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1.
Adv Exp Med Biol ; 1185: 227-231, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31884616

RESUMO

Pre-mRNA splicing is a critical step in RNA processing in all eukaryotic cells. It consists of introns removal and requires the assembly of a large RNA-protein complex called the spliceosome. This complex of small nuclear ribonucleoproteins is associated with accessory proteins from the pre-mRNA processing factor (PRPF) family. Mutations in different splicing factor-encoding genes were identified in retinitis pigmentosa (RP) patients. A surprising feature of these ubiquitous factors is that the outcome of their alteration is restricted to the retina. Because of their high metabolic demand, most studies focused on photoreceptors dysfunction and associated degeneration. However, cells from the retinal pigment epithelium (RPE) are also crucial to maintaining retinal homeostasis and photoreceptor function. Moreover, mutations in RPE-specific genes are associated with some RP cases. Indeed, we identified major RPE defects in Prpf31-mutant mice: circadian rhythms of both photoreceptor outer segments (POS) phagocytosis and retinal adhesion were attenuated or lost, leading to ultrastructural anomalies and vacuoles. Taken together, our published and ongoing data suggest that (1) similar molecular events take place in human and mouse cells and (2) these functional defects generate various stress processes.


Assuntos
Células Epiteliais/patologia , Proteínas do Olho/genética , Retinite Pigmentosa/genética , Animais , Ritmo Circadiano , Células Epiteliais/ultraestrutura , Humanos , Camundongos , Fagocitose , Células Fotorreceptoras de Vertebrados/patologia , Fatores de Processamento de RNA/genética , Epitélio Pigmentado da Retina/citologia , Retinite Pigmentosa/patologia
2.
Toxicol Lett ; 317: 92-101, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31593750

RESUMO

Cigarette smoke (CS) is known to cause mitochondrial dysfunction leading to cellular senescence in lung cells. We determined the mechanism of mitochondrial dysfunction by CS in lung epithelial cells. CS extract (CSE) treatment differentially affected mitochondrial function, such as membrane potential, mitochondrial reactive oxygen species (mtROS) and mitochrondrial mass as analyzed by FACS, and were associated with altered oxidative phosphorylation (OXPHOS) protein levels (Complexes I-IV) in primary lung epithelial cells (SAEC and NHBE), and (complexes I and II) in BEAS2B cells. There were dose- and time-dependent changes in mitochondrial respiration (oxygen consumption rate parameters i.e. maximal respiration, ATP production and spare capacity, measured by the Seahorse analyzer) in control vs. CSE treated BEAS2B and NHBE/DHBE cells. Electron microscopy (EM) analysis revealed perinuclear clustering by localization and increased mitochondrial fragmentation by fragement length analysis. Immunoblot analysis revealed CS-mediated increase in Drp1 and decrease in Mfn2 levels that are involved in mitochondrial fission/fusion process. CSE treatment reduced Miro1 and Pink1 abundance that play a crucial role in the intercellular transfer mechanism and mitophagy process. Overall, these findings highlight the role of Miro1 in context of CS-induced mitochondrial dysfunction in lung epithelial cells that may contribute to the pathogenesis of chronic inflammatory lung diseases.


Assuntos
Fumar Cigarros/efeitos adversos , Células Epiteliais/metabolismo , Pulmão/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Doença Pulmonar Obstrutiva Crônica/etiologia , Fumaça/efeitos adversos , Proteínas rho de Ligação ao GTP/metabolismo , Estudos de Casos e Controles , Células Cultivadas , Regulação para Baixo , Metabolismo Energético , Células Epiteliais/ultraestrutura , Humanos , Pulmão/ultraestrutura , Mitocôndrias/ultraestrutura , Estresse Oxidativo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Transdução de Sinais
3.
Toxicol Lett ; 317: 1-12, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31562913

