NR5A1 Gene Research Articles

Parkinson’s disease-associated, sex-specific changes in DNA methylation at PARK7 (DJ-1), SLC17A6 (VGLUT2), PTPRN2 (IA-2β), and NR4A2 (NURR1) in cortical neurons

Evidence for epigenetic regulation playing a role in Parkinson’s disease (PD) is growing, particularly for DNA methylation. Approximately 90% of PD cases are due to a complex interaction between age, genes, and environmental factors, and epigenetic marks are thought to mediate the relationship between aging, genetics, the environment, and disease risk. To date, there are a small number of published genome-wide studies of DNA methylation in PD, but none accounted for cell type or sex in their analyses. Given the heterogeneity of bulk brain tissue samples and known sex differences in PD risk, progression, and severity, these are critical variables to account for. In this genome-wide analysis of DNA methylation in an enriched neuronal population from PD postmortem parietal cortex, we report sex-specific PD-associated methylation changes in PARK7 (DJ-1), SLC17A6 (VGLUT2), PTPRN2 (IA-2β), NR4A2 (NURR1), and other genes involved in developmental pathways, neurotransmitter packaging and release, and axon and neuron projection guidance.

Cocaine regulation of Nr4a1 chromatin bivalency and mRNA in male and female mice

Cocaine epigenetically regulates gene expression via changes in histone post-translational modifications (HPTMs). We previously found that the immediate early gene Nr4a1 is epigenetically activated by cocaine in mouse brain reward regions. However, few studies have examined multiple HPTMs at a single gene. Bivalent gene promoters are simultaneously enriched in both activating (H3K4me3 (K4)) and repressive (H3K27me3 (K27)) HPTMs. As such, bivalent genes are lowly expressed but poised for activity-dependent gene regulation. In this study, we identified K4&K27 bivalency at Nr4a1 following investigator-administered cocaine in male and female mice. We applied sequential chromatin immunoprecipitation and qPCR to define Nr4a1 bivalency and expression in striatum (STR), prefrontal cortex (PFC), and hippocampus (HPC). We used Pearson’s correlation to quantify relationships within each brain region across treatment conditions for each sex. In female STR, cocaine increased Nr4a1 mRNA while maintaining Nr4a1 K4&K27 bivalency. In male STR, cocaine enriched repressive H3K27me3 and K4&K27 bivalency at Nr4a1 and maintained Nr4a1 mRNA. Furthermore, cocaine epigenetically regulated a putative NR4A1 target, Cartpt, in male PFC. This study defined the epigenetic regulation of Nr4a1 in reward brain regions in male and female mice following cocaine, and, thus, shed light on the biological relevance of sex to cocaine use disorder.

Novel Genomic Variants, Atypical Phenotypes and Evidence of a Digenic/Oligogenic Contribution to Disorders/Differences of Sex Development in a Large North African Cohort.

In a majority of individuals with disorders/differences of sex development (DSD) a genetic etiology is often elusive. However, new genes causing DSD are routinely reported and using the unbiased genomic approaches, such as whole exome sequencing (WES) should result in an increased diagnostic yield. Here, we performed WES on a large cohort of 125 individuals all of Algerian origin, who presented with a wide range of DSD phenotypes. The study excluded individuals with congenital adrenal hypoplasia (CAH) or chromosomal DSD. Parental consanguinity was reported in 36% of individuals. The genetic etiology was established in 49.6% (62/125) individuals of the total cohort, which includes 42.2% (35/83) of 46, XY non-syndromic DSD and 69.2% (27/39) of 46, XY syndromic DSD. No pathogenic variants were identified in the 46, XX DSD cases (0/3). Variants in the AR, HSD17B3, NR5A1 and SRD5A2 genes were the most common causes of DSD. Other variants were identified in genes associated with congenital hypogonadotropic hypogonadism (CHH), including the CHD7 and PROKR2. Previously unreported pathogenic/likely pathogenic variants (n = 30) involving 25 different genes were identified in 22.4% of the cohort. Remarkably 11.5% of the 46, XY DSD group carried variants classified as pathogenic/likely pathogenic variant in more than one gene known to cause DSD. The data indicates that variants in PLXNA3, a candidate CHH gene, is unlikely to be involved in CHH. The data also suggest that NR2F2 variants may cause 46, XY DSD.

