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1.
Gene ; 806: 145920, 2022 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-34455026

RESUMO

Depression is deemed a mood disorder characterized by a high rate of relapse. Therefore, overcoming of the recurrent depression is globally expecting. Kososan, a traditional Japanese herbal medicine, has been clinically used for mild depressive mood, and our previous studies have shown some evidence for its antidepressive-like efficacy in experimental animal models of depression. However, it remains unclear whether kososan has beneficial effects on recurrent depression. Here, we examined its effect using a mouse model of modified repeated social defeat stress (SDS) paradigm. Male BALB/c mice were exposed to a 5-min SDS from unfamiliar aggressive CD-1 mice for 5 days. Kososan extract (1.0 kg/kg/day) or an antidepressant milnacipran (60 mg/kg/day) was administered orally for 26 days (days 7-32) to depression-like mice with social avoidant behaviors on day 6. Single 5 min of SDS was subjected to mice recovered from the social avoidance on day 31, and then the recurrence of depression-like behaviors was evaluated on day 32. Hippocampal gene expression patterns were also assayed by DNA microarray analysis. Water- or milnacipran-administered mice resulted in a recurrence of depression-like behaviors by re-exposure of single SDS, whereas kososan-administered mice did not recur depression-like behaviors. Distinct gene expression patterns were also found for treating kososan and milnacipran. Collectively, this finding suggests that kososan exerts a preventive effect on recurrent depression-like behaviors in mice. Pretreatment of kososan is more useful for recurrent depression than that of milnacipran.


Assuntos
Antidepressivos/farmacologia , Depressão/prevenção & controle , Medicamentos de Ervas Chinesas/farmacologia , Proteínas do Tecido Nervoso/genética , Derrota Social , Estresse Psicológico/tratamento farmacológico , Administração Oral , Animais , Depressão/genética , Depressão/fisiopatologia , Depressão/psicologia , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ontologia Genética , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipocampo/fisiopatologia , Japão , Masculino , Medicina Kampo/métodos , Camundongos , Camundongos Endogâmicos BALB C , Milnaciprano/farmacologia , Anotação de Sequência Molecular , Proteínas do Tecido Nervoso/classificação , Proteínas do Tecido Nervoso/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Recidiva , Estresse Psicológico/genética , Estresse Psicológico/fisiopatologia
2.
Nat Commun ; 12(1): 4087, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34471112

RESUMO

We utilized forebrain organoids generated from induced pluripotent stem cells of patients with a syndromic form of Autism Spectrum Disorder (ASD) with a homozygous protein-truncating mutation in CNTNAP2, to study its effects on embryonic cortical development. Patients with this mutation present with clinical characteristics of brain overgrowth. Patient-derived forebrain organoids displayed an increase in volume and total cell number that is driven by increased neural progenitor proliferation. Single-cell RNA sequencing revealed PFC-excitatory neurons to be the key cell types expressing CNTNAP2. Gene ontology analysis of differentially expressed genes (DEgenes) corroborates aberrant cellular proliferation. Moreover, the DEgenes are enriched for ASD-associated genes. The cell-type-specific signature genes of the CNTNAP2-expressing neurons are associated with clinical phenotypes previously described in patients. The organoid overgrowth phenotypes were largely rescued after correction of the mutation using CRISPR-Cas9. This CNTNAP2-organoid model provides opportunity for further mechanistic inquiry and development of new therapeutic strategies for ASD.


Assuntos
Transtorno do Espectro Autista/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Organoides/metabolismo , Prosencéfalo/metabolismo , Adolescente , Transtorno do Espectro Autista/genética , Diferenciação Celular , Proliferação de Células , Criança , Feminino , Predisposição Genética para Doença/genética , Humanos , Células-Tronco Pluripotentes Induzidas , Proteínas de Membrana/genética , Mutação , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Fenótipo , Análise de Sequência de RNA
3.
Int J Mol Sci ; 22(16)2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34445257

RESUMO

The production of pancreatic ß cells is the most challenging step for curing diabetes using next-generation treatments. Adult pancreatic endocrine cells are thought to be maintained by the self-duplication of differentiated cells, and pancreatic endocrine neogenesis can only be observed when the tissue is severely damaged. Experimentally, this can be performed using a method named partial duct ligation (PDL). As the success rate of PDL surgery is low because of difficulties in identifying the pancreatic duct, we previously proposed a method for fluorescently labeling the duct in live animals. Using this method, we performed PDL on neurogenin3 (Ngn3)-GFP transgenic mice to determine the origin of endocrine precursor cells and evaluate their potential to differentiate into multiple cell types. Ngn3-activated cells, which were marked with GFP, appeared after PDL operation. Because some GFP-positive cells were aligned proximally to the duct, we hypothesized that Ngn3-positive cells arise from the pancreatic duct. Therefore, we next developed an in vitro pancreatic duct culture system using Ngn3-GFP mice and examined whether Ngn3-positive cells emerge from this duct. We observed GFP expressions in ductal organoid cultures. GFP expressions were correlated with Ngn3 expressions and endocrine cell lineage markers. Interestingly, tuft cell markers were also correlated with GFP expressions. Our results demonstrate that in adult mice, Ngn3-positive endocrine precursor cells arise from the pancreatic ducts both in vivo and in vitro experiments indicating that the pancreatic duct could be a potential donor for therapeutic use.