RESUMO

During extrusion of some polymers, fused filament fabrication (FFF) 3-D printers emit billions of particles per minute and numerous organic compounds. The scope of this study was to evaluate FFF 3-D printer emission-induced toxicity in human small airway epithelial cells (SAEC). Emissions were generated from a commercially available 3-D printer inside a chamber, while operating for 1.5 h with acrylonitrile butadiene styrene (ABS) or polycarbonate (PC) filaments, and collected in cell culture medium. Characterization of the culture medium revealed that repeat print runs with an identical filament yield various amounts of particles and organic compounds. Mean particle sizes in cell culture medium were 201 ±â€¯18 nm and 202 ±â€¯8 nm for PC and ABS, respectively. At 24 h post-exposure, both PC and ABS emissions induced a dose dependent significant cytotoxicity, oxidative stress, apoptosis, necrosis, and production of pro-inflammatory cytokines and chemokines in SAEC. Though the emissions may not completely represent all possible exposure scenarios, this study indicate that the FFF could induce toxicological effects. Further studies are needed to quantify the detected chemicals in the emissions and their corresponding toxicological effects.


Assuntos
Resinas Acrílicas/toxicidade , Butadienos/toxicidade , Células Epiteliais/efeitos dos fármacos , Nanopartículas/toxicidade , Cimento de Policarboxilato/toxicidade , Poliestirenos/toxicidade , Impressão Tridimensional , Mucosa Respiratória/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Humanos , Mediadores da Inflamação/metabolismo , Necrose , Estresse Oxidativo/efeitos dos fármacos , Tamanho da Partícula , Mucosa Respiratória/metabolismo , Mucosa Respiratória/ultraestrutura , Medição de Risco , Fatores de Tempo
4.
Toxicol Lett ; 316: 49-59, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31520698

RESUMO

Epidemiological studies have established the correlations between PM2.5 and a wide variety of pulmonary diseases. However, their underlying pathogeneses have not been clearly elucidated yet. In the present study, the epithelial-mesenchymal transition (EMT) phenotype with enhanced proliferation and migration activity of human pulmonary epithelial cell line BEAS-2B was observed after exposure to low dose PM2.5 exposure (50 µg/ml) for 30 passages. Then, epithelial cells derived-exosomal micro-RNA (miRNA) and intracellular total RNA were extracted, and the differentially expressed exosomal miRNAs (DE-Exo-MiRs) as well as differentially expressed protein coding genes (DEGs) were identified by RNA sequencing (RNA-seq) and transcriptome analysis. We found that chronic PM2.5 exposure stimulated the release of pulmonary epithelium derived exosomes. 45 DE-Exo-MiRs including 32 novelly predicted miRNAs and 843 DEGs between PM2.5 exposed group and the normal control were detected. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that DEGs were significantly enriched in extracellular matrix organization, focal adhesion and cancer related terms. Besides, the enrichment analyses on 7774 mRNA targets of 27 DE-Exo-MiRs predicted by MiRanda software also revealed the potential regulatory role of exosomal miRNAs in pathways in cancer, Wingless/Integrated (Wnt) signaling pathway, focal adhesion related genes and other multiple pathogenic pathways. Moreover, the interactive exosomal miRNA-mRNA pair networks were constructed using Cytoscape software. Our results provided a novel basis for a better understanding of the mechanisms of chronic PM2.5 exposure induced pulmonary disorders including pulmonary fibrosis and cancer, in which exosomal miRNAs (Exo-MiRs) potentially functions by dynamically regulating gene expressions.


Assuntos
Células Epiteliais/efeitos dos fármacos , Exossomos/efeitos dos fármacos , Perfilação da Expressão Gênica/métodos , Pulmão/efeitos dos fármacos , MicroRNAs/genética , Material Particulado/toxicidade , RNA Mensageiro/genética , Transcriptoma/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Biologia Computacional , Bases de Dados Genéticas , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Exossomos/genética , Exossomos/metabolismo , Exossomos/patologia , Redes Reguladoras de Genes , Humanos , Pulmão/metabolismo , Pulmão/ultraestrutura , MicroRNAs/metabolismo , Tamanho da Partícula , RNA Mensageiro/metabolismo , Medição de Risco , Fatores de Tempo , Testes de Toxicidade Crônica
5.
J Recept Signal Transduct Res ; 39(3): 235-242, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31538845