Adeno-Associated Virus-Mediated Gene Therapy for Patients' Fibroblasts, Induced Pluripotent Stem Cells, and a Mouse Model of Congenital Adrenal Hyperplasia.

Congenital adrenal hyperplasia (CAH) is an autosomal recessive disorder caused by steroidogenic enzymes containing monogenetic defects. Most steroidogenic enzymes are cytochrome P450 groups that can be categorized as microsomal P450s, including 21-hydroxylase and 17α-hydroxylase/17,20 lyase, and mitochondrial P450s, including 11β-hydroxylase. It has been shown that ectopic administration of Cyp21a1 ameliorates steroid metabolism in 21-hydroxylase-deficient mice. However, the effectiveness of this approach for mitochondrial P450 has not yet been evaluated. In this study, primary fibroblasts from patients with 21-hydroxylase deficiency (CYP21A2D) (n = 4), 17α-hydroxylase/17,20 lyase deficiency (CYP17A1D) (n = 1), and 11β-hydroxylase deficiency (CYP11B1D) (n = 1) were infected with adeno-associated virus type 2 (AAV2) vectors. Steroidogenic enzymatic activity was not detected in the AAV2-infected CYP11B1D fibroblasts. Induced pluripotent stem cells (iPSCs) of CYP11B1D were established and differentiated into adrenocortical cells by induction of the NR5A1 gene. Adrenocortical cells established from iPSCs of CYP11B1D (CYP11B1D-iPSCs) were infected with an AAV type 9 (AAV9) vector containing CYP11B1 and exhibited 11β-hydroxylase activity. For an in vivo evaluation, we knocked out Cyp11b1 in mice by using the CRISPR/Cas9 method. Direct injection of Cyp11b1-containing AAV9 vectors into the adrenal gland of Cyp11b1-deficient mice significantly reduced serum 11-deoxycorticosterone/corticosterone ratios at 4 weeks after injection and the effect was prolonged for up to 12 months. This study indicated that CYP11B1D could be ameliorated by gene induction in the adrenal glands, which suggests that a defective-enzyme-dependent therapeutic strategy for CAH would be required. Defects in microsomal P450, including CYP21A2D and CYP17A1D, can be treated with extra-adrenal gene induction. However, defects in mitochondrial P450, as represented by CYP11B1D, may require adrenal gene induction.

Case Report: Severe Gonadal Dysgenesis Causing 46,XY Disorder of Sex Development Due to a Novel NR5A1 Variant

Mutations in the nuclear receptor subfamily 5 group A member 1 (NR5A1) are the underlying cause of 10–20% of 46,XY disorders of sex development (DSDs). We describe a young girl with 46,XY DSD due to a unique novel mutation of the NR5A1 gene. An 11-year-old subject, raised as a female, was noticed to have clitromegly. She looked otherwise normal. However, her evaluation revealed a 46,XY karyotype, moderate clitromegly but otherwise normal female external genitalia, undescended atrophied testes, rudimentary uterus, no ovaries, and lack of breast development. Serum testosterone and estradiol were low, and gonadotropins were elevated. Adrenocortical function was normal. DNA was isolated from the peripheral leucocytes and used for whole exome sequencing. The results were confirmed by Sanger sequencing. We identified a novel mutation in NR5A1 changing the second nucleotide of the translation initiation codon (ATG>ACG) and resulting in a change of the first amino acid, methionine to threonine (p.Met1The). This led to severe gonadal dysgenesis with deficiency of testosterone and anti-Müllerian hormone (AMH) secretion. Lack of the former led to the development of female external genitalia, and lack of the latter allowed the Müllerian duct to develop into the uterus and the upper vagina. The patient has a female gender identity. Bilateral orchidectomy was performed and showed severely atrophic testes. Estrogen/progesterone therapy was initiated with excellent breast development and normal cyclical menses. In summary, we describe a severely affected case of 46,XY DSD due to a novel NR5A1 mutation involving the initiation codon that fully explains the clinical phenotype in this subject.

