RESUMEN
BACKGROUND: The nucleus incertus (NI) was originally described by Streeter in 1903, as a midline region in the floor of the fourth ventricle of the human brain with an 'unknown' function. More than a century later, the neuroanatomy of the NI has been described in lower vertebrates, but not in humans. Therefore, we examined the neurochemical anatomy of the human NI using markers, including the neuropeptide, relaxin-3 (RLN3), and began to explore the distribution of the NI-related RLN3 innervation of the hippocampus. METHODS: Histochemical staining of serial, coronal sections of control human postmortem pons was conducted to reveal the presence of the NI by detection of immunoreactivity (IR) for the neuronal markers, microtubule-associated protein-2 (MAP2), glutamic acid dehydrogenase (GAD)-65/67 and corticotrophin-releasing hormone receptor 1 (CRHR1), and RLN3, which is highly expressed in NI neurons in diverse species. RLN3 and vesicular GABA transporter 1 (vGAT1) mRNA were detected by fluorescent in situ hybridization. Pons sections containing the NI from an AD case were immunostained for phosphorylated-tau, to explore potential relevance to neurodegenerative diseases. Lastly, sections of the human hippocampus were stained to detect RLN3-IR and somatostatin (SST)-IR. RESULTS: In the dorsal, anterior-medial region of the human pons, neurons containing RLN3- and MAP2-IR, and RLN3/vGAT1 mRNA-positive neurons were observed in an anatomical pattern consistent with that of the NI in other species. GAD65/67- and CRHR1-immunopositive neurons were also detected within this area. Furthermore, RLN3- and AT8-IR were co-localized within NI neurons of an AD subject. Lastly, RLN3-IR was detected in neurons within the CA1, CA2, CA3 and DG areas of the hippocampus, in the absence of RLN3 mRNA. In the DG, RLN3- and SST-IR were co-localized in a small population of neurons. CONCLUSIONS: Aspects of the anatomy of the human NI are shared across species, including a population of stress-responsive, RLN3-expressing neurons and a RLN3 innervation of the hippocampus. Accumulation of phosphorylated-tau in the NI suggests its possible involvement in AD pathology. Further characterization of the neurochemistry of the human NI will increase our understanding of its functional role in health and disease.
Asunto(s)
Puente , Humanos , Puente/metabolismo , Masculino , Hipocampo/química , Hipocampo/metabolismo , Femenino , Relaxina/metabolismo , Relaxina/genética , Anciano , Neuronas/química , Memoria/fisiología , Proteínas Asociadas a Microtúbulos/metabolismo , Persona de Mediana Edad , Anciano de 80 o más Años , Inmunohistoquímica , Hibridación Fluorescente in Situ , Glutamato Descarboxilasa/metabolismo , Glutamato Descarboxilasa/genética , Receptores de Hormona Liberadora de CorticotropinaRESUMEN
Healthy aging results in cardiac structural and electrical remodeling that increase susceptibility to cardiovascular diseases. Relaxin has shown broad cardioprotective effects including anti-fibrotic, anti-arrhythmic and anti-inflammatory outcomes in multiple models. This paper focuses on the cardioprotective effects of Relaxin in a rat model of aging. Sustained atrial or ventricular fibrillation are readily induced in the hearts of aged but not young control animals. Treatment with Relaxin suppressed this arrhythmogenic response by increasing conduction velocity, decreasing fibrosis and promoting substantial cardiac remodeling. Relaxin treatment resulted in a significant increase in the levels of: Nav1.5, Cx43, ßcatenin and Wnt1 in rat hearts. In isolated cardiomyocytes, Relaxin increased Nav1.5 expression. These effects were mimicked by CHIR 99021, a pharmacological activator of canonical Wnt signaling, but blocked by the canonical Wnt inhibitor Dickkopf1. Relaxin prevented TGF-ß-dependent differentiation of cardiac fibroblasts into myofibroblasts while increasing the expression of Wnt1; the effects of Relaxin on cardiac fibroblast differentiation were blocked by Dickkopf1. RNASeq studies demonstrated reduced expression of pro-inflammatory cytokines and an increase in the expression of α- and ß-globin in Relaxin-treated aged males. Relaxin reduces arrhythmogenicity in the hearts of aged rats by reduction of fibrosis and increased conduction velocity. These changes are accompanied by substantial remodeling of the cardiac tissue and appear to be mediated by increased canonical Wnt signaling. Relaxin also exerts significant anti-inflammatory and anti-oxidant effects in the hearts of aged rodents. The mechanisms by which Relaxin increases the expression of Wnt ligands, promotes Wnt signaling and reprograms gene expression remain to be determined.
