RESUMEN
Triple-negative breast cancer (TNBC) is a subtype of breast cancer with aggressive behavior and poor prognosis. Current therapeutic options available for TNBC patients are primarily chemotherapy. With our evolving understanding of this disease, novel targeted therapies, including poly ADP-ribose polymerase (PARP) inhibitors, antibody-drug conjugates, and immune-checkpoint inhibitors, have been developed for clinical use. Previous reports have demonstrated the essential role of estrogen receptor ß (ERß) in TNBC, but the detailed molecular mechanisms downstream ERß activation in TNBC are still far from elucidated. In this study, we demonstrated that a specific ERß agonist, LY500307, potently induces R-loop formation and DNA damage in TNBC cells. Subsequent interactome experiments indicated that the residues 151 to 165 of U2 small nuclear RNA auxiliary factor 1 (U2AF1) and the Trp439 and Lys443 of ERß were critical for the binding between U2AF1 and ERß. Combined RNA sequencing and ribosome sequencing analysis demonstrated that U2AF1-regulated downstream RNA splicing of 5-oxoprolinase (OPLAH) could affect its enzymatic activity and is essential for ERß-induced R-loop formation and DNA damage. In clinical samples including 115 patients from The Cancer Genome Atlas (TCGA) and 32 patients from an in-house cohort, we found a close correlation in the expression of ESR2 and U2AF1 in TNBC patients. Collectively, our study has unraveled the molecular mechanisms that explain the therapeutic effects of ERß activation in TNBC, which provides rationale for ERß activation-based single or combined therapy for patients with TNBC.
Asunto(s)
Empalme Alternativo , Benzopiranos , Receptor beta de Estrógeno , Estructuras R-Loop , Factor de Empalme U2AF , Neoplasias de la Mama Triple Negativas , Humanos , Receptor beta de Estrógeno/agonistas , Receptor beta de Estrógeno/metabolismo , Factor de Empalme U2AF/química , Factor de Empalme U2AF/genética , Factor de Empalme U2AF/metabolismo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismo , Terapia Combinada , Células MDA-MB-231 , Empalme Alternativo/efectos de los fármacos , Benzopiranos/farmacología , Benzopiranos/uso terapéutico , Unión Proteica , Sitios de UniónRESUMEN
Knockout of ERß in the mouse leads to nuclear expression of epidermal growth factor receptor (EGFR) in the prostate. To examine whether ERß plays a similar role in the human prostate, we used four cohorts of men: 1) a Swedish cohort of normal prostates and PCa (prostate cancer) of different Gleason grades; 2) men with benign prostatic hyperplasia (BPH) treated with the 5α-reductase inhibitor, finasteride, and finasteride together with the ERß agonists, soy isoflavones; 3) men with PCa above Gleason grade 4 (GG4), treated with ADT (androgen deprivation therapy) and abiraterone (AA), the blocker of androgen synthesis for different durations; and 4) men with GG4 PCa on ADT or ADT with the AR (androgen receptor) blocker, enzalutamide, for 4 mo to 6 mo. In men with BPH, finasteride treatment induced EGFR nuclear expression, but, when finasteride was combined with isoflavones, EGFR remained on the cell membrane. In GG4 patients, blocking of AR for 4 mo to 6 mo resulted in loss of ERß and PTEN expression and increase in patients with nuclear EGFR from 10 to 40%. In the men with GG4 PCa, blocking of adrenal synthesis of testosterone for 2 mo to 7 mo had the beneficial effect of increasing ERß expression, but, on treatment longer than 8 mo, ERß was lost and EGFR moved to the nucleus. Since nuclear EGFR is a predictor of poor outcome in PCa, addition of ERß agonists together with abiraterone should be considered as a treatment that might sustain expression of ERß and offer some benefit to patients.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Receptor beta de Estrógeno/agonistas , Hiperplasia Prostática/tratamiento farmacológico , Neoplasias de la Próstata/tratamiento farmacológico , Transporte Activo de Núcleo Celular/efectos de los fármacos , Antagonistas de Andrógenos/farmacología , Antagonistas de Andrógenos/uso terapéutico , Androstenos/farmacología , Androstenos/uso terapéutico , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Benzamidas/farmacología , Benzamidas/uso terapéutico , Biopsia , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Estudios de Cohortes , Receptores ErbB/metabolismo , Receptor beta de Estrógeno/metabolismo , Finasterida/farmacología , Finasterida/uso terapéutico , Humanos , Masculino , Ratones , Ratones Noqueados , Clasificación del Tumor , Nitrilos/farmacología , Nitrilos/uso terapéutico , Fosfohidrolasa PTEN/metabolismo , Feniltiohidantoína/farmacología , Feniltiohidantoína/uso terapéutico , Fitoestrógenos/farmacología , Fitoestrógenos/uso terapéutico , Próstata/citología , Próstata/efectos de los fármacos , Próstata/patología , Hiperplasia Prostática/patología , Neoplasias de la Próstata/patología , Receptores Androgénicos/metabolismo , Receptores de Estrógenos/genética , Receptores de Estrógenos/metabolismoRESUMEN