RESUMO

Renal tubular epithelial cell (RTEC) injury is the main cause and common pathological process of various renal diseases. Mitochondrial dysfunction (MtD) is a pathological process after renal injury. Mitophagy is vital for mitochondrial function. Hypoxia is a common cause of RTEC injury. Peroxisome proliferator-activated receptor γ (PPARγ) is involved in cell proliferation, apoptosis, and inflammation. Previous studies have shown that the low expression of PPARγ might be involved in hypoxia-induced RTEC injury. The present study aimed to investigate the correlation between PPARγ and mitophagy in damaged RTEC in the hypoxia/reoxygenation (HR) model. The results showed that HR inhibited the expression of PPARγ, but increased the expression of LC3II, Atg5, SQSTM1/P62, and PINK1 in a time-dependent manner. Moreover, mitochondrial DNA (mt DNA) copy number, mitochondria membrane potential (MMP) levels, ATP content, and cell viability were decreased in hypoxic RTECs, the expression of SQSTM1/P62 and PINK1, the release of cytochrome c (cyt C), and production of reactive oxygen species (ROS) were increased. Mitochondrial-containing autophagosomes (APs) were detected using transmission election microscope (TEM) and laser scanning confocal microscope (LSCM). Furthermore, PPARγ protein expression was negatively correlated with that of LC3II, PINK1, and the positive rate of RTEC-containing mitochondrial-containing APs (all p < .05), but positively correlated with cell viability, MMP level, and ATP content (all p < .05). These data suggested that PPARγ and mitophagy are involved in the RTEC injury process. Thus, a close association could be detected between PPARγ and mitophagy in HR-induced RTEC injury, albeit additional investigation is imperative.


Assuntos
Células Epiteliais/patologia , Túbulos Renais/patologia , PPAR gama/metabolismo , Animais , Autofagia , Hipóxia Celular , Linhagem Celular , Sobrevivência Celular , Células Epiteliais/ultraestrutura , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Mitocôndrias/ultraestrutura , Oxigênio , PPAR gama/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos
6.
Physiol Int ; 106(3): 225-235, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31560236

RESUMO

OBJECTIVES: Impaired intestinal barrier function has been demonstrated in the pathophysiology of diarrhea-predominant irritable bowel syndrome (IBS-D). This study aimed to describe the intestinal ultrastructural findings in the intestinal mucosal layer of IBS-D patients. METHODS: In total, 10 healthy controls and 10 IBS-D patients were analyzed in this study. The mucosa of each patient's rectosigmoid colon was first assessed by confocal laser endomicroscopy (CLE); next, biopsied specimens of these sites were obtained. Intestinal tissues of IBS-D patients and healthy volunteers were examined to observe cellular changes by transmission electron microscopy (TEM). RESULTS: CLE showed no visible epithelial damage or inflammatory changes in the colonic mucosa of IBS-D compared with healthy volunteers. On transmission electron microscopic examination, patients with IBS-D displayed a larger apical intercellular distance with a higher proportion of dilated (>20 nm) intercellular junctional complexes, which was indicative of impaired mucosal integrity. In addition, microvillus exfoliation, extracellular vesicle as well as increased presence of multivesicular bodies were visible in IBS-D patients. Single epithelial cells appeared necrotic, as characterized by cytoplasmic vacuolization, cytoplasmic swelling, and presence of autolysosome. A significant association between bowel habit, frequency of abdominal pain, and enlarged intercellular distance was found. CONCLUSION: This study showed ultrastructural alterations in the architecture of intestinal epithelial cells and intercellular junctional complexes in IBS-D patients, potentially representing a pathophysiological mechanism in IBS-D.


Assuntos
Diarreia/patologia , Mucosa Intestinal/ultraestrutura , Síndrome do Intestino Irritável/patologia , Dor Abdominal/patologia , Colo Sigmoide/ultraestrutura , Citoplasma/patologia , Citoplasma/ultraestrutura , Células Epiteliais/patologia , Células Epiteliais/ultraestrutura , Feminino , Humanos , Junções Intercelulares/ultraestrutura , Masculino , Pessoa de Meia-Idade , Reto/patologia , Reto/ultraestrutura
7.
Biomed Pharmacother ; 118: 109363, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31545277

