Asymmetric Adrenals: Sexual Dimorphism of Adrenal Tumors.

CONTEXT: Sexual dimorphism has direct consequences on the incidence and survival of cancer. Early and accurate diagnosis is crucial to improve prognosis. OBJECTIVE: This work aimed to characterized the influence of sex and adrenal asymmetry on the emergence of adrenal tumors. METHODS: We conducted a multicenter, observational study involving 8037 patients with adrenal tumors, including adrenocortical carcinoma (ACC), aldosterone-producing adenoma (APA), cortisol-secreting adrenocortical adenomas (CSAs), non-aldosterone-producing adrenal cortical adenoma (NAPACA), pheochromocytoma (PCC), and neuroblastoma (NB), and investigated tumor lateralization according to sex. Human adrenal tissues (n = 20) were analyzed with a multiomics approach that allows determination of gene expression, catecholamine, and steroid contents in a single sample. In addition, we performed a literature review of computed tomography and magnetic resonance imaging-based studies examining adrenal gland size. RESULTS: ACC (n = 1858); CSA (n = 68), NAPACA (n = 2174), and PCC (n = 1824) were more common in females than in males (female-to-male ratio: 1.1:1-3.8:1), whereas NBs (n = 2320) and APAs (n = 228) were less prevalent in females (0.8:1). ACC, APA, CSA, NAPACA, and NB occurred more frequently in the left than in the right adrenal (left-to-right ratio: 1.1:1-1.8:1), whereas PCC arose more often in the right than in the left adrenal (0.8:1). In both sexes, the left adrenal was larger than the right adrenal; females have smaller adrenals than males. CONCLUSION: Adrenal asymmetry in both sexes may be related to the pathogenesis of adrenal tumors and should be considered during the diagnosis of these tumors.

The -KTS splice variant of WT1 is essential for ovarian determination in mice.

Sex determination in mammals depends on the differentiation of the supporting lineage of the gonads into Sertoli or pregranulosa cells that govern testis and ovary development, respectively. Although the Y-linked testis-determining gene Sry has been identified, the ovarian-determining factor remains unknown. In this study, we identified -KTS, a major, alternatively spliced isoform of the Wilms tumor suppressor WT1, as a key determinant of female sex determination. Loss of -KTS variants blocked gonadal differentiation in mice, whereas increased expression, as found in Frasier syndrome, induced precocious differentiation of ovaries independently of their genetic sex. In XY embryos, this antagonized Sry expression, resulting in male-to-female sex reversal. Our results identify -KTS as an ovarian-determining factor and demonstrate that its time of activation is critical in gonadal sex differentiation.

Crosstalk between androgen receptor and WNT/ß-catenin signaling causes sex-specific adrenocortical hyperplasia in mice.

Female bias is highly prevalent in conditions such as adrenal cortex hyperplasia and neoplasia, but the reasons behind this phenomenon are poorly understood. In this study, we show that overexpression of the secreted WNT agonist R-spondin 1 (RSPO1) leads to ectopic activation of WNT/ß-catenin signaling and causes sex-specific adrenocortical hyperplasia in mice. Although female adrenals show ectopic proliferation, male adrenals display excessive immune system activation and cortical thinning. Using a combination of genetic manipulations and hormonal treatment, we show that gonadal androgens suppress ectopic proliferation in the adrenal cortex and determine the selective regulation of the WNT-related genes Axin2 and Wnt4. Notably, genetic removal of androgen receptor (AR) from adrenocortical cells restores the mitogenic effect of WNT/ß-catenin signaling. This is the first demonstration that AR activity in the adrenal cortex determines susceptibility to canonical WNT signaling-induced hyperplasia.

Single-cell transcriptomic profiling redefines the origin and specification of early adrenogonadal progenitors.

Adrenal cortex and gonads represent the two major steroidogenic organs in mammals. Both tissues are considered to share a common developmental origin characterized by the expression of Nr5a1/Sf1. The precise origin of adrenogonadal progenitors and the processes driving differentiation toward the adrenal or gonadal fate remain, however, elusive. Here, we provide a comprehensive single-cell transcriptomic atlas of early mouse adrenogonadal development including 52 cell types belonging to twelve major cell lineages. Trajectory reconstruction reveals that adrenogonadal cells emerge from the lateral plate rather than the intermediate mesoderm. Surprisingly, we find that gonadal and adrenal fates have already diverged prior to Nr5a1 expression. Finally, lineage separation into gonadal and adrenal fates involves canonical versus non-canonical Wnt signaling and differential expression of Hox patterning genes. Thus, our study provides important insights into the molecular programs of adrenal and gonadal fate choice and will be a valuable resource for further research into adrenogonadal ontogenesis.

