The cytokine ciliary neurotrophic factor (CNTF) activates hypothalamic urocortin-expressing neurons both in vitro and in vivo.
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ABSTRACT:
SUBMITTER: Purser MJ
PROVIDER: S-EPMC3633986 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Neurotrophic factors are agents with a promising ability to retard progression of neurodegenerative diseases and are effective in slowing photoreceptor degeneration in animal models of retinitis pigmentosa. Here we report a human clinical trial of a neurotrophic factor for retinal neurodegeneration. In this Phase I safety trial, human ciliary neurotrophic factor (CNTF) was delivered by cells transfected with the human CNTF gene and sequestered within capsules that were surgically implanted into the vitreous of the eye. The outer membrane of the encapsulated cell implant is semipermeable to allow CNTF to reach the retina. Ten participants received CNTF implants in one eye. When the implants were removed after 6 months, they contained viable cells with minimal cell loss and gave CNTF output at levels previously shown to be therapeutic for retinal degeneration in rcd1 dogs. Although the trial was not powered to form a judgment as to clinical efficacy, of seven eyes for which visual acuity could be tracked by conventional reading charts, three eyes reached and maintained improved acuities of 10-15 letters, equivalent to two- to three-line improvement on standard Snellen acuity charts. A surgically related choroidal detachment in one eye resulted in a transient acuity decrease that resolved with conservative management. This Phase I trial indicated that CNTF is safe for the human retina even with severely compromised photoreceptors. The approach to delivering therapeutic proteins to degenerating retinas using encapsulated cell implants may have application beyond disease caused by genetic mutations.
| S-EPMC1383495 | biostudies-literature
Project description:In humans and experimental animals, the administration of ciliary neurotrophic factor (CNTF) reduces food intake and body weight. To gain further insights into the mechanism(s) underlying its satiety effect, we: (i) evaluated the CNTF-dependent activation of the Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) pathway in mouse models where neuropeptide Y (NPY) and pro-opiomelanocortin (POMC) neurons can be identified by green fluorescent protein (GFP); and (ii) assessed whether CNTF promotes leptin signaling in hypothalamic feeding centers. Immunohistochemical experiments enabled us to establish that intraperitoneal injection of mouse recombinant CNTF activated the JAK2-STAT3 pathway in a substantial proportion of arcuate nucleus (ARC) NPY neurons (18.68% ± 0.60 in 24-h fasted mice and 25.50% ± 1.17 in fed mice) but exerted a limited effect on POMC neurons (4.15% ± 0.33 in 24-h fasted mice and 2.84% ± 0.45 in fed mice). CNTF-responsive NPY neurons resided in the ventromedial ARC, facing the median eminence (ME), and were surrounded by albumin immunoreactivity, suggesting that they are located outside the blood-brain barrier (BBB). In both normally fed and high-fat diet (HFD) obese animals, CNTF activated extracellular signal-regulated kinase signaling in ME ?1- and ?2-tanycytes, an effect that has been linked to the promotion of leptin entry into the brain. Accordingly, compared to the animals treated with leptin, mice treated with leptin/CNTF showed: (i) a significantly greater leptin content in hypothalamic protein extracts; (ii) a significant increase in phospho-STAT3 (P-STAT3)-positive neurons in the ARC and the ventromedial hypothalamic nucleus of normally fed mice; and (iii) a significantly increased number of P-STAT3-positive neurons in the ARC and dorsomedial hypothalamic nucleus of HFD obese mice. Collectively, these data suggest that exogenously administered CNTF reduces food intake by exerting a leptin-like action on distinctive NPY ARC neurons and by promoting leptin signaling in hypothalamic feeding centers.