Asunto(s)
Envejecimiento , Fibrilación Atrial , Fibrosis , Relaxina , Animales , Relaxina/farmacología , Masculino , Ratas , Fibrilación Atrial/metabolismo , Fibrilación Atrial/tratamiento farmacológico , Fibrilación Atrial/prevención & control , Envejecimiento/efectos de los fármacos , Inflamación/metabolismo , Inflamación/tratamiento farmacológico , Ratas Endogámicas F344RESUMEN
OBJECTIVE: To explore the effect and mechanism of relaxin (RLX) in the growth and metastasis of livercancer after combination treatment with transarterial chemoembolization (TACE). MATERIALS AND METHODS: HCCLM3 and Huh-7 cells were adopted to evaluate the effect of tumor proliferation, migration, and invasion after RLX administration in vitro. The rabbit VX2 model was used to evaluate the biosafety, doxorubicin penetration, local tumor response, tumor metastasis, and survival benefit of RLX combined with TACE treatment. RESULTS: RLX did not affect the proliferation, migration, or invasion of HCCLM3 and Huh-7 cells, and the expression of E-cadherin and HIF-1α also remained unchanged while the MMP-9 protein was upregulated in vitro. In the rabbit VX2 model, compared to the normal saline group (NS), RLX group (RLX) and TACE mono-therapy group (TACE), the group that received TACE combined with RLX (TACE + RLX) showed an improved local tumor response and survival benefit. Furthermore, TACE combined with RLX was found to reduce tumor metastasis. This combination therapy reduced the fibrotic extracellular matrix in the tumor microenvironment, allowing for better penetration of doxorubicin, improved infiltration of CD8+ T cells and affected the secretion of cytokines. Additionally, RLX combined with TACE was able to decrease the expression of HIF-1α and PD-L1. The biosafety of TACE combined with RLX was also confirmed. CONCLUSION: RLX synergized with TACE by mitigating the fibrotic extracellular matrix and tumor hypoxic microenvironment, improving the therapeutic effect and inhibiting metastasis during the treatment of liver cancer.
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Quimioembolización Terapéutica , Neoplasias Hepáticas , Relaxina , Animales , Quimioembolización Terapéutica/métodos , Conejos , Relaxina/administración & dosificación , Neoplasias Hepáticas/secundario , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/tratamiento farmacológico , Doxorrubicina/administración & dosificación , Humanos , Terapia Combinada , Proliferación Celular/efectos de los fármacos , Línea Celular Tumoral , Modelos Animales de Enfermedad , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/tratamiento farmacológico , Metástasis de la NeoplasiaRESUMEN
BACKGROUND: Heart failure mortality remains high despite recent progress in pharmacological treatment. AZD3427 is a selective long-acting analog of relaxin, a vasodilatory hormone with antifibrotic effects. We assessed the safety, pharmacokinetics, and pharmacodynamics of AZD3427 in healthy volunteers and patients with heart failure on standard-of-care therapy. METHODS AND RESULTS: In this first-in-human, phase 1a/b, randomized, single-blind, placebo-controlled study, healthy volunteers were randomized 6:2 to receive a single dose of AZD3427 or placebo by subcutaneous injection in 5 mixed-ethnicity cohorts (5, 10, 30, 90, or 270 mg) and 1 Japanese-descent cohort (270 mg), or by intravenous injection in 1 cohort (15 mg). After confirming safety and tolerability in healthy volunteers, 3 cohorts of patients with heart failure and left ventricular ejection fraction ≤40% and 3 cohorts with ejection fraction ≥41% were randomized 6:2 to receive 5 weekly doses of AZD3427 (5, 15, or 45 mg) or placebo by subcutaneous injection. In total, 56 healthy volunteers and 48 patients with heart failure were randomized. AZD3427 was well tolerated at all doses. After subcutaneous administration, AZD3427 was absorbed slowly, and exposure was approximately linear across the dose range. In patients with heart failure, AZD3427 terminal half-life was 13 to 14 days and there were numerical increases in stroke volume and estimated glomerular filtration rate. No treatment-emergent antidrug antibodies were detected. CONCLUSIONS: AZD3427 had favorable safety and pharmacokinetic profiles. Hemodynamic changes in patients with heart failure were consistent with the anticipated effects of a relaxin analog. These findings support further development of AZD3427 as a novel long-term treatment for patients with heart failure. REGISTRATION: URL: https://www.clinicaltrials.gov; Unique Identifier: NCT04630067.