Sexual dimorphism in COVID-19: potential clinical and public health implications.

Current evidence suggests that severity and mortality of COVID-19 is higher in men than in women, whereas women might be at increased risk of COVID-19 reinfection and development of long COVID. Differences between sexes have been observed in other infectious diseases and in the response to vaccines. Sex-specific expression patterns of proteins mediating virus binding and entry, and divergent reactions of the immune and endocrine system, in particular the hypothalamic-pituitary-adrenal axis, in response to acute stress might explain the higher severity of COVID-19 in men. In this Personal View, we discuss how sex hormones, comorbidities, and the sex chromosome complement influence these mechanisms in the context of COVID-19. Due to its role in the severity and progression of SARS-CoV-2 infections, we argue that sexual dimorphism has potential implications for disease treatment, public health measures, and follow-up of patients predisposed to the development of long COVID. We suggest that sex differences could be considered in future pandemic surveillance and treatment of patients with COVID-19 to help to achieve better disease stratification and improved outcomes.

A mosaic of induced and non-induced branches promotes variation in leaf traits, predation and insect herbivore assemblages in canopy trees.

Forest canopies are complex and highly diverse environments. Their diversity is affected by pronounced gradients in abiotic and biotic conditions, including variation in leaf chemistry. We hypothesised that branch-localised defence induction and vertical stratification in mature oaks constitute sources of chemical variation that extend across trophic levels. To test this, we combined manipulation of plant defences, predation monitoring, food-choice trials with herbivores and sampling of herbivore assemblages. Both induction and vertical stratification affected branch chemistry, but the effect of induction was stronger. Induction increased predation in the canopy and reduced herbivory in bioassays. The effects of increased predation affected herbivore assemblages by decreasing their abundance, and indirectly, their richness. In turn, we show that there are multiple factors contributing to variation across canopies. Branch-localised induction, variation between tree individuals and predation may be the ones with particularly strong effects on diverse assemblages of insects in temperate forests.

Retinoic acid signaling is directly activated in cardiomyocytes and protects mouse hearts from apoptosis after myocardial infarction.

Retinoic acid (RA) is an essential signaling molecule for cardiac development and plays a protective role in the heart after myocardial infarction (MI). In both cases, the effect of RA signaling on cardiomyocytes, the principle cell type of the heart, has been reported to be indirect. Here we have developed an inducible murine transgenic RA-reporter line using CreERT2 technology that permits lineage tracing of RA-responsive cells and faithfully recapitulates endogenous RA activity in multiple organs during embryonic development. Strikingly, we have observed a direct RA response in cardiomyocytes during mid-late gestation and after MI. Ablation of RA signaling through deletion of the Aldh1a1/a2/a3 genes encoding RA-synthesizing enzymes leads to increased cardiomyocyte apoptosis in adults subjected to MI. RNA sequencing analysis reveals Tgm2 and Ace1, two genes with well-established links to cardiac repair, as potential targets of RA signaling in primary cardiomyocytes, thereby providing novel links between the RA pathway and heart disease.

Mitotic WNT signalling orchestrates neurogenesis in the developing neocortex.

The role of WNT/ß-catenin signalling in mouse neocortex development remains ambiguous. Most studies demonstrate that WNT/ß-catenin regulates progenitor self-renewal but others suggest it can also promote differentiation. Here we explore the role of WNT/STOP signalling, which stabilizes proteins during G2/M by inhibiting glycogen synthase kinase (GSK3)-mediated protein degradation. We show that mice mutant for cyclin Y and cyclin Y-like 1 (Ccny/l1), key regulators of WNT/STOP signalling, display reduced neurogenesis in the developing neocortex. Specifically, basal progenitors, which exhibit delayed cell cycle progression, were drastically decreased. Ccny/l1-deficient apical progenitors show reduced asymmetric division due to an increase in apical-basal astral microtubules. We identify the neurogenic transcription factors Sox4 and Sox11 as direct GSK3 targets that are stabilized by WNT/STOP signalling in basal progenitors during mitosis and that promote neuron generation. Our work reveals that WNT/STOP signalling drives cortical neurogenesis and identifies mitosis as a critical phase for neural progenitor fate.