Can Non-Coding NR5A1 Gene Variants Explain Phenotypes of Disorders of Sex Development?

Introduction: NR5A1 is an essential transcription factor that regulates several target genes involved in reproduction and endocrine function. Pathogenic variants in this gene are responsible for a wide spectrum of disorders/differences of sex development (DSD). Methods: The molecular study involved Sanger sequencing, in vitro assays, and whole exome sequencing (WES). Results: Four variants were identified within the NR5A1 non-coding region in 3 patients with 46,XY DSD. In vitro analyses showed that promoter activity was affected in all cases. WES revealed variants in SRA1, WWOX, and WDR11 genes. Discussion/Conclusion: Evaluation of clinical and phenotypic significance of variants located in a non-coding region of a gene can be complex, and little is known regarding their association with DSD. Nevertheless, based on the important region for interaction with cofactors essential to promote appropriated sex development and on our in vitro results, it is feasible to say that an impact on gene expression can be expected and that this may be correlated with the DSD pathophysiology presented in our patients. Considering the number of cases that remain elusive after screening for the well-known DSD related genes, we emphasize the importance of a careful molecular analysis of NR5A1 non-coding region which is commonly neglected and might explain some idiopathic DSD cases.

Abstract 4239: Development of multi-targeting immunomodulatory SeekR RNA therapeutics to overcome resistance to PD-1 blockade

Abstract T-cell exhaustion describes the dysfunctional state of effector T cells in the tumor microenvironment that results from chronic and prolonged exposure to antigen. It also represents an important mechanism of clinical resistance to immune checkpoint blockade. Recently, the transcription factor NR4A1 has been shown to be upregulated in tumor-specific T cells and plays a role in driving exhaustion during chronic antigen stimulation of T cells. Pro-oncogenic activities of NR4A1 have long been observed in various solid tumors including colorectal and breast cancers, and co-expression of NR4A1 with known immune checkpoint markers such as PD-L1 have also been observed in various cancer cell lines. These findings have identified NR4A1 as an important target for overcoming resistance to cancer immunotherapy. We hypothesize that multi-targeting to simultaneously block PD-1 signaling and silence NR4A1 gene expression can reverse T-cell exhaustion and expand the clinical benefit of PD-1 blockade in immune therapy resistant solid tumors. Using our SeekR™ RNA therapeutics platform, we combined aptamer binders to PD-1 with siRNA against NR4A1, andNR4A1 and evaluated their ability to reverse T-cell exhaustion and re-activate T-cell based anticancer immunity in solid tumors. Specifically, we created SeekR™ therapeutic RNA molecules with dual flanking RNA aptamers binders to PD-1 and other checkpoint inhibitors, connected by a double stranded bridge that encodes for siRNA silencers that target multiple immunomodulatory genes, such as NR4A1. The novel anti-PD-1 RNA aptamer component serve to: A) direct and deliver the SeekR™ RNA oligo therapeutics to exhausted T-cells that express PD-1 receptors; and B) internalize the siRNA component into the T-cells. The siRNA components of the SeekR™ serve to silence NR4A1 which lead to reversal of the cytotoxic T-cell exhausted phenotype following chronic in-vitro stimulation. These results demonstrate that dual targeting of PD-1 and NR4A1 has the potential to reverse T-cell exhaustion and the potential to overcome cancer immunotherapy resistance in the clinic. Citation Format: Marvin O'ketch, Victoria A. Love, Jeffrey A. Kiefer, David W. Lee, Lizette A. Castaño, Spyro Mousses, David O. Azorsa. Development of multi-targeting immunomodulatory SeekR RNA therapeutics to overcome resistance to PD-1 blockade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4239.