To identify regulators of triple-negative breast cancer (TNBC), gene expression profiles of malignant parts of TNBC (mTNBC) and normal adjacent (nadj) parts of the same breasts have been compared. We are interested in the roles of estrogen receptor ß (ERß) and the cytochrome P450 family (CYPs) as drivers of TNBC. We examined by RNA sequencing the mTNBC and nadj parts of five women. We found more than a fivefold elevation in mTNBC of genes already known to be expressed in TNBC: BIRC5/survivin, Wnt-10A and -7B, matrix metalloproteinases (MMPs), chemokines, anterior gradient proteins, and lysophosphatidic acid receptor and the known basal characteristics of TNBC, sox10, ROPN1B, and Col9a3. There were two unexpected findings: 1) a strong induction of CYPs involved in activation of fatty acids (CYP4), and in inactivation of calcitriol (CYP24A1) and retinoic acid (CYP26A1); and 2) a marked down-regulation of FOS, FRA1, and JUN, known tethering partners of ERß. ERß is expressed in 20 to 30% of TNBCs and is being evaluated as a target for treating TNBC. We used ERß+ TNBC patient-derived xenografts in mice and found that the ERß agonist LY500703 had no effect on growth or proliferation. Expression of CYPs was confirmed by immunohistochemistry in formalin-fixed and paraffin-embedded (FFPE) TNBC. In TNBC cell lines, the CYP4Z1-catalyzed fatty acid metabolite 20-hydroxyeicosatetraenoic acid (20-HETE) increased proliferation, while calcitriol decreased proliferation but only after inhibition of CYP24A1. We conclude that CYP-mediated pathways can be drivers of TNBC but that ERß is unlikely to be a tumor suppressor because the absence of its main tethering partners renders ERß functionless on genes involved in proliferation and inflammation.
Asunto(s)
Sistema Enzimático del Citocromo P-450/metabolismo , Receptor beta de Estrógeno/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Neoplasias de la Mama Triple Negativas/metabolismo , Proteínas Anfibias/genética , Proteínas Anfibias/metabolismo , Animales , Benzopiranos/farmacología , Calcitriol/farmacología , Sistema Enzimático del Citocromo P-450/genética , Regulación hacia Abajo , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Receptor beta de Estrógeno/genética , Ácidos Grasos/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/fisiología , Humanos , Ratones , Neoplasias Experimentales , Distribución Aleatoria , Survivin/genética , Survivin/metabolismo , Transcriptoma , Tretinoina/farmacología , Neoplasias de la Mama Triple Negativas/genética , Proteínas Wnt/genética , Proteínas Wnt/metabolismoRESUMEN
Members of the nuclear receptor (NR) superfamily orchestrate cellular processes that can impact on numerous cancer hallmarks. NR activity plays important roles in the tumor microenvironment by controlling inflammation and immune responses. We summarize recent insights into the diverse mechanisms by which NR activity can control tumor inflammation, the roles of different NRs in modulating tumor immunity, and the biological features of immune cells that express specific NRs in the context of cancer. NR-dependent alterations in tumor inflammation and immunity may be amenable to pharmacological manipulation and offer new clues regarding the development of novel cancer therapeutic regimens.
Asunto(s)
Neoplasias , Receptores Citoplasmáticos y Nucleares , Humanos , Inflamación/inmunología , Neoplasias/inmunología , Receptores Citoplasmáticos y Nucleares/inmunología , Transducción de Señal , Microambiente Tumoral/inmunologíaRESUMEN
BACKGROUND: Cholesterol loaded macrophage foam cells are a prominent feature of atherosclerotic plaques. Single-cell RNA sequencing has identified foam cells as TREM2 (triggering receptor expressed on myeloid cells 2) positive populations with low expression of inflammatory genes, resembling the TREM2 positive microglia of neurodegenerative diseases. Cholesterol loading of macrophages in vitro results in activation of LXR (liver X receptor) transcription factors and suppression of inflammatory genes. METHODS: To test the hypothesis that LXRs mediate anti-inflammatory effects in Trem2 expressing atherosclerotic plaque foam cells, we performed RNA profiling on plaque cells from hypercholesterolemic mice with myeloid LXR deficiency. RESULTS: Myeloid LXR deficiency led to a dramatic increase in atherosclerosis with increased monocyte entry, foam cell formation, and plaque inflammation. Bulk cell-RNA profiling of plaque myeloid cells showed prominent upregulation of inflammatory mediators including oxidative, chemokine, and chemotactic genes. Single-cell RNA sequencing revealed increased numbers of foamy TREM2-expressing macrophages; however, these cells had reduced expression of the Trem2 gene expression module, including phagocytic and cholesterol efflux genes, and had switched to a proinflammatory and proliferative phenotype. Expression of Trem2 was suppressed by inflammatory signals but not directly affected by LXR activation in bone marrow-derived macrophages. CONCLUSIONS: Our current studies reveal the key role of macrophage LXRs in promoting the Trem2 gene expression program and in suppressing inflammation in foam cells of atherosclerotic plaques.