Assuntos
Antígenos de Diferenciação/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Células Secretoras de Insulina/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Ductos Pancreáticos/metabolismo , Células-Tronco/metabolismo , Animais , Antígenos de Diferenciação/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células Secretoras de Insulina/citologia , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Organoides/citologia , Organoides/metabolismo , Ductos Pancreáticos/citologia , Células-Tronco/citologia
4.
Int J Mol Sci ; 22(16)2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34445283

RESUMO

Botulinum neurotoxins (BoNTs) are the most poisonous substances in nature. Currently, the only therapy for botulism is antitoxin. This therapy suffers from several limitations and hence new therapeutic strategies are desired. One of the limitations in discovering BoNT inhibitors is the absence of an in vitro assay that correlates with toxin neutralization in vivo. In this work, a high-throughput screening assay for receptor-binding inhibitors against BoNT/A was developed. The assay is composed of two chimeric proteins: a receptor-simulating protein, consisting of the fourth luminal loop of synaptic vesicle protein 2C fused to glutathione-S-transferase, and a toxin-simulating protein, consisting of the receptor-binding domain of BoNT/A fused to beta-galactosidase. The assay was applied to screen the LOPAC1280 compound library. Seven selected compounds were evaluated in mice exposed to a lethal dose of BoNT/A. The compound aurintricarboxylic acid (ATA) conferred 92% protection, whereas significant delayed time to death (p < 0.005) was observed for three additional compounds. Remarkably, ATA was also fully protective in mice challenged with a lethal dose of BoNT/E, which also uses the SV2 receptor. This study demonstrates that receptor-binding inhibitors have the potential to serve as next generation therapeutics for botulism, and therefore the assay developed may facilitate discovery of new anti-BoNT countermeasures.


Assuntos
Ácido Aurintricarboxílico/farmacologia , Toxinas Botulínicas Tipo A/toxicidade , Toxinas Botulínicas/toxicidade , Botulismo/tratamento farmacológico , Botulismo/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Animais , Botulismo/genética , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/genética , Camundongos , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , beta-Galactosidase/genética , beta-Galactosidase/metabolismo
5.
Nat Commun ; 12(1): 4872, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381052

RESUMO

The Netrin-1 receptor UNC5B is an axon guidance regulator that is also expressed in endothelial cells (ECs), where it finely controls developmental and tumor angiogenesis. In the absence of Netrin-1, UNC5B induces apoptosis that is blocked upon Netrin-1 binding. Here, we identify an UNC5B splicing isoform (called UNC5B-Δ8) expressed exclusively by ECs and generated through exon skipping by NOVA2, an alternative splicing factor regulating vascular development. We show that UNC5B-Δ8 is a constitutively pro-apoptotic splicing isoform insensitive to Netrin-1 and required for specific blood vessel development in an apoptosis-dependent manner. Like NOVA2, UNC5B-Δ8 is aberrantly expressed in colon cancer vasculature where its expression correlates with tumor angiogenesis and poor patient outcome. Collectively, our data identify a mechanism controlling UNC5B's necessary apoptotic function in ECs and suggest that the NOVA2/UNC5B circuit represents a post-transcriptional pathway regulating angiogenesis.


Assuntos
Apoptose , Vasos Sanguíneos/crescimento & desenvolvimento , Receptores de Netrina/metabolismo , Isoformas de RNA/metabolismo , Processamento Alternativo , Animais , Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/metabolismo , Células Endoteliais , Humanos , Morfogênese , Neovascularização Patológica/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Receptores de Netrina/genética , Netrina-1/metabolismo , Isoformas de RNA/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Análise de Sobrevida , Peixe-Zebra
6.
Adv Protein Chem Struct Biol ; 127: 249-270, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34340769

RESUMO

We present an overview of current state of proteomic approaches as applied to optic nerve regeneration in the historical context of nerve regeneration particularly central nervous system neuronal regeneration. We present outlook pertaining to the optic nerve regeneration proteomics that the latter can extrapolate information from multi-systems level investigations. We present an account of the current need of systems level standardization for comparison of proteome from various models and across different pharmacological or biophysical treatments that promote adult neuron regeneration. We briefly overview the need for deriving knowledge from proteomics and integrating with other omics to obtain greater biological insight into process of adult neuron regeneration in the optic nerve and its potential applicability to other central nervous system neuron regeneration.