RESUMO

OBJECTIVE: Alveolar epithelial barrier dysfunction in response to inflammatory reaction contributes to pulmonary edema in acute lung injury(ALI).Irisin,a newly-found myokine,exerts the anti-inflammatory effects. This study aims to investigate the protective effects of irisin on lipopolysaccharide (LPS)-induced ALIin vivo and in vitro, and to explore its underlying mechanism. METHODS: Male SD rats and A549 cells were divided into 4 groups: control group, LPS group, Irisin pretreated group, and Irisin/Compound C(a special inhibitor of AMPK)-treated group. The ALI model was established by intravenous injection of LPS in rats, and LPS challenge in A549 cells. Pulmonary specimens were harvested for microscopic examination of the pathological changes, and the expression of AMPK,SIRT1,NF-κB, p66Shc and caspase-3 in lung tissues. The pulmonary permeability were examined by wet/dry lung weight ratio(W/D) and lung permeability index(LPI). The apoptotic index, and the expression of tumor necrosis factor-α(TNF-α), interleukin-1ß(IL-1ß), monocyte chemoattractant activating protein-1 (MCP-1), tight junctions (occludin,ZO-1) were determined both in lung tissue and A549 cells. RESULTS: Irisin alleviated lung histological changes and decreased pulmonary microvascular permeability in LPS-induced rats. Irisin up-regulated the expression of occludin, ZO-1,AMPK,SIRT1, down-regulated the expression of TNF-α,IL-1ß,MCP-1,NF-κB, p66Shc caspase-3, and decreased the apoptotic index in LPS-induced rats and A549 cells. All these protective effects of irisin could be reversed by Compound C. CONCLUSION: Irisin improved LPS-induced alveolar epithelial barrier dysfunction via suppressing inflammation and apoptosis, and this protective effect might be mediated by activating AMPK/SIRT1 pathways.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Lesão Pulmonar Aguda/fisiopatologia , Epitélio/fisiopatologia , Fibronectinas/uso terapêutico , Pulmão/fisiopatologia , Transdução de Sinais , Sirtuína 1/metabolismo , Células A549 , Lesão Pulmonar Aguda/patologia , Animais , Apoptose/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Células Epiteliais/ultraestrutura , Epitélio/ultraestrutura , Fibronectinas/farmacologia , Humanos , Inflamação/patologia , Pulmão/patologia , Pulmão/ultraestrutura , Masculino , Permeabilidade , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo , Regulação para Cima/efeitos dos fármacos
8.
Toxicol Lett ; 316: 10-19, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31476341

RESUMO

Rapid risk assessment models for different types of cigarette smoke extract (CSE) exposure are critical to understanding the etiology of chronic obstructive pulmonary disease. The present study investigated inflammation of cultured tracheal tissues with CSE exposure. Rat trachea rings were isolated, cultured, then exposed to various concentrations of CSE from 3R4 F reference cigarettes for 4 h. Tissue/cellular morphology, ultrastructure, viability and damage, inflammatory cell infiltration, and inflammatory protein levels were measured and compared to untreated controls. Human bronchial epithelial cells (BEAS-2B) exposed to 0 or 300 µg/mL CSE were cocultured with macrophages to assess extent of mobilization and phagocytosis. Endotracheal epithelium cilia densities were significantly reduced with increasing CSE concentrations, while mucous membranes became increasingly disordered; both eventually disappeared. Macrophages became larger as the CSE concentration increased, with microvilli and extended pseudopodium covering their surface, and many primary and secondary lysosomes present in the cytoplasm. Inflammatory cell infiltration also increased with increasing CSE dose, as did intracellular adhesion molecule-1(ICAM-1), interleukin-6(IL-6). The method described here may be useful to qualitatively characterized the effects of the compound under study. Then, we use BEAS-2B cell line system to strength the observation made in the cultured tissues. Probably, an approach to integrate results from both experiments will facilitate its application. These results demonstrate that cultured rat tracheal rings have a whole-tissue structure that undergoes inflammatory processes similar to in vivo tissues upon CSE exposure.