Asunto(s)
Aterosclerosis , Placa Aterosclerótica , Animales , Aterosclerosis/genética , Aterosclerosis/metabolismo , Colesterol/metabolismo , Células Espumosas/metabolismo , Expresión Génica , Inflamación/genética , Inflamación/metabolismo , Receptores X del Hígado/genética , Receptores X del Hígado/metabolismo , Macrófagos/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Ratones , Placa Aterosclerótica/metabolismo , ARN , Receptores Inmunológicos/genética , Receptores Inmunológicos/metabolismoRESUMEN
Loss of the tumor suppressor, PTEN, is one of the most common findings in prostate cancer (PCa). This loss leads to overactive Akt signaling, which is correlated with increased metastasis and androgen independence. However, another tumor suppressor, inositol-polyphosphate 4-phosphatase type II (INPP4B), can partially compensate for the loss of PTEN. INPP4B is up-regulated by androgens, and this suggests that androgen-deprivation therapy (ADT) would lead to hyperactivity of AKT. However, in the present study, we found that in PCa, samples from men treated with ADT, ERß, and INPP4B expression were maintained in some samples. To investigate the role of ERß1 in regulation of INPPB, we engineered the highly metastatic PCa cell line, PC3, to express ERß1. In these cells, INPP4B was induced by ERß ligands, and this induction was accompanied by inhibition of Akt activity and reduction in cell migration. These findings reveal that, in the absence of androgens, ERß1 induces INPP4B to dampen AKT signaling. Since the endogenous ERß ligand, 3ß-Adiol, is lost upon long-term ADT, to obtain the beneficial effects of ERß1 on AKT signaling, an ERß agonist should be added along with ADT.
Asunto(s)
Receptor beta de Estrógeno/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Neoplasias de la Próstata/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Antagonistas de Andrógenos/metabolismo , Andrógenos/farmacología , Línea Celular Tumoral , Movimiento Celular/fisiología , Proliferación Celular/fisiología , Humanos , Masculino , Células PC-3 , Fosfohidrolasa PTEN/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Neoplasias de la Próstata/patología , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismo , Transducción de SeñalRESUMEN
Disagreements about the phenotype of estrogen receptor ß (ERß) knockout mouse, created by removing the DNA-binding domain of the ERß gene or interruption of the gene with a neocassette (Oliver Smithies ERß knockout mice [ERßOS-/-]), prompted us to create an ERß knockout mouse by deleting the ERß gene with the use of CRISPR/Cas9 technology. We confirmed that the ERß gene was eliminated from the mouse genome and that no ERß mRNA or protein was detectable in tissues of this mouse. Overall the phenotype of the ventral prostate (VP) and mammary gland (MG) in ERßcrispr-/- mice was similar to, but more severe than, that in the ERßOS-/-mice. In the VP of 6-mo-old ERßcrispr-/- mice there was epithelial hyperplasia, fibroplasia, inflammation, stromal overgrowth, and intraductal cancer-like lesions. This was accompanied by an increase in Ki67 and P63 and loss in DACH1 and PURα, two androgen receptor (AR) repressors. In the MG there was overexpression of estrogen receptor α and progesterone receptor, loss of collagen, increase in proliferation and expression of metalloproteases, and invasive epithelium. Surprisingly, by 18 mo of age, the number of hyperplastic foci was reduced, the ducts of the VP and MG became atrophic, and, in the VP, there was massive immune infiltration and massive desquamation of the luminal epithelial cells. These changes were coincident with reduced levels of androgens in males and estrogens in females. We conclude that ERß is a tumor suppressor gene in the VP and MG where its loss increases the activity AR and ERα, respectively.
Asunto(s)
Receptor beta de Estrógeno/genética , Glándulas Mamarias Animales/metabolismo , Fenotipo , Próstata/metabolismo , Receptores Androgénicos/metabolismo , Eliminación de Secuencia , Andrógenos/metabolismo , Animales , Sistemas CRISPR-Cas , Quimiocinas/metabolismo , Proteínas de Unión al ADN/metabolismo , Epitelio/metabolismo , Receptor alfa de Estrógeno/metabolismo , Estrógenos/metabolismo , Proteínas del Ojo , Femenino , Hiperplasia/patología , Inflamación , Antígeno Ki-67/metabolismo , Masculino , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/metabolismo , Próstata/patología , ARN Mensajero/metabolismo , Transducción de Señal , Células del Estroma , Transactivadores , Factores de Transcripción/metabolismo , Transcriptoma , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Liver X receptors (LXRα and LXRß) are oxysterol-activated nuclear receptors that play key roles in cholesterol homeostasis, the central nervous system, and the immune system. We have previously reported that LXRαß-deficient mice are more susceptible to dextran sodium sulfate (DSS)-induced colitis than their WT littermates, and that an LXR agonist protects against colitis in mice mainly via the regulation of the immune system in the gut. We now report that both LXRα and LXRß are expressed in the colonic epithelium and that in aging LXRαß-/- mice there is a reduction in the intensity of goblet cells, mucin (MUC2), TFF3, and estrogen receptor ß (ERß) levels. The cytoplasmic compartment of the surface epithelial cells was markedly reduced and there was a massive invasion of macrophages in the lamina propria. The expression and localization of ß-catenin, α-catenin, and E-cadherin were not changed, but the shrinkage of the cytoplasm led to an appearance of an increase in staining. In the colonic epithelium there was a reduction in the expression of plectin, a hemidesmosome protein whose loss in mice leads to spontaneous colitis, ELOVL1, a fatty acid elongase protein coding gene whose overexpression is found in colorectal cancer, and non-neuronal choline acetyltransferase (ChAT) involved in the regulation of epithelial cell adhesion. We conclude that in aging LXRαß-/- mice, the phenotype in the colon is due to loss of ERß expression.