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Insuficiencia Cardíaca , Receptores Acoplados a Proteínas G , Humanos , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/fisiopatología , Masculino , Femenino , Método Simple Ciego , Persona de Mediana Edad , Receptores Acoplados a Proteínas G/agonistas , Adulto , Anciano , Resultado del Tratamiento , Receptores de Péptidos/agonistas , Volumen Sistólico/efectos de los fármacos , Inyecciones Subcutáneas , Función Ventricular Izquierda/efectos de los fármacos , Adulto Joven , Relaxina/farmacocinética , Relaxina/administración & dosificación , Relaxina/efectos adversos , Relaxina/uso terapéutico , Relación Dosis-Respuesta a Droga , Inyecciones IntravenosasRESUMEN
The current literature suggests that relaxin-3/relaxin/insulin-like family peptide receptor 3 (RLN-3/RXFP-3) system is involved in the pathophysiology of affective disorders because the results of anatomical and pharmacological studies have shown that the RLN-3 signaling pathway plays a role in modulating the stress response, anxiety, arousal, depression-like behavior, and neuroendocrine homeostasis. The risk of developing mental illnesses in adulthood is increased by exposure to stress in early periods of life. The available data indicate that puberty is especially characterized by the development of the neural system and emotionality and is a "stress-sensitive" period. The presented study assessed the short-term changes in the expression of RLN-3 and RXFP-3 mRNA in the stress-dependent brain regions in male pubertal Wistar rats that had been subjected to acute stress. Three stressors were applied from 42 to 44 postnatal days (first day: a single forced swim; second day: stress on an elevated platform that was repeated three times; third day: restraint stress three times). Anxiety (open field, elevated plus maze test) and anhedonic-like behavior (sucrose preference test) were estimated during these tests. The corticosterone (CORT) levels and blood morphology were estimated. We found that the RXFP-3 mRNA expression decreased in the brainstem, whereas it increased in the hypothalamus 72 h after acute stress. These molecular changes were accompanied by the increased levels of CORT and anxiety-like behavior detected in the open field test that had been conducted earlier, that is, 24 h after the stress procedure. These findings shed new light on the neurochemical changes that are involved in the compensatory response to adverse events in pubertal male rats and support other data that suggest a regulatory interplay between the RLN-3 pathway and the hypothalamus-pituitary-adrenal axis activity in the mechanisms of anxiety-like behavior.
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Ansiedad , Encéfalo , ARN Mensajero , Ratas Wistar , Receptores Acoplados a Proteínas G , Estrés Psicológico , Animales , Masculino , Ratas , Estrés Psicológico/metabolismo , Estrés Psicológico/fisiopatología , Ansiedad/metabolismo , Ansiedad/fisiopatología , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Encéfalo/metabolismo , ARN Mensajero/metabolismo , Conducta Animal/fisiología , Relaxina/metabolismo , Relaxina/genética , Receptores de Péptidos/metabolismo , Receptores de Péptidos/genética , Maduración Sexual/fisiología , Proteínas del Tejido NerviosoRESUMEN
Heart failure (HF) is a complex, progressive disorder that is associated with substantial morbidity and mortality on a global scale. Relaxin-2 is a naturally occurring hormone that may have potential therapeutic benefit for patients with HF. To investigate the therapeutic potential of relaxin in the treatment of patients with HF, mRNA-0184, a novel, investigational, lipid nanoparticle (LNP)-encapsulated mRNA therapy that encodes for human relaxin-2 fused to variable light chain kappa (Rel2-vlk) was developed. A translational semi-mechanistic population pharmacokinetic (PK)/pharmacodynamic (PD) model was developed using data from non-human primates at dose levels ranging from 0.15 to 1 mg/kg. The PK/PD model was able to describe the PK of Rel2-vlk mRNA and translated Rel2-vlk protein in non-human primates adequately with relatively precise estimates. The preclinical PK/PD model was then scaled allometrically to determine the human mRNA-0184 dose that would achieve therapeutic levels of Rel2-vlk protein expression in patients with stable HF with reduced ejection fraction. Model-based simulations derived from the scaled PK/PD model support the selection of 0.025 mg/kg as an appropriate starting human dose of mRNA-0184 to achieve average trough relaxin levels between 1 and 2.5 ng/mL, which is the potential exposure for cardioprotective action of relaxin.
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Insuficiencia Cardíaca , ARN Mensajero , Relaxina , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/genética , Animales , Humanos , Relaxina/farmacocinética , Relaxina/farmacología , Relaxina/administración & dosificación , Relaxina/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Nanopartículas/química , Investigación Biomédica Traslacional , Modelos Biológicos , Masculino , Proteínas Recombinantes de Fusión/farmacocinética , Proteínas Recombinantes de Fusión/administración & dosificación , Proteínas Recombinantes de Fusión/farmacología , Lípidos/química , LiposomasRESUMEN
The peptide hormone relaxin plays a critical role in tissue remodeling in a variety of tissues through activation of its cognate receptor, RXFP1. Relaxin's ability to modify extracellular matrices has provided a strong rationale for treating fibrosis in a variety of tissues. Treatment with recombinant relaxin peptides in clinical studies of heart failure has not yet proven useful, likely due to the short half-life of infused peptide. To circumvent this particular pharmacokinetic pitfall we have used a Protein-in-Protein (PiP) antibody technology described previously, to insert a single-chain human relaxin construct into the complementarity-determining region (CDR) of an immunoglobulin G (IgG) backbone, creating a relaxin molecule with a half-life of â¼4-5 days in mice. Relaxin-PiP biologics displaced Europium-labeled human relaxin in RXFP1-expressing cells and demonstrated full agonist activity on both human and mouse RXFP1 receptors. Relaxin-PiPs did not show signal transduction bias, as they activated cAMP in THP-1 cells, and cGMP and pERK signaling in primary human cardiac fibroblasts. In an induced carbon tetrachloride mouse model of liver fibrosis one relaxin-PiP, R2-PiP, caused reduction of liver lesions, ameliorated collagen accumulation in the liver with the corresponding reduction of Collagen1a1 gene expression, and increased cell proliferation in hepatic parenchyma. These relaxin biologics represent a novel approach to the design of a long-acting RXFP1 agonist to probe the clinical utility of relaxin/RXFP1 signaling to treat a variety of human fibrotic diseases.