| S-EPMC7253709 | biostudies-literature
Project description:Defensive behaviors are evolved responses to threat stimuli, and a potential threat elicits risk assessment (RA) behavior. However, neural mechanisms underlying RA behavior are hardly understood. Urocortin-3 (Ucn3) is a member of corticotropin-releasing factor peptide family and here, we report that Ucn3 neurons in the hypothalamic perifornical area (PeFA) are involved in RA of a novel object, a potential threat stimulus, in mice. Histological and in vivo fiber photometry studies revealed that the activity of PeFA Ucn3 neurons was associated with novel object investigation involving the stretch-attend posture, a behavioral marker for RA. Chemogenetic activation of these neurons increased RA and burying behaviors toward a novel object without affecting anxiety and corticosterone levels. Ablation of these neurons caused the abnormal behaviors of gnawing and direct contacts with novel objects, especially in a home-cage. These results suggest that PeFA Ucn3 neurons modulate defensive responses to a potential threat stimulus.
| S-EPMC7770982 | biostudies-literature
Project description:In animal models, the administration of ciliary neurotrophic factor (CNTF) was demonstrated to reduce bone mass and to participate in bone remodeling. Cementoblasts, a cell type embedded in the cementum, are the main cells to produce and mineralize the extracellular matrix. The effect of CNTF on cementoblasts has not yet been addressed. Thus, the goal of this in vitro study was to investigate possible influences of exogenous CNTF on cementogenesis, as well as autophagy regulation and subsequent mechanisms in cementoblasts. Cementoblasts (OCCM-30) were stimulated with exogenous CNTF. Alizarin Red staining was performed to analyze the functional differentiation (mineralization) of OCCM-30 cells. The release of OPG was quantified by ELISA. The expression of cementogenesis markers (RUNX-2, OCN, BMP-7, BSP, and SPON-2) was evaluated by RT-qPCR. Western blotting (WB) was performed for the protein expression of STAT3, COX-2, SHP-2, cPLAα, cPLAβ; ERK1/2, P38, and JNK. The autophagic flux was assessed using WB and RT-qPCR analysis of LC3A/B, Beclin-1, and Atg-5, and the autophagosome was investigated by immunofluorescence staining (IF). The ERK1/2 (FR180204) or STAT3 (sc-202818) antagonist was added, and the cellular response was analyzed using flow cytometry. Exogenous CNTF significantly attenuated mineralized nodule formation, impaired OPG release, and downregulated the mRNA levels of RUNX-2, OCN, BMP-7, and BSP. Moreover, CNTF induced the phosphorylation of STAT3 and activated a transient activation of SHP-2, cPLAβ, ERK1/2, P38, and JNK protein. CNTF also induced autophagosome formation and promoted autophagy-associated gene and protein expressions. Additionally, the inhibition of ERK1/2 or STAT3 reversed a CNTF-induced mineralization impairment and had regulatory effects on CNTF-induced autophagosome formation. Our data revealed that CNTF acts as a potent inhibitor of cementogenesis, and it can trigger autophagy, in part by ERK1/2 and STAT3 commitment in the cementoblasts. Thus, it may play an important role in inducing or facilitating inflammatory root resorption during orthodontic tooth movement.
| S-EPMC9408951 | biostudies-literature
Project description:Ciliary neurotrophic factor, cardiotrophin-like cytokine, and neuropoietin are members of the four-helix bundle cytokine family. These proteins signal through a common tripartite receptor composed of leukemia inhibitory factor receptor, gp130, and ciliary neurotrophic factor receptor alpha. Binding to ciliary neurotrophic factor receptor alpha occurs through an interaction site located at the C terminus of the cytokine AB loop and alphaD helix, known as site 1. In the present study, we have generated a model of neuropoietin and identified a conserved binding site for the three cytokines interacting with ciliary neurotrophic factor receptor alpha. To identify the counterpart of this site on ciliary neurotrophic factor receptor alpha, its cytokine binding domain was modeled, and the physicochemical properties of its surface were analyzed. This analysis revealed an area displaying properties complementary to the site 1 of ciliary neurotrophic factor, cardiotrophin-like cytokine, and neuropoietin. Based on our computational predictions, residues were selected for their potential involvement in the ciliary neurotrophic factor receptor alpha binding epitope, and site-directed mutagenesis was carried out. Biochemical, cell proliferation, and cell signaling analyses showed that Phe(172) and Glu(286) of ciliary neurotrophic factor receptor alpha are key interaction residues. Our results demonstrated that ciliary neurotrophic factor, cardiotrophin-like cytokine, and neuropoietin share a conserved binding site on ciliary neurotrophic factor receptor alpha.