Asunto(s)
Colitis , Receptor beta de Estrógeno , Ratones , Animales , Receptor beta de Estrógeno/metabolismo , Ratones Noqueados , Colitis/inducido químicamente , Colitis/genética , Colitis/metabolismo , Colon/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Mucosa Intestinal/metabolismo , Sulfato de Dextran/toxicidad , Ratones Endogámicos C57BLRESUMEN
Triple Negative Breast Cancer (TNBC) has the worst prognosis among all breast cancers, and survival in patients with recurrence is rarely beyond 12 months due to acquired resistance to chemotherapy, which is the standard of care for these patients. Our hypothesis is that Estrogen Receptor ß1 (ERß1) increases response to chemotherapy but is opposed by ERß4, which it preferentially dimerizes with. The role of ERß1 and ERß4 in influencing chemotherapy sensitivity has never been studied before. CRISPR/CAS9 was used to truncate ERß1 Ligand Binding Domain (LBD) and knock down the exon unique to ERß4. We show that the truncated ERß1 LBD in a variety of mutant p53 TNBC cell lines, where ERß1 ligand dependent function was inactivated, had increased resistance to Paclitaxel, whereas the ERß4 knockdown cell line was sensitized to Paclitaxel. We further show that ERß1 LBD truncation, as well as treatment with ERß1 antagonist 2-phenyl-3-(4-hydroxyphenyl)-5,7-bis(trifluoromethyl)-pyrazolo[1,5-a] pyrimidine (PHTPP), leads to increase in the drug efflux transporters. Hypoxia Inducible Factors (HIFs) activate factors involved in pluripotency and regulate the stem cell phenotype, both in normal and cancer cells. Here we show that the ERß1 and ERß4 regulate these stem cell markers like SOX2, OCT4, and Nanog in an opposing manner; and we further show that this regulation is mediated by HIFs. We show the increase of cancer cell stemness due to ERß1 LBD truncation is attenuated when HIF1/2α is knocked down by siRNA. Finally, we show an increase in the breast cancer stem cell population due to ERß1 antagonist using both ALDEFLUORTM and SOX2/OCT4 response element (SORE6) reporters in SUM159 and MDA-MB-231 cell lines. Since most TNBC cancers are ERß4 positive, while only a small proportion of TNBC patients are ERß1 positive, we believe that simultaneous activation of ERß1 with agonists and inactivation of ERß4, in combination with paclitaxel, can be more efficacious and yield better outcome for chemotherapy resistant TNBC patients.
Asunto(s)
Neoplasias de la Mama Triple Negativas , Humanos , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismo , Receptores de Estrógenos , Ligandos , Receptor beta de Estrógeno/genética , Receptor beta de Estrógeno/metabolismo , Paclitaxel/farmacología , Paclitaxel/uso terapéutico , Células Madre Neoplásicas/metabolismo , Línea Celular TumoralRESUMEN
Because exposure to bisphenol A (BPA) has been linked to health problems in humans and wildlife, BPA analogues have been synthesized to be considered as replacement molecules. We here have examined estrogenic activity of BPA and five of its analogues, BPAF, BPE, BPC, BPC-Cl, and BPS by a combination of zebrafish-based in vivo and in vitro assays. We used transgenic estrogen reporter (5xERE:GFP) fish to study agonistic effects of bisphenols. Exposures to BPA, BPAF, BPE, and BPC, induced GFP expression in estrogen reporter fish at low exposure concentrations in the heart valves and at higher concentrations in the liver, whereas BPC-Cl activated GFP expression mainly in the liver, and BPS faintly in the heart only. The in vivo response was compared to in vitro estrogenicity of bisphenol exposure using reporter cells that express the zebrafish estrogen receptors driving expression of an estrogen response element (ERE)-luciferase reporter. In these cells, BPA, BPAF, BPC, BPE and BPS preferentially activated Esr1, whereas BPC-Cl preferentially activated Esr2a. By quantitative PCR we found that exposure to BPAF induced expression of the classical estrogen target genes vtg1, esr1, and cyp19a1b in a concentration response manner, but the most responsive target gene was f13a1a. Exposure to BPC-Cl resulted in a different expression pattern of vtg1 and f13a1a with an activation at low concentrations, followed by a declining expression at higher concentrations. Because expression of f13a1a was strongly activated by all compounds tested, we suggest including this mRNA as a biomarker for estrogenicity in larval fish. We further showed that exposure to BPAF and BPC-Cl increased E2 levels in zebrafish larvae, indicating that bisphenol exposures result in a feed-forward response that can further augment the estrogenic activity of these compounds.
