RESUMEN
Beta amyloid cleaving enzyme 1 (BACE1) is largely expressed by neurons and is the sole ß-secretase for initiating the production of neuronal ß-amyloid peptides (Aß). To fully understand the physiological functions of neuronal BACE1, we used mouse genetic approach coupled with unbiased single nucleus RNA sequencing (snRNAseq) to investigate how targeted deletion of Bace1 in neurons, driven by Thy-1-Cre recombinase, would affect functions in the nervous system. Our transcriptome results revealed that BACE1 is essential for maturation of neural precursor cells and oligodendrocytes in mice. RNA velocity analysis confirmed deficit in the trajectory of neuroblasts in reaching the immature granule neuron state in young Bace1fl/fl; Thy1-cre mice. Further analysis of differential gene expression indicated changes in genes important for SNARE signaling, tight junction signaling, synaptogenesis and insulin secretion pathways. Morphological studies revealed a hypomyelination in Bace1fl/fl;Thy1-cre sciatic nerves, but no detectable myelination changes in the corpus callosum, despite clear reduction in myelination proteins in the brain. Functional studies showed reduction in long-term potential, defects in synaptogenesis and learning behavioral. Altogether, our results show that neuronal BACE1 is critical for optimal development of central and peripheral nervous system, and inhibition of neuronal BACE1 will result in deficits in synaptic functions and cognitive behaviors.
Asunto(s)
Enfermedad de Alzheimer , Células-Madre Neurales , Ratones , Animales , Secretasas de la Proteína Precursora del Amiloide/genética , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/genética , Ácido Aspártico Endopeptidasas/metabolismo , Células-Madre Neurales/metabolismo , Neuronas/metabolismo , Péptidos beta-Amiloides/metabolismo , Oligodendroglía/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Enfermedad de Alzheimer/metabolismoRESUMEN
Reticulon (RTN) proteins are a family of proteins biochemically identified for shaping tubular endoplasmic reticulum, a subcellular structure important for vesicular transport and cell-to-cell communication. In our recent study of mice with knockout of both reticulon 1 (Rtn1) and Rtn3, we discovered that Rtn1-/-;Rtn3-/- (brief as R1R3dKO) mice exhibited neonatal lethality, despite the fact that mice deficient in either RTN1 or RTN3 alone exhibit no discernible phenotypes. This has been the first case to find early lethality in animals with deletion of partial members of RTN proteins. The complete penetrance for neonatal lethality can be attributed to multiple defects including the impaired neuromuscular junction found in the diaphragm. We also observed significantly impaired axonal growth in a regional-specific manner, detected by immunohistochemical staining with antibodies to neurofilament light chain and neurofilament medium chain. Ultrastructural examination by electron microscopy revealed a significant reduction in synaptic active zone length in the hippocampus. Mechanistic exploration by unbiased proteomic assays revealed reduction of proteins such as FMR1, Staufen2, Cyfip1, Cullin-4B and PDE2a, which are known components in the fragile X mental retardation pathway. Together, our results reveal that RTN1 and RTN3 are required to orchestrate neurofilament organization and intact synaptic structure of the central nervous system.
Asunto(s)
Axones , Citoesqueleto , Hipocampo , Proteínas del Tejido Nervioso , Animales , Ratones , Genes Letales , Ratones Noqueados , Axones/metabolismo , Axones/patología , Citoesqueleto/metabolismo , Citoesqueleto/patología , Proteínas del Tejido Nervioso/metabolismo , Retículo Endoplásmico/metabolismo , Sinapsis , Hipocampo/metabolismo , Hipocampo/patologíaRESUMEN
Alzheimer's disease (AD) increases the risk for seizures and sleep disorders. We show here that germline deletion of ß-site amyloid precursor protein (APP) cleaving enzyme-1 (BACE1) in neurons, but not in astrocytes, increased epileptiform activity. However, Bace1 deletion at adult ages did not alter the normal EEG waveform, indicating less concern for BACE1 inhibition in patients. Moreover, we showed that deletion of Bace1 in the adult was able to reverse epileptiform activity in 5xFAD mice. Intriguingly, treating 5xFAD and APPNL-G-F/NL-G-F (APP KI) mice of either sex with one BACE1 inhibitor Lanabecestat (AZD3293) dramatically increased epileptiform spiking, likely resulting from an off-target effect. We also monitored sleep-wake pathologies in these mice and showed increased wakefulness, decreased non-rapid eye movement sleep, and rapid eye movement sleep in both 5xFAD and APP KI mice; BACE1 inhibition in the adult 5xFAD mice reversed plaque load and sleep disturbances, but this was not seen in APP KI mice. Further studies with and without BACE1 inhibitor treatment showed different levels of plaque-associated microgliosis and activated microglial proteins in 5xFAD mice compared with APP KI mice. Together, BACE1 inhibition should be developed to avoid off-target effect for achieving benefits in reducing epileptic activity and sleep disturbance in Alzheimer's patients.SIGNIFICANCE STATEMENT BACE1 is widely recognized as a therapeutic target for treating Alzheimer's disease patients. However, BACE1 inhibitors failed in clinical trials because of inability to show cognitive improvement in patients. Here we show that BACE1 inhibition actually reduces sleep disturbances and epileptic seizures; both are seen in AD patients. We further showed that one of clinically tested BACE1 inhibitors does have off-target effects, and development of safer BACE1 inhibitors will be beneficial to AD patients. Results from this study will provide useful guidance for additional drug development.