| S-EPMC2662086 | biostudies-literature
Project description:IL-27, consisting of the subunits IL-27p28 and Epstein-Barr virus-induced gene 3 (EBI3), is a heterodimeric cytokine belonging to the IL-6/IL-12 family of cytokines. IL-27p28 is a four-helical cytokine requiring association with the soluble receptor EBI3 to be efficiently secreted and functionally active. Computational and biological analyses of the IL-27 binding site 1 to its receptor revealed important structural proximities with the ciliary neurotrophic factor group of cytokines and highlighted the contribution of p28 Trp(97), as well as of EBI3 Phe(97), Asp(210), and Glu(159), as key residues in the interactions between both cytokine subunits. WSX-1 (IL-27R) and gp130 compose the IL-27 receptor-signaling complex, recruiting the STAT-1 and STAT-3 pathways. A study of IL-27 binding site 3 showed that Trp(197) was crucial for the cytokine's interaction with gp130, but that the mutated cytokine still recognized IL-27R on the cell surface. IL-27 exerts both pro- and anti-inflammatory functions, promoting proliferation and differentiation of Th1 and inhibiting Th17 differentiation. Our results led us to develop mutated forms of human and mouse IL-27 with antagonistic activities. Using an in vivo mouse model of concanavalin A-induced Th1-cell-mediated hepatitis, we showed that the murine IL-27 antagonist W195A decreased liver inflammation by downregulating the synthesis of CXCR3 ligands and several acute phase proteins. Together, these data suggest that IL-27 antagonism could be of interest in down-modulating acute IL-27-driven Th1-cell-mediated immune response.
| S-EPMC2984150 | biostudies-literature
Project description:Previous studies indicate that the activity of hypothalamic POMC neurons can be regulated by glucose via intracellular mechanisms, but its regulation by lactate is poorly understood. In addition to its energetic role, lactate acts as a signaling molecule. In this study, we evaluated the function and location of the lactate receptor, hydroxycarboxylic acid receptor 1 (HCAR1). We used a conditional genetic approach to label POMC neurons and evaluated their sensitivity to lactate using patch-clamp recordings. L-Lactate and 3-chloro-5-hydroxybenzoic acid (3Cl-HBA), HCAR1 specific agonist depolarized POMC neurons and the increase in excitability was abolished by pertussis toxin (PTX), indicating the involvement of Gαi/o-protein-coupled receptors. In addition, the depolarization of a subset of POMC neurons was sensitive to α-cyano-4-hydroxycinnamate (4-CIN), a lactate transporter blocker, suggesting that the depolarization induced by L-lactate can also occur by direct intracellular action. Surprisingly, HCAR1 was not detected in POMC neurons, but instead localized in astrocytes. These results suggest a new lactate-mediated mechanism for astrocyte-neuron intercellular communication.