Asunto(s)
Receptores de Estrógenos , Pez Cebra , Animales , Humanos , Pez Cebra/genética , Receptores de Estrógenos/genética , Receptores de Estrógenos/metabolismo , Compuestos de Bencidrilo/toxicidad , Estrona , Estrógenos/toxicidad , Estrógenos/metabolismo , Larva/metabolismo , Luciferasas , ARN MensajeroRESUMEN
Astrocytes are integral components of synaptic transmission, and their dysfunction leads to neuropsychiatric disorders such as anxiety and depression. Liver X receptor ß (LXRß) is expressed in astrocytes, and LXRß global knockout mice shows impaired synaptic formation. In order to define the role of LXRß in astrocytes, we used a conditional Cre-loxP system to specifically remove LXRß from astrocytes. We found that this deletion caused anxiety-like but not depressive-like behaviors in adult male mice. This behavioral phenotype could be completely reproduced by selective deletion of LXRß in astrocytes in the medial prefrontal cortex (mPFC). Pyramidal neurons in layer V of mPFC are involved in mood behaviors. We found that there was an increased spontaneous excitatory synaptic transmission in layer V pyramidal neurons of the mPFC of these mice. This was concurrent with increased dendritic complexity, despite normal appearance and number of dendritic spines. In addition, gene ontology analysis of RNA sequencing revealed that deletion of astrocytic LXRß led to the enrichment of the process of synaptic transmission in mPFC. Finally, we also confirmed that renormalized excitatory synaptic transmission in layer V pyramidal neurons alleviated the anxiety in mice with astrocytic LXRß deletion in mPFC. Together, our findings reveal that astrocytic LXRß in mPFC is critical in the regulation of synaptic transmission, and this provides a potential new target for treatment of anxiety-like behavior.
Asunto(s)
Astrocitos , Corteza Prefrontal , Animales , Ansiedad/genética , Astrocitos/fisiología , Receptores X del Hígado/genética , Masculino , Ratones , Ratones Noqueados , Transmisión Sináptica/fisiologíaRESUMEN
BACKGROUND: Depression is one of the most common psychiatric diseases. The monoamine transmitter theory suggests that neurotransmitters are involved in the mechanism of depression; however, the regulation on serotonin production is still unclear. We previously showed that Ahi1 knockout (KO) mice exhibited depression-like behavior accompanied by a significant decrease in brain serotonin. METHODS: In the present study, western blot, gene knockdown, immunofluorescence, dual-luciferase reporter assay, and rescue assay were used to detect changes in the Ahi1/GR/ERß/TPH2 pathway in the brains of male stressed mice and male Ahi1 KO mice to explain the pathogenesis of depression-like behaviors. In addition, E2 levels in the blood and brain of male and female mice were measured to investigate the effect on the ERß/TPH2 pathway and to reveal the mechanisms for the phenomenon of gender differences in depression-like behaviors. RESULTS: We found that the serotonin-producing pathway-the ERß/TPH2 pathway was inhibited in male stressed mice and male Ahi1 KO mice. We further demonstrated that glucocorticoid receptor (GR) as a transcription factor bound to the promoter of ERß that contains glucocorticoid response elements and inhibited the transcription of ERß. Our recent study had indicated that Ahi1 regulates the nuclear translocation of GR upon stress, thus proposing the Ahi1/GR/ERß/TPH2 pathway for serotonin production. Interestingly, female Ahi1 KO mice did not exhibit depressive behaviors, indicating sexual differences in depressive behaviors compared with male mice. Furthermore, we found that serum 17ß-estradiol (E2) level was not changed in male and female mice; however, brain E2 level significantly decreased in male but not female Ahi1 KO mice. Further, ERß agonist LY-500307 increased TPH2 expression and 5-HT production. Therefore, both Ahi1 and E2 regulate the ERß/TPH2 pathway and involve sexual differences in brain serotonin production and depressive behaviors. CONCLUSIONS: In conclusion, although it is unclear how Ahi1 controls E2 secretion in the brain, our findings demonstrate that Ahi1 regulates serotonin production by the GR/ERß/TPH2 pathway in the brain and possibly involves the regulation on sex differences in depressive behaviors. Video Abstract.
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Receptores de Glucocorticoides , Serotonina , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Encéfalo/metabolismo , Receptor beta de Estrógeno/metabolismo , Femenino , Masculino , Ratones , Ratones Noqueados , Receptores de Glucocorticoides/metabolismo , Triptófano Hidroxilasa/genética , Triptófano Hidroxilasa/metabolismoRESUMEN
The retina is an extension of the brain. Like the brain, neurodegeneration of the retina occurs with age and is the cause of several retinal diseases including optic neuritis, macular degeneration, and glaucoma. Liver X receptors (LXRs) are expressed in the brain where they play a key role in maintenance of cerebrospinal fluid and the health of dopaminergic neurons. Herein, we report that LXRs are expressed in the retina and optic nerve and that loss of LXRß, but not LXRα, leads to loss of ganglion cells in the retina. In the retina of LXRß-/- mice, there is an increase in amyloid A4 and deposition of beta-amyloid (Aß) aggregates but no change in the level of apoptosis or autophagy in the ganglion cells and no activation of microglia or astrocytes. However, in the optic nerve there is a loss of aquaporin 4 (AQP4) in astrocytes and an increase in activation of microglia. Since loss of AQP4 and microglial activation in the optic nerve precedes the loss of ganglion cells, and accumulation of Aß in the retina, the cause of the neuronal loss appears to be optic nerve degeneration. In patients with optic neuritis there are frequently AQP4 autoantibodies which block the function of AQP4. LXRß-/- mouse is another model of optic neuritis in which AQP4 antibodies are not detectable, but AQP4 function is lost because of reduction in its expression.