Assuntos
Células Epiteliais/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Doença Pulmonar Obstrutiva Crônica/etiologia , Fumaça/efeitos adversos , Fumar/efeitos adversos , Tabaco/efeitos adversos , Traqueia/efeitos dos fármacos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Humanos , Mediadores da Inflamação/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-6/metabolismo , Macrófagos/metabolismo , Macrófagos/ultraestrutura , Masculino , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Ratos Sprague-Dawley , Medição de Risco , Fatores de Tempo , Técnicas de Cultura de Tecidos , Traqueia/metabolismo , Traqueia/ultraestrutura
9.
Micron ; 126: 102747, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31505373

RESUMO

Despite the exploration of mitochondria-rich cells (MRCs) in different animal classes, very limited information has been documented about MRCs in reptiles. The present study was designed to investigate the effect of seasonal variation on the cell ultrastructure and ion transport protein activity of MRCs during hibernation and non-hibernation of Chinese soft-shelled turtle's intestine. Transmission electron microscopy revealed that, during hibernation the high-density cytoplasm of MRCs occupied large cross-sectional area and showed heterogeneous abundance of mitochondria and an expanded extensive tubular system as compared to non-hibernation. During hibernation the cytoplasm of MRCs exhibited more mitochondrial vacuolization, autophagosomes, phagophore formation and well-structured endoplasmic reticulum. During hibernation, MRCs connected with absorptive cells through wide interdigitation, and created tight junction and more desmosomes as compared to non-hibernation. Immunohistochemistry and immunofluorescence showed, the strong immunopositive reactions and immunosignaling of Na+/K+-ATPase (NKA) and Na+/K+/2Cl- cotransporter (NKCC) at basolateral region of mucosal surface of intestine during hibernation. However, weak immunopositive reactions and immunosignaling of NKA and NKCC during non-hibernation. The statistical analysis showed that the number and size of MRCs with NKA-associated immunoreactivity were significantly increased during hibernation. NKA and NKCC mRNA expression was significantly increased during hibernation via qPCR. Further confirmed, the intensity of NKA and NKCC proteins was more elevated during hibernation than non-hibernation shown by immunobloting. However, the concentrations of the plasma ions Na+ and Cl- were significantly higher during hibernation; conversely, K+ concentration was significantly higher during non-hibernation. The findings suggest that the potential role of MRCs is affected by seasonal fluctuations, during which intestinal homeostasis and hydromineral balance are essential for turtles.


Assuntos
Células Epiteliais/ultraestrutura , Intestino Delgado/citologia , Mitocôndrias/enzimologia , Estações do Ano , ATPase Trocadora de Sódio-Potássio/química , Tartarugas , Animais , Hibernação , Microscopia Eletrônica de Transmissão
10.
Environ Toxicol Pharmacol ; 71: 103217, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31284173

RESUMO

Ultrastructural and histopathological reponses in the organs of living organisms are important and useful tools to determine the health condition and the effects of pollutants, such as pesticides, on the organisms. The aim of this study is to determine possible histopathological, cytopathological and ultrastructural alterations in gills of Oreochromis niloticus individuals exposed to 850 µg/L carbaryl standart at 7th, 14th and 21st days with light and electron microscopes. The fish were exposed to carbaryl for 21 days and the histopatological, ultrastructural and cytopathological alterations occuring in the gill tissues of organisms were determined by light, Scanning and Transmission Electron Microscopes (SEM and TEM). At the end of the study, it was observed that carbaryl caused both histopathological and cytopathological changes in the gills of O. niloticus. It has been determined that the most of the pathological changes in the exposed organisms are the metabolic defence reactions.


Assuntos
Carbaril/toxicidade , Ciclídeos , Células Epiteliais/efeitos dos fármacos , Brânquias/efeitos dos fármacos , Inseticidas/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Células Epiteliais/ultraestrutura , Brânquias/ultraestrutura , Hiperplasia , Microscopia Eletroquímica de Varredura , Microscopia Eletrônica de Transmissão , Membrana Mucosa/efeitos dos fármacos , Membrana Mucosa/ultraestrutura
11.
Int J Mol Sci ; 20(14)2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31340547