Assuntos
Modelos Neurológicos , Regeneração Nervosa , Proteínas do Tecido Nervoso/metabolismo , Nervo Óptico/fisiologia , Proteoma/metabolismo , Proteômica , Animais , Humanos
7.
Int J Mol Sci ; 22(15)2021 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-34360710

RESUMO

A presynaptic active zone organizer protein Bassoon orchestrates numerous important functions at the presynaptic active zone. We previously showed that the absence of Bassoon exclusively in forebrain glutamatergic presynapses (BsnEmx1cKO) in mice leads to developmental disturbances in dentate gyrus (DG) affecting synaptic excitability, morphology, neurogenesis and related behaviour during adulthood. Here, we demonstrate that hyperexcitability of the medial perforant path-to-DG (MPP-DG) pathway in BsnEmx1cKO mice emerges during adolescence and is sustained during adulthood. We further provide evidence for a potential involvement of tropomyosin-related kinase B (TrkB), the high-affinity receptor for brain-derived neurotrophic factor (BDNF), mediated signalling. We detect elevated TrkB protein levels in the dorsal DG of adult mice (~3-5 months-old) but not in adolescent (~4-5 weeks-old) mice. Electrophysiological analysis reveals increased field-excitatory-postsynaptic-potentials (fEPSPs) in the DG of the adult, but not in adolescent BsnEmx1cKO mice. In line with an increased TrkB expression during adulthood in BsnEmx1cKO, blockade of TrkB normalizes the increased synaptic excitability in the DG during adulthood, while no such effect was observed in adolescence. Accordingly, neurogenesis, which has previously been found to be increased in adult BsnEmx1cKO mice, was unaffected at adolescent age. Our results suggest that Bassoon plays a crucial role in the TrkB-dependent postnatal maturation of the hippocampus.


Assuntos
Hipocampo/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Prosencéfalo/metabolismo , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais , Transmissão Sináptica , Animais , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Proteínas Tirosina Quinases/genética
8.
Int J Mol Sci ; 22(15)2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34360775

RESUMO

Coupling glycolysis and mitochondrial tricarboxylic acid cycle, pyruvate dehydrogenase (PDH) complex (PDHC) is highly responsive to cellular demands through multiple mechanisms, including PDH phosphorylation. PDHC also produces acetyl-CoA for protein acetylation involved in circadian regulation of metabolism. Thiamine (vitamin B1) diphosphate (ThDP) is known to activate PDH as both coenzyme and inhibitor of the PDH inactivating kinases. Molecular mechanisms integrating the function of thiamine-dependent PDHC into general redox metabolism, underlie physiological fitness of a cell or an organism. Here, we characterize the daytime- and thiamine-dependent changes in the rat brain PDHC function, expression and phosphorylation, assessing their impact on protein acetylation and metabolic regulation. Morning administration of thiamine significantly downregulates both the PDH phosphorylation at Ser293 and SIRT3 protein level, the effects not observed upon the evening administration. This action of thiamine nullifies the daytime-dependent changes in the brain PDHC activity and mitochondrial acetylation, inducing diurnal difference in the cytosolic acetylation and acetylation of total brain proteins. Screening the daytime dependence of central metabolic enzymes and proteins of thiol/disulfide metabolism reveals that thiamine also cancels daily changes in the malate dehydrogenase activity, opposite to those of the PDHC activity. Correlation analysis indicates that thiamine abrogates the strong positive correlation between the total acetylation of the brain proteins and PDHC function. Simultaneously, thiamine heightens interplay between the expression of PDHC components and total acetylation or SIRT2 protein level. These thiamine effects on the brain acetylation system change metabolic impact of acetylation. The changes are exemplified by the thiamine enhancement of the SIRT2 correlations with metabolic enzymes and proteins of thiol-disulfide metabolism. Thus, we show the daytime- and thiamine-dependent changes in the function and phosphorylation of brain PDHC, contributing to regulation of the brain acetylation system and redox metabolism. The daytime-dependent action of thiamine on PDHC and SIRT3 may be of therapeutic significance in correcting perturbed diurnal regulation.