Asunto(s)
Receptores X del Hígado/deficiencia , Degeneración Nerviosa/patología , Nervio Óptico/patología , Retina/patología , Péptidos beta-Amiloides/metabolismo , Animales , Acuaporina 4/metabolismo , Astrocitos/metabolismo , Astrocitos/patología , Femenino , Receptores X del Hígado/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Degeneración Nerviosa/metabolismo , Neuroglía/metabolismo , Neuroglía/patología , Oligodendroglía/metabolismo , Nervio Óptico/metabolismo , Retina/metabolismo , Células Ganglionares de la Retina/metabolismo , Células Ganglionares de la Retina/patologíaRESUMEN
Prostaglandin E2 (PGE2) plays an important role in vascular homeostasis. Its receptor, E-prostanoid receptor 4 (EP4) is essential for physiological remodeling of the ductus arteriosus (DA). However, the role of EP4 in pathological vascular remodeling remains largely unknown. We found that chronic angiotensin II (AngII) infusion of mice with vascular smooth muscle cell (VSMC)-specific EP4 gene knockout (VSMC-EP4-/-) frequently developed aortic dissection (AD) with severe elastic fiber degradation and VSMC dedifferentiation. AngII-infused VSMC-EP4-/- mice also displayed more profound vascular inflammation with increased monocyte chemoattractant protein-1 (MCP-1) expression, macrophage infiltration, matrix metalloproteinase-2 and -9 (MMP2/9) levels, NADPH oxidase 1 (NOX1) activity, and reactive oxygen species production. In addition, VSMC-EP4-/- mice exhibited higher blood pressure under basal and AngII-infused conditions. Ex vivo and in vitro studies further revealed that VSMC-specific EP4 gene deficiency significantly increased AngII-elicited vasoconstriction of the mesenteric artery, likely by stimulating intracellular calcium release in VSMCs. Furthermore, EP4 gene ablation and EP4 blockade in cultured VSMCs were associated with a significant increase in MCP-1 and NOX1 expression and a marked reduction in α-SM actin (α-SMA), SM22α, and SM differentiation marker genes myosin heavy chain (SMMHC) levels and serum response factor (SRF) transcriptional activity. To summarize, the present study demonstrates that VSMC EP4 is critical for vascular homeostasis, and its dysfunction exacerbates AngII-induced pathological vascular remodeling. EP4 may therefore represent a potential therapeutic target for the treatment of AD.
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Angiotensina II/metabolismo , Disección Aórtica/metabolismo , Presión Sanguínea/fisiología , Inflamación/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E , Animales , Aorta/química , Aorta/metabolismo , Aneurisma de la Aorta/metabolismo , Dinoprostona/metabolismo , Femenino , Hipertensión/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Músculo Liso Vascular/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/genética , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Remodelación Vascular/genéticaRESUMEN
The initiation and progression of autoimmune disorders such as multiple sclerosis (MS) is linked to aberrant cholesterol metabolism and overt inflammation. Liver X receptors (LXR) are nuclear receptors that function at the crossroads of cholesterol metabolism and immunity, and their activation is considered a promising therapeutic strategy to attenuate autoimmunity. However, despite clear functional heterogeneity and cell-specific expression profiles, the impact of the individual LXR isoforms on autoimmunity remains poorly understood. Here, we show that LXRα and LXRß have an opposite impact on immune cell function and disease severity in the experimental autoimmune encephalomyelitis model, an experimental MS model. While Lxrα deficiency aggravated disease pathology and severity, absence of Lxrß was protective. Guided by flow cytometry and by using cell-specific knockout models, reduced disease severity in Lxrß-deficient mice was primarily attributed to changes in peripheral T cell physiology and occurred independent from alterations in microglia function. Collectively, our findings indicate that LXR isoforms play functionally non-redundant roles in autoimmunity, potentially having broad implications for the development of LXR-based therapeutic strategies aimed at dampening autoimmunity and neuroinflammation.