Asunto(s)
Enfermedad de Alzheimer , Trastornos del Sueño-Vigilia , Ratones , Animales , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/genética , Ratones Transgénicos , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/genética , Ácido Aspártico Endopeptidasas/metabolismo , Placa Amiloide , Convulsiones , Trastornos del Sueño-Vigilia/etiología , Trastornos del Sueño-Vigilia/genética , Sueño , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Péptidos beta-Amiloides/metabolismo , Modelos Animales de EnfermedadRESUMEN
BACE1 initiates production of ß-amyloid peptides (Aß), which is associated with cognitive dysfunction in Alzheimer's disease (AD) due to abnormal oligomerization and aggregation. While BACE1 inhibitors show strong reduction in Aß deposition, they fail to improve cognitive function in patients, largely due to its role in synaptic function. We show that BACE1 is required for optimal release of synaptic vesicles. BACE1 deficiency or inhibition decreases synaptic vesicle docking in the synaptic active zones. Consistently, BACE1-null mice or mice treated with clinically tested BACE1 inhibitors Verubecestat and Lanabecestat exhibit severe reduction in hippocampal LTP and learning behaviors. To counterbalance this synaptic deficit, we discovered that BACE1-null mice treated with positive allosteric modulators (PAMs) of metabotropic glutamate receptor 1 (mGluR1), whose levels were reduced in BACE1-null mice and significantly improved long-term potentiation and cognitive behaviors. Similarly, mice treated with mGluR1 PAM showed significantly mitigated synaptic deficits caused by BACE1 inhibitors. Together, our data suggest that a therapy combining BACE1 inhibitors for reducing amyloid deposition and an mGluR1 PAM for counteracting BACE1-mediated synaptic deficits appears to be an effective approach for treating AD patients.
Asunto(s)
Enfermedad de Alzheimer , Secretasas de la Proteína Precursora del Amiloide , Péptidos beta-Amiloides , Animales , Ácido Aspártico Endopeptidasas , Humanos , Ratones , Vesículas SinápticasRESUMEN
Neurofibrillary tangles likely cause neurodegeneration in Alzheimer's disease (AD). We demonstrate that the CX3CL1 C-terminal domain can upregulate neurogenesis, which may ameliorate neurodegeneration. Here we generated transgenic (Tg-CX3CL1) mice by overexpressing CX3CL1 in neurons. Tg-CX3CL1 mice exhibit enhanced neurogenesis in both subgranular and subventricular zones. This enhanced neurogenesis correlates well with elevated expression of TGF-ß2 and TGF-ß3, and activation of their downstream signaling molecule Smad2. Intriguingly, the enhanced adult neurogenesis was mitigated when Smad2 expression was deleted in neurons, supporting a role for the CX3CL1-TGF-ß2/3-Smad2 pathway in the control of adult neurogenesis. When Tg-CX3CL1 mice were crossed with Alzheimer's PS19 mice, which overexpress a tau P301S mutation and exhibit age-dependent neurofibrillary tangles and neurodegeneration, overexpressed CX3CL1 in both male and female mice was sufficient to rescue the neurodegeneration, increase survival time, and improve cognitive function. Hence, we provide in vivo evidence that CX3CL1 is a strong activator of adult neurogenesis, and that it reduces neuronal loss and improves cognitive function in AD.SIGNIFICANCE STATEMENT This study will be the first to demonstrate that enhanced neurogenesis by overexpressed CX3CL1 is mitigated by disruption of Smad2 signaling and is independent of its interaction with CX3CR1. Overexpression of CX3CL1 lengthens the life span of PS19 tau mice by enhancing adult neurogenesis while having minimal effect on tau pathology. Enhancing neuronal CX3CL1, mainly the C-terminal fragment, is a therapeutic strategy for blocking or reversing neuronal loss in Alzheimer's disease or related neurodegenerative disease patients.