Arrest of WNT/ß-catenin signaling enables the transition from pluripotent to differentiated germ cells in mouse ovaries.

Germ cells form the basis for sexual reproduction by producing gametes. In ovaries, primordial germ cells exit the cell cycle and the pluripotency-associated state, differentiate into oogonia, and initiate meiosis. Despite the importance of germ cell differentiation for sexual reproduction, signaling pathways regulating their fate remain largely unknown. Here, we show in mouse embryonic ovaries that germ cell-intrinsic ß-catenin activity maintains pluripotency and that its repression is essential to allow differentiation and meiosis entry in a timely manner. Accordingly, in ß-catenin loss-of-function and gain-of-function mouse models, the germ cells precociously enter meiosis or remain in the pluripotent state, respectively. We further show that interaction of ß-catenin and the pluripotent-associated factor POU5F1 in the nucleus is associated with germ cell pluripotency. The exit of this complex from the nucleus correlates with germ cell differentiation, a process promoted by the up-regulation of Znrf3, a negative regulator of WNT/ß-catenin signaling. Together, these data identify the molecular basis of the transition from primordial germ cells to oogonia and demonstrate that ß-catenin is a central gatekeeper in ovarian differentiation and gametogenesis.

The Sexually Dimorphic Adrenal Cortex: Implications for Adrenal Disease.

Many adrenocortical diseases are more prevalent in women than in men, but the reasons underlying this sex bias are still unknown. Recent studies involving gonadectomy and sex hormone replacement experiments in mice have shed some light onto the molecular basis of sexual dimorphism in the adrenal cortex. Indeed, it has been shown that gonadal hormones influence many aspects of adrenal physiology, ranging from stem cell-dependent tissue turnover to steroidogenesis and X-zone dynamics. This article reviews current knowledge on adrenal cortex sexual dimorphism and the potential mechanisms underlying sex hormone influence of adrenal homeostasis. Both topics are expected to contribute to personalized and novel therapeutic approaches in the future.

Adrenal cortex renewal in health and disease.

Resident progenitor and/or stem cell populations in the adult adrenal cortex enable cortical cells to undergo homeostatic renewal and regeneration after injury. Renewal occurs predominantly in the outer layers of the adrenal gland but newly formed cells undergo centripetal migration, differentiation and lineage conversion in the process of forming the different functional steroidogenic zones. Over the past 10 years, advances in the genetic characterization of adrenal diseases and studies of mouse models with altered adrenal phenotypes have helped to elucidate the molecular pathways that regulate adrenal tissue renewal, several of which are fine-tuned via complex paracrine and endocrine influences. Moreover, the adrenal gland is a sexually dimorphic organ, and testicular androgens have inhibitory effects on cell proliferation and progenitor cell recruitment in the adrenal cortex. This Review integrates these advances, including the emerging role of sex hormones, into existing knowledge on adrenocortical cell renewal. An in-depth understanding of these mechanisms is expected to contribute to the development of novel therapies for severe endocrine diseases, for which current treatments are unsatisfactory.

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.

Pituitary stem cells produce paracrine WNT signals to control the expansion of their descendant progenitor cells.

In response to physiological demand, the pituitary gland generates new hormone-secreting cells from committed progenitor cells throughout life. It remains unclear to what extent pituitary stem cells (PSCs), which uniquely express SOX2, contribute to pituitary growth and renewal. Moreover, neither the signals that drive proliferation nor their sources have been elucidated. We have used genetic approaches in the mouse, showing that the WNT pathway is essential for proliferation of all lineages in the gland. We reveal that SOX2+ stem cells are a key source of WNT ligands. By blocking secretion of WNTs from SOX2+ PSCs in vivo, we demonstrate that proliferation of neighbouring committed progenitor cells declines, demonstrating that progenitor multiplication depends on the paracrine WNT secretion from SOX2+ PSCs. Our results indicate that stem cells can hold additional roles in tissue expansion and homeostasis, acting as paracrine signalling centres to coordinate the proliferation of neighbouring cells.

Insulin and obesity transform hypothalamic-pituitary-adrenal axis stemness and function in a hyperactive state.