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Encefalomielitis Autoinmune Experimental/inmunología , Receptores X del Hígado/metabolismo , Microglía/patología , Esclerosis Múltiple/inmunología , Linfocitos T/inmunología , Animales , Autoinmunidad , Colesterol/metabolismo , Modelos Animales de Enfermedad , Humanos , Receptores X del Hígado/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Inflamación NeurogénicaRESUMEN
Sexually dimorphic bone structure emerges largely during puberty. Sex steroids are critical for peak bone mass acquisition in both genders. In particular, the biphasic effects of estrogens mediate the skeletal sexual dimorphism. However, so far the stimulatory vs inhibitory actions of estrogens on bone mass are not fully explained by direct effects on bone cells. Recently, it has become evident that there is possible neuroendocrine action of estrogen receptor alpha (ERα) on the skeleton. Based on these considerations, we hypothesized that neuronal ERα-signaling may contribute to the skeletal growth during puberty. Here, we generated mice with tamoxifen-inducible Thy1-Cre mediated ERα inactivation during late puberty specifically in extrahypothalamic neurons (N-ERαKO). Inactivation of neuronal ERα did not alter the body weight in males, whereas N-ERαKO females exhibited a higher body weight and increased body and bone length compared to their control littermates at 16 weeks of age. Ex vivo microCT analysis showed increased radial bone expansion of the midshaft femur in female N-ERαKO along with higher serum levels of insulin-like growth factor (IGF)-1 as well as IGF-binding protein (IGFBP)-3. Furthermore, the 3-point bending test revealed increased bone strength in female N-ERαKO. In contrast, inactivation of neuronal ERα had no major effect on bone growth in males. In conclusion, we demonstrate that central ERα-signaling limits longitudinal bone growth and radial bone expansion specifically in females potentially by interacting with the GH/IGF-1 axis.
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Desarrollo Óseo/fisiología , Receptor alfa de Estrógeno/metabolismo , Neuronas/metabolismo , Maduración Sexual/fisiología , Animales , Fenómenos Biomecánicos , Densidad Ósea/genética , Densidad Ósea/fisiología , Desarrollo Óseo/genética , Huesos/anatomía & histología , Huesos/diagnóstico por imagen , Huesos/fisiología , Receptor alfa de Estrógeno/deficiencia , Receptor alfa de Estrógeno/genética , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/genética , ARN Mensajero/metabolismo , Caracteres Sexuales , Maduración Sexual/genética , Transducción de Señal , Microtomografía por Rayos XRESUMEN
OBJECTIVE: Fetal exposure to the anticonvulsant drug valproic acid (VPA), used to treat certain types of epilepsy, increases the risk for birth defects, including neural tube defects, as well as learning difficulties and behavioral problems. Here, we investigated neurotoxic effects of VPA exposure using zebrafish as a model organism. The capacity of folic acid (FA) supplementation to rescue the VPA-induced neuronal and behavioral perturbations was also examined. METHODS: Zebrafish embryos of different transgenic lines with neuronal green fluorescent protein expression were exposed to increasing concentrations of VPA with or without FA supplementation. Fluorescence microscopy was used to visualize alterations in brain structures and neural progenitor cells, as well as motor neurons and neurite sprouting. A twitching behavioral assay was used to examine the functional consequences of VPA and FA treatment. RESULTS: In zebrafish embryos, VPA exposure caused a decrease in the midbrain size, an increase in the midline gap of the hindbrain, and perturbed neurite sprouting of secondary motor neurons, in a concentration-dependent manner. VPA exposure also decreased the fluorescence intensity of neuronal progenitor cells in early developmental stages, indicating fewer cells. Furthermore, VPA exposure significantly altered embryonic twitching activity, causing hyperactivity in dark and hypoactivity in light. Supplementation of FA rescued the VPA-induced smaller midbrain size and hindbrain midline gap defects. FA treatment also increased the number of neuronal progenitor cells in VPA-treated embryos and salvaged neurite sprouting of the secondary motor neurons. FA rescued the VPA-induced alterations in twitching activity in light but not in dark. SIGNIFICANCE: We conclude that VPA exposure induces specific neurotoxic perturbations in developing zebrafish embryos, and that FA reversed most of the identified defects. The results demonstrate that zebrafish is a promising model to study VPA-induced teratogenesis and to screen for countermeasures.
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Anticonvulsivantes/toxicidad , Conducta Animal/efectos de los fármacos , Ácido Fólico/uso terapéutico , Síndromes de Neurotoxicidad/prevención & control , Síndromes de Neurotoxicidad/psicología , Ácido Valproico/toxicidad , Vitaminas/uso terapéutico , Pez Cebra , Animales , Animales Modificados Genéticamente , Suplementos Dietéticos , Desarrollo Embrionario/efectos de los fármacos , Larva , Iluminación , Mesencéfalo/anatomía & histología , Mesencéfalo/efectos de los fármacos , Neuronas Motoras/efectos de los fármacos , Células-Madre Neurales/efectos de los fármacos , Defectos del Tubo Neural/inducido químicamente , Neuritas/efectos de los fármacos , Rombencéfalo/anatomía & histología , Rombencéfalo/efectos de los fármacos , Ácido Valproico/antagonistas & inhibidoresRESUMEN
7α,25-dihydroxycholesterol (7α,25-OHC) is a ligand for the G protein-coupled receptor EBI2; however, the cellular sources of this oxysterol are undefined. 7α,25-OHC is synthesized from cholesterol by the stepwise actions of two enzymes, CH25H and CYP7B1, and is metabolized to a 3-oxo derivative by HSD3B7. We showed that all three enzymes control EBI2 ligand concentration in lymphoid tissues. Lymphoid stromal cells were the main CH25H- and CYP7B1-expressing cells required for positioning of B cells, and they also mediated 7α,25-OHC inactivation. CH25H and CYP7B1 were abundant at the follicle perimeter, whereas CH25H expression by follicular dendritic cells was repressed. CYP7B1, CH25H, and HSD3B7 deficiencies each resulted in defective T cell-dependent plasma cell responses. These findings establish that CYP7B1 and HSD3B7, as well as CH25H, have essential roles in controlling oxysterol production in lymphoid tissues, and they suggest that differential enzyme expression in stromal cell subsets establishes 7α,25-OHC gradients required for B cell responses.