Asunto(s)
Enfermedad de Alzheimer , Quimiocina CX3CL1/metabolismo , Neurogénesis , Neuronas/metabolismo , Proteína Smad2/metabolismo , Memoria Espacial/fisiología , Proteínas tau/metabolismo , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/psicología , Animales , Modelos Animales de Enfermedad , Femenino , Masculino , Ratones Transgénicos , Neuronas/patologíaRESUMEN
BACKGROUND: Reticulon 3 (RTN3) is an endoplasmic reticulum protein that has previously been shown to play a role in neurodegenerative diseases, but little is known about its role in lipid metabolism. METHODS: Obese patients (n=149), hypertriglyceridemic patients (n=343), and healthy control subjects (n=84) were enrolled to assess their levels of RTN3. To explore the pathophysiological roles of RTN3 in the control of lipid metabolism, we used transgenic mice overexpressing the wild-type human RTN3 gene, the RTN3-null transgenic mouse model, and multiple Caenorhabditis legans strains for molecular characterization. The underlying mechanisms were studied with 3T3L1 cell cultures in vitro. RESULTS: We report that overexpressed RTN3 in mice induces obesity and higher accumulation of triglycerides. Increased RTN3 expression is also found in patients with obesity and hypertriglyceridemia. We reveal that RTN3 plays critical roles in regulating the biosynthesis and storage of triglycerides and in controlling lipid droplet expansion. Mechanistically, RTN3 regulates these events through its interactions with heat shock protein family A (Hsp70) member 5, and this enhanced interaction increases sterol regulatory element-binding protein 1c and AMP-activated kinase activity. CONCLUSIONS: This study provides evidence for a role of RTN3 in inducing obesity and triglyceride accumulation and suggests that inhibiting the expression of RTN3 in fat tissue may be a novel therapeutic approach to treat obesity and hypertriglyceridemia.
Asunto(s)
Tejido Adiposo/metabolismo , Proteínas Portadoras/sangre , Proteínas de Choque Térmico/metabolismo , Hipertrigliceridemia/sangre , Proteínas de la Membrana/sangre , Proteínas del Tejido Nervioso/sangre , Proteínas del Tejido Nervioso/metabolismo , Obesidad/sangre , Triglicéridos/sangre , Células 3T3-L1 , Proteínas Quinasas Activadas por AMP , Adolescente , Adulto , Animales , Biomarcadores/sangre , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas Portadoras/genética , Estudios de Casos y Controles , Chaperón BiP del Retículo Endoplásmico , Femenino , Predisposición Genética a la Enfermedad , Proteínas de Choque Térmico/genética , Humanos , Hipertrigliceridemia/diagnóstico , Hipertrigliceridemia/genética , Gotas Lipídicas/metabolismo , Masculino , Proteínas de la Membrana/genética , Ratones , Ratones Noqueados , Persona de Mediana Edad , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Obesidad/diagnóstico , Obesidad/genética , Fenotipo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Regulación hacia Arriba , Adulto JovenRESUMEN
Reticulon 3 (RTN3) is a neuronally-expressed reticulon family protein that was previously shown to negatively regulate BACE1, a protease that is required for the generation of ß-amyloid peptides (Aß) from amyloid precursor protein. Despite biochemical and morphological evidence that supports a role of RTN3 in the formation of neuritic amyloid plaques, no systematic analyses of RTN3 mutations in patients with Alzheimer's disease (AD) have yet been reported. RTN3 were targeted sequenced in 154 sporadic early-onset and 285 late-onset AD patients. Luciferase reporter assay and kymographs were performed to analysis the expression of RNT3 and BACE1-RFP particle mobility on cells transfected with wild-type or variants RTN3 constructs. We identified heterozygous variants such as c.-8G > T, c.17C > A, c.42C > T, and c.116C > T from patients in the early-onset AD group and c.-8G > T, c.17C > A, from patients in the late-onset AD group. Such variants of RTN3 were not observed in control individuals. Further biochemical studies show that the RTN3 c.-8G > T variant in the 5'-untranslated region appears to cause reduced expression of RTN3. The RTN3 c.116 C > T variant causes a change of codon T39 to M39 (T39 M). Overexpression of RTN3 T39 M in cultured neurons led to impaired axonal transport of BACE1. The variants found in this study are likely genetic modifiers for RTN3-mediated formation of neuritic plaques in AD.
