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1.
Gene ; 716: 144036, 2019 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-31381952

RESUMO

Nebulin is a 770 kDa protein that is localized along the thin filaments of skeletal muscles in vertebrates. It is also present in the striated muscles of Amphioxus, an invertebrate cephalochordate that is phylogenetically close to vertebrates. However, the nebulin of urochordate ascidians or its expression in invertebrate hearts has not been investigated. In this study, we investigated the structure and cardiac expression of the nebulin gene in Ciona intestinalis, a urochordate whose phylogeny lies between cephalochordates and vertebrates. As a result of the gene structure analysis, we found that the Ciona nebulin gene predicted to be 62 kb and consists of 143 exons. The nebulin was expected to consist of a unique N-terminal region, followed by 155 nebulin repeats, another unique region, a Ser-rich region and a C-terminal SH3 domain. Whole-mount in situ hybridization experiments showed that the Ciona nebulin gene was expressed in a variety of muscles, including hearts. However, Western blot analysis using antibody to Ciona nebulin did not detect the presence of full-length nebulin. Alternatively, RT-PCR experiments on samples of Ciona heart detected the expression of nebulette-like and nrap-like isoforms from the Ciona nebulin gene. These results indicate that, similarly to vertebrate hearts, Ciona hearts do not express nebulin, but rather nrap- and nebulette-like isoforms. These results also imply that the nebulin, nebulette and nrap genes in vertebrates were separated from an ancestral invertebrate nebulin gene during vertebrate evolution.


Assuntos
Ciona intestinalis/genética , Família Multigênica , Proteínas Musculares/genética , Miocárdio/metabolismo , Animais , Ciona intestinalis/metabolismo , Evolução Molecular , Éxons , Íntrons , Proteínas Musculares/química , Proteínas Musculares/metabolismo , Domínios Proteicos , RNA Mensageiro/metabolismo
2.
Mol Med Rep ; 19(6): 4955-4963, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059031

RESUMO

In most cases, exogenous oestradiol benzoate (EB) inhibits spermatogenesis, however, the mechanism underlying this process has not been fully elucidated. The present study investigated the effect of EB on redox equilibrium and glycometabolism in mouse testes. Male Kunming mice were divided into 3 groups and injected with 0, 5 and 10 mg/kg EB, respectively. Histological analysis revealed no sperm and far fewer spermatogenic cells in the testes of EB­treated mice. Additionally, transmission electron microscopy revealed that mitochondria in Sertoli cells were transformed to vacuoles with irregular cristae in the EB­treated group. EB also significantly decreased the activities and mRNA expression of catalase, superoxide dismutase, and glutathione peroxidase and increased the activity of nitric oxide synthase and nitric oxide concentration in the testes compared with the control. These results indicated that oxidative damage was caused by EB treatment. With regard to glycometabolism, ATP content and activities of hexokinase and pyruvate kinase were significantly reduced in the EB­treated group. Although glucose and pyruvate concentrations were significantly increased by EB treatment, levels of lactate, the main energy source of spermatogenic cells, were unchanged. Monocarboxylate transporter 2 (MCT2) and MCT4, which are responsible for lactate transportation, were downregulated by EB. In conclusion, the results of the present study indicated that azoospermia induced by EB in male mice was associated with oxidative damage and the disorder of testicular metabolic cooperation.


Assuntos
Azoospermia/patologia , Estradiol/análogos & derivados , Metaboloma/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Testículo/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Azoospermia/induzido quimicamente , Azoospermia/veterinária , Cromatografia Líquida de Alta Pressão , Regulação para Baixo/efeitos dos fármacos , Estradiol/farmacologia , Transportador de Glucose Tipo 3/genética , Transportador de Glucose Tipo 3/metabolismo , Hexoquinase/genética , Hexoquinase/metabolismo , Masculino , Camundongos , Microscopia Eletrônica , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Fosforilação/efeitos dos fármacos , Células de Sertoli/ultraestrutura , Espermatogênese/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Testículo/metabolismo , Testículo/ultraestrutura
3.
Nat Rev Dis Primers ; 5(1): 30, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-31048702

RESUMO

Myasthenia gravis (MG) is an autoimmune disease caused by antibodies against the acetylcholine receptor (AChR), muscle-specific kinase (MuSK) or other AChR-related proteins in the postsynaptic muscle membrane. Localized or general muscle weakness is the predominant symptom and is induced by the antibodies. Patients are grouped according to the presence of antibodies, symptoms, age at onset and thymus pathology. Diagnosis is straightforward in most patients with typical symptoms and a positive antibody test, although a detailed clinical and neurophysiological examination is important in antibody-negative patients. MG therapy should be ambitious and aim for clinical remission or only mild symptoms with near-normal function and quality of life. Treatment should be based on MG subgroup and includes symptomatic treatment using acetylcholinesterase inhibitors, thymectomy and immunotherapy. Intravenous immunoglobulin and plasma exchange are fast-acting treatments used for disease exacerbations, and intensive care is necessary during exacerbations with respiratory failure. Comorbidity is frequent, particularly in elderly patients. Active physical training should be encouraged.