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Receptores Acoplados a Proteínas G , Receptores de Péptidos , Relaxina , Relaxina/farmacología , Animales , Humanos , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/metabolismo , Ratones , Receptores de Péptidos/agonistas , Receptores de Péptidos/metabolismo , Receptores de Péptidos/genética , Ingeniería de Proteínas/métodos , Masculino , Ratones Endogámicos C57BL , Cricetulus , Células HEK293 , Células CHORESUMEN
Background: Female athletes are at increased risk for anterior cruciate ligament (ACL) injuries. The influence of hormonal variation on female ACL injury risk remains ill-defined. Recent data suggests that the collagen-degrading menstrual hormone relaxin may cyclically impact female ACL tissue quality. This review aims to identify any correlation between menstrual relaxin peaks and rates of female ACL injury. Methods: A systematic review was performed, utilizing the MEDLINE, EMBASE, and CINAHL databases. Included studies had to directly address relaxin/female ACL interactions. The primary outcome variable was relaxin proteolysis of the ACL, at cellular, tissue, joint, and whole-organism levels. The secondary outcome variable was any discussed method of moderating relaxin levels, and the clinical results if available. Results: AllThe numerous relaxin receptors on female ACLs upregulate local collagenolysis and suppress local collagen production. Peak serum relaxin concentrations (SRC) occur during menstrual cycle days 21-24; a time phase associated with greater risk of ACL injury. Oral contraceptives (OCPs) reduce SRC, with a potential ACLprotective effect. Conclusion: A reasonable correlative and plausible causative relationship exists between peak relaxin levels and increased risk of ACL injury in females, and further investigation is warranted. Level of Evidence: III.
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Lesiones del Ligamento Cruzado Anterior , Ciclo Menstrual , Relaxina , Humanos , Relaxina/sangre , Relaxina/metabolismo , Femenino , Ciclo Menstrual/fisiología , Traumatismos en Atletas , AtletasRESUMEN
Relaxin3 (rln3) has been associated with various emotional and cognitive processes, including stress, anxiety, learning, memory, motivational behavior, and circadian rhythm. Notably, previous report revealed that Rln3a played an indispensable role in testicular development and male fertility in Nile tilapia (Oreochromis niloticus). However, the underlying molecular mechanisms remain largely unknown. We found that Rln3a is expressed exclusively in the diencephalon* (Di*) of the brain. Deficiency of Rln3a resulted in a significant increase in serum dopamine level and an upregulation of gene expression of gnrh1 and kisspeptin2. To further elucidate the role of Rln3a in fish fertility, we collected two different regions of Di* and hypothalamus (Hyp) tissues for subsequent RNA-seq analysis of both wild-type (rln3a+/+) and rln3a-/- male tilapia. Upon the transcriptomic data, 1136 and 755 differentially expressed genes (DEGs) were identified in the Di* and Hyp tissues, respectively. In Di*, the up-regulated genes were enriched in circadian rhythm, chemical carcinogenesis, while the down-regulated genes were enriched in type II diabetes mellitus, dopaminergic synapse, and other pathways. In Hyp, the up-regulated genes were enriched in circadian rhythm, pyrimidine metabolism, while the down-regulated genes were enriched in type I diabetes mellitus, autoimmune thyroid disease, and other pathways. Subsequently, the results of both qRT-PCR and FISH assays highlighted a pronounced up-regulation of core circadian rhythm genes, cry1b and per3, whereas genes such as clocka, clockb, and arntl exhibited down-regulation. Furthermore, the genes associated with dopamine biosynthesis were significantly increased in the Hyp. In summary, the mutation of rln3a in male tilapia resulted in notable changes in circadian rhythm and disease-linked signaling pathways in the Di* and Hyp. These changes might account for the fertility defects observed in rln3a-/- male mutants in tilapia.