Effects and related mechanisms of Huang's SiYu pills on Leydig cell function in kidney-yin deficiency rat models

The prescription “Huang's SiYu Pill” (HSYP) was founded by professor Huang Haibo, a famous traditional Chinese medicine in Inner Mongolia, based on long-term clinical experience. In this study we observed the effects of HYSP on sperm quality, hormone level and the expression level of reproductive related genes and proteins in testicular stromal cells of rats with kidney-yin deficiency to explore the therapeutic mechanism of HSYP in the treatment of infertility with kidney-yin deficiency, Liuwei Dihuang Pill group (LDP) and Tamoxifen Citrate Tablets combined with Pancreatic Kallidinogenase Enteric Coated Tablets were used to be the positive control drug. After 25 days of medicine administration, the animals were anesthetized and killed, the serum and testicular tissues were retained for further experiments. The ELISA results showed that HSYP had a better effect on increasing serum FSH level than the conventional Chinese medicine or western medicine, while there was no remarkable difference in the regulation of serum LH in each administration group. The results of RT-qPCR showed that the transcription levels of Cyp11a1, Hsd3b1, Lhcgr and Star genes in the medium and high dose groups of HSYP were significantly upregulated compared with the model group of kidney-yin deficiency, while the expression of Nr5a1 gene was not significantly affected. The results of HE and immunohistochemistry showed that HSYP significantly improved the development level of testicular stromal cells and promoted the expression levels of CYP11A1 and HSD11B1 in testicular stromal cells, compared with the model group of kidney-yin deficiency. Together, our results showed that Huang's SiYu Pills can significantly improve the development and function of Leydig cells in SD rats with kidney-yin deficiency model, which provided a good research basis for deep research on relevant pharmacological mechanisms of the medicine.

Whole exome sequencing applied to 42 Han Chinese patients with posterior hypospadias

Hypospadias, a malformation of male external genitalia, is characterized by an aberrant opening of the urethra on the ventral side of the penis. It is considered a complex disorder with both environmental and genetic factors involved in its pathogenesis. To identify the genetic abnormality involved in the pathogenesis of hypospadias, we performed whole exome sequencing (WES) analysis in 42 hypospadias patients with karyotype 46, XY in the Nanhai Meternity&Child Health Hospital of Foshan. All the likely pathogenic variants were confirmed by Sanger sequencing and assessed by Sorting Intolerant from Tolerant (SIFT), PROVEAN, PolyPhen2, ClinPred, LRT, Mutation Assessor, FATHMM, and GERP software. We discovered 27 gene mutations in 20 patients, including eight cases of the SRD5A2 gene, 4 cases of the AR gene, 3 cases of the CYP17A1 gene, 1 case of the WT1 gene, 1 case of the ANOS1 gene, 1 case of the NR5A1 gene, 1 case of the FGFR1 gene, and one case of the DHX37 gene. Our study is the first to describe six novel missense mutations, AR(c.302G > A, c.2593G > T, and c.1705G > T), CYP17A1(c.1298 T > C), FGFR1 (c.995C > T) and DHX37(c.923G > A). In summary, genetic defect detection was useful for early diagnosis of severe hypospadias in the Han Chinese population. Nevertheless, most cases remain unexplained, and the exact pathogenesis of hypospadias still needs further study.

Association study of NR5A1 rs1110061 with infertile male in 401 Vietnamese individuals

Male infertility is a reproductive issue involving defects in the quantity and quality of sperm. Besides the exogenous elements such as harmful habits, pollution, genetic factors including disorders in a single gene, group of genes, or chromosomes, are the etiology causing infertility in men. NR5A1 is well-known as a candidate gene involved in male infertility, including 46, XY disorders of sex development (DSD), cryptorchidism, micropenis, spermatogenic failure, non-obstructive azoospermia, and oligozoospermia. This study aimed to identify the single nucleotide polymorphism (SNP) associated with male infertility in the NR5A1 gene in a Vietnamese cohort of 202 infertile men and 199 healthy controls. By directly sequencing the coding region of the NR5A1 gene in 56 cases and 21 controls, only one missense mutation c.437G>C (p.Gly146Ala; rs1110061) in exon 4 was found. The variant was detected in 34 patients, of which 27 were heterozygous (GC) and 7 were homozygous (CC). In the control group, 15 individuals carried this mutation, including 10 with heterozygous (GC) and 5 with homozygous (CC). To further investigate the association of the polymorphism NR5A1 rs11100061 with male infertility disease, we performed PCR-RFLP in 202 infertile and 199 healthy men to assess the genotypes and alleles. The results indicated that the distribution of the genotypes of the polymorphism was in accordance with Hardy-Weinberg equilibrium (p-values > 0.05). However, no association was detected between the polymorphism and male infertility (p-values > 0.05).