Assuntos
Miastenia Gravis/diagnóstico , Miastenia Gravis/terapia , Acetilcolinesterase/genética , Acetilcolinesterase/fisiologia , Corticosteroides/uso terapêutico , Agrina/genética , Agrina/fisiologia , Anti-Inflamatórios não Esteroides/uso terapêutico , Autoanticorpos/análise , Autoanticorpos/sangue , Biomarcadores/análise , Biomarcadores/sangue , Blefaroptose/etiologia , Colágeno/genética , Colágeno/fisiologia , Cortactina/genética , Cortactina/fisiologia , Eletromiografia/métodos , Humanos , Canal de Potássio Kv1.4/genética , Canal de Potássio Kv1.4/fisiologia , Proteínas Relacionadas a Receptor de LDL/genética , Proteínas Relacionadas a Receptor de LDL/fisiologia , Proteínas Musculares/genética , Proteínas Musculares/fisiologia , Miastenia Gravis/fisiopatologia , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/fisiologia , Receptores Colinérgicos/genética , Receptores Colinérgicos/fisiologia , Receptores Nicotínicos/genética , Fatores de Risco , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/fisiologia
4.
BMC Genomics ; 20(1): 396, 2019 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-31113376

RESUMO

BACKGROUND: Phenotypic plasticity is a common and highly adaptive phenomenon where the same genotype produces different phenotypes in response to environmental cues. Sogatella furcifera, a migratory pest of rice exhibits wing dimorphism, is a model insect for studying phenotypic plasticity of wing size. The Insullin-PI3K-Akt-FOXO signaling pathway plays a crucial role in the manipulation of wing size in the migratory insects. However, the regulatory mechanism via the pathway involved in wing dimorphism are still unexplored. RESULTS: Accompanied by special alternative splicing, genes involved in muscle contraction and energy metabolism were highly expressed in the wing hinges of macropters, demonstrating their adaptation for energy-demanding long-distance flights. Based on FOXO ChIP-Seq analysis, a total of 1259 putative target genes were observed in the wing hinges, including wing morph development, flight muscle and energy metabolism genes. An integrated gene interaction network was built by combining four heterogeneous datasets, and the IIS-PI3K-Akt-FOXO pathway was clustered in a divided functional module. In total, 45 genes in the module directly interacting with the IIS-PI3K-Akt-FOXO pathway showed differential expression levels between the two wing hinges, thus are regarded as potential Insulin pathway mediated wing dimorphism related genes (IWDRGs). Of the 45 IWDRGs, 5 were selected for verification by gene knockdown experiments, and played significant roles in the insect wing size regulation. CONCLUSIONS: We provided valuable insights on the genetic basis of wing dimorphism, and also demonstrated that network analysis is a powerful approach to identify new genes regulating wing dimorphic development via insulin signaling pathway in the migratory insect.


Assuntos
Genes de Insetos , Hemípteros/genética , Insulina/fisiologia , Asas de Animais/metabolismo , Processamento Alternativo , Animais , Ácidos Graxos/metabolismo , Feminino , Fatores de Transcrição Forkhead/metabolismo , Expressão Gênica , Redes Reguladoras de Genes , Hemípteros/anatomia & histologia , Hemípteros/metabolismo , Proteínas de Insetos/metabolismo , Proteínas Musculares/genética , Fenótipo , Transdução de Sinais , Asas de Animais/anatomia & histologia
5.
Int J Mol Sci ; 20(10)2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31096563

RESUMO

High immunogenicity and systemic toxicity are the main obstacles limiting the clinical use of the therapeutic agents based on Pseudomonas aeruginosa exotoxin A. In this work, we studied the immunogenicity, general toxicity and antitumor effect of the targeted toxin DARPin-LoPE composed of HER2-specific DARPin and a low immunogenic exotoxin A fragment lacking immunodominant human B lymphocyte epitopes. The targeted toxin has been shown to effectively inhibit the growth of HER2-positive human ovarian carcinoma xenografts, while exhibiting low non-specific toxicity and side effects, such as vascular leak syndrome and liver tissue degradation, as well as low immunogenicity, as was shown by specific antibody titer. This represents prospects for its use as an agent for targeted therapy of HER2-positive tumors.


Assuntos
Epitopos de Linfócito B/imunologia , Xenoenxertos , Imunotoxinas/imunologia , Imunotoxinas/farmacologia , Proteínas Musculares/imunologia , Proteínas Nucleares/imunologia , Neoplasias Ovarianas/tratamento farmacológico , Receptor ErbB-2/imunologia , ADP Ribose Transferases/imunologia , ADP Ribose Transferases/farmacologia , Sequência de Aminoácidos , Animais , Antineoplásicos/imunologia , Antineoplásicos/farmacologia , Toxinas Bacterianas/imunologia , Toxinas Bacterianas/farmacologia , Biomarcadores Tumorais , Carcinoma/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Epitopos de Linfócito B/genética , Exotoxinas/imunologia , Exotoxinas/farmacologia , Feminino , Humanos , Concentração Inibidora 50 , Fígado/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Terapia de Alvo Molecular , Proteínas Musculares/genética , Proteínas Nucleares/genética , Neoplasias Ovarianas/patologia , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/uso terapêutico , Baço/patologia , Fatores de Virulência/imunologia , Fatores de Virulência/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Muscle Nerve ; 60(2): 192-201, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31093982