| S-EPMC8569171 | biostudies-literature
Project description:Ciliary neurotrophic factor (CNTF) is a neurotrophic factor with therapeutic potential for neurodegenerative diseases. Moreover, therapeutic application of CNTF reduced body weight in mice and humans. CNTF binds to high or low affinity receptor complexes consisting of CNTFR·gp130·LIFR or IL-6R·gp130·LIFR, respectively. Clinical studies of the CNTF derivative Axokine revealed intolerance at higher concentrations, which may rely on the low-affinity binding of CNTF to the IL-6R. Here, we aimed to generate a CNTFR-selective CNTF variant (CV). CV-1 contained the single amino acid exchange R28E. Arg(28) is in close proximity to the CNTFR binding site. Using molecular modeling, we hypothesized that Arg(28) might contribute to IL-6R/CNTFR plasticity of CNTF. CV-2 to CV-5 were generated by transferring parts of the CNTFR-binding site from cardiotrophin-like cytokine to CNTF. Cardiotrophin-like cytokine selectively signals via the CNTFR·gp130·LIFR complex, albeit with a much lower affinity compared with CNTF. As shown by immunoprecipitation, all CNTF variants retained the ability to bind to CNTFR. CV-1, CV-2, and CV-5, however, lost the ability to bind to IL-6R. Although all variants induced cytokine-dependent cellular proliferation and STAT3 phosphorylation via CNTFR·gp130·LIFR, only CV-3 induced STAT3 phosphorylation via IL-6R·gp130·LIFR. Quantification of CNTF-dependent proliferation of CNTFR·gp130·LIFR expressing cells indicated that only CV-1 was as biologically active as CNTF. Thus, the CNTFR-selective CV-1 will allow discriminating between CNTFR- and IL-6R-mediated effects in vivo.
| S-EPMC4140248 | biostudies-literature
Project description:The TrkB receptor is critical for the control of energy balance, as mutations in its gene (NTRK2) lead to hyperphagia and severe obesity. The main neural substrate mediating the appetite-suppressing activity of TrkB, however, remains unknown. Here, we demonstrate that selective Ntrk2 deletion within paraventricular hypothalamus (PVH) leads to severe hyperphagic obesity. Furthermore, chemogenetic activation or inhibition of TrkB-expressing PVH (PVHTrkB) neurons suppresses or increases food intake, respectively. PVHTrkB neurons project to multiple brain regions, including ventromedial hypothalamus (VMH) and lateral parabrachial nucleus (LPBN). We find that PVHTrkB neurons projecting to LPBN are distinct from those to VMH, yet Ntrk2 deletion in PVH neurons projecting to either VMH or LPBN results in hyperphagia and obesity. Additionally, TrkB activation with BDNF increases firing of these PVH neurons. Therefore, TrkB signaling is a key regulator of a previously uncharacterized neuronal population within the PVH that impinges upon multiple circuits to govern appetite.
| S-EPMC7138837 | biostudies-literature
Project description:Animals must ingest water via drinking to maintain fluid homeostasis, yet the neurons that specifically promote drinking behavior are incompletely characterized. The lateral hypothalamic area (LHA) as a whole is essential for drinking behavior but most LHA neurons indiscriminately promote drinking and feeding. By contrast, activating neurotensin (Nts)-expressing LHA neurons (termed LHA Nts neurons) causes mice to immediately drink water with a delayed suppression of feeding. We therefore hypothesized that LHA Nts neurons are sufficient to induce drinking behavior and that these neurons specifically bias for fluid intake over food intake. To test this hypothesis we used designer receptors exclusively activated by designer drugs (DREADDs) to selectively activate LHA Nts neurons and studied the impact on fluid intake, fluid preference and feeding. Activation of LHA Nts neurons stimulated drinking in water-replete and dehydrated mice, indicating that these neurons are sufficient to promote water intake regardless of homeostatic need. Interestingly, mice with activated LHA Nts neurons drank any fluid that was provided regardless of its palatability, but if given a choice they preferred water or palatable solutions over unpalatable (quinine) or dehydrating (hypertonic saline) solutions. Notably, acute activation of LHA Nts neurons robustly promoted fluid but not food intake. Overall, our study confirms that activation of LHA Nts neurons is sufficient to induce drinking behavior and biases for fluid intake. Hence, LHA Nts neurons may be important targets for orchestrating the appropriate ingestive behavior necessary to maintain fluid homeostasis. This article is part of the Special Issue entitled 'Hypothalamic Control of Homeostasis'.
| S-EPMC6433557 | biostudies-literature