Asunto(s)
Enfermedad de Alzheimer/genética , Secretasas de la Proteína Precursora del Amiloide/genética , Péptidos beta-Amiloides/genética , Ácido Aspártico Endopeptidasas/genética , Proteínas Portadoras/genética , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Adulto , Anciano , Enfermedad de Alzheimer/fisiopatología , Precursor de Proteína beta-Amiloide/genética , Animales , Transporte Axonal/genética , Encéfalo/metabolismo , Encéfalo/patología , Femenino , Regulación de la Expresión Génica/genética , Humanos , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Mutación , Neuronas/metabolismo , Neuronas/fisiologíaRESUMEN
BACE1 is an indispensable enzyme for generating ß-amyloid peptides, which are excessively accumulated in brains of Alzheimer's patients. However, BACE1 is also required for proper myelination of peripheral nerves, as BACE1-null mice display hypomyelination. To determine the precise effects of BACE1 on myelination, here we have uncovered a role of BACE1 in the control of Schwann cell proliferation during development. We demonstrate that BACE1 regulates the cleavage of Jagged-1 and Delta-1, two membrane-bound ligands of Notch. BACE1 deficiency induces elevated Jag-Notch signaling activity, which in turn facilitates proliferation of Schwann cells. This increase in proliferation leads to shortened internodes and decreased Schmidt-Lanterman incisures. Functionally, evoked compound action potentials in BACE1-null nerves were significantly smaller and slower, with a clear decrease in excitability. BACE1-null nerves failed to effectively use lactate as an alternative energy source under conditions of increased physiological activity. Correlatively, BACE1-null mice showed reduced performance on rotarod tests. Collectively, our data suggest that BACE1 deficiency enhances proliferation of Schwann cell due to the elevated Jag1/Delta1-Notch signaling, but fails to myelinate axons efficiently due to impaired the neuregulin1-ErbB signaling, which has been documented.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Proliferación Celular/fisiología , Células de Schwann/metabolismo , Secretasas de la Proteína Precursora del Amiloide/genética , Animales , Ácido Aspártico Endopeptidasas/genética , Axones/metabolismo , Proliferación Celular/genética , Ratones Noqueados , Vaina de Mielina/metabolismo , Neurogénesis/genética , Neurogénesis/fisiología , Células de Schwann/citología , Nervio Ciático/metabolismo , Transducción de Señal/fisiologíaRESUMEN
Through sol-gel and dip-coating processes, commercial polyurethane sponge modified by polysiloxane was fabricated under low temperature (60 °C) and atmosphere. The contact angle of the obtained polysiloxane/polyurethane sponge is 145 ± 5°. Hence, the polysiloxane/polyurethane sponge could float on water and selectively absorb organics from the surface of the water, indicating simultaneous properties of hydrophobicity and oleophilicity. The absorbent maximum value is 50-150 times of its own weight. The polysiloxane/polyurethane sponge exhibited excellent recyclability, which could be reused by squeezing the sponge due to its high mechanical stability and flexibility. Thermogravimetry-differential thermal analysis test indicated that the polysiloxane/polyurethane sponge exhibited good thermal stability and the stable contact angle of samples tested under increasing temperature indicated its good weather resistance. Due to the commercial property of polyurethane sponge and easy-handling of polysiloxane, the polysiloxane/polyurethane sponge can be easily scaled up to recover a large-area oil spill in water and further work based on the designed equipment has been under consideration.
Asunto(s)
Contaminación por Petróleo/prevención & control , Poliuretanos/química , Reciclaje , Siloxanos/química , Eliminación de Residuos Líquidos/métodos , Adsorción , Interacciones Hidrofóbicas e Hidrofílicas , Eliminación de Residuos Líquidos/economíaRESUMEN
Oil-water separation has recently become a worldwide challenge due to the frequent occurrence of oil spill accidents and increasing industrial oily wastewater. In this work, the multifunctional mesh films with underwater oleophobicity and certain bacteriostatic effects are prepared by layer-by-layer assembly of graphene oxide-silica coatings on stainless steel mesh. The mesh film exhibits excellent environmental stability under a series of harsh conditions. The new, facile and reusable separation system is proposed to achieve deep treatment of oily wastewater, and the oil collection rate can reach over 99%.