RESUMO

INTRODUCTION: We recently demonstrated the beneficial effects of 4-aminopyridine (4-AP), a potassium channel blocker, in enhancing remyelination and recovery of nerve conduction velocity and motor function after sciatic nerve crush injury in mice. Although muscle atrophy occurs very rapidly after nerve injury, the effect of 4-AP on muscle atrophy and intrinsic muscle contractile function is largely unknown. METHODS: Mice were assigned to sciatic nerve crush injury and no-injury groups and were followed for 3, 7, and 14 days with/without 4-AP or saline treatment. Morphological, functional, and transcriptional properties of skeletal muscle were assessed. RESULTS: In addition to improving in vivo function, 4-AP significantly reduced muscle atrophy with increased muscle fiber diameter and contractile force. Reduced muscle atrophy was associated with attenuated expression of atrophy-related genes and increased expression of proliferating stem cells. DISCUSSION: These findings provide new insights into the potential therapeutic benefits of 4-AP against nerve injury-induced muscle atrophy and dysfunction. Muscle Nerve 60: 192-201, 2019.


Assuntos
4-Aminopiridina/farmacologia , Lesões por Esmagamento/fisiopatologia , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/patologia , Traumatismos dos Nervos Periféricos/fisiopatologia , Bloqueadores dos Canais de Potássio/farmacologia , Remielinização/efeitos dos fármacos , Nervo Isquiático/efeitos dos fármacos , Animais , Lesões por Esmagamento/metabolismo , Lesões por Esmagamento/patologia , Proteína Forkhead Box O1/efeitos dos fármacos , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O3/efeitos dos fármacos , Proteína Forkhead Box O3/genética , Camundongos , Proteínas Musculares/efeitos dos fármacos , Proteínas Musculares/genética , Músculo Esquelético/inervação , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Atrofia Muscular/genética , Traumatismos dos Nervos Periféricos/genética , Traumatismos dos Nervos Periféricos/patologia , Regeneração/efeitos dos fármacos , Nervo Isquiático/lesões , Nervo Isquiático/patologia , Nervo Isquiático/fisiopatologia , Proteínas com Motivo Tripartido/efeitos dos fármacos , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/efeitos dos fármacos , Ubiquitina-Proteína Ligases/genética
7.
Anim Sci J ; 90(8): 1018-1025, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31132809

RESUMO

Dietary fish oil intake improves muscle atrophy in several atrophy models however the effect on denervation-induced muscle atrophy is not clear. Thus, the aim of this study was to investigate the effects of dietary fish oil intake on muscle atrophy and the expression of muscle atrophy markers induced by sciatic nerve denervation in mice. We performed histological and quantitative mRNA expression analysis of muscle atrophy markers in mice fed with fish oil with sciatic nerve denervation. Histological analysis indicated that dietary fish oil intake slightly prevented the decrease of muscle fiber diameter induced by denervation treatment. In addition, dietary fish oil intake suppressed the MuRF1 (tripartite motif-containing 63) expression up-regulated by denervation treatment, and this was due to decreased tumor necrosis factor-alpha (TNF-α) production in skeletal muscle. We concluded that dietary fish oil intake suppressed MuRF1 expression by decreasing TNF-α production during muscle atrophy induced by sciatic nerve denervation in mice.


Assuntos
Denervação/efeitos adversos , Gorduras Insaturadas na Dieta/farmacologia , Óleos de Peixe/farmacologia , Expressão Gênica/efeitos dos fármacos , Proteínas Musculares/metabolismo , Atrofia Muscular/etiologia , Nervo Isquiático , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Masculino , Camundongos Endogâmicos C57BL , Fibras Musculares Esqueléticas/patologia , Proteínas Musculares/genética , Músculo Esquelético/metabolismo , Atrofia Muscular/prevenção & controle , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas com Motivo Tripartido/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Ubiquitina-Proteína Ligases/genética , Regulação para Cima/efeitos dos fármacos
8.
Mol Med Rep ; 19(6): 4919-4926, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30942422

RESUMO

Cervical cancer is a malignancy that threatens female health. The present study aimed to investigate the role of transgelin 2 (TAGLN2) in cervical cancer. Reverse transcription­quantitative polymerase chain reaction and western blotting were conducted to detect the mRNA and protein expression levels of particular factors in HeLa cells. Cell Counting kit­8, wound healing and Transwell assays were conducted to determine cell viability, and migratory and invasive abilities, respectively. The results demonstrated that the expression levels of TAGLN2 were decreased in cervical cancer tissues and were associated with the survival time of patients with cervical cancer. In addition, the expression of TAGLN2 was significantly reduced in three cervical cancer cell lines (HeLa, SiHa and C­33A) compared with in a normal cervical cell line. The present study also demonstrated that TAGLN2 overexpression in HeLa cells could inhibit cell viability, migration and invasion, and it was suggested that this may occur via upregulation of the expression levels of E­cadherin and inhibitor of nuclear factor κ­light­chain­enhancer of activated B cells (NF­κB) (IκB), and downregulation of C­X­C chemokine receptor type 4, matrix metalloproteinase (MMP)­2, MMP­9, p50 and transcription factor p65. In conclusion, TAGLN2 was revealed to inhibit cell viability, and the migratory and invasive abilities of HeLa cervical cancer cells via regulating the expression of metastasis­associated factors and the NF­κB signaling pathway. The present study proposed a novel target gene for the diagnosis, treatment and prognosis of cervical cancer.