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Encéfalo , Cíclidos , Fertilidad , Animales , Masculino , Cíclidos/genética , Cíclidos/metabolismo , Encéfalo/metabolismo , Fertilidad/genética , Relaxina/genética , Relaxina/metabolismo , Proteínas de Peces/genética , Proteínas de Peces/metabolismoRESUMEN
Relaxin's role in differentiated thyroid cancer (DTC) has been suggested but its characterization in a large clinical sample remains limited. We performed immunohistochemistry for relaxin-2 (RLN2), CD68 (total macrophages), CD163 (M2 macrophages) on tissue microarrays from 181 subjects with non-distant metastatic DTC, and 185 subjects with benign thyroid tissue. Mean pixels/area for each marker was compared between tumor and adjacent tissue via paired-t test and between DTC and benign subjects via t-test assuming unequal variances. RNA qPCR was performed for expression of RLN2, RLN1, and RXFP1 in cell lines. Amongst 181 cases, the mean age was 46 years, 75 % were females. Tumoral tissue amongst the DTC cases demonstrated higher mean expression of RLN2 (53.04 vs. 9.79; p < 0.0001) compared to tumor-adjacent tissue. DTC tissue also demonstrated higher mean expression of CD68 (14.46 vs. 4.79; p < 0.0001), and CD163 (23.13 vs. -0.73; p < 0.0001) than benign thyroid. These markers did not differ between tumor-adjacent and benign thyroid tissue groups; and amongst cases, did not differ by demographic or clinicopathologic features. RLN1 and RXFP1 expression was detected in a minority of the cell lines, while RLN2 was expressed by 6/7 cell lines. In conclusion, widespread RLN2 expression in DTC tissue and most cell lines demonstrates that RLN2 acts in a paracrine manner, and that RLN1 and RXFP1 are probably not involved in thyroid cancer cell signaling. RLN2 is a biomarker for thyroid carcinogenesis, being associated with but not secreted by immunosuppressive macrophages. These findings will guide further investigations for therapeutic avenues against thyroid cancer.
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Biomarcadores de Tumor , Relaxina , Neoplasias de la Tiroides , Humanos , Relaxina/metabolismo , Relaxina/genética , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/metabolismo , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/diagnóstico , Femenino , Persona de Mediana Edad , Masculino , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Adulto , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Línea Celular Tumoral , Antígenos CD/genética , Antígenos CD/metabolismo , Anciano , Receptores de Péptidos/metabolismo , Receptores de Péptidos/genética , Antígenos de Diferenciación Mielomonocítica/metabolismo , Antígenos de Diferenciación Mielomonocítica/genéticaRESUMEN
The neuropeptide relaxin-3 and its cognate receptor, relaxin family peptide-3 receptors (RXFP3), have been implicated in modulating learning and memory processes, but their specific roles remain unclear. This study utilized behavioral and molecular approaches to investigate the effects of putatively reversible blockade of RXFP3 in the ventral dentate gyrus (vDG) of the hippocampus on spatial and fear memory formation in rats. Male Wistar rats received bilateral vDG cannula implantation and injections of the RXFP3 antagonist, R3(BΔ23-27)R/I5 (400â¯ng/0.5⯵L per side), or vehicle at specific time points before acquisition, consolidation, or retrieval phases of the Morris water maze and passive avoidance learning tasks. RXFP3 inhibition impaired acquisition in the passive avoidance task but not the spatial learning task. However, both memory consolidation and retrieval were disrupted in both tasks following RXFP3 antagonism. Ventral hippocampal levels of the consolidation-related kinase p70-S6 kinase (p70S6K) were reduced RXFP3 blockade. These findings highlight a key role for ventral hippocampal RXFP3 signaling in the acquisition, consolidation, and retrieval of spatial and emotional memories, extending previous work implicating this neuropeptide system in hippocampal memory processing.
Asunto(s)
Giro Dentado , Miedo , Ratas Wistar , Receptores Acoplados a Proteínas G , Animales , Giro Dentado/metabolismo , Ratas , Receptores Acoplados a Proteínas G/metabolismo , Masculino , Miedo/fisiología , Reacción de Prevención/fisiología , Reacción de Prevención/efectos de los fármacos , Memoria/fisiología , Relaxina/metabolismo , Memoria Espacial/fisiología , Memoria Espacial/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Aprendizaje por Laberinto/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/efectos de los fármacos , Receptores de Péptidos/metabolismoRESUMEN
The retrosplenial cortex (RSC) plays a central role in processing contextual fear conditioning. In addition to corticocortical and thalamocortical projections, the RSC receives subcortical inputs, including a substantial projection from the nucleus incertus in the pontine tegmentum. This GABAergic projection contains the neuropeptide, relaxin-3 (RLN3), which inhibits target neurons via its Gi/o-protein-coupled receptor, RXFP3. To assess this peptidergic system role in contextual fear conditioning, we bilaterally injected the RSC of adult rats with an adeno-associated-virus (AAV), expressing the chimeric RXFP3 agonist R3/I5 or a control AAV, and subjected them to contextual fear conditioning. The R3/I5 injected rats did not display any major differences to control-injected and naïve rats but displayed a significantly delayed extinction. Subsequently, we employed acute bilateral injections of the specific RXFP3 agonist peptide, RXFP3-Analogue 2 (A2), into RSC. While the administration of A2 before each extinction trial had no impact on the extinction process, treatment with A2 before each acquisition trial resulted in delayed extinction. In related anatomical studies, we detected an enrichment of RLN3-immunoreactive nerve fibers in deep layers of the RSC, and a higher level of co-localization of RXFP3 mRNA with vesicular GABA transporter (vGAT) mRNA than with vesicular glutamate transporter-1 (vGLUT1) mRNA across the RSC, consistent with an effect of RLN3/RXFP3 signalling on the intrinsic, inhibitory circuits within the RSC. These findings suggest that contextual conditioning processes in the RSC involve, in part, RLN3 afferent modulation of local inhibitory neurons that provides a stronger memory acquisition which, in turn, retards the extinction process.