Assuntos
Encéfalo/metabolismo , Cetona Oxirredutases/metabolismo , Mitocôndrias/enzimologia , Proteínas Mitocondriais/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Sirtuínas/metabolismo , Tiamina/farmacologia , Acetilação/efeitos dos fármacos , Animais , Ciclo do Ácido Cítrico/efeitos dos fármacos , Masculino , Fosforilação/efeitos dos fármacos , Ratos , Ratos Wistar , Fatores de Tempo
9.
Int J Mol Sci ; 22(15)2021 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-34361063

RESUMO

BACKGROUND: Induced tooth movement during orthodontic therapy requires mechano-induced bone remodeling. Besides various cytokines and growth-factors, neuronal guidance molecules gained attention for their roles in bone homeostasis and thus, potential roles during tooth movement. Several neuronal guidance molecules have been implicated in the regulation of bone remodeling. Amongst them, Semaphorin 3A is particular interesting as it concurrently induces osteoblast differentiation and disturbs osteoclast differentiation. METHODS: Mechano-regulation of Sema3A and its receptors PlexinA1 and Neuropilin (RT-qPCR, WB) was evaluated by applying compressive and tension forces to primary human periodontal fibroblasts (hPDLF) and alveolar bone osteoblasts (hOB). The association of the transcription factor Osterix (SP7) and SEMA3A was studied by RT-qPCR. Mechanisms involved in SEMA3A-mediated osteoblast differentiation were assessed by Rac1GTPase pull-downs, ß-catenin expression analyses (RT-qPCR) and nuclear translocation assays (IF). Osteogenic markers were analyzed by RT-qPCR. RESULTS: SEMA3A, PLXNA1 and NRP1 were differentially regulated by tension or compressive forces in hPDLF. Osterix (SP7) displayed the same pattern of regulation. Recombinant Sema3A induced the activation of Rac1GTPase, the nuclear translocation of ß-catenin and the expression of osteogenic marker genes. CONCLUSION: Sema3A, its receptors and Osterix are regulated by mechanical forces in hPDLF. SEMA3A upregulation was associated with Osterix (SP7) modulation. Sema3A-enhanced osteogenic marker gene expression in hOB might be dependent on a pathway involving Rac1GTPase and ß-catenin. Thus, Semaphorin 3A might contribute to bone remodeling during induced tooth movement.


Assuntos
Fibroblastos/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Neuropilinas/metabolismo , Osteoblastos/fisiologia , Ligamento Periodontal/fisiologia , Receptores de Superfície Celular/metabolismo , Semaforina-3A/metabolismo , Técnicas de Movimentação Dentária/métodos , Adolescente , Adulto , Remodelação Óssea , Diferenciação Celular , Células Cultivadas , Criança , Fibroblastos/citologia , Humanos , Proteínas do Tecido Nervoso/genética , Neuropilinas/genética , Osteoblastos/citologia , Osteogênese , Ligamento Periodontal/citologia , Receptores de Superfície Celular/genética , Semaforina-3A/genética , Adulto Jovem
10.
Int J Mol Sci ; 22(15)2021 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-34361042

RESUMO

Various neurodegenerative disorders are associated with human NTE/PNPLA6 dysfunction. Mechanisms of neuropathogenesis in these diseases are far from clearly elucidated. Hereditary spastic paraplegia belongs to a type of neurodegeneration associated with NTE/PNLPLA6 and is implicated in neuron death. In this study, we used Drosophila melanogaster to investigate the consequences of neuronal knockdown of swiss cheese (sws)-the evolutionarily conserved ortholog of human NTE/PNPLA6-in vivo. Adult flies with the knockdown show longevity decline, locomotor and memory deficits, severe neurodegeneration progression in the brain, reactive oxygen species level acceleration, mitochondria abnormalities and lipid droplet accumulation. Our results suggest that SWS/NTE/PNPLA6 dysfunction in neurons induces oxidative stress and lipid metabolism alterations, involving mitochondria dynamics and lipid droplet turnover in neurodegeneration pathogenesis. We propose that there is a complex mechanism in neurological diseases such as hereditary spastic paraplegia, which includes a stress reaction, engaging mitochondria, lipid droplets and endoplasmic reticulum interplay.