Asunto(s)
Grafito/química , Membranas Artificiales , Petróleo/análisis , Dióxido de Silicio/química , Aguas Residuales/química , Antibacterianos , Residuos Industriales , Acero Inoxidable , Eliminación de Residuos Líquidos/métodos , Contaminantes Químicos del Agua/químicaRESUMEN
Reticulon 3 (RTN3) has previously been shown to interact with BACE1 and negatively regulate BACE1 activity. To what extent RTN3 deficiency affects BACE1 activity is an intriguing question. In this study, we aimed to address this by generating RTN3-null mice. Mice with complete deficiency of RTN3 grow normally and have no obviously discernible phenotypes. Morphological analyses of RTN3-null mice showed no significant alterations in cellular structure, although RTN3 is recognized as a protein contributing to the shaping of tubular endoplasmic reticulum. Biochemical analysis revealed that RTN3 deficiency increased protein levels of BACE1. This elevation of BACE1 levels correlated with enhanced processing of amyloid precursor protein at the ß-secretase site. We also demonstrated that RTN3 deficiency in Alzheimer's mouse models facilitates amyloid deposition, further supporting an in vivo role of RTN3 in the regulation of BACE1 activity. Since it has been shown that RTN3 monomer is reduced in brains of Alzheimer's patients, our results suggest that long-lasting reduction of RTN3 levels has adverse effects on BACE1 activity and may contribute to Alzheimer's pathogenesis.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/biosíntesis , Ácido Aspártico Endopeptidasas/biosíntesis , Regulación Enzimológica de la Expresión Génica , Proteínas del Tejido Nervioso/deficiencia , Placa Amiloide/metabolismo , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Animales , Encéfalo/metabolismo , Encéfalo/patología , Femenino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Placa Amiloide/patologíaRESUMEN
BACE1 is a type I transmembrane aspartyl protease that cleaves amyloid precursor protein at the ß-secretase site to initiate the release of ß-amyloid peptide. As a secretase, BACE1 also cleaves additional membrane-bound molecules by exerting various cellular functions. In this study, we showed that BACE1 can effectively shed the membrane-anchored signaling molecule Jagged 1 (Jag1).Wealso mapped the cleavage sites of Jag1 by ADAM10 and ADAM17. Although Jag1 shares a high degree of homology with Jag2 in the ectodomain region, BACE1 fails to cleave Jag2 effectively, indicating a selective cleavage of Jag1. Abolished cleavage of Jag1 in BACE1-null mice leads to enhanced astrogenesis and, concomitantly, reduced neurogenesis. This characterization provides biochemical evidence that the Jag1-Notch pathway is under the control of BACE1 activity
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Proteínas de Unión al Calcio/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteínas de la Membrana/metabolismo , Proteolisis , Receptores Notch/metabolismo , Transducción de Señal/fisiología , Proteínas ADAM/genética , Proteínas ADAM/metabolismo , Proteína ADAM10 , Proteína ADAM17 , Secretasas de la Proteína Precursora del Amiloide/genética , Animales , Ácido Aspártico Endopeptidasas/genética , Proteínas de Unión al Calcio/genética , Células HEK293 , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Proteína Jagged-1 , Proteína Jagged-2 , Proteínas de la Membrana/genética , Ratones , Ratones Mutantes , Ratas , Receptores Notch/genética , Proteínas Serrate-JaggedRESUMEN
Alkaline deoxygenated graphene oxide (aGO) was prepared through alkaline hydrothermal treatment and used as adsorbent to remove Cd(II) ions from aqueous solutions for the first time. The characterization results of transmission electron microscopy, X-ray diffraction, Raman spectroscopy, and Fourier transform infrared (FT-IR) spectra indicate that aGO was successfully synthesized. The batch adsorption experiments showed that the adsorption kinetics could be described by the pseudo-second-order kinetic model, and the isotherms equilibrium data were well fitted with the Langmuir model. The maximum adsorption capacity of Cd(II) on aGO was 156 mg/g at pH 5 and T=293 K. The adsorption thermodynamic parameters indicated that the adsorption process was a spontaneous and endothermic reaction. The mainly adsorption mechanism speculated from FT-IR results may be attributed to the electrostatic attraction between Cd2+ and negatively charged groups (-CO-) of aGO and cation-π interaction between Cd2+ and the graphene planes. The findings of this study demonstrate the potential utility of the nanomaterial aGO as an effective adsorbent for Cd(II) removal from aqueous solutions.