OBJECTIVE: Metabolic diseases are an increasing problem in society with the brain-metabolic axis as a master regulator of the human body for sustaining homeostasis under metabolic stress. However, metabolic inflammation and disease will trigger sustained activation of the hypothalamic-pituitary-adrenal axis. In this study, we investigated the role of metabolic stress on progenitor cells in the hypothalamic-pituitary-adrenal axis. METHODS: In vitro, we applied insulin and leptin to murine progenitor cells isolated from the pituitary and adrenal cortex and examined the role of these hormones on proliferation and differentiation. In vivo, we investigated two different mouse models of metabolic disease, obesity in leptin-deficient ob/ob mice and obesity achieved via feeding with a high-fat diet. RESULTS: Insulin was shown to lead to enhanced proliferation and differentiation of both pituitary and adrenocortical progenitors. No alterations in the progenitors were noted in our chronic metabolic stress models. However, hyperactivation of the hypothalamic-pituitary-adrenal axis was observed and the expression of the appetite-regulating genes Npy and Agrp changed in both the hypothalamus and adrenal. CONCLUSIONS: It is well-known that chronic stress and stress hormones such as glucocorticoids can induce metabolic changes including obesity and diabetes. In this article, we show for the first time that this might be based on an early sensitization of stem cells of the hypothalamic-pituitary-adrenal axis. Thus, pituitary and adrenal progenitor cells exposed to high levels of insulin are metabolically primed to a hyper-functional state leading to enhanced hormone production. Likewise, obese animals exhibit a hyperactive hypothalamic-pituitary-adrenal axis leading to adrenal hyperplasia. This might explain how stress in early life can increase the risk for developing metabolic syndrome in adulthood.

Local retinoic acid signaling directs emergence of the extraocular muscle functional unit.

Coordinated development of muscles, tendons, and their attachment sites ensures emergence of functional musculoskeletal units that are adapted to diverse anatomical demands among different species. How these different tissues are patterned and functionally assembled during embryogenesis is poorly understood. Here, we investigated the morphogenesis of extraocular muscles (EOMs), an evolutionary conserved cranial muscle group that is crucial for the coordinated movement of the eyeballs and for visual acuity. By means of lineage analysis, we redefined the cellular origins of periocular connective tissues interacting with the EOMs, which do not arise exclusively from neural crest mesenchyme as previously thought. Using 3D imaging approaches, we established an integrative blueprint for the EOM functional unit. By doing so, we identified a developmental time window in which individual EOMs emerge from a unique muscle anlage and establish insertions in the sclera, which sets these muscles apart from classical muscle-to-bone type of insertions. Further, we demonstrate that the eyeballs are a source of diffusible all-trans retinoic acid (ATRA) that allow their targeting by the EOMs in a temporal and dose-dependent manner. Using genetically modified mice and inhibitor treatments, we find that endogenous local variations in the concentration of retinoids contribute to the establishment of tendon condensations and attachment sites that precede the initiation of muscle patterning. Collectively, our results highlight how global and site-specific programs are deployed for the assembly of muscle functional units with precise definition of muscle shapes and topographical wiring of their tendon attachments.

New Horizons: Novel Adrenal Regenerative Therapies.

Adrenal insufficiency requires lifelong corticoid replacement therapies. However, current therapies are not able to replace the physiological circadian pattern of the adrenal cortex and are associated with many metabolic, vascular, neuroendocrine, and mental perturbations. Therefore, regenerative and more curative strategies would be desirable. In the current perspective, we describe emerging new regenerative therapies for the treatment of adrenal insufficiency. In particular, we discuss gene therapy and cell replacement strategies. Furthermore, we discuss how adrenal cells might be used as a source for regenerative therapies of nonadrenal neurodegenerative diseases such as Parkinson disease.

Retinoic acid synthesis by ALDH1A proteins is dispensable for meiosis initiation in the mouse fetal ovary.

In mammals, the timing of meiosis entry is regulated by signals from the gonadal environment. All-trans retinoic acid (ATRA) signaling is considered the key pathway that promotes Stra8 (stimulated by retinoic acid 8) expression and, in turn, meiosis entry. This model, however, is debated because it is based on analyzing the effects of exogenous ATRA on ex vivo gonadal cultures, which not accurately reflects the role of endogenous ATRA. Aldh1a1 and Aldh1a2, two retinaldehyde dehydrogenases synthesizing ATRA, are expressed in the mouse ovaries when meiosis initiates. Contrary to the present view, here, we demonstrate that ATRA-responsive cells are scarce in the ovary. Using three distinct gene deletion models for Aldh1a1;Aldh1a2;Aldh1a3, we show that Stra8 expression is independent of ATRA production by ALDH1A proteins and that germ cells progress through meiosis. Together, these data demonstrate that ATRA signaling is dispensable for instructing meiosis initiation in female germ cells.