RESUMO
Identification of factors regulating renal development is important to understand the pathogenesis of congenital kidney diseases. Little is known about the molecular mechanism of renal development and functions triggered by the angiogenic hormone prokineticin-2 and its receptor, PKR1. Utilizing the Gata5 (G5)-Cre and Wilms tumor 1 (Wt1)(GFP)cre transgenic lines, we generated mutant mice with targeted PKR1 gene disruptions in nephron progenitors. These mutant mice exhibited partial embryonic and postnatal lethality. Kidney developmental defects in PKR(G5-/-) mice are manifested in the adult stage as renal atrophy with glomerular defects, nephropathy, and uremia. PKR1(Wt1-/-) embryos exhibit hypoplastic kidneys with premature glomeruli and necrotic nephrons as a result of impaired proliferation and increased apoptosis in Wt1(+) renal mesenchymal cells. PKR1 regulates renal mesenchymal-epithelial transition (MET) that is involved in formation of renal progenitors, regulating glomerulogenesis toward forming nephrons during kidney development. In the isolated embryonic Wt1(+) renal cells, overexpression or activation of PKR1 promotes MET defined by the transition from elongated cell to octagonal cell morphology, and alteration of the expression of MET markers via activating NFATc3 signaling. Together, these results establish PKR1 via NFATc3 as a crucial modifier of MET processing to the development of nephron. Our study should facilitate new therapeutic opportunities in human renal disorders.-Arora, H., Boulberdaa, M., Qureshi, R., Bitirim, V., Messadeq, N., Dolle, P., Nebigil, C. G. Prokineticin receptor 1 is required for mesenchymal-epithelial transition in kidney development.
Assuntos
Transição Epitelial-Mesenquimal/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Receptores Acoplados a Proteínas G/metabolismo , Animais , Apoptose , Proliferação de Células , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário , Transição Epitelial-Mesenquimal/genética , Camundongos , Camundongos Knockout , Mutação , Neovascularização Fisiológica , Receptores Acoplados a Proteínas G/genéticaRESUMO
Mutations in the DYNC1H1 gene encoding for dynein heavy chain cause two closely related human motor neuropathies, dominant spinal muscular atrophy with lower extremity predominance (SMA-LED) and axonal Charcot-Marie-Tooth (CMT) disease, and lead to sensory neuropathy and striatal atrophy in mutant mice. Dynein is the molecular motor carrying mitochondria retrogradely on microtubules, yet the consequences of dynein mutations on mitochondrial physiology have not been explored. Here, we show that mouse fibroblasts bearing heterozygous or homozygous point mutation in Dync1h1, similar to human mutations, show profoundly abnormal mitochondrial morphology associated with the loss of mitofusin 1. Furthermore, heterozygous Dync1h1 mutant mice display progressive mitochondrial dysfunction in muscle and mitochondria progressively increase in size and invade sarcomeres. As a likely consequence of systemic mitochondrial dysfunction, Dync1h1 mutant mice develop hyperinsulinemia and hyperglycemia and progress to glucose intolerance with age. Similar defects in mitochondrial morphology and mitofusin levels are observed in fibroblasts from patients with SMA-LED. Last, we show that Dync1h1 mutant fibroblasts show impaired perinuclear clustering of mitochondria in response to mitochondrial uncoupling. Our results show that dynein function is required for the maintenance of mitochondrial morphology and function with aging and suggest that mitochondrial dysfunction contributes to dynein-dependent neurological diseases, such as SMA-LED.
Assuntos
Envelhecimento/patologia , Dineínas do Citoplasma/genética , Mitocôndrias/patologia , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patologia , Mutação/genética , Animais , Células Cultivadas , Embrião de Mamíferos , Feminino , Glucagon/sangue , Ácido Glutâmico/genética , Humanos , Insulina/sangue , Lisina/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mitocôndrias/ultraestrutura , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1 , TransfecçãoRESUMO
Barded-Biedl syndrome (BBS) is a rare genetic disorder with an unmet medical need for retinal degeneration. Small-molecule drugs were previously identified to slow down the apoptosis of photoreceptors in BBS mouse models. Clinical translation was not practical due to the necessity of repetitive invasive intravitreal injections for pediatric populations. Non-invasive methods of retinal drug targeting are a prerequisite for acceptable adaptation to the targeted pediatric patient population. Here, we present the development and functional testing of a non-invasive, topical, magnetically assisted delivery system, harnessing the ability of magnetic nanoparticles (MNPs) to cargo two drugs (guanabenz and valproic acid) with anti-unfolded protein response (UPR) properties towards the retina. Using magnetic resonance imaging (MRI), we showed the MNPs' presence in the retina of Bbs wild-type mice, and their photoreceptor localization was validated using transmission electron microscopy (TEM). Subsequent electroretinogram recordings (ERGs) demonstrated that we achieved beneficial biological effects with the magnetically assisted treatment translating the maintained light detection in Bbs-/- mice (KO). To our knowledge, this is the first demonstration of efficient magnetic drug targeting in the photoreceptors in vivo after topical administration. This non-invasive, needle-free technology expands the application of SMDs for the treatment of a vast spectrum of retinal degenerations and other ocular diseases.
