Endocrinology最新文献

Missense mutations in nuclear receptors (NR) transcription factors (TF) cause a number of genetic disorders, including PPARG mutations that result in familial partial lipodystrophy type 3 (FPLD3). Experimental assessment is essential to establish a newly identified mutation as disease-causing, as accurately predicting the effect of new mutation in silico remains challenging due to the multifunctional and modular nature of these proteins. However, deep structure-function characterisation often requires specialised and technically demanding approaches, which may not be readily available. Therefore, we established a simple and robust experimental framework, based on four complementary reporter assays that independently assess: (1) ability of the full-length receptor to activate transcription; (2) integrity of the ligand-binding domain (LBD); (3) heterodimerization potential; and (4) DNA binding capacity. As a proof-of-concept we analysed 3 uncharacterized FPLD3-associated loss-of-function (LOF) variants and two bladder cancer-associated gain-of-function (GOF) variants. Together, the four complementary assays showed unique functional phenotypes for all 5 mutants, that were further supported by co-regulator profiling. We therefore conclude that this framework provides a simple and robust first line approach to identify functional alterations in PPARγ mutants with mechanistic resolution. This framework is broadly applicable across NRs and offers a scalable path to systematic variant interpretation in both research and clinical contexts.

The circadian clock plays a critical role in coordinating energy metabolism across tissues, including brown adipose tissue (BAT), a major site of nonshivering thermogenesis. This study aimed to elucidate the cell-autonomous role of the peripheral circadian clock in brown adipocyte thermogenesis using an in vitro model independent of extrinsic cues. Primary brown adipocytes were differentiated from the stromal vascular fraction of interscapular BAT isolated from C57BL/6J mice. An in vitro model of BAT clock disruption was established by siRNA-mediated knockdown of the core clock gene Bmal1. Thermogenic function was assessed via measurement of oxygen consumption rate (OCR) using an extracellular flux analyzer. To further assess the thermogenic process, protein expression levels of lipolytic enzymes and mitochondrial oxidative phosphorylation (OXPHOS) complexes were analyzed by Western blotting. Bmal1 knockdown markedly reduced both basal and β-adrenergic-stimulated OCR, indicating impaired thermogenic function, despite comparable cellular differentiation, preserved β-adrenergic responsiveness, and elevated uncoupling protein 1 (UCP1) expression. Notably, Bmal1-deficient cells exhibited decreased protein expression of key lipolytic enzymes, adipose triglyceride lipase and hormone-sensitive lipase, as well as multiple mitochondrial OXPHOS subunits, suggesting decreased free fatty acid supply and reduced mitochondrial ability to generate the proton gradient required for UCP1-mediated thermogenesis. The peripheral circadian clock in brown adipocytes supports thermogenic function by regulating lipid mobilization and mitochondrial oxidative function; thus its disruption may lead to decreased energy expenditure and increased susceptibility to metabolic disorders.

Water movement across cell membranes through aquaporin water channels creates osmotic equilibrium between extracellular and intracellular fluid compartments. Plasma osmolality is tightly regulated by the kidneys and brain through the process of osmoregulation. The antidiuretic hormone, arginine vasopressin (AVP), is normally released from the posterior pituitary in response to increased osmolality or decreased intravascular volume. Defects in the synthesis or release of AVP result in AVP deficiency (AVP-D) and the syndrome of central diabetes insipidus, characterized by inappropriate aquaresis leading to hyperosmolality and insatiable thirst. While most cases of AVP-D are due to local mechanical, infiltrative, compressive, infectious, or inflammatory processes, some recreational and pharmacological substances can cause AVP-D. In this review, we discuss the history and current knowledge about these substances, including cannabinoids, ethanol, κ opioid receptor agonists, phenytoin, and anesthetic agents.