Assuntos
Encéfalo/metabolismo , Proteínas de Drosophila/metabolismo , Gotículas Lipídicas/metabolismo , Mitocôndrias/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Encéfalo/citologia , Proteínas de Drosophila/deficiência , Proteínas de Drosophila/genética , Drosophila melanogaster , Metabolismo dos Lipídeos , Mitocôndrias/ultraestrutura , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Estresse Oxidativo
11.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360929

RESUMO

Complexins (Cplxs) 1 to 4 are components of the presynaptic compartment of chemical synapses where they regulate important steps in synaptic vesicle exocytosis. In the retina, all four Cplxs are present, and while we know a lot about Cplxs 3 and 4, little is known about Cplxs 1 and 2. Here, we performed in situ hybridization experiments and bioinformatics and exploited Cplx 1 and Cplx 2 single-knockout mice combined with immunocytochemistry and light microscopy to characterize in detail the cell type and synapse-specific distribution of Cplx 1 and Cplx 2. We found that Cplx 2 and not Cplx 1 is the main isoform expressed in normal and displaced amacrine cells and ganglion cells in mouse retinae and that amacrine cells seem to operate with a single Cplx isoform at their conventional chemical synapses. Surprising was the finding that retinal function, determined with electroretinographic recordings, was altered in Cplx 1 but not Cplx 2 single-knockout mice. In summary, the results provide an important basis for future studies on the function of Cplxs 1 and 2 in the processing of visual signals in the mammalian retina.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Células Amácrinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Células Fotorreceptoras/metabolismo , Células Bipolares da Retina/metabolismo , Células Ganglionares da Retina/metabolismo , Células Horizontais da Retina/metabolismo , Proteínas SNARE/metabolismo , Sinapses/metabolismo , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Células Cultivadas , Biologia Computacional/métodos , Eletrorretinografia/métodos , Feminino , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/genética
12.
Biomed Res Int ; 2021: 6647734, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34307664

RESUMO

Pancreatic enzyme replacement therapy (PERT) and fat predigestion are key in ensuring the optimal growth of patients with cystic fibrosis. Our study attempted to highlight differences between fat predigestion and conventional PERT on body composition of young pigs with exocrine pancreatic insufficiency (EPI). EPI and healthy pigs were fed with high-fat diet for six weeks. During the last two weeks of the study, all pigs received additional nocturnal alimentation with Peptamen AF (PAF) and were divided into three groups: H-healthy pigs receiving PAF; P-EPI pigs receiving PAF+PERT; and L-EPI pigs receiving PAF predigested with an immobilized microbial lipase. Additional nocturnal alimentation increased the body weight gain of EPI pigs with better efficacy in P pigs. Humerus length and area in pigs in groups L and P were lower than that observed in pigs in group H (p value 0.005-0.088). However, bone mineral density and strength were significantly higher in P and L as compared to that of H pigs (p value 0.0026-0.0739). The gut structure was improved in P pigs. The levels of neurospecific proteins measured in the brain were mainly affected in P and less in L pigs as compared to H pigs. The beneficial effects of the nocturnal feeding with the semielemental diet in the prevention of EPI pigs' growth/development retardation are differently modified by PERT or fat predigestion in terms of growth, bone properties, neurospecific protein distribution, and gut structure.


Assuntos
Dieta , Terapia de Reposição de Enzimas , Insuficiência Pancreática Exócrina/terapia , Comportamento Alimentar , Lipase/uso terapêutico , Pancrelipase/uso terapêutico , Animais , Astrócitos/metabolismo , Composição Corporal , Osso e Ossos/patologia , Trato Gastrointestinal/patologia , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Suínos , Ganho de Peso
13.
Int J Mol Sci ; 22(12)2021 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-34203049

RESUMO

The present study aims to determine the neuroprotective effect of Bergenin against spatial memory deficit associated with neurodegeneration. Preliminarily, the protective effect of Bergenin was observed against H2O2-induced oxidative stress in HT-22 and PC-12 cells. Further studies were performed in 5xFAD Tg mouse model by administering Bergenin (1, 30 and 60 mg/kg; orally), whereas Bergenin (60 mg/kg) significantly attenuated the memory deficit observed in the Y-maze and Morris water maze (MWM) test. Fourier transform-infrared (FT-IR) spectroscopy displayed restoration of lipids, proteins and their derivatives compared to the 5xFAD Tg mice group. The differential scanning calorimeter (DSC) suggested an absence of amyloid beta (Aß) aggregation in Bergenin-treated mice. The immunohistochemistry (IHC) analysis suggested the neuroprotective effect of Bergenin by increasing Reelin signaling (Reelin/Dab-1) and attenuated Aß (1-42) aggregation in hippocampal regions of mouse brains. Furthermore, IHC and western blot results suggested antioxidant (Keap-1/Nrf-2/HO-1), anti-inflammatory (TLR-4/NF-kB) and anti-apoptotic (Bcl-2/Bax/Caspase-3) effect of Bergenin. Moreover, a decrease in Annexin V/PI-stained hippocampal cells suggested its effect against neurodegeneration. The histopathological changes were reversed significantly by Bergenin. In addition, a remarkable increase in antioxidant level with suppression of pro-inflammatory cytokines, oxidative stress and nitric oxide production were observed in specific regions of the mouse brains.