Assuntos
Proteínas dos Microfilamentos/metabolismo , Proteínas Musculares/metabolismo , Neoplasias do Colo do Útero/patologia , Adulto , Idoso , Caderinas/metabolismo , Linhagem Celular , Movimento Celular , Sobrevivência Celular , Feminino , Células HeLa , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Proteínas dos Microfilamentos/genética , Pessoa de Meia-Idade , Proteínas Musculares/genética , NF-kappa B/metabolismo , Estadiamento de Neoplasias , Receptores CXCR4/metabolismo , Transdução de Sinais , Regulação para Cima , Neoplasias do Colo do Útero/metabolismo
9.
Biomed Pharmacother ; 112: 108681, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30970510

RESUMO

Acute kidney injury (AKI) is a significant medical problem worldwide. Ischemia-reperfusion (I/R) injury of the kidney is a major cause of AKI. However, the pathogenesis that contributes to renal I/R injury is still unclear. Apoptosis repressor with caspase recruitment domain (ARC) is abundantly expressed in various tissues, and has been reported to play a strong protective role during pathological processes. Our results indicated that ARC expression was decreased in the reperfused kidneys. ARC deficiency markedly accelerated renal dysfunction, promoted reperfusion-regulated tubular epithelial cell apoptosis, and enhanced the vulnerability of kidney to I/R damage. Furthermore, in the kidney samples of mice underwent renal I/R injury, ARC knockout significantly accelerated the expression levels of inflammatory factors, including interleukin (IL)-1ß, IL-6, tumor necrosis factor a (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and IL-2. In addition, renal I/R injury-induced apoptosis was further exacerbated in ARC-deficient mice through promoting the expression of cleaved Caspase-3 and poly (ADP-ribose) polymerase (PARP). From the molecular level, ARC deletion obviously accelerated mitochondrial injury, as evidenced by the further decreased adenosine triphosphate (ATP) levels and mitochondrial potential in hypoxia-reoxygenation (H/R)-treated cells. Moreover, ARC knockout exacerbated AKI through activating phosphorylated protein kinase B (AKT), mammalian target of Rapamycin (mTOR) and p53, whereas reducing phosphorylated glycogen synthase kinase 3ß (GSK3ß). Of note, blocking AKT/mTOR signaling markedly attenuated inflammation, mitochondrial damage and apoptosis stimulated by H/R in ARC knockdown cells. In summary, our results suggested that ARC played a pivotal role in the pathogenesis of AKI induced by renal I/R operation through regulating AKT/mTOR signaling.


Assuntos
Lesão Renal Aguda/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Apoptose , Rim/metabolismo , Proteínas Musculares/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Traumatismo por Reperfusão/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Lesão Renal Aguda/imunologia , Lesão Renal Aguda/patologia , Animais , Apoptose/imunologia , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular , Sobrevivência Celular , Citocinas/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Inflamação , Rim/imunologia , Rim/patologia , Túbulos Renais Proximais/citologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Musculares/genética , Traumatismo por Reperfusão/imunologia , Traumatismo por Reperfusão/patologia , Transdução de Sinais
10.
Gene ; 707: 36-43, 2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-30930226

RESUMO

Muscle LIM protein (MLP/CSRP3/CRP3) is a microtubule-associated protein preferentially expressed in cardiac and skeletal muscle and has a central role during muscle development and for architectural maintenance of muscle cells. LIM-domain proteins act as both modulators and downstream targets of TGF-ß signaling, which is well documented to negatively regulate differentiation of myogenic precursor cells or myoblasts. Herein, we determined whether CSRP3 regulates chicken satellite cell proliferation and differentiation in vitro, and examined its mechanism of action by focusing on the TGF-ß signaling pathway. Interference of CSRP3 mRNA expression had no effect on the proliferation of satellite cells, but significantly inhibited satellite cell differentiation into myotubes at 24, 48, and 72 h after initiation of differentiation. However, CSRP3 overexpression did not affect the proliferation or differentiation of satellite cells. Moreover, knockdown of CSRP3 caused up-regulation of TGF-ß and Smad3 mRNA and protein levels. The phosphorylation of Smad3 in CSRP3-knockdown cells was greater than that in wild-type cells at 24, 48, and 72 h after initiation of differentiation. Collectively, knockdown of CSRP3 suppressed chicken satellite cell differentiation by regulating Smad3 phosphorylation in the TGF-ß signaling pathway. Our results indicate that CSRP3 might play an important role in promoting satellite cell differentiation in chicken.


Assuntos
Proteínas com Domínio LIM/genética , Proteínas Musculares/genética , Células Satélites de Músculo Esquelético/citologia , Proteína Smad3/genética , Proteína Smad3/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Galinhas , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Fosforilação , Células Satélites de Músculo Esquelético/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo
11.
Nat Commun ; 10(1): 1791, 2019 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-30996251