WT1, NR0B1, NR5A1, LHX9, ZFP92, ZNF275, INSL3, and NRIP1 Genetic Variants in Patients with Premature Ovarian Insufficiency in a Mexican Cohort.

Premature ovarian insufficiency (POI) is one of the main causes of female premature infertility. POI is a genetically heterogeneous disorder with a complex molecular etiology; as such, the genetic causes remain unknown in the majority of patients. Therefore, this study aimed to identify mutations and characterize the associated molecular contribution of gonadogenesis-determinant genes to POI. Genomic assays, including PCR-SSCP and Sanger sequencing, followed by in silico analyses were used to investigate the underpinnings of ovarian deficiency in 11 women affected by POI. Large deletions and nucleotide insertions and duplications were excluded by PCR. Thirteen genetic variants were identified in the WT1 (c.213G>T, c.609T>C, c.873A>G, c.1122G>A), NR0B1 (c.353C>T, c.425G>A), NR5A1 (c.437G>C, IVS4-20C>T), LHX9 (IVS2-12G>C, IVS3+13C>T, c.741T>C), ZNF275 (c.969C>T), and NRIP1 (c.3403C>T) genes. Seven novel genetic variants and five unpublished substitutions were identified. No genetic aberrations were detected in the ZFP92 and INSL3 genes. Each variant was genotyped using PCR-SSCP in 100 POI-free subjects, and their allelic frequencies were similar to the patients. These analyses indicated that allelic variation in the WT1, NR0B1, NR5A1, LHX9, ZFP92, ZNF275, INSL3, and NRIP1 genes may be a non-disease-causing change or may not contribute significantly to the genetics underlying POI disorders. Findings support the polygenic nature of this clinical disorder, with the SNVs identified representing only a probable contribution to the variability of the human genome.

Disorders of Sex Development in a Large Ukrainian Cohort: Clinical Diversity and Genetic Findings.

BackgroundThe clinical profile and genetics of individuals with Disorders/Differences of Sex Development (DSD) has not been reported in Ukraine.Materials and MethodsWe established the Ukrainian DSD Register and identified 682 DSD patients. This cohort includes, 357 patients (52.3% [303 patients with Turner syndrome)] with sex chromosome DSD, 119 (17.5%) with 46,XY DSD and 206 (30.2%) with 46,XX DSD. Patients with sex chromosome DSD and congenital adrenal hyperplasia (CAH, n=185) were excluded from further studies. Fluorescence in situ hybridization (FISH) was performed for eight 46,XX boys. 79 patients underwent Whole Exome Sequencing (WES).ResultsThe majority of patients with 46,XY and 46,XX DSD (n=140), were raised as female (56.3% and 61.9% respectively). WES (n=79) identified pathogenic (P) or likely pathogenic (LP) variants in 43% of the cohort. P/LP variants were identified in the androgen receptor (AR) and NR5A1 genes (20.2%). Variants in other DSD genes including AMHR2, HSD17B3, MYRF, ANOS1, FGFR11, WT1, DHX37, SRD5A1, GATA4, TBCE, CACNA1A and GLI2 were identified in 22.8% of cases. 83.3% of all P/LP variants are novel. 35.3% of patients with a genetic diagnosis had an atypical clinical presentation. A known pathogenic variant in WDR11, which was reported to cause congenital hypogonadotropic hypogonadism (CHH), was identified in individuals with primary hypogonadism.ConclusionsWES is a powerful tool to identify novel causal variants in patients with DSD, including a significant minority that have an atypical clinical presentation. Our data suggest that heterozygous variants in the WDR11 gene are unlikely to cause of CHH.