Assuntos
Benzopiranos/farmacologia , Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Transtornos da Memória/etiologia , Transtornos da Memória/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Serina Endopeptidases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Antioxidantes/farmacologia , Benzopiranos/química , Biomarcadores , Linhagem Celular , Citocinas/metabolismo , Modelos Animais de Doenças , Ligação de Hidrogênio , Mediadores da Inflamação/metabolismo , Aprendizagem em Labirinto/efeitos dos fármacos , Transtornos da Memória/diagnóstico , Transtornos da Memória/tratamento farmacológico , Camundongos , Camundongos Transgênicos , Modelos Moleculares , Estrutura Molecular , Atividade Motora/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Relação Estrutura-Atividade , Resultado do Tratamento
14.
Cardiovasc Ther ; 2021: 5554569, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34257705

RESUMO

Ginkgolide B (GB) is an active ingredient extracted from Ginkgo biloba leaves. However, the effects of GB on cardiac hypertrophy remain unclear. The study is aimed at determining whether GB could alleviate cardiac hypertrophy and exploring its underlying molecular mechanism. Rat cardiomyocyte cell line H9c2 cells were pretreated with GB and incubated with angiotensin II (Ang II) to simulate an in vitro cardiac hypertrophy model. Cell viability, cell size, hypertrophy markers, and autophagy were determined in H9c2 cells after Ang II treatment. Proteins involved in autophagy and the SIRT1 pathway were determined by western blot. Our data demonstrated that GB attenuated Ang II-induced cardiac hypertrophy and reduced the mRNA expressions of hypertrophy marker, atrial natriuretic peptide (ANP), and ß-myosin heavy chain (ß-MHC). GB further increased Ang II-induced autophagy in H9c2 cells and modulated expressions of autophagy-related proteins Beclin1 and P62. Modulation of autophagy using autophagy inhibitor 3-methyladenine (3-MA) could abrogate GB-downregulated transcription of NPPA. We then showed that GB attenuated Ang II-induced oxidative stress and reduction in SIRT1 and FoxO1 protein expression. Finally, the effect of GB on autophagy and cardiac hypertrophy could be reversed by SIRT1 inhibitor EX-527. GB inhibits Ang II-induced cardiac hypertrophy by enhancing autophagy via the SIRT1-FoxO1 signaling pathway and might be a potential agent in treating pathological cardiac hypertrophy.


Assuntos
Angiotensina II/toxicidade , Autofagia/efeitos dos fármacos , Ginkgolídeos/farmacologia , Lactonas/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Proteínas do Tecido Nervoso/metabolismo , Sirtuína 1/metabolismo , Animais , Fator Natriurético Atrial/genética , Cardiomegalia/tratamento farmacológico , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Linhagem Celular , Miócitos Cardíacos/patologia , Substâncias Protetoras/farmacologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Transcrição Genética/efeitos dos fármacos , Miosinas Ventriculares/genética
15.
Int J Mol Sci ; 22(14)2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-34299206

RESUMO

Despite the intensive investigation of the molecular mechanism of skeletal muscle hypertrophy, the underlying signaling processes are not completely understood. Therefore, we used an overload model, in which the main synergist muscles (gastrocnemius, soleus) of the plantaris muscle were surgically removed, to cause a significant overload in the remaining plantaris muscle of 8-month-old Wistar male rats. SIRT1-associated pro-anabolic, pro-catabolic molecular signaling pathways, NAD and H2S levels of this overload-induced hypertrophy were studied. Fourteen days of overload resulted in a significant 43% (p < 0.01) increase in the mass of plantaris muscle compared to sham operated animals. Cystathionine-ß-synthase (CBS) activities and bioavailable H2S levels were not modified by overload. On the other hand, overload-induced hypertrophy of skeletal muscle was associated with increased SIRT1 (p < 0.01), Akt (p < 0.01), mTOR, S6 (p < 0.01) and suppressed sestrin 2 levels (p < 0.01), which are mostly responsible for anabolic signaling. Decreased FOXO1 and SIRT3 signaling (p < 0.01) suggest downregulation of protein breakdown and mitophagy. Decreased levels of NAD+, sestrin2, OGG1 (p < 0.01) indicate that the redox milieu of skeletal muscle after 14 days of overloading is reduced. The present investigation revealed novel cellular interactions that regulate anabolic and catabolic processes in the hypertrophy of skeletal muscle.