RESUMO

Apoptotic death of cells damaged by genotoxic stress requires regulatory input from surrounding tissues. The C. elegans scaffold protein KRI-1, ortholog of mammalian KRIT1/CCM1, permits DNA damage-induced apoptosis of cells in the germline by an unknown cell non-autonomous mechanism. We reveal that KRI-1 exists in a complex with CCM-2 in the intestine to negatively regulate the ERK-5/MAPK pathway. This allows the KLF-3 transcription factor to facilitate expression of the SLC39 zinc transporter gene zipt-2.3, which functions to sequester zinc in the intestine. Ablation of KRI-1 results in reduced zinc sequestration in the intestine, inhibition of IR-induced MPK-1/ERK1 activation, and apoptosis in the germline. Zinc localization is also perturbed in the vasculature of krit1-/- zebrafish, and SLC39 zinc transporters are mis-expressed in Cerebral Cavernous Malformations (CCM) patient tissues. This study provides new insights into the regulation of apoptosis by cross-tissue communication, and suggests a link between zinc localization and CCM disease.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/fisiologia , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Hemangioma Cavernoso do Sistema Nervoso Central/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Zinco/metabolismo , Animais , Animais Geneticamente Modificados , Apoptose/efeitos da radiação , Proteínas Reguladoras de Apoptose/genética , Encéfalo/patologia , Encéfalo/cirurgia , Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/efeitos da radiação , Proteínas de Caenorhabditis elegans/genética , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Hemangioma Cavernoso do Sistema Nervoso Central/genética , Hemangioma Cavernoso do Sistema Nervoso Central/cirurgia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteína KRIT1/genética , Proteína KRIT1/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 7 Ativada por Mitógeno/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Mutagênese , Mutação , Fosforilação/fisiologia , Alinhamento de Sequência , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
12.
Skelet Muscle ; 9(1): 9, 2019 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-30992050

RESUMO

BACKGROUND: Critical illness myopathy (CIM) is associated with severe skeletal muscle wasting and impaired function in intensive care unit (ICU) patients. The mechanisms underlying CIM remain incompletely understood. To elucidate the biological activities occurring at the transcriptional level in the skeletal muscle of ICU patients with CIM, the gene expression profiles, potential upstream regulators, and enrichment pathways were characterized using RNA sequencing (RNA-seq). We also compared the skeletal muscle gene signatures in ICU patients with CIM and genes perturbed by mechanical loading in one leg of the ICU patients, with an aim of reducing the loss of muscle function. METHODS: RNA-seq was used to assess gene expression changes in tibialis anterior skeletal muscle samples from seven critically ill, immobilized, and mechanically ventilated ICU patients with CIM and matched control subjects. We also examined skeletal muscle gene expression for both legs of six ICU patients with CIM, where one leg was mechanically loaded for 10 h/day for an average of 9 days. RESULTS: In total, 6257 of 17,221 detected genes were differentially expressed (84% upregulated; p < 0.05 and fold change ≥ 1.5) in skeletal muscle from ICU patients with CIM when compared to control subjects. The differentially expressed genes were highly associated with gene changes identified in patients with myopathy, sepsis, long-term inactivity, polymyositis, tumor, and repeat exercise resistance. Upstream regulator analysis revealed that the CIM signature could be a result of the activation of MYOD1, p38 MAPK, or treatment with dexamethasone. Passive mechanical loading only reversed expression of 0.74% of the affected genes (46 of 6257 genes). CONCLUSIONS: RNA-seq analysis revealed that the marked muscle atrophy and weakness observed in ICU patients with CIM were associated with the altered expression of genes involved in muscle contraction, newly identified E3 ligases, autophagy and calpain systems, apoptosis, and chaperone expression. In addition, MYOD1, p38 MAPK, and dexamethasone were identified as potential upstream regulators of skeletal muscle gene expression in ICU patients with CIM. Mechanical loading only marginally affected the skeletal muscle transcriptome profiling of ICU patients diagnosed with CIM.


Assuntos
Apoptose , Autofagia , Chaperonas Moleculares/metabolismo , Contração Muscular , Músculo Esquelético/metabolismo , Doenças Musculares/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Estado Terminal , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Chaperonas Moleculares/genética , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Debilidade Muscular/genética , Debilidade Muscular/metabolismo , Atrofia Muscular/etiologia , Atrofia Muscular/metabolismo , Doenças Musculares/etiologia , Análise de Sequência de RNA , Transcriptoma , Ubiquitina-Proteína Ligases/genética
13.
Mol Biol (Mosk) ; 53(1): 74-83, 2019.
Artigo em Russo | MEDLINE | ID: mdl-30895954

RESUMO

This work studied the changes in the levels of the main proteins of the calpain system (µ-calpain, Ca^(2+)-dependent protease, and fragments of its autolysis, inhibitor calpastatin) and µ-calpain substrates (giant proteins of the sarcomere cytoskeleton, titin and nebulin) in skeletal muscle (m. gastrocnemius, m. soleus, m. longissimus dorsi) of rats alcoholized for three months by different methods using agar containing 30% ethanol and nutrient-balanced liquid feed containing 5% ethanol using gel electrophoresis methods under denaturing conditions and immunoblotting. No decrease in the muscle mass/body weight ratio, indicating the development of atrophy, no increase in autolysis of µ-calpain, indicating an increase in the activity of this enzyme, no changes in the content of intact titin (T1), nebulin, µ-calpain and calpastatin, as well as the total calpain activity measured using Calpain Activity Assay Kit were detected in alcoholized rats of both groups. No changes in the total level of titin phosphorylation in the rat muscles of alcoholized groups were detected using Pro-Q Diamond fluorescent dye for phosphate groups of proteins. No statistically significant differences in the content of titin and nebulin mRNA in skeletal muscles of control rats and rats alcoholized using agar were detected. In rats, alcoholized by the method of liquid feed, the levels of titin and nebulin mRNA were increased 1.5-2.5 times possibly due to a higher fat content in such a diet. The presented data may be useful for choosing a chronic alcoholization model for animals.