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Extinción Psicológica , Miedo , Receptores Acoplados a Proteínas G , Animales , Masculino , Miedo/fisiología , Miedo/efectos de los fármacos , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Ratas , Extinción Psicológica/fisiología , Extinción Psicológica/efectos de los fármacos , Relaxina/metabolismo , Corteza Cerebral/metabolismo , Corteza Cerebral/efectos de los fármacos , Giro del Cíngulo/metabolismo , Giro del Cíngulo/efectos de los fármacos , Giro del Cíngulo/fisiología , Receptores de PéptidosRESUMEN
Heart failure (HF) prevalence is rising due to reduced early mortality and demographic change. Relaxin (RLN) mediates protective effects in the cardiovascular system through Relaxin-receptor 1 (RXFP1). Cardiac overexpression of RXFP1 with additional RLN supplementation attenuated HF in the pressure-overload transverse aortic constriction (TAC) model. Here, we hypothesized that robust transgenic RXFP1 overexpression in cardiomyocytes (CM) protects from TAC-induced HF even in the absence of RLN. Hence, transgenic mice with a CM-specific overexpression of human RXFP1 (hRXFP1tg) were generated. Receptor functionality was demonstrated by in vivo hemodynamics, where the administration of RLN induced positive inotropy strictly in hRXFP1tg. An increase in phospholamban-phosphorylation at serine 16 was identified as a molecular correlate. hRXFP1tg were protected from TAC without additional RLN administration, presenting not only less decline in systolic left ventricular (LV) function but also abrogated LV dilation and pulmonary congestion compared to WT mice. Molecularly, transgenic hearts exhibited not only a significantly attenuated fetal and fibrotic gene activation but also demonstrated less fibrotic tissue and CM hypertrophy in histological sections. These protective effects were evident in both sexes. Similar cardioprotective effects of hRXFP1tg were detectable in a RLN-knockout model, suggesting an alternative mechanism of receptor activation through intrinsic activity, alternative endogenous ligands or crosstalk with other receptors. In summary, CM-specific RXFP1 overexpression provides protection against TAC even in the absence of endogenous RLN. This suggests RXFP1 overexpression as a potential therapeutic approach for HF, offering baseline protection with optional RLN supplementation for specific activation.
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Miocitos Cardíacos , Receptores Acoplados a Proteínas G , Receptores de Péptidos , Relaxina , Animales , Humanos , Masculino , Ratones , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/prevención & control , Insuficiencia Cardíaca/genética , Ratones Endogámicos C57BL , Ratones Transgénicos , Miocitos Cardíacos/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Péptidos/genética , Receptores de Péptidos/metabolismo , Relaxina/genética , Relaxina/metabolismoRESUMEN
Fibrotic changes in musculoskeletal diseases arise from the abnormal buildup of fibrotic tissue around the joints, leading to limited mobility, compromised joint function, and diminished quality of life. Relaxin (RLX) attenuates fibrosis by accelerating collagen degradation and inhibiting excessive extracellular matrix (ECM) production. Further, RLX disrupts myofibroblast activation by modulating the TGF-ß/Smads signaling pathways, which reduces connective tissue fibrosis. However, the mechanisms and effects of RLX in musculoskeletal pathologies are emerging as increasing research focuses on relaxin's impact on skin, ligaments, tendons, cartilage, joint capsules, connective tissues, and muscles. This review delineates the actions of relaxin within the musculoskeletal system and the challenges to its clinical application. Relaxin shows significant potential in both in vivo and in vitro studies for broadly managing musculoskeletal fibrosis; however, challenges such as short biological half-life and sex-specific responses may pose hurdles for clinical use.