RESUMO
Tumor Necrosis Factor Receptor-Associated Factors (TRAFs) are major signal transducers for the TNF and interleukin-1/Toll-like receptor superfamilies. However, TRAF4 does not fit the paradigm of TRAF function in immune and inflammatory responses. Its physiological and molecular functions remain poorly understood. Behavorial analyses show that TRAF4-deficient mice (TRAF4-KO) exhibit altered locomotion coordination typical of ataxia. TRAF4-KO central nervous system (CNS) ultrastructure shows strong myelin perturbation including disorganized layers and disturbances in paranode organization. TRAF4 was previously reported to be expressed by CNS neurons. Using primary cell culture, we now show that TRAF4 is also expressed by oligodendrocytes, at all stages of their differentiation. Moreover, histology and electron microscopy show degeneration of a high number of Purkinje cells in TRAF4-KO mice, that was confirmed by increased expression of the Bax pro-apoptotic marker (immunofluorescence), TUNEL analysis, and caspase-3 activation and PARP1 cleavage (western blotting). Consistent with this phenotype, MAG and NogoA, two myelin-induced neurite outgrowth inhibitors, and their neuron partners, NgR and p75NTR were overexpressed (Q-RT-PCR and western blotting). The strong increased phosphorylation of Rock2, a RhoA downstream target, indicated that the NgR/p75NTR/RhoA signaling pathway, known to induce actin cytoskeleton rearrangement that favors axon regeneration inhibition and neuron apoptosis, is activated in the absence of TRAF4 (western blotting). Altogether, these results provide conclusive evidence for the pivotal contribution of TRAF4 to myelination and to cerebellar homeostasis, and link the loss of TRAF4 function to demyelinating or neurodegenerative diseases.
Assuntos
Sistema Nervoso Central/metabolismo , Homeostase , Bainha de Mielina/metabolismo , Fator 4 Associado a Receptor de TNF/metabolismo , Envelhecimento/patologia , Animais , Comportamento Animal/fisiologia , Western Blotting , Peso Corporal/fisiologia , Células Cultivadas , Sistema Nervoso Central/patologia , Sistema Nervoso Central/fisiopatologia , Imunofluorescência , Proteínas Ligadas por GPI/metabolismo , Locomoção/fisiologia , Camundongos , Camundongos Knockout , Proteínas da Mielina/metabolismo , Bainha de Mielina/patologia , Glicoproteína Associada a Mielina/metabolismo , Degeneração Neural/metabolismo , Degeneração Neural/patologia , Degeneração Neural/fisiopatologia , Neuritos/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Proteínas Nogo , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Células de Purkinje/metabolismo , Células de Purkinje/patologia , Receptores de Fator de Crescimento Neural/metabolismo , Transdução de Sinais , Fator 4 Associado a Receptor de TNF/deficiência , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
This is the first in vivo study of plasma-sprayed carbon nanotube (CNT)-reinforced hydroxyapatite (HA) coating on titanium implants embedded in rodents' bone. No adverse effect or cytotoxicity of CNT addition on bone tissues and cells was observed. Normal bone growth was observed around HA-CNT-coated implants. CNT addition induces higher osseointegration as compared to HA. Elastic modulus of new bone was compared with the modulus of HA-CNT/bone interface to understand the mechanical integrity of the implant.
Assuntos
Materiais Revestidos Biocompatíveis/química , Durapatita/química , Nanotecnologia/métodos , Osseointegração/efeitos dos fármacos , Titânio/química , Animais , Fenômenos Biomecânicos , Substitutos Ósseos , Fêmur/patologia , Teste de Materiais , Microscopia Eletrônica de Varredura , Próteses e Implantes , Desenho de Prótese , RatosRESUMO
Diminished mitochondrial oxidative phosphorylation and aerobic capacity are associated with reduced longevity. We tested whether resveratrol (RSV), which is known to extend lifespan, impacts mitochondrial function and metabolic homeostasis. Treatment of mice with RSV significantly increased their aerobic capacity, as evidenced by their increased running time and consumption of oxygen in muscle fibers. RSV's effects were associated with an induction of genes for oxidative phosphorylation and mitochondrial biogenesis and were largely explained by an RSV-mediated decrease in PGC-1alpha acetylation and an increase in PGC-1alpha activity. This mechanism is consistent with RSV being a known activator of the protein deacetylase, SIRT1, and by the lack of effect of RSV in SIRT1(-/-) MEFs. Importantly, RSV treatment protected mice against diet-induced-obesity and insulin resistance. These pharmacological effects of RSV combined with the association of three Sirt1 SNPs and energy homeostasis in Finnish subjects implicates SIRT1 as a key regulator of energy and metabolic homeostasis.