Assuntos
Cistationina beta-Sintase/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/patologia , Animais , Hipertrofia/genética , Hipertrofia/metabolismo , Hipertrofia/patologia , Masculino , Proteínas Musculares/genética , Músculo Esquelético/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar , Proteínas Quinases S6 Ribossômicas/genética , Proteínas Quinases S6 Ribossômicas/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Sirtuínas/antagonistas & inibidores , Sirtuínas/genética , Sirtuínas/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
16.
Nat Commun ; 12(1): 4601, 2021 07 29.
Artigo em Inglês | MEDLINE | ID: mdl-34326322

RESUMO

Genomic sequencing of thousands of tumors has revealed many genes associated with specific types of cancer. Similarly, large scale CRISPR functional genomics efforts have mapped genes required for cancer cell proliferation or survival in hundreds of cell lines. Despite this, for specific disease subtypes, such as metastatic prostate cancer, there are likely a number of undiscovered tumor specific driver genes that may represent potential drug targets. To identify such genetic dependencies, we performed genome-scale CRISPRi screens in metastatic prostate cancer models. We then created a pipeline in which we integrated pan-cancer functional genomics data with our metastatic prostate cancer functional and clinical genomics data to identify genes that can drive aggressive prostate cancer phenotypes. Our integrative analysis of these data reveals known prostate cancer specific driver genes, such as AR and HOXB13, as well as a number of top hits that are poorly characterized. In this study we highlight the strength of an integrated clinical and functional genomics pipeline and focus on two top hit genes, KIF4A and WDR62. We demonstrate that both KIF4A and WDR62 drive aggressive prostate cancer phenotypes in vitro and in vivo in multiple models, irrespective of AR-status, and are also associated with poor patient outcome.


Assuntos
Proteínas de Ciclo Celular/genética , Cinesina/genética , Proteínas do Tecido Nervoso/genética , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Animais , Sistemas CRISPR-Cas , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular/metabolismo , Movimento Celular/fisiologia , Células Cultivadas , Bases de Dados Genéticas , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Cinesina/metabolismo , Masculino , Camundongos Endogâmicos NOD , Camundongos SCID , Metástase Neoplásica , Estadiamento de Neoplasias , Proteínas do Tecido Nervoso/metabolismo , Neoplasias da Próstata/metabolismo , Taxa de Sobrevida
17.
Nat Commun ; 12(1): 4611, 2021 07 29.
Artigo em Inglês | MEDLINE | ID: mdl-34326333

RESUMO

Hedgehog signaling is essential for bone formation, including functioning as a means for the growth plate to drive skeletal mineralization. However, the mechanisms regulating hedgehog signaling specifically in bone-forming osteoblasts are largely unknown. Here, we identified SLIT and NTRK-like protein-5(Slitrk5), a transmembrane protein with few identified functions, as a negative regulator of hedgehog signaling in osteoblasts. Slitrk5 is selectively expressed in osteoblasts and loss of Slitrk5 enhanced osteoblast differentiation in vitro and in vivo. Loss of SLITRK5 in vitro leads to increased hedgehog signaling and overexpression of SLITRK5 in osteoblasts inhibits the induction of targets downstream of hedgehog signaling. Mechanistically, SLITRK5 binds to hedgehog ligands via its extracellular domain and interacts with PTCH1 via its intracellular domain. SLITRK5 is present in the primary cilium, and loss of SLITRK5 enhances SMO ciliary enrichment upon SHH stimulation. Thus, SLITRK5 is a negative regulator of hedgehog signaling in osteoblasts that may be attractive as a therapeutic target to enhance bone formation.


Assuntos
Cílios/metabolismo , Proteínas Hedgehog/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Osteoblastos/metabolismo , Osteogênese/fisiologia , Receptor Patched-1/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Proteínas Hedgehog/genética , Humanos , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Osteoblastos/citologia , Receptor Patched-1/genética , Transdução de Sinais
18.
Nat Commun ; 12(1): 4578, 2021 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-34321481

RESUMO

Mitochondria are transported along microtubules by opposing kinesin and dynein motors. Kinesin-1 and dynein-dynactin are linked to mitochondria by TRAK proteins, but it is unclear how TRAKs coordinate these motors. We used single-molecule imaging of cell lysates to show that TRAK2 robustly activates kinesin-1 for transport toward the microtubule plus-end. TRAK2 is also a novel dynein activating adaptor that utilizes a conserved coiled-coil motif to interact with dynein to promote motility toward the microtubule minus-end. However, dynein-mediated TRAK2 transport is minimal unless the dynein-binding protein LIS1 is present at a sufficient level. Using co-immunoprecipitation and co-localization experiments, we demonstrate that TRAK2 forms a complex containing both kinesin-1 and dynein-dynactin. These motors are functionally linked by TRAK2 as knockdown of either kinesin-1 or dynein-dynactin reduces the initiation of TRAK2 transport toward either microtubule end. We propose that TRAK2 coordinates kinesin-1 and dynein-dynactin as an interdependent motor complex, providing integrated control of opposing motors for the proper transport of mitochondria.