Effector kinases of the lipid second messenger diacylglycerol (DAG), including protein kinase C (PKC) and protein kinase D (PKD) isozymes, have been widely implicated in the development and progression of prostate cancer. By acting as central hubs of growth factor-mediated signaling, these kinases integrate oncogenic signals with the androgen receptor (AR) pathway, contributing to prostate tumor growth. Distinct members of the DAG-regulated kinases contribute to the acquisition of castration-resistant prostate cancer (CRPC) and bypass AR dependence, promoting the proliferative, migratory, and invasive competencies of androgen-independent prostate cancer cells. As predicted from their coupling to signaling cascades that impact gene expression, PKC/PKD isozymes control the activation of transcription factors such as NF-κB, E2F, and STAT3, and additionally regulate epithelial-to-mesenchymal transition (EMT) transcription factors in prostate cancer cells, providing an additional layer of control in invasive signaling. The aberrant expression/activation of DAG-regulated kinases during prostate cancer progression results in pronounced deregulation and rewiring of transcriptional networks associated with cell cycle control, invasiveness, and cancer cell interactions with the tumor microenvironment (TME). The multifaceted regulation of nuclear functions by these pleiotropic kinases underscores their convoluted roles in prostate cancer development and progression, offering new opportunities for therapeutic targeting.

Anti-Müllerian hormone (AMH) is produced by granulosa cells within growing ovarian follicles and limits the number of follicles reaching ovulation. AMH is synthesized as a precursor protein comprising N-terminal prodomains and C-terminal mature domains, separated by a furin-like cleavage motif (RXXR). Proteolytic maturation of AMH (140 kDa) is required to release the bioactive mature dimer (25 kDa), which potentiates signaling via AMH receptors (AMHR2 and ALK2/3). However, the abundance of unprocessed AMH in human follicular fluid suggests that cleavage within the ovary is inefficient. This study hypothesized that enhancing AMH maturation would increase AMH activity in vitro and in vivo. Using targeted mutagenesis, we optimized the murine AMH cleavage site (from wild-type (WT) 443RTGR445 to 443RKKR445) and showed in vitro that this favored production of bioactive AMH. We then introduced this mutation into the Amh gene in C57Bl6/J mice using CRISPR/Cas9 and assessed the consequences for female reproduction. Analyses of 12-week-old AmhRKKR/RKKR mice revealed that the ovaries were significantly lower in mass (-25%, P < .05) relative to AmhWT/WT controls. Despite differences in ovarian masses, estrous cyclicity, and fertility were unaltered. Although maturing follicle numbers did not differ, ovaries from 12- and 24-week-old AmhRKKR/RKKR females contained a greater proportion of atretic secondary follicles (1.6- to 4-fold more, P < .05), underscoring AMH's role in preantral follicle survival. Analyses of adult male AmhRKKR/RKKR mice indicated that testis mass and morphology were unaltered. These findings support a physiological role for ovarian AMH in limiting preantral follicle survival and indicate that enhancing AMH maturation is otherwise nondisruptive to female reproduction.

About 12% of couples worldwide are infertile. Male factor infertility causes or is contributory to a couple's ability to conceive in approximately 50% of cases. Evidence has emerged that infertile men have poor overall health and increased morbidity and mortality, yet the causes for this are poorly understood. Although these men may appear healthy, research shows that they can harbor a wide variety of systemic diseases and illnesses that may share common links with the causes of their infertility. In fact, as semen parameters decline, their risks of several health conditions increase. In the early 1990s to the present, studies revealed that 1% to 6% of unselected infertile men seeking clinical evaluation have significant and (sometimes) life-threatening pathologies ranging from endocrine abnormalities to malignancies, developmental anomalies, and genetic diseases. Yet, despite this knowledge, for couples seeking treatment of their infertility, the female partner undergoes extensive clinical evaluation but the male partner frequently is only asked to provide a specimen for a routine semen analysis. This review focuses on the current understanding of the association of the genetic causes of male infertility and a multitude of diseases that affect these men's overall health and their increased risk of mortality.

