Glucocorticoids are an important signal for the differentiation of many types of cells. Consistent with this, several studies have demonstrated that glucocorticoids promote the somatotrope differentiation program and functionality. Interestingly, we previously found that loss of the forkhead factor, FOXO1, results in delayed emergence of somatotropes and prevents glucocorticoid-induced premature differentiation of somatotropes. In the current study, we find that pituitary-specific deletion of Nr3c1, the gene encoding the glucocorticoid receptor, impairs somatotrope differentiation and increases lactotrope numbers embryonically and at five days after birth. The number of somatotropes remains reduced at seven weeks of age in females and males, but lactotrope cell numbers are only increased in females at this age. FOXO1 is nearly undetectable in pituitary glands from mouse embryos lacking NR3C1 and continues to be reduced in adults. These findings suggest that glucocorticoids are an important signal for determining the balance between somatotropes and lactotropes and that FOXO1 may mediate NR3C1 induction of somatotrope differentiation.

The preovulatory luteinizing hormone (LH) surge is a defining neuroendocrine event that triggers ovulation through estradiol (E2)-dependent positive feedback on gonadotropin-releasing hormone (GnRH) secretion. Mechanistic studies of this process have relied predominantly on ovariectomized rodents supplemented with exogenous hormones, a paradigm that disrupts ovarian function and limits physiological and translational relevance. Here, we establish a robust and reproducible model of LH surge induction in intact female mice that preserves the hypothalamic-pituitary-gonadal axis. We show that a single low-dose injection of estradiol benzoate (EB; 0.5 µg) administered during diestrus reliably induces a temporally synchronized LH surge in most intact females, whereas EB treatment during proestrus or estrus is ineffective. EB-induced LH surges were accompanied by a significant increase in circulating progesterone, confirming the engagement of ovarian luteinizing pathways. The LH surge occurrence was independent of proestrus detection by vaginal cytology, accentuating the limited predictive value of cytology in mice. At the neuroendocrine level, EB treatment markedly increased cFOS expression in GnRH neurons across key preoptic regions and modestly activated kisspeptin neurons in the rostral periventricular area of the third ventricle, consistent with activation of E2-positive feedback circuitry. In contrast, arcuate kisspeptin neurons remained inactive, supporting their role in negative feedback. Together, these findings define a physiologically relevant, intact mouse model for studying E2-induced LH surges that maintains endogenous ovarian signaling.

Growth hormone (GH) controls sexual dimorphism in hepatocyte gene expression programs governing lipid metabolism, bile acid synthesis and xenobiotic processing, which contribute to sex differences in metabolic dysfunction-associated steatotic liver disease (MASLD) risk. Although GH-regulated sex-specific transcription is well-studied, the functional cis-regulatory hepatocyte enhancers that orchestrate these sex-dependent metabolic programs remain largely unknown. Here, we integrated single-nucleus multiomic profiling of hepatocyte chromatin accessibility with in vivo functional enhancer assays to identify and validate GH-responsive, sex-biased hepatocyte enhancers in intact mouse liver. We constructed a tiled HDI-STARR-seq library of 23 912 reporters spanning 1839 liver ATAC regions and delivered it to liver by hydrodynamic injection, enabling enhancer activity assessment across different biological conditions. Reporters representing 840 ATAC regions showed sex-biased and/or GH-regulated enhancer activity, in many cases mirroring regulation of their accessibility in hepatocyte chromatin, validating them as functional, physiologically regulated enhancers. The regulated enhancer sequences were enriched for activating histone marks (H3K27ac, H3K4me1), and for binding sites for the STAT5-dependent, sex-specific repressors BCL6 and CUX2; whereas, STAT5 binding was enriched at both regulated and non-regulated enhancers. Motifs for HNF4A and for several novel factors identified de novo were specifically enriched at the regulated enhancers. Sex-biased and GH-regulated enhancers were linked to both MASLD-enabling and MASLD-protective genes, suggesting that GH-dependent chromatin remodeling at these loci contributes to sex-differential metabolic disease susceptibility. This integrated in vivo approach defines a validated set of GH-regulated hepatocyte enhancers through which chromatin accessibility and transcription factor binding drive sexual dimorphism in hepatic metabolism and MASLD risk.