Assuntos
Alcoolismo/genética , Conectina/genética , Proteínas Musculares/genética , Músculo Esquelético/metabolismo , Animais , Modelos Animais de Doenças , Ratos
14.
Pain ; 160(5): 1166-1174, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30913166

RESUMO

Recent studies have made significant progress in identifying distinct populations of peripheral neurons involved in itch transmission, whereas the cellular identity of spinal interneurons that contribute to itch processing is still a debate. Combining genetic and pharmacological ablation of spinal excitatory neuronal subtypes and behavioral assays, we demonstrate that spinal somatostatin-positive (SOM) excitatory interneurons transmit pruritic sensation. We found that the ablation of spinal SOM/Lbx1 (SOM) neurons caused significant attenuation of scratching responses evoked by various chemical pruritogens (chemical itch). In an attempt to identify substrates of spinal itch neural circuit, we observed that spinal SOM neurons partially overlapped with neurons expressing natriuretic peptide receptor A (Npra), the receptor of peripheral itch transmitter B-type natriuretic peptide. Spinal SOM neurons, however, did not show any overlap with itch transmission neurons expressing gastrin-releasing peptide receptor in the dorsal spinal cord, and the gastrin-releasing peptide-triggered scratching responses were intact after ablating spinal SOM neurons. Dual ablation of SOM and Npra neurons in the spinal cord reduced chemical itch responses to a greater extent than ablation of SOM or Npra neurons alone, suggesting the existence of parallel spinal pathways transmitting chemical itch. Furthermore, we showed that SOM peptide modulated itch processing through disinhibition of somatostatin receptor 2A-positive inhibitory interneuron. Together, our findings reveal a novel spinal mechanism for sensory encoding of itch perception.


Assuntos
Interneurônios/metabolismo , Prurido/induzido quimicamente , Prurido/patologia , Somatostatina/metabolismo , Medula Espinal/patologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/genética , Inibidores da Angiogênese/farmacologia , Animais , Cloroquina/toxicidade , Modelos Animais de Doenças , Técnicas In Vitro , Interneurônios/fisiologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Lisina/análogos & derivados , Lisina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Nitrobenzoatos/farmacologia , Técnicas de Patch-Clamp , Proteínas Proto-Oncogênicas c-fos/metabolismo , Somatostatina/genética , Medula Espinal/efeitos dos fármacos , Medula Espinal/metabolismo , p-Metoxi-N-metilfenetilamina/toxicidade , Proteínas tau/genética , Proteínas tau/metabolismo
15.
Arch Anim Nutr ; 73(2): 75-87, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30821190

RESUMO

The present study investigated the hypothesis that dietary concentrations of leucine (Leu) in excess of the breeder´s recommendations activates protein synthesis and decreases protein degradation in muscle of broilers. Day-old male Ross 308 broilers (n = 450) were phase-fed corn-soybean meal-based diets during starter (d 1-10), grower (d 11-22), and finisher (d 23-34) period. The basal diets fed to the control group (L0) met the broilers' requirements for nutrients and amino acids, and contained Leu, Leu:isoleucine (Ile) and Leu:valine (Val) ratios, close to those recommended by the breeder (Leu:Ile: 100:54, 100:52, 100:51; Leu:Val 100:64, 100:61, 100:58; in starter, grower and finisher diet, resp.). Basal diets were supplemented with Leu to exceed the breeder's recommendations by 35% (group L35) and 60% (group L60). Growth performance during 34 d, and carcass weights, and breast and thigh muscle weights on d 34 were similar among groups. Hepatic and muscle mRNA levels of genes involved in the somatotropic axis [growth hormone receptor, insulin-like growth factor (IGF)-1, IGF binding protein 2, IGF receptor] on d 34 were not influenced by Leu. In the breast muscle, relative mRNA abundances of genes involved in the mammalian target of rapamycin (mTOR) pathway of protein synthesis (mTOR, ribosomal p70 S6 kinase) and the ubiquitin-proteasome pathway of protein degradation (F-box only protein 32, Forkhead box protein O1, Muscle RING-finger protein-1) on d 34 were largely similar among groups. Likewise, relative phosphorylation and thus activation of mTOR and ribosomal protein S6 involved in the mTOR pathway, and of eukaryotic translation initiation factor 2A (eIF2a) involved in the general control nonderepressible 2 (GCN2)/eIF2a pathway of protein synthesis inhibition, were not influenced. These data indicate that dietary Leu concentrations exceeding the broiler´s requirements up to 60% neither influence protein synthesis nor degradation pathways nor muscle growth in growing broilers.