Patients with diabetes are disproportionately affected by cardiovascular and kidney disease. Mineralocorticoid receptor antagonists (MRA) show organ protection against cardiovascular and renal injury; however, major side effects including hyperkalaemia and reduced renal function limit their use in individuals with diabetic complications. The non-steroidal MR modulator, balcinrenone, may offer end-organ protection with fewer side effects. We compared responses to balcinrenone and eplerenone delivered from 8 weeks post-induction of streptozotocin (STZ)-induced type 1 diabetes in male mice. RNA-sequencing revealed diabetes induced modulation of immune function, and metabolic and vascular targets in the kidney, which were similarly attenuated by balcinrenone or eplerenone treatment. Urine K+ excretion was lower following eplerenone treatment, but not balcinrenone treatment, compared to diabetes without treatment. We identified a 5.90-fold increase in the expression of K+ transporter G protein-activated inward rectifier potassium channel 1 (GIRK-1) in eplerenone-, but not balcinrenone-treated diabetic mice. Balcinrenone and eplerenone similarly attenuated the diabetes-induced reduction in peak E-wave/A-wave velocity (E/A) compared to mice without treatment at 15 weeks post-STZ. Gene markers of cardiac injury, B-type natriuretic peptide (Bnp) and beta-myosin heavy chain protein (Myh7), were higher in diabetic versus non-diabetic left ventricles (LV). Conversely, gene expression of Ca2+ ion channel subunits, voltage-dependent L type, calcium channel subunit alpha 1C (Cav1.2) and ryanodine receptor 2 (Ryr2), in LV was lower in diabetic but not eplerenone- or balcinrenone-treated diabetic mice. Although balcinrenone and eplerenone similarly modified cardiac changes, potassium excretion was greater with balcinrenone, consistent with a reduced risk of hyperkalemia with the non-steroidal MR modulator.

Bone remodeling, mediated by bone-forming osteoblasts and bone-resorbing osteoclasts, is a physiologically relevant process controlled by several local and systemic regulatory mechanisms. Recent evidence in mice has suggested that it is also affected by housing temperature, which provides a basis to identify novel molecular regulators of bone remodeling. Here, we compared the skeletal phenotype of mice housed at thermoneutral (30 °C), room (22 °C), or cold (6 °C) temperature for 1 or 4 weeks. We observed that cold exposure for 1 week differentially affected osteoclastogenesis and osteoblast activity, which caused a significant reduction of trabecular number and cortical thickness after 4 weeks. Cold exposure is known to induce type II iodothyronine deiodinase (DIO2) expression in thermogenic adipose tissues. Because this enzyme catalyzes the deiodination of T4 to the active thyroid hormone T3, we addressed the question whether the cold-induced bone loss depends on DIO2. Importantly, however, we found that Dio2 deficiency in male and female mice did not affect the cold-induced reduction of trabecular and cortical bone mass, demonstrating that this process does not depend on thyroid hormone activation. To identify potential metabolic differences between the different groups of mice, we additionally performed lipidomic analyses. Here, we observed a remarkable reduction of specific lipid species after cold exposure, suggesting that either the systemic catabolic metabolism or the decrease of specific lipid species cause cold-induced bone loss. Taken together, although our data demonstrate that sustained cold exposure has a remarkable negative impact on bone mass, future studies are needed to identify causative molecules.

African American men experience a higher incidence and severity of prostate cancer relative to European American men, and there is a range of risk factors that may contribute to this disparity. Prostate adenocarcinoma originates from the epithelium, which is significantly influenced by signaling from the surrounding fibromuscular stroma. To identify ancestry-associated differences in the stroma, gene expression profiling was compared between laser-capture microdissected prostate cancer stroma from patients of African descent and those of European descent. Estrogen receptor signaling was the top differential pathway between the groups, with the steroid hormone dehydrogenase HSD17B7 identified as the most differentially expressed gene. In a separate cohort of patients, protein expression of HSD17B7 was higher in African American patients relative to European American patients in a radical prostatectomy tissue microarray, validating the transcriptional findings. African American patients also exhibited significantly increased levels of HSD17B7 protein in the stroma surrounding benign areas compared to the stroma near tumors. These studies provide important evidence of ancestry-associated differences in stromal estrogen signaling.

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.