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Fibrosis , Relaxina , Relaxina/uso terapéutico , Relaxina/metabolismo , Humanos , Fibrosis/tratamiento farmacológico , Animales , Enfermedades Musculoesqueléticas/tratamiento farmacológico , Enfermedades Musculoesqueléticas/metabolismoRESUMEN
Relaxin 3 is a neuropeptide that plays a crucial role in reproductive functions of mammals. Previous studies have confirmed that rln3a plays an important role in the male reproduction of tilapia. To further understand the significance of its paralogous gene rln3b in male fertility, we generated a homozygous mutant line of rln3b in Nile tilapia. Our findings indicated that rln3b mutation delayed spermatogenesis and led to abnormal testes structure. Knocking out rln3b gene resulted in a decrease in sperm count, sperm motility and male fish fertility. TUNEL detection revealed a small amount of apoptosis in the testes of rln3b-/- male fish at 390 days after hatching (dah). RT-qPCR analysis demonstrated that mutation of rln3b gene caused a significant downregulation of steroid synthesis-related genes such as cyp17a1, cyp11b2, germ cell marker gene, Vasa, and gonadal somatic cell marker genes of amh and amhr2. Furthermore, we found a significant down-regulation of hypothalamic-pituitary-gonadal (HPG) axis-related genes, while a significantly up-regulation of the dopamine synthetase gene in the rln3b-/- male fish. Taken together, our data strongly suggested that Rln3b played a crucial role in the fertility of XY tilapia by regulating HPG axis genes.
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Hipogonadismo , Espermatogénesis , Testículo , Tilapia , Animales , Masculino , Tilapia/genética , Hipogonadismo/genética , Espermatogénesis/genética , Testículo/metabolismo , Relaxina/genética , Relaxina/metabolismo , Fertilidad/genética , Motilidad Espermática/genética , Mutación , Proteínas de Peces/genética , Proteínas de Peces/metabolismoRESUMEN
The human relaxins belong to the Insulin/IGF/Relaxin superfamily of peptide hormones, and their physiological function is primarily associated with reproduction. In this study, we focused on a prostate tissue-specific relaxin RLN1 (REL1_HUMAN protein) and a broader tissue specificity RLN2 (REL2_HUMAN protein). Due to their structural similarity, REL1 and REL2 proteins were collectively named a 'human relaxin protein' in previous studies and were exclusively measured by immunoassays. We hypothesized that the highly selective and sensitive immunoaffinity-selected reaction monitoring (IA-SRM) assays would reveal the identity and abundance of the endogenous REL1 and REL2 in biological samples and facilitate the evaluation of these proteins for diagnostic applications. High levels of RLN1 and RLN2 transcripts were found in prostate and breast cancer cell lines by RT-PCR. However, no endogenous prorelaxin-1 or mature REL1 were detected by IA-SRM in cell lines, seminal plasma, or blood serum. The IA-SRM assay of REL2 demonstrated its undetectable levels (<9.4 pg/mL) in healthy control female and male sera and relatively high levels of REL2 in maternal sera across different gestational weeks (median 331 pg/mL; N = 120). IA-SRM assays uncovered potential cross-reactivity and nonspecific binding for relaxin immunoassays. The developed IA-SRM assays will facilitate the investigation of the physiological and pathological roles of REL1 and REL2 proteins.
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Relaxina , Humanos , Relaxina/metabolismo , Relaxina/genética , Masculino , Femenino , Línea Celular Tumoral , Inmunoensayo/métodos , Espectrometría de Masas/métodos , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/diagnóstico , Semen/química , Semen/metabolismoRESUMEN
The nucleus incertus (NI), a conserved hindbrain structure implicated in the stress response, arousal, and memory, is a major site for production of the neuropeptide relaxin-3. On the basis of goosecoid homeobox 2 (gsc2) expression, we identified a neuronal cluster that lies adjacent to relaxin 3a (rln3a) neurons in the zebrafish analogue of the NI. To delineate the characteristics of the gsc2 and rln3a NI neurons, we used CRISPR/Cas9 targeted integration to drive gene expression specifically in each neuronal group, and found that they differ in their efferent and afferent connectivity, spontaneous activity, and functional properties. gsc2 and rln3a NI neurons have widely divergent projection patterns and innervate distinct subregions of the midbrain interpeduncular nucleus (IPN). Whereas gsc2 neurons are activated more robustly by electric shock, rln3a neurons exhibit spontaneous fluctuations in calcium signaling and regulate locomotor activity. Our findings define heterogeneous neurons in the NI and provide new tools to probe its diverse functions.
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Neuronas , Pez Cebra , Animales , Neuronas/fisiología , Neuronas/metabolismo , Relaxina/metabolismo , Relaxina/genética , Proteínas de Pez Cebra/metabolismo , Proteínas de Pez Cebra/genética , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Sistemas CRISPR-Cas , Rombencéfalo/fisiología , Rombencéfalo/metabolismoRESUMEN
INTRODUCTION: The neuropeptide relaxin-3/RXFP3 system belongs to the relaxin/insulin superfamily and is involved in many important physiological processes, such as stress responses, appetite control, and motivation for reward. Although relaxin-3 is the endogenous agonist for RXFP3, it can also bind to and activate RXFP1 and RXFP4. Consequently, research has been focused on the development of RXFP3-specific peptides and small-molecule ligands to validate the relaxin-3/RXFP3 system as a novel drug target. AREAS COVERED: This review provides an overview of patents on the relaxin-3/RXFP3 system covering ligand development and pharmacological studies since 2003. Related patents and literature reports were obtained from established sources including SciFinder, Google Patents, and Espacenet for patents and SciFinder, PubMed, and Google Scholar for literature reports. EXPERT OPINION: There has been an increasing amount of patent activities around relaxin-3/RXFP3, highlighting the importance of this novel neuropeptide system for drug discovery. The development of relaxin-3 derived peptides and small-molecule modulators, as well as behavioral studies in rodents, have shown that the relaxin-3/RXFP3 system is a promising drug target for treating various metabolic and neuropsychiatric diseases including obesity, anxiety, and alcohol addiction.