Influence of CPM-dependent sorting on the multi-omics profile of hepatocyte-like cells matured in microscale biochips

Liver modeling in disease via advanced in vitro physiological tissues remains a challenging issue, yet crucial for the future development of relevant tools for drug screening. In that regard, advanced technics such as human induced Pluripotent Stem Cells (hiPSCs) and organ-on-chip are promising technologies for the relevant reproduction of the in vivo micro physiology in in vitro culture conditions. In the present work, the maturation of carboxypeptidase M positive (CPM+) Hepatocyte-Like-Cells (HLCs) in microfluidic culture conditions was investigated. hiPSCs were differentiated in culture dishes until sorting and further amplified until seeding in biochips. After 2 weeks of maturation in biochips, evaluation of the cell metabolism was performed, and samples were collected for characterization via RNA sequencing, proteomics and metabolomics. The omics profile of biochips loaded with CPM+ HLCs was then compared with the one of previously cultured biochips loaded with unsorted HLCs. CPM+ HLCs presented advanced liver characteristics in terms of a higher activity of important Transcriptions Factors (TFs) such as HNF1, HNF4A, and CEBP/A, of the upregulation of steroids/corticoids-related nuclear receptors (FXR, NR3C2, NR4A1 genes), of phase I, and phase II metabolism genes (CYP1A1, CYP1B1, CYP2C18, CYP27A1, several UGT, SULT and GST genes). Further differences in the cellular reorganization, in the lipids and steroids metabolisms, in the OXPHOS respiration and in the expression of TGFβ signaling were also observed. The present study introduces a novel protocol using advances hiPSCs differentiation and sorting methods as well as the organ-on-chip technology to highlight important in vitro liver regeneration and hepatic maturation processes.

The Selective Agonist for Sphingosine-1-Phosphate Receptors Siponimod Increases the Expression Level of NR4A Genes in Microglia Cell Line.

Fingolimod (FTY720) and siponimod (BAF312) are selective agonists for sphingosine-1-phosphate (S1P) receptors approved for the treatment of relapsing–remitting (RR) and secondary progressive (SP) multiple sclerosis (MS), respectively. BAF312 exerts pro-myelination and neuro-protective functions on CNS resident cells, although the underlying molecular mechanism is not yet fully understood. NR4A2 is an anti-inflammatory gene, belonging to the NR4A family, whose expression is reduced in blood from treatment-naïve patients with RRMS, but is restored in patients treated with FTY720 for more than two years. We performed an in vitro study to investigate the potential involvement of the NR4A genes in the protective and restorative effects of BAF312. We showed that BAF312 enhances the expression of NR4A1 and NR4A2 in the N9 microglial cell line, but has no effect in the peripheral blood mononuclear cells and oligodendrocytes. This study suggests a novel molecular mechanism of action for the selective agonists for S1P receptors within the CNS.

Structural, molecular hybridization and network based identification of miR-373-3p and miR-520e-3p as regulators of NR4A2 human gene involved in neurodegeneration

MicroRNAs (miRNAs) are short non-coding RNAs with a 22 nucleotide sequence length and docks to the 3′UTR/5’UTR of the gene to regulate their mRNA translation to play a vital role in neurodegenerative diseases. The Nuclear Receptor gene (NR4A2), a transcription factor, and a steroid-thyroid hormone retinoid receptor is involved in neural development, memory formation, dopaminergic neurotransmission, and cellular protection from inflammatory damage. Therefore, recognizing the miRNAs is essential to efficiently target the 3′UTR/5’UTR of the NR4A2 gene and regulate neurodegeneration. Highly stabilized top miRNA-mRNA hybridized structures, their homologs, and identification of the best structures based on their least free energy were evaluated using in silico techniques. The miR-gene, gene-gene network analysis, miR-disease association, and transcription factor binding sites were also investigated. Results suggest top 166 miRNAs targeting the NR4A2 mRNA, but with a total of 10 miRNAs bindings with 100% seed sequence identity (both at 3’ and 5’UTR) at the same position on the NR4A2 mRNA region. The miR-373-3p and miR-520e-3p are considered the best candidate miRNAs hybridizing with high efficiency at both 3’ and 5’UTR of NR4A2 mRNA. This could be due to the most significant seed sequence length complementary, supplementary pairing, and absence of non-canonical base pairs. Furthermore, the miR-gene network, target gene-gene interaction analysis, and miR-disease association provide an understanding of the molecular, cellular, and biological processes involved in various pathways regulated by four transcription factors (PPARG, ZNF740, NRF1, and RREB1). Therefore, miR-373-3p, 520e-3p, and four transcription factors can regulate the NR4A2 gene involved in the neurodegenerative process.