RESUMO

BACKGROUND: Lung cancer cells are known to change proliferation and migration under simulated microgravity. In this study, we sought to evaluate cell adherence, apoptosis, cytoskeleton arrangement, and gene expression under simulated microgravity. METHODS: Human lung cancer cells were exposed to simulated microgravity in a random-positioning machine (RPM). Cell morphology and adherence were observed under phase-contrast microscopy, cytoskeleton staining was performed, apoptosis rate was determined, and changes in gene and protein expression were detected by real-time PCR with western blot confirmation. RESULTS: Three-dimensional (3D)-spheroid formation was observed under simulated microgravity. Cell viability was not impaired. Actin filaments showed a shift in alignment from longitudinal to spherical. Apoptosis rate was significantly increased in the spheroids compared to the control. TP53, CDKN2A, PTEN, and RB1 gene expression was significantly upregulated in the adherent cells under simulated microgravity with an increase in corresponding protein production for p14 and RB1. SOX2 expression was significantly upregulated in the adherent cells, but protein was not. Gene expressions of AKT3, PIK3CA, and NFE2L2 remained unaltered. CONCLUSION: Simulated microgravity induces alteration in cell adherence, increases apoptosis rate, and leads to upregulation of tumor suppressor genes in human lung cancer cells.


Assuntos
Apoptose/genética , Adesão Celular/genética , Células Epiteliais/metabolismo , Regulação Neoplásica da Expressão Gênica , Ausência de Peso , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestrutura , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Classe I de Fosfatidilinositol 3-Quinases/genética , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Células Epiteliais/ultraestrutura , Humanos , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Proteínas de Ligação a Retinoblastoma/genética , Proteínas de Ligação a Retinoblastoma/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transdução de Sinais , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Simulação de Ausência de Peso/instrumentação , Simulação de Ausência de Peso/métodos
13.
BMC Vet Res ; 15(1): 180, 2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31146764

RESUMO

BACKGROUND: Breast cancer resistance protein (BCRP) and multidrug resistance protein 4 (MRP4) are involved in uric acid excretion in humans and mice. Despite evidence suggesting that renal proximal tubular epithelial cells participate in uric acid excretion in chickens, the roles of BCRP and MRP4 therein remain unclear. This study evaluated the relationship between BCRP and MRP4 expression and renal function in chickens. RESULTS: Sixty laying hens were randomly divided into four treatment groups: a control group (NC) fed a basal diet; a sulfonamide-treated group (SD) fed the basal diet and supplemented with sulfamonomethoxine sodium via drinking water (8 mg/L); a fish meal group (FM) fed the basal diet supplemented with 16% fishmeal; and a uric acid injection group (IU) fed the basal diet and intraperitoneally injected with uric acid (250 mg/kg body weight). The results showed that serum uric acid, creatinine, and blood urea nitrogen levels were significantly higher in the SD and IU, but not FM, than in the NC groups. Renal tubular epithelial cells in the SD and IU groups were damaged. Liver BCRP and MRP4 mRNA and protein levels were significantly decreased in the SD and IU groups, but slightly increased in the FM group. In the SD group, BCRP and MRP4 were significantly increased in the ileum and slightly increased in the kidney. In the FM group, BCRP and MRP4 were significantly increased in the kidney and slightly increased in the ileum. In the IU group, BCRP and MRP4 were significantly increased in the kidney and ileum. BCRP and MRP4 expression in the jejunum was not affected by the treatments. CONCLUSION: Together, these results demonstrate that BCRP and MRP4 are involved in renal and intestinal uric acid excretion in chickens and that BCRP is positively related to MRP4 expression. Further, impairment of renal function results in an increase in serum uric acid as well as a compensatory increase in BCRP and MRP4 in the ileum; however, under normal renal function, renal BCRP and MRP4 are the main regulators of uric acid excretion.


Assuntos
Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Galinhas/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Ácido Úrico/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Animais , Nitrogênio da Ureia Sanguínea , Galinhas/sangue , Células Epiteliais/ultraestrutura , Feminino , Mucosa Intestinal/metabolismo , Rim/metabolismo , Rim/ultraestrutura , Túbulos Renais/ultraestrutura , Fígado/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , RNA Mensageiro/metabolismo , Ácido Úrico/sangue
14.
Int J Nanomedicine ; 14: 3995-4005, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31213811