The clinical characteristics of type 2 diabetes (T2D) differ between the sexes. For example, the risk of T2D is higher in males than in premenopausal females, whereas the risk of T2D-associated cardiovascular disease is higher in females. Notwithstanding, the sex-dependent mechanisms of T2D pathogenesis remain incompletely understood. Publicly available human islet datasets, such as HPAP and Humanislets.com, are valuable tools for uncovering the impact of biological sex on islet structure, gene expression, and function at a scale that was not previously possible. We performed integrated analyses of data from public sources to identify sex differences in baseline islet characteristics in donors without diabetes and also in donors who lived with T2D. Among donors without diabetes, female islets had a greater proportion of alpha-cells compared with male islets and showed enriched expression of ribosomal and mitochondrial pathways in both beta-cells and alpha-cells. Measurements of mitochondrial function in female islets revealed lower spare respiratory capacity compared with male islets. Male and female islets had distinct changes in gene and protein expression in the context of T2D with female islets having greater preservation of insulin content and fewer defects in islet function. Together, these data show female islets have fewer impairments in T2D. This highlights the need for detailed mechanistic studies in both sexes to support effective and sex-informed interventions for T2D.

Circulating estradiol is predominantly protein-bound, with human serum albumin (HSA) serving as its major carrier. While traditionally considered a carrier with low affinity and readily reversible binding at a single site, the molecular details and kinetics of estradiol-HSA interactions remain incompletely understood. We employed equilibrium dialysis, steady-state and time-resolved fluorescence spectroscopy to characterize estradiol-HSA interactions. Surface plasmon resonance (SPR) was used to elucidate the kinetics of estradiol's association and dissociation with HSA. Structural and energetic features of binding were investigated using molecular docking and structure network analyses. Binding isotherms generated using equilibrium dialysis, steady-state and time-resolved fluorescence spectroscopy revealed nonlinear asymmetric binding with apparent Kd that varied as a function of estradiol and HSA concentrations, inconsistent with canonical model of low-affinity, single-site interaction characterized by a fixed Kd. Kinetic analyses by SPR revealed initial rapid association dynamics followed by a slower second phase. Molecular modeling identified a high-affinity estradiol-binding pocket in Sudlow's Site I and two additional low-affinity sites within a highly interconnected hub of structural blocks. Spatially coordinated conformational rearrangements accompanying estradiol partitioning into the high-affinity pocket of Sudlow's Site I and two additional moderate-affinity sites suggest an allosterically-coupled binding architecture that enables albumin to actively regulate estradiol bioavailability across a broad, physiologically relevant concentration range. Estradiol's binding to HSA is a dynamic, multi-equilibrium process driven by ligand-induced conformational rearrangements within HSA; the binding data are inconsistent with canonical model of estradiol-HSA interaction with 1:1 stoichiometry and a fixed Kd.

The binding of human growth hormone (hGH) to the human growth hormone receptor (hGHR) is a key endocrinological process that controls critical aspects of cell growth, proliferation, and differentiation. Mechanistically, this sequential, asymmetric binding event involves the interaction between a single hGH molecule and distinct sites (sites 1 and 2) on the extracellular domain of a preformed hGHR homodimer. Our group recently identified S1H, a rationally designed peptide sequence mimetic of the hGH site 1-binding helix (residues 36-51) that disrupts the hGH-hGHR interaction and inhibits hGH-mediated phosphorylation of signal transducer and activator of transcription 5 (STAT5) in hGHR-positive cell lines. Structure-activity relationship studies revealed a positive correlation between helical propensity and inhibitory potency of the S1H peptide, prompting the design of structurally "stabilized" S1H variants (SS1H) with improved biological activity. In this study, we employed a chemical strategy, termed hydrocarbon stapling, to generate a series of SS1H peptides that proved to be more helical, proteolytically stable, and biologically active compared to linear (unstructured) S1H. Notably, one SS1H derivative (SS1HB) inhibited hGH-induced STAT5 phosphorylation in hGHR-positive human bladder cancer cells more effectively than pegvisomant, the only hGHR antagonist currently approved by the FDA. Collectively, our results demonstrate that hydrocarbon stapling improves the antagonistic effects of S1H peptides and elevates their potential as chemical probes to study the molecular mechanisms of hGH signaling. It is also anticipated that SS1H peptides will serve as potent lead compounds for developing next-generation therapeutics designed to treat endocrine disorders that manifest along the hGH-hGHR signaling axis.

The mineralocorticoid receptor (MR), classically known for its role in electrolyte homeostasis, has emerged since the early 2000s as a key regulator of adipocyte biology. MR activation induces oxidative stress and inflammation, promotes white adipogenesis, and suppresses brown fat differentiation, potentially contributing to obesity and metabolic syndrome (MetS). In mice, MR antagonists (MRAs) have shown beneficial metabolic effects by reversing adipose tissue dysfunctions, counteracting excessive expansion of fat depots, and enhancing brown adipose tissue (BAT) thermogenesis. Clinical translation remains limited, with inconsistent outcomes in individuals with overweight or obesity, and concerns about side effects of steroidal MRAs (sMRAs), particularly hyperkalemia. Nevertheless, recent data showed that MRAs are able to activate BAT in healthy individuals and in patients with familial partial lipodystrophy type 2 (FPLD2). Nonsteroidal MR antagonists (nsMRAs), such as finerenone, exhibit anti-inflammatory and antifibrotic properties in the kidney and heart, with improved safety profiles compared to sMRAs, and are currently being explored in metabolic disorders beyond cardiorenal disease. This review summarizes novel insights into MR function in adipose tissue and critically evaluates whether MR can still be considered a valid target to treat obesity and associated metabolic disorders.