Evaluation of animal model congruence to human depression based on large-scale gene expression patterns of the CNS

Depression is a complex mental health disorder that is difficult to study. A wide range of animal models exist and for many of these data on large-scale gene expression patterns in the CNS are available. The goal of this study was to evaluate how well animal models match human depression by evaluating congruence and discordance of large-scale gene expression patterns in the CNS between almost 300 animal models and a portrait of human depression created from male and female datasets. Multiple approaches were used, including a hypergeometric based scoring system that rewards common gene expression patterns (e.g., up-up or down-down in both model and human depression), but penalizes opposing gene expression patterns. RRHO heat maps, Uniform Manifold Approximation Plot (UMAP), and machine learning were used to evaluate matching of models to depression. The top ranked model was a histone deacetylase (HDAC2) conditional knockout in forebrain neurons. Also highly ranked were various models for Alzheimer’s, including APPsa knock-in (2nd overall), APP knockout, and an APP/PS1 humanized double mutant. Other top models were the mitochondrial gene HTRA2 knockout (that is lethal in adulthood), a modified acetylcholinesterase, a Huntington’s disease model, and the CRTC1 knockout. Over 30 stress related models were evaluated and while some matched highly with depression, others did not. In most of the top models, a consistent dysregulation of MAP kinase pathway was identified and the genes NR4A1, BDNF, ARC, EGR2, and PDE7B were consistently downregulated as in humans with depression. Separate male and female portraits of depression were also evaluated to identify potential sex specific depression matches with models. Individual human depression datasets were also evaluated to allow for comparisons across the same brain regions. Heatmap, UMAP, and machine learning results supported the hypergeometric ranking findings. Together, this study provides new insights into how large-scale gene expression patterns may be similarly dysregulated in some animals models and humans with depression that may provide new avenues for understanding and treating depression.

Nur77-Tempo mice reveal T cell steady state antigen recognition.

In lymphocytes, Nr4a gene expression is specifically regulated by antigen receptor signalling, making them ideal targets for use as distal T cell receptor (TCR) reporters. Nr4a3-Timer of cell kinetics and activity (Tocky) mice are a ground-breaking tool to report TCR-driven Nr4a3 expression using Fluorescent Timer protein (FT). FT undergoes a time-dependent shift in its emission spectrum following translation, allowing for the temporal reporting of transcriptional events. Our recent work suggested that Nr4a1/Nur77 may be a more sensitive gene to distal TCR signals compared to Nr4a3, so we, therefore, generated Nur77-Timer-rapidly-expressed-in-lymphocytes (Tempo) mice that express FT under the regulation of Nur77. We validated the ability of Nur77-Tempo mice to report TCR and B cell receptor signals and investigated the signals regulating Nur77-FT expression. We found that Nur77-FT was sensitive to low-strength TCR signals, and its brightness was graded in response to TCR signal strength. Nur77-FT detected positive selection signals in the thymus, and analysis of FT expression revealed that positive selection signals are often persistent in nature, with most thymic Treg expressing FT Blue. We found that active TCR signals in the spleen are low frequency, but CD69+ lymphoid T cells are enriched for FT Blue+ Red+ T cells, suggesting frequent TCR signalling. In non-lymphoid tissue, we saw a dissociation of FT protein from CD69 expression, indicating that tissue residency is not associated with tonic TCR signals. Nur77-Tempo mice, therefore, combine the temporal dynamics from the Tocky innovation with increased sensitivity of Nr4a1 to lower TCR signal strengths.