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Neuropéptidos , Relaxina , Humanos , Receptores Acoplados a Proteínas G/metabolismo , Relaxina/metabolismo , Patentes como Asunto , Insulina/metabolismo , Receptores de Péptidos/agonistas , Receptores de Péptidos/metabolismoRESUMEN
Deregulation of the relaxin family peptide system (RFPS) appears to increase the risk of range of cancers, including epithelial ovarian cancers (EOC). The present study examines the effect of relaxin family peptide receptor 1 (RXFP1) level on the biological properties of human epithelial ovarian adenocarcinoma cells (OVCAR4 and SKOV3). RXFP1 was downregulated (RXFP1↓) in the cells using the RXFP1 sgRNA CRISPR All-in-One Lentivirus set (pLenti-U6-sgRNA-SFFV-Cas9-2A-Puro), and upregulated (RXFP1↑) using the RXFP1 CRISPRa sgRNA Lentivector (pLenti-U6-sgRNA-PGK-Neo) kit, which activates the RXFP1 gene when paired with dCas9-SAM. The changes taking place during adhesion to extracellular matrix (ECM) proteins were assessed in multi-well plates coated with collagen, fibronectin, laminin and gelatin. Cellular viability was monitored based on mitochondrial metabolic activity (MTT Assay, Alamar Blue Assay) and adenosine triphosphate production (ATP Assay). The rate of cell proliferation was determined based on the percentage of Ki67 immunoreactive cells and the numbers of cells in particular cell-cycle phases. The mesenchymal-like (Boyden Chamber Assay) and amoeboid-like movements (Wound Healing Assay) of ovarian cancer cells were also analyzed after transfection. RXFP1 downregulation decreased the adhesion properties of ovarian cancer cells and increased the tendency for apoptosis under stressful conditions. In contrast, RXFP1 upregulation had pro-proliferative, pro-survival and promigratory effects. Our findings confirm that the relaxin-2/RXFP1 signaling pathway plays a role in the promotion of growth and progression of ovarian cancer.
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Proliferación Celular , Neoplasias Ováricas , Receptores Acoplados a Proteínas G , Humanos , Femenino , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Neoplasias Ováricas/metabolismo , Línea Celular Tumoral , Receptores de Péptidos/metabolismo , Receptores de Péptidos/genética , Carcinoma Epitelial de Ovario/metabolismo , Carcinoma Epitelial de Ovario/genética , Regulación Neoplásica de la Expresión Génica , Movimiento Celular , Relaxina/metabolismo , Adhesión CelularRESUMEN
PURPOSE: High-grade serous ovarian cancer (HGSC) is the most common ovarian cancer subtype. Parity is an important risk-reducing factor, but the underlying mechanism behind the protective effect is unclear. Our aim was to study if the expression of hormones and proteins involved in pregnancy were affected by the woman's parity status, and if they may be associated with tumor stage and survival. METHODS: We evaluated expression of progesterone receptor (PR), progesterone receptor membrane component 1 (PGRMC1), relaxin-2, and transforming growth factor beta 1 (TGFß1) in tumor tissue from 92 women with HGSC parous (n = 73) and nulliparous (n = 19). Key findings were then evaluated in an independent expansion cohort of 49 patients. Survival rates by hormone/protein expression were illustrated using the Kaplan-Meier method. The independent prognostic value was tested by Cox regression, using models adjusted for established poor-prognostic factors (age at diagnosis, FIGO stage, type of surgery, and macroscopic residual tumor after surgery). RESULTS: HGSC tumors from parous women were PR positive (≥ 1% PR expression in tumor cells) more often than tumors from nulliparous women (42% vs. 16%; p-value 0.04), and having more children was associated with developing PR positive tumors [i.e., ≥ 3 children versus nulliparity, adjusted for age at diagnosis and stage: OR 4.31 (95% CI 1.12-19.69)]. A similar result was seen in the expansion cohort. Parity status had no impact on expression of PGRMC1, relaxin-2 and TGFß1. No associations were seen with tumor stage or survival. CONCLUSION: Tumors from parous women with HGSC expressed PR more often than tumors from nulliparous women, indicating that pregnancies might possibly have a long-lasting impact on ovarian cancer development.