Assuntos
Dineínas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Cinesina/metabolismo , Mitocôndrias/metabolismo , Proteínas do Tecido Nervoso/metabolismo , 1-Alquil-2-acetilglicerofosfocolina Esterase , Proteínas de Transporte/metabolismo , Complexo Dinactina/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Associadas aos Microtúbulos , Microtúbulos/metabolismo , Proteínas do Tecido Nervoso/genética , Transporte Proteico/fisiologia , Transcriptoma
19.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201747

RESUMO

Rett syndrome (RTT) is a rare neurological disorder caused by mutations in the X-linked MECP2 gene and a major cause of intellectual disability in females. No cure exists for RTT. We previously reported that the behavioural phenotype and brain mitochondria dysfunction are widely rescued by a single intracerebroventricular injection of the bacterial toxin CNF1 in a RTT mouse model carrying a truncating mutation of the MeCP2 gene (MeCP2-308 mice). Given the heterogeneity of MECP2 mutations in RTT patients, we tested the CNF1 therapeutic efficacy in a mouse model carrying a null mutation (MeCP2-Bird mice). CNF1 selectively rescued cognitive defects, without improving other RTT-related behavioural alterations, and restored brain mitochondrial respiratory chain complex activity in MeCP2-Bird mice. To shed light on the molecular mechanisms underlying the differential CNF1 effects on the behavioural phenotype, we compared treatment effects on relevant signalling cascades in the brain of the two RTT models. CNF1 provided a significant boost of the mTOR activation in MeCP2-308 hippocampus, which was not observed in the MeCP2-Bird model, possibly explaining the differential effects of CNF1. These results demonstrate that CNF1 efficacy depends on the mutation beared by MeCP2-mutated mice, stressing the need of testing potential therapeutic approaches across RTT models.


Assuntos
Toxinas Bacterianas/farmacologia , Encéfalo/efeitos dos fármacos , Proteínas de Escherichia coli/farmacologia , Proteína 2 de Ligação a Metil-CpG/genética , Mitocôndrias/efeitos dos fármacos , Síndrome de Rett/tratamento farmacológico , Animais , Toxinas Bacterianas/administração & dosagem , Encéfalo/metabolismo , Modelos Animais de Doenças , Proteínas de Escherichia coli/administração & dosagem , Medo/efeitos dos fármacos , Feminino , Infusões Intraventriculares , Mutação com Perda de Função , Masculino , Transtornos da Memória/tratamento farmacológico , Transtornos da Memória/etiologia , Camundongos Mutantes , Proteínas dos Microfilamentos/metabolismo , Mitocôndrias/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Síndrome de Rett/etiologia , Serina-Treonina Quinases TOR/metabolismo
20.
Commun Biol ; 4(1): 870, 2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34267322

RESUMO

The role of oligodendrocyte lineage cells, the largest glial population in the adult central nervous system (CNS), in the pathogenesis of Alzheimer's disease (AD) remains elusive. Here, we developed a culture method for adult oligodendrocyte progenitor cells (aOPCs). Fibroblast growth factor 2 (FGF2) promotes survival and proliferation of NG2+ aOPCs in a serum-free defined medium; a subpopulation (~5%) of plexin-B3+ aOPCs was also found. FGF2 withdrawal decreased NG2+, but increased plexin-B3+ aOPCs and Aß1-42 secretion. Plexin-B3+ aOPCs were distributed throughout the adult rat brain, although less densely than NG2+ aOPCs. Spreading depolarization induced delayed cortical plexin-B3+ aOPC gliosis in the ipsilateral remote cortex. Furthermore, extracellular Aß1-42 accumulation was occasionally found around plexin-B3+ aOPCs near the lesions. In AD brains, virtually all cortical SPs were immunostained for plexin-B3, and plexin-B3 levels increased significantly in the Sarkosyl-soluble fractions. These findings suggest that plexin-B3+ aOPCs may play essential roles in AD pathogenesis, as natural Aß-secreting cells.


Assuntos
Doença de Alzheimer/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Células Precursoras de Oligodendrócitos/metabolismo , Oligodendroglia/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Animais , Antígenos/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Células Cultivadas , Feminino , Humanos , Masculino , Moléculas de Adesão de Célula Nervosa/metabolismo , Células Precursoras de Oligodendrócitos/citologia , Oligodendroglia/citologia , Fragmentos de Peptídeos/metabolismo , Proteoglicanas/metabolismo , Ratos Sprague-Dawley
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