NR4A1 agonist cytosporone B attenuates neuroinflammation in a mouse model of multiple sclerosis.

Nuclear receptor subfamily 4 group A1 (NR4A1) is an orphan nuclear receptor, which is expressed in the majority of cells. NR4A1 expression in peripheral blood mononuclear cells is low during the preclinical stage of multiple sclerosis. Knockout of the Nr4a1 gene in mice can aggravate the symptoms of experimental autoimmune encephalomyelitis (EAE), which is an animal model of multiple sclerosis. In this study, we intragastrically administered the NR4A1 agonist cytosporone B (Csn-B) to mice after inducing EAE. After treatment with Csn-B, the clinical symptoms in the EAE mice were substantially attenuated compared with that in PBS-treated control mice. The percentages of CD4+ T cells and F4/80+ cells in the central nervous system were decreased. In addition, interferon-γ and interleukin-17 production by proinflammatory Th1/Th17 cells in the central nervous system and interferon-γ levels in splenocytes were decreased after Csn-B treatment. These findings suggest that the NR4A1 agonist Csn-B can alleviate nerve injury after EAE induction, and, therefore, may be useful as a potential treatment for multiple sclerosis.

Feminizing genitoplasty for prepubertal children and teenagers female

The aim of this study was to report our experience after 10 years of practice of feminizing genitoplasty in prepubertal and adolescent patients with disorders of sex development (DSD) assigned females as females in a developing country. This was a cross-sectional, descriptive and retrospective study over a period of 9 years. All pre-pubertal (8-12 years) and adolescent patients female sex assigned with DSD who had willfully consented to the surgery with their guardians and underwent feminizing genital surgery were enrolled in the study. Data collection included: age at presentation, precise diagnosis, surgical procedures, complications, cosmetic result and duration of follow-up. Each patient had a precise diagnosis and the surgery was planned after discussion with the multidisciplinary team. Cosmetic results were assessed based on: appearance of the clitoris and separation of the vaginal and urethral openings. Nine patients raised as females with a median age of 8 years (IR: 10.75) were recorded. Surgery was performed at a median age of 11 years (IR: 9.5). In this series, 6 had a 46, XY karyotype with varying diagnoses: partial androgen insensitivity syndrome (n=2); 5-alphareductase insufficiency (n=2); 17-ketoreductase insufficiency (n=2); gonadal dysgenesis with a mutation in the NR5A1 gene (n=2), 2 had ovostesticular DSD, (karyotypes 46, XX), and 1 had mixed gonadal dysgenesis (karyotype 45, X/46, XY). Partial or total gonad(s) removal in accordance with assigned gender was the most common associated procedure. It was bilateral in 7 cases and unilateral in 2 cases. Follow-up ranged from 3 months to 4.5 years (median: 26 months, IR:18.25). One patient had acute urinary retention in the early follow-up. No other complication such as incision bleeding was recorded. The cosmetic appearance of the external genitalia was satisfactory in all patients. Feminizing genital surgery in Cameroon remains a major challenge and should seldom be realized without a precise diagnosis. Late age at presentation is peculiar to our setting; however, it gives room for the patients' participation and input to decisions that will have a life-long personal impact on their lives in terms of psychosocial development and fertility. 3.

Functional heterogeneity and adaptation of naive T cells in response to tonic TCR signals.

Mature CD4+ and CD8+ T cells constitutively experience weak T cell receptor (TCR) stimulation in response to self-antigens, termed tonic (or basal) signaling. How tonic TCR signal strength impacts T cell responses to foreign antigens is an active area of investigation. Such studies rely on surrogate markers of tonic signal strength, including CD5, Ly6C, and transgenic reporters of Nr4a genes. Recent research indicates that strong tonic TCR signal strength influences basal T cell metabolism, effector differentiation, and TCR signal transduction. T cells that experience the strongest tonic TCR signaling exhibit features of T cell activation and negative regulation. These data suggest a model whereby adaptation to tonic signaling has lasting effects that alter T cell activation and differentiation.