Assuntos
Galinhas/fisiologia , Isoleucina/farmacologia , Leucina/farmacologia , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Valina/farmacologia , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Animais , Composição Corporal , Dieta/veterinária , Suplementos Nutricionais , Regulação da Expressão Gênica/efeitos dos fármacos , Isoleucina/administração & dosagem , Leucina/administração & dosagem , Masculino , Proteínas Musculares/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Valina/administração & dosagem , Ganho de Peso
16.
J Biol Chem ; 294(16): 6364-6374, 2019 04 19.
Artigo em Inglês | MEDLINE | ID: mdl-30819805

RESUMO

The formation of new myofibers in vertebrates occurs by myoblast fusion and requires fusogenic activity of the muscle-specific membrane protein myomaker. Here, using in silico (BLAST) genome analyses, we show that the myomaker gene from trout includes 14 minisatellites, indicating that it has an unusual structure compared with those of other animal species. We found that the trout myomaker gene encodes a 434-amino acid (aa) protein, in accordance with its apparent molecular mass (∼40 kDa) observed by immunoblotting. The first half of the trout myomaker protein (1-220 aa) is similar to the 221-aa mouse myomaker protein, whereas the second half (222-234 aa) does not correspond to any known motifs and arises from two protein extensions. The first extension (∼70 aa) apparently appeared with the radiation of the bony fish clade Euteleostei, whereas the second extension (up to 236 aa) is restricted to the superorder Protacanthopterygii (containing salmonids and pike) and corresponds to the insertion of minisatellites having a length of 30 nucleotides. According to gene expression analyses, trout myomaker expression is consistently associated with the formation of new myofibers during embryonic development, postlarval growth, and muscle regeneration. Using cell-mixing experiments, we observed that trout myomaker has retained the ability to drive the fusion of mouse fibroblasts with C2C12 myoblasts. Our work reveals that trout myomaker has fusogenic function despite containing two protein extensions.


Assuntos
Proteínas de Peixes , Regulação da Expressão Gênica/fisiologia , Proteínas de Membrana , Repetições Minissatélites , Proteínas Musculares , Oncorhynchus mykiss , Animais , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Miofibrilas/metabolismo , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/metabolismo
17.
Zhonghua Zhong Liu Za Zhi ; 41(2): 91-96, 2019 Feb 23.
Artigo em Chinês | MEDLINE | ID: mdl-30862136

RESUMO

Objective: To investigate the expression of microRNA-133b (miR-133b) in esophageal squamous cell carcinoma (ESCC), and explore its effect and the underlying molecular mechanisms on cell proliferation and invasion. Methods: Real-time quantitative PCR (qPCR) was used to examine miR-133b expression in 63 ESCC tissues and paired adjacent non-cancerous tissues, several ESCC cells (Eca109, EC9706, EC1, TE1, KYSE70) and normal esophageal epithelial cell Het-1A. MiR-133b mimic, inhibitor and negative control (NC) were transfected into TE1 cells. The effect of miR-133b on cell proliferation and invasion were determined by CCK-8 and Transwell assays, respectively. Subsequently, the target gene of miR-133b was predicted by online tools TargetScan and miRDB, which was verified by dual luciferase reporter assays. Finally, Western blot was utilized to detect the effects of miR-133b overexpression on expression of target gene TAGLN2 as well as EMT-related proteins E-cadherin, N-cadherin, Snail, Slug and Vimentin. Results: Relative levels of miR-133b in ESCC tissues (0.295±0.040) were significantly lower than those in adjacent non-cancerous tissues (1.002±0.011, P<0.001). The expression of miR-133b was tightly associated with clinical staging, lymph node metastasis and prognosis. Moreover, relative levels of miR-133b in ESCC cells Eca109, EC9706, EC1, TE1 and KYSE70 (0.679±0.031, 0.391±0.008, 0.236±0.016, 0.031±0.005 and 0.099±0.020) were evidently lower than that in normal esophageal epithelial cell Het-1A (1.005±0.016, all P<0.001). In TE1 cells, miR-133b mimic significantly increased the level of miR-133b to 6.199±0.627, and suppressed cell proliferation and invasion, whereas miR-133b inhibitor obviously decreased its expression to 0.182±0.023, and promoted cell proliferation and invasion. Most notably, the relative luciferase activities of miR-133b-mimic group (0.320±0.018) in TE1 cells transfected with TAGLN-3'UTR-WT were markedly lower than that in NC group (1.010±0.036, P<0.001), whereas those in TAGLN-3'UTR-MUT transfection cells were 1.019±0.056 and 1.008±0.021, respectively, showing no significantly statistical difference (P>0.05). Furthermore, miR-133b overexpression markedly downregulated TAGLN2, N-cadherin, Snail, Slug and Vimentin levels, and increased E-cadherin expression. Conclusion: MiR-133b plays an important role in the proliferation and invasion of ESCC cells by regulating TAGLN2 expression, and it may be a potential therapeutic target for ESCC patients.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/patologia , MicroRNAs/metabolismo , Proteínas dos Microfilamentos/metabolismo , Proteínas Musculares/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas dos Microfilamentos/genética , Proteínas Musculares/genética , Invasividade Neoplásica , Sincalida/metabolismo
18.
Mol Vis ; 25: 93-105, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30820145

