knockout (DKO) model mimics the cancer incidence observed in humans, where male mice are more susceptible to spontaneous tumorigenesis than females at 12 months of age. The circulating bile acid profiles of the male DKO mice are more hydrophobic and have a greater proportion of secondary bile acids than the females. Since we observed changes in the bile acid composition, we next investigated if fecal and cecal microbial profiles of male and female DKO mice differed, along with their wild-type counterparts. 16s rRNA full-length sequencing was performed on fecal and cecal samples from 12-month-old mice, and analyses are currently ongoing. We expect that the microbes identified in the male DKO mice will reflect their bile acid compositions, as well as spontaneous tumorigenesis. Additionally, we are analyzing the 16s rRNA gene from mice treated with cholestyramine (CHR), a bile acid binding resin, that exhibit decreased tumorigenesis at 12 months of age. In this manner, we can correlate microbiota changes to the bile acid composition, as well as the extent of tumor burden. Thus, this study will be able to integrate how sex differences in liver cancer incidence can be secondary to microbial modification of bile acids.

Presentation: Saturday, June 17, 2023