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Linfocitos B/inmunología , Movimiento Celular/inmunología , Hidroxicolesteroles/inmunología , Inmunidad Humoral/inmunología , Células del Estroma/inmunología , 3-Hidroxiesteroide Deshidrogenasas/genética , 3-Hidroxiesteroide Deshidrogenasas/inmunología , 3-Hidroxiesteroide Deshidrogenasas/metabolismo , Animales , Células de la Médula Ósea/inmunología , Células de la Médula Ósea/metabolismo , Células Cultivadas , Familia 7 del Citocromo P450 , Femenino , Citometría de Flujo , Expresión Génica , Células HEK293 , Humanos , Hidroxicolesteroles/metabolismo , Activación de Linfocitos/inmunología , Tejido Linfoide/inmunología , Tejido Linfoide/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/inmunología , Receptores Acoplados a Proteínas G/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Esteroide Hidroxilasas/genética , Esteroide Hidroxilasas/inmunología , Esteroide Hidroxilasas/metabolismo , Células del Estroma/metabolismoRESUMEN
BACKGROUND: Male estrogen receptor beta (ERß) knockout (BERKO) mice display anxiety and aggression linked to, among others, altered serotonergic signaling in the basolateral amygdala and dorsal raphe, impaired cortical radial glia migration, and reduced GABAergic signaling. The effects on primary motor cortex (M1 cortex) and locomotor activity as a consequence of ERß loss have not been investigated. OBJECTIVE: The aim of this study was to determine whether locomotor activity is altered as a consequence of the changes in the M1 cortex. METHODS: The locomotor activity of male wild-type (WT) and BERKO mice was evaluated using the open-field and rotarod tests. Molecular changes in the M1 cortex were analyzed by RNA sequencing, electron microscopy, electrophysiology, and immunohistological techniques. In addition, we established oligodendrocyte (OL) cultures from WT and BERKO mouse embryonic stem cells to evaluate OL function. RESULTS: Locomotor profiling revealed that BERKO mice were more active than WT mice but had impaired motor coordination. Analysis of the M1 cortex pointed out differences in synapse function and myelination. There was a reduction in GABAergic signaling resulting in imbalanced excitatory and inhibitory neurotransmission as well as a defective OL differentiation accompanied by myelin defects. The effects of ERß loss on OL differentiation were confirmed in vitro. CONCLUSION: ERß is an important regulator of GABAergic interneurons and OL differentiation, which impacts on adult M1 cortex function and may be linked to increased locomotor activity and decreased motor coordination in BERKO mice.
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Receptor beta de Estrógeno/genética , Locomoción/genética , Corteza Motora/fisiopatología , Vaina de Mielina/fisiología , Desempeño Psicomotor , Transmisión Sináptica , Animales , Expresión Génica , Masculino , Ratones , Ratones Noqueados , Corteza Motora/metabolismo , Oligodendroglía/patologíaRESUMEN
Hypertonicity in renal medulla is critical for the kidney to produce concentrated urine. Renal medullary cells have to survive high medullary osmolarity during antidiuresis. Previous study reported that farnesoid X receptor (FXR), a nuclear receptor transcription factor activated by endogenous bile acids, increases urine concentrating ability by up-regulating aquaporin 2 expression in medullary collecting duct cells (MCDs). However, whether FXR is also involved in the maintenance of cell survival of MCDs under dehydration condition and hypertonic stress remains largely unknown. In the present study, we demonstrate that 24-hours water restriction selectively up-regulated renal medullary expression of FXR with little MCD apoptosis in wild-type mice. In contrast, water deprivation caused a massive apoptosis of MCDs in both global FXR gene-deficient mice and collecting duct-specific FXR knockout mice. In vitro studies showed that hypertonicity significantly increased FXR and tonicity response enhancer binding protein (TonEBP) expression in mIMCD3 cell line and primary cultured MCDs. Activation and overexpression of FXR markedly increased cell viability and decreased cell apoptosis under hyperosmotic conditions. In addition, FXR can increase gene expression and nuclear translocation of TonEBP. We conclude that FXR protects MCDs from hypertonicity-induced cell injury very likely via increasing TonEBP expression and nuclear translocation. This study provides insights into the molecular mechanism by which FXR enhances urine concentration via maintaining cell viability of MCDs under hyperosmotic condition.