RESUMO

Purpose: To investigate the genetic basis of primary closed angle glaucoma (PCAG) in European Basset Hounds using genome-wide association and RNA sequencing strategies. Methods: DNA samples from 119 European Basset Hounds were genotyped on the 170 K SNP CanineHD BeadChip array (Illumina) comprising 37 with normal iridocorneal angles (controls), 57 with pectinate ligament abnormality (PLA cases), and 25 with PCAG (PCAG cases). Genome-wide association studies (GWASs) of the PLA and PCAG cases were conducted. Whole transcriptome sequences of iridocorneal angle tissues from five Basset Hounds with PCAG were compared with those from four dogs with normal eyes to investigate differences in gene expression between the affected and unaffected eyes in GWAS-associated loci. A variant in NEB, previously reported to be associated with PCAG in American Basset Hounds, was genotyped in cohorts of European Basset Hounds and non-Basset Hounds. Results: The GWASs revealed 1.4 and 0.2 Mb regions, on chromosomes 24 and 37, respectively, that are statistically associated with PCAG. The former locus has previously been associated with glaucoma in humans. Whole transcriptome analysis revealed differential gene expression of eight genes within these two loci. The NEB variant was not associated with PLA or PCAG in this set of European Basset Hounds. Conclusions: We identified two novel loci for canine PCAG. Further investigation is required to elucidate candidate variants that underlie canine PCAG.


Assuntos
Doenças do Cão/genética , Proteínas do Olho/genética , Predisposição Genética para Doença , Genoma , Glaucoma de Ângulo Fechado/veterinária , Transcriptoma , Animais , Estudos de Casos e Controles , Doenças do Cão/patologia , Cães , Europa (Continente) , Proteínas do Olho/metabolismo , Feminino , Ontologia Genética , Loci Gênicos , Estudo de Associação Genômica Ampla , Glaucoma de Ângulo Fechado/genética , Glaucoma de Ângulo Fechado/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Masculino , Anotação de Sequência Molecular , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Polimorfismo de Nucleotídeo Único , Análise de Sequência de RNA , Estados Unidos
19.
Int J Mol Sci ; 20(4)2019 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-30795533

RESUMO

BACKGROUND: Skin cancer represents the most common human malignancy, and it includes BCC, SCC, and melanoma. Since melanoma is one of the most aggressive types of cancer, we have herein attempted to develop a gene-specific intron retention signature that can distinguish BCC and SCC from melanoma biopsy tumors. METHODS: Intron retention events were examined through RT-sqPCR protocols, using total RNA preparations derived from BCC, SCC, and melanoma Greek biopsy specimens. Intron-hosted miRNA species and their target transcripts were predicted via the miRbase and miRDB bioinformatics platforms, respectively. Ιntronic ORFs were recognized through the ORF Finder application. Generation and visualization of protein interactomes were achieved by the IntAct and Cytoscape softwares, while tertiary protein structures were produced by using the I-TASSER online server. RESULTS: c-MYC and Sestrin-1 genes proved to undergo intron retention specifically in melanoma. Interaction maps of proteins encoded by genes being potentially targeted by retained intron-accommodated miRNAs were generated and SRPX2 was additionally delivered to our melanoma-specific signature. Novel ORFs were identified in MCT4 and Sestrin-1 introns, with potentially critical roles in melanoma development. CONCLUSIONS: The property of c-MYC, Sestrin-1, and SRPX2 genes to retain specific introns could be clinically used to molecularly differentiate non-melanoma from melanoma tumors.


Assuntos
Testes Genéticos/métodos , Melanoma/genética , Processamento de RNA , Neoplasias Cutâneas/genética , Idoso , Idoso de 80 Anos ou mais , Diagnóstico Diferencial , Feminino , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Íntrons , Masculino , Melanoma/patologia , Pessoa de Meia-Idade , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Neoplasias Cutâneas/patologia
20.
Nat Commun ; 10(1): 845, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30783087

RESUMO

Cell metabolism is strongly influenced by mechano-environment. We show here that a fraction of kindlin-2 localizes to mitochondria and interacts with pyrroline-5-carboxylate reductase 1 (PYCR1), a key enzyme for proline synthesis. Extracellular matrix (ECM) stiffening promotes kindlin-2 translocation into mitochondria and its interaction with PYCR1, resulting in elevation of PYCR1 level and consequent increase of proline synthesis and cell proliferation. Depletion of kindlin-2 reduces PYCR1 level, increases reactive oxygen species (ROS) production and apoptosis, and abolishes ECM stiffening-induced increase of proline synthesis and cell proliferation. In vivo, both kindlin-2 and PYCR1 levels are markedly increased in lung adenocarcinoma. Ablation of kindlin-2 in lung adenocarcinoma substantially reduces PYCR1 and proline levels, and diminishes fibrosis in vivo, resulting in marked inhibition of tumor growth and reduction of mortality rate. Our findings reveal a mechanoresponsive kindlin-2-PYCR1 complex that links mechano-environment to proline metabolism and signaling, and suggest a strategy to inhibit tumor growth.


Assuntos
Adenocarcinoma de Pulmão/metabolismo , Proteínas do Citoesqueleto/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Musculares/metabolismo , Proteínas de Neoplasias/metabolismo , Prolina/biossíntese , Células A549 , Adenocarcinoma de Pulmão/patologia , Animais , Proliferação de Células/fisiologia , Sobrevivência Celular/fisiologia , Proteínas do Citoesqueleto/genética , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Proteínas de Membrana/genética , Camundongos Transgênicos , Mitocôndrias/metabolismo , Proteínas Musculares/genética , Proteínas de Neoplasias/genética , Pirrolina Carboxilato Redutases/genética , Pirrolina Carboxilato Redutases/metabolismo
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