Asunto(s)
Cadmio/química , Grafito/química , Óxidos/química , Eliminación de Residuos Líquidos/métodos , Contaminantes Químicos del Agua/química , Adsorción , CinéticaRESUMEN
Neuritic dystrophy is one of the important pathological features associated with amyloid plaques in Alzheimer's disease (AD) and age-dependent neuronal dysfunctions. We reported previously that reticulon-3 (RTN3) immunoreactive dystrophic neurites (RIDNs) are abundantly present in the hippocampus of AD patients, in AD mouse models, and in aged wild-type mice. Transgenic mice overexpressing the human RTN3 transgene spontaneously develop RIDNs in their hippocampi, and the formation of RIDNs correlates with the appearance of RTN3 aggregation. To further elucidate whether the formation of RIDNs is reversible, we generated transgenic mice expressing wild-type human RTN3 under the control of a tetracycline-responsive promoter. Treatment with doxycycline for 2 months effectively turned off expression of the human RTN3 transgene, confirming the inducible nature of the system. However, the formation of hippocampal RIDNs was dependent on whether the transgene was turned off before or after the formation of RTN3 aggregates. When transgenic human RTN3 expression was turned off at young age, formation of RIDNs was essentially eliminated compared with the vehicle-treated transgenic mice. More importantly, a fear conditioning study demonstrated that contextual associative learning and memory in inducible transgenic mice was improved if the density of RIDNs was lowered. Additional mechanistic study suggested that a reduction in BDNF levels in transgenic mice might contribute to the reduced learning and memory in transgenic mice overexpressing RTN3. Hence, we conclude that age-dependent RIDNs cannot be effectively cleared once they have formed, and we postulate that successful prevention of RIDN formation should be initiated before RTN3 aggregation.
Asunto(s)
Cognición/fisiología , Proteínas del Tejido Nervioso/fisiología , Neuritas/patología , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animales , Ácido Aspártico Endopeptidasas/metabolismo , Western Blotting , Factor Neurotrófico Derivado del Encéfalo/análisis , Factor Neurotrófico Derivado del Encéfalo/biosíntesis , Condicionamiento Psicológico , Señales (Psicología) , Ensayo de Inmunoadsorción Enzimática , Miedo/psicología , Técnica del Anticuerpo Fluorescente , Hipocampo/patología , Humanos , Inmunohistoquímica , Indicadores y Reactivos , Memoria/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Confocal , Proteínas del Tejido Nervioso/inmunologíaRESUMEN
BACE1 is the sole enzyme responsible for cleaving amyloid precursor protein at the ß-secretase site, and this cleavage initiates the generation of ß-amyloid peptide (Aß). Because amyloid precursor protein is predominantly expressed by neurons and deposition of Aß aggregates in the human brain is highly correlated with the Aß released at axonal terminals, we focused our investigation of BACE1 localization on the neuritic region. We show that BACE1 was not only enriched in the late Golgi, trans-Golgi network, and early endosomes but also in both axons and dendrites. BACE1 was colocalized with the presynaptic vesicle marker synaptophysin, indicating the presence of BACE1 in synapses. Because the excessive release of Aß from synapses is attributable to an increase in amyloid deposition, we further explored whether the presence of BACE1 in synapses was regulated by reticulon 3 (RTN3), a protein identified previously as a negative regulator of BACE1. We found that RTN3 is not only localized in the endoplasmic reticulum but also in neuritic regions where no endoplasmic reticulum-shaping proteins are detected, implicating additional functions of RTN3 in neurons. Coexpression of RTN3 with BACE1 in cultured neurons was sufficient to reduce colocalization of BACE1 with synaptophysin. This reduction correlated with decreased anterograde transport of BACE1 in axons in response to overexpressed RTN3. Our results in this study suggest that altered RTN3 levels can impact the axonal transport of BACE1 and demonstrate that reducing axonal transport of BACE1 in axons is a viable strategy for decreasing BACE1 in axonal terminals and, perhaps, reducing amyloid deposition.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuritas/metabolismo , Red trans-Golgi/metabolismo , Amiloide/genética , Secretasas de la Proteína Precursora del Amiloide/genética , Precursor de Proteína beta-Amiloide/genética , Animales , Ácido Aspártico Endopeptidasas/genética , Axones/metabolismo , Transporte Biológico Activo/genética , Humanos , Ratones , Proteínas del Tejido Nervioso/genética , Sinapsis/metabolismo , Red trans-Golgi/genéticaRESUMEN