Disruption of lymphatic function underlies a broad spectrum of inflammatory and metabolic disorders, yet the hormonal pathways that regulate lymphatic biology remain poorly defined. GH, which is implicated in similar disease states, has an unclear role in lymphatic homeostasis. To address this gap, we investigated how chronic alterations in GH signaling alter lymphatic structure and function. Using transgenic mouse lines with increased, decreased, or absent GH action, we quantified the effect of GH on lymphatic pumping rate and lymphangiogenic remodeling during wound healing using near-infrared fluorescent imaging. We also measured markers of lymphatic endothelial cells using Western blot and immunohistochemistry across multiple mouse organs. Lymphatic pumping rate positively correlated with GH action, whereas both elevated and absent GH signaling delayed wound healing. In contrast, the lymphatic vascular density and the expression of protein markers of lymphatic endothelial cells were inversely correlated with GH activity. Additionally, we showed that primary human dermal lymphatic endothelial cells express the GH receptor and exhibit acute GH-activated signaling and that this activation can be blocked with new and Food and Drug Administration-approved GH receptor antagonists. Together, these findings identify GH as a regulator of the lymphatic system and suggest that GH receptor antagonism could be a potential strategy to address lymphatic dysfunction.

Maternal obesity before and during pregnancy causes maladaptive fetal development with long-term effects on offspring's metabolic health, including a higher risk of metabolic dysfunction-associated steatotic liver disease. Treatment with metformin during obese pregnancy has been suggested to prevent adverse fetal programming, but its long-term effects on offspring liver metabolism remain uncertain. In wild-type C57BL/6NCrl mice, obesity was induced by feeding a high-fat/high-sucrose Western-style diet before and throughout gestation and lactation. A subset of obese dams received metformin during gestation. Offspring from control, obese (OB), and obese with metformin-treated (OB + M) dams were analyzed at postnatal days (P) 21 and 56 for their metabolic phenotype, hepatic histomorphology, and key metabolic proteins. At P21, maternal metformin treatment worsened obesity-related traits in male OB + M offspring, including increased body weight, length, and fat volume, higher plasma leptin, insulin, and resistin levels, and impaired glucose tolerance. Female OB + M offspring also showed a worsening of obesity traits, though less pronounced. Hepatic lipid accumulation displayed sex-specific patterns; male OB + M offspring exhibited reduced lipid accumulation, whereas female OB + M offspring demonstrated increased lipid accumulation. By P56, phenotypic parameters returned to normal, but molecular alterations persisted, involving shifts in hepatic fatty acid metabolism and mitochondrial respiratory chain complexes. Maternal metformin during obese pregnancy has age- and sex-specific effects on offspring, aggravating early obesity traits in a sex-dependent manner and prompting adaptations in hepatic metabolism during adolescence. These findings highlight the controversy surrounding metformin use during obese pregnancy, given its potential to induce sex-specific obesity and metabolic disturbances in offspring.

Despite the widespread use of mammography as the standard of care for breast cancer screening, its accuracy remains limited for select patient populations, such as women with high breast density. Liquid biopsy-based tests offer an accessible complement to conventional screening methods. Here, we conducted a case-control study to develop a plasma-based protein classifier to distinguish between those with early-stage breast cancer and healthy individuals. A total of 335 women, comprising 116 patients with newly diagnosed, treatment-naïve breast cancer (stage 0-2) and 219 healthy controls, had plasma samples collected and processed in a blinded manner using a sample preparation method coupled with semiquantitative, label-free mass spectrometry-based analysis. The median number of proteins detected per patient across breast cancer and healthy individuals was 6991 and 6818, respectively. A machine learning-based classifier was trained and validated on patient proteome profiles using a leave-one-out cross-validation approach to identify patients with breast cancer. The classifier achieved an area under the curve of 0.96 (95% CI, 0.93-0.97), with a sensitivity of 86.2% (95% CI, 78.8-91.3%) and a specificity of 90.4% (95% CI, 85.8-93.6%). In patients with breast cancer, the classifier retained >85% sensitivity regardless of breast density (low density: 87.2%, high density: 90.2%) at 90% specificity. Our workflow demonstrates the potential of plasma proteomics as a potent diagnostic tool in early-stage breast cancer screening.