ß-Site amyloid precursor protein convertase enzyme 1 (BACE1), a type I transmembrane aspartyl protease required to cleave amyloid precursor protein for releasing a toxic amyloid peptide, also cleaves type I and type III neuregulin-1 (Nrg-1). BACE1 deficiency in mice causes hypomyelination during development and impairs remyelination if injured. In BACE1-null mice, the abolished cleavage of neuregulin-1 by BACE1 is speculated to cause reduced myelin sheath thickness in both the central nervous system and peripheral nervous system because reduced cleavage of Nrg-1 correlates with reduced Akt phosphorylation, a downstream signaling molecule of the Nrg-1/ErbB pathway. Here we tested specifically whether increasing Akt activity alone in oligodendrocytes would be sufficient to reverse the hypomyelination phenotype in BACE1-null mice. BACE1-null mice were bred with transgenic mice expressing constitutively active Akt (Akt-DD; mutations with D(308)T and D(473)S) in oligodendrocytes. Relative to littermate BACE1-null controls, BACE1(-/-)/Akt-DD mice exhibited enhanced expression of myelin basic protein and promoter of proteolipid protein. The elevated expression of myelin proteins correlated with a thicker myelin sheath in optic nerves; comparison of quantified g ratios with statistic significance was used to confirm this reversion. However, it appeared that myelin sheath thickness in the sciatic nerves was not increased in BACE1(-/-)/Akt-DD mice, as the g ratio was not significantly different from the control. Hence, increased Akt activity in BACE1-null myelinating cells only compensates for the loss of BACE1 activity in the central nervous system, which is consistent with the observation that overexpression of Akt-DD in Schwann cells did not induce hypermyelination. Our results suggest that signaling activity other than Akt may also contribute to proper myelination in peripheral nerves.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/deficiencia , Ácido Aspártico Endopeptidasas/deficiencia , Proteína Básica de Mielina/biosíntesis , Proteína Proteolipídica de la Mielina/biosíntesis , Vaina de Mielina/fisiología , Proteínas Proto-Oncogénicas c-akt/biosíntesis , Animales , Femenino , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Vaina de Mielina/patología , Oligodendroglía/metabolismo , Fenotipo , Proteínas Proto-Oncogénicas c-akt/genéticaRESUMEN
Alzheimer's disease (AD) is characterized by the formation ß-amyloid (Aß) deposited neuritic plaques. Recent evidence suggests that abnormal lipid metabolism and accumulation could serve as biomarkers for neurodegenerative diseases, including AD. Tubular endoplasmic reticulum protein, reticulon 3 (RTN3), plays a crucial role in the development of neuritic plaque and lipid metabolism in AD brains. In present study, we sought to investigate a potential association between neutral lipid accumulation and AD pathology. BODIPY 500/510 dye was used to label neutral lipid surrounding Aß plaques in APPNL-G-F mouse and AD postmortem brains samples. Immunofluorescent images were captured using confocal microscope and co-localization between lipid metabolism proteins and neutral lipids were evaluated. Lipid accumulation in Aß plaque surrounding dystrophic neurites (DNs) was observed in the cortical region of AD mouse models and human AD brain samples. The neutral lipid staining was not co-localized with IBA1-labeled microglia or GFAP-labeled astrocytes, but it was co-labeled with VAMP2 and neurofilament. We further showed that neutral lipids were accumulated in RTN3 immunoreactive DNs. Both the neutral lipids accumulation and RIDNs formation showed age-dependent patterns in surrounding amyloid plaques. Mechanistic studies revealed that RTN3 likely contributes to the enrichment of neutral lipids near plaques by interacting with heat shock cognate protein 70 (HSC70) and diminishing its function in chaperone-mediated lipophagy. Our study provides immunohistochemical evidence of neutral lipids being enriched in DNs near amyloid plaques. Our findings shed light on RTN3-mediaed lipid accumulation in AD neuropathology and provide fresh insights into the role of RTN3 in neurodegenerative diseases.
Asunto(s)
Enfermedad de Alzheimer , Ratones , Animales , Humanos , Enfermedad de Alzheimer/metabolismo , Neuritas/patología , Placa Amiloide/metabolismo , Ratones Transgénicos , Proteínas del Tejido Nervioso/metabolismo , LípidosRESUMEN
BACKGROUND: While symptoms related to lower urinary tract dysfunction (LUTD) are common in individuals with Alzheimer's disease (AD), pathophysiological links between AD and LUTD remain unclear. OBJECTIVE: This study aimed to investigate whether AD neuropathology would cause autonomic dysfunction along the spinal cord-bladder axis, which could result in alterations in bladder muscle kinetics. METHODS: We utilized APPNL-G-F/NL-G-F knock-in (APP KI) and APPwt/wt (wild-type) mice at two different ages, 4- and 10-month-old, to investigate how AD impacts bladder tissue function by immunohistochemistry, western blotting, and pharmacomyography. RESULTS: We showed that the mucosal layer partially separated from the detrusor in 10-month-old APP KI mouse bladders. Although there was no detectable amyloid deposition in the APP KI bladder, we found amyloid plaques in APP KI lumbar spinal cord. Further immunoblot analysis revealed that tyrosine hydroxylase protein levels were significantly reduced in both 4- and 10-month-old bladder tissues, suggesting reduction of norepinephrine synthesis in APP KI mouse bladders. In contrast, the level of ß2 adrenergic receptor was increased in 4-month-old but not 10-month-old APP KI bladders. In bladder strips, the adrenergic agonist isoproterenol induced increased relaxation in 4- but not 10-month-old APP KI bladders. With 10âHz electrical field stimulation, 10-month-old APP KI bladder strips were more responsive than wild-type controls, with no differences observed in 4-month-old APP KI bladders. CONCLUSIONS: APP KI mice exhibit LUTD, which is likely arising from amyloid pathology in the spinal cord, and results in maturational declines in presynaptic activity combined with compensatory postsynaptic upregulation.
Asunto(s)
Enfermedad de Alzheimer , Ratones , Animales , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Vejiga Urinaria/metabolismo , Vejiga Urinaria/patología , Modelos Animales de Enfermedad , Ratones Transgénicos , Placa Amiloide/patologíaRESUMEN
A cultivable endophytic fungus, Piriformospora indica, improves growth and enhances stress tolerance of host plants, but the underlying mechanisms remain unknown. We hypothesized that P. indica enhanced the drought tolerance of the host by regulating the antioxidant defense system and composition of fatty acids. Trifoliate orange (Poncirus trifoliata) seedlings were inoculated with P. indica under ample water and drought stress to analyze the change in plant growth, reactive oxygen species (ROS) levels, antioxidant enzyme activities, non-enzymatic antioxidant concentrations, fatty acid compositions, and expressions of both antioxidant enzyme genes and fatty acid desaturase (FAD) genes. The 9-week soil water deficit significantly increased the colonization of P. indica to roots, and P. indica promoted the increase of shoot biomass under drought. Soil drought triggered an elevation of hydrogen peroxide in roots, while the inoculated plants had lower levels of ROS (hydrogen peroxide and superoxide anion radicals) and lower degree of membrane lipid peroxidation (based on malondialdehyde levels) under drought. Drought treatment also elevated ascorbic acid and glutathione concentrations, and the elevation was further amplified after P. indica inoculation. Inoculated plants under drought also recorded significantly higher iron-superoxide dismutase (Fe-SOD), manganese-superoxide dismutase (Mn-SOD), peroxidases, catalase, glutathione reductase and ascorbate peroxidase activities, accompanied by up-regulation of PtFe-SOD and PtCu/Zn-SOD expressions. Inoculation with P. indica significantly increased total saturated fatty acids (e.g., C6:0, C15:0, C16:0, C23:0 and C24:0) concentration and reduced total unsaturated fatty acids (e.g., C18:1N9C, C18:2N6, C18:3N3, C18:1N12 and C19:1N9T) concentrations, leading to a decrease in the unsaturation index of fatty acids, which may be associated with the up-regulation of PtFAD2 and PtFAD6 and down-regulation of PtΔ9. It was concluded that the colonization of P. indica can activate enzyme and non-enzyme defense systems and regulate the composition of fatty acids under drought, thus alleviating the oxidative damage to the host caused by drought.
Asunto(s)
Basidiomycota , Poncirus , Antioxidantes/metabolismo , Poncirus/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Resistencia a la Sequía , Peróxido de Hidrógeno/metabolismo , Ácidos Grasos/metabolismo , Basidiomycota/fisiología , Superóxido Dismutasa/metabolismo , Sequías , Agua/metabolismoRESUMEN
Neuregulin-1 (Nrg1) is encoded by a single gene and exists in naturally secreted and transmembrane isoforms. Nrg1 exerts its signaling activity through interaction with its cognate ErbB receptors. Multiple membrane-anchored Nrg1 isoforms, present in six different membrane topologies, must be processed by a protease to initiate a signaling cascade. Here, we demonstrate that BACE1 and ADAM10 can process type I and III Nrg1 at two adjacent sites. Our cleavage site mapping experiments showed that the BACE1 cleavage site is located eight amino acids downstream of the ADAM10 cleavage site, and this order of cleavage is the opposite of amyloid precursor protein cleavage by these two enzymes. Cleavages were further confirmed via optimized electrophoresis. Cleavage of type I or III Nrg1 by ADAM10 and BACE1 released a signaling-capable N-terminal fragment (ntf), either Nrg1-ntfα or Nrg1-ntfß, which could similarly activate an ErbB receptor as evidenced by increased phosphorylation of Akt and ERK, two downstream signaling molecules. Although both Nrg1-ntfα and Nrg1-ntfß could initiate a common signaling cascade, inhibition or down-regulation of ADAM10 alone in a co-culture system did not affect normal myelination, whereas specific inhibition of BACE1 impaired normal myelination. Thus, processing of Nrg1 by BACE1 appears to be more critical for regulating myelination. Our results imply that a significant inhibition of BACE1 could potentially impair Nrg1 signaling activity in vivo.