RESUMO

Purpose: Since nanoparticles (NPs) are beginning to be introduced in medicine and industry, it is mendatory to evaluate their biological side-effects, among other things. The present study aimed to investigate the pathways by which nickel nanoparticles (NiNPs) enter nephrons and to evaluate their localization and effects on cellular ultrastructure. Methods: Rats were injected intraperitoneally with 20 nm NiNPs (20 mg/Kg/b.w./day) for 28 consecutive days. Transmission electron microscope technique was used to detect localization of NiNPs and their effects on cellular ultrastructure in rat kidneys. Additionally, measurements of certain biochemical parameters such as creatinine, urea, uric acid and phosphorus for investigating renal function following NiNPs treatment were taken. Results: The presence of NiNPs in the nephrons in treated rats was confirmed by transmission electron microscopy. NiNPs entered the renal tubules cells via various pathways. The results indicated that NiNPs administration induced ultrastructural changes in the proximal cells of renal tubules and certain glomerular cells (podocytes and mesangial cells). Additionally, NiNPs were found to be localized in the mitochondria, which led to a significant decrease in their density and morphology. Furthermore, cell death was induced in the glomerular cells as found with a Terminal deoxynucleotidyl transferase dUTP Nick End Labeling (TUNEL) assay and through detection of p35 using immunohistochemical staining. Conclusion: Herein, NiNPs were found to induce various cellular ultrastructural changes in the kidneys of rats. NiNPs used diverse pathways to internalize into the cytoplasm of the proximal convoluted tubules (PT) cells across the basement membrane, and also through the plasma membrane of two adjacent PT cells. NiNPs internalization, accumulation and their alterations of the cellular ultrastructure affected rat renal function.


Assuntos
Endocitose , Rim/ultraestrutura , Nanopartículas Metálicas/ultraestrutura , Animais , Células Epiteliais/ultraestrutura , Masculino , Células Mesangiais/ultraestrutura , Mitocôndrias/ultraestrutura , Níquel/química , Tamanho da Partícula , Podócitos/ultraestrutura , Ratos Wistar
15.
Acta Histochem ; 121(5): 595-603, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31109687

RESUMO

Due to the broad toxic relevance of acrylamide, many measures have been taken since the 1900s. These measures increased day by day when acrylamide was discovered in foods in 2002, and its toxic spectrum was found to be wider than expected. Therefore, in some countries, the products with higher acrylamide content were restricted. On the other hand, the effects of acrylamide on the respiratory system cells have yet to be well understood. In this study, we aimed at investigating the effect of acrylamide on lung epithelial BEAS-2B cells. Initially, the cytotoxic effect of acrylamide on BEAS-2B was determined by MTT assay. Then, cellular oxidative stress was measured. Flow cytometry analysis was conducted for Annexin-V and caspase 3/7. Furthermore, Bax, Bcl-2 and Nrf-2 proteins were evaluated by immunocytochemistry. Finally, acrylamide-induced cellular morphological changes were observed under confocal and TEM microscopes. According to MTT results, the IC50 concentration of acrylamide was 2.00 mM. After acrylamide treatment, oxidative stress increased dose-dependently. Annexin V-labelled apoptotic cells and caspase 3/7 activity were higher than untreated cells in acrylamide-treated cells. Immunocytochemical examination revealed a marked decrease in Bcl-2, an increase in Bax and Nrf-2 protein staining upon acrylamide treatment. Furthermore, in confocal and TEM microscopy, apoptotic hallmarks were pronounced. In the present study, acrylamide was suggested to display anti-proliferative activity, decrease viability, induce apoptosis and oxidative stress and cause morphological changes in BEAS-2B cells.


Assuntos
Acrilamida/toxicidade , Apoptose/efeitos dos fármacos , Citotoxinas/toxicidade , Pulmão/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Linhagem Celular , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Células Epiteliais/ultraestrutura , Humanos , Pulmão/patologia , Pulmão/ultraestrutura , Microscopia Confocal , Microscopia Eletrônica de Transmissão
16.
Discov Med ; 27(148): 153-160, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-31095924

RESUMO

Amyloid-ß (Aß) accumulation has been reported in patients with age-related macular degeneration (AMD), and it has been demonstrated to play an important role in the development of AMD. This study was performed to detect the activation of autophagy in retinal pigment epithelial (RPE) cells treated with Aß, aiming to investigate the potential protective mechanism of RPE cells against Aß stress. Human ARPE-19 cells were treated with soluble Aß1-42 oligomer. Transmission electron microscopy was used to assess the formation of autophagic compartments; immunofluorescence was used to detect the subcellular localization of LC3; and western blot was used to detect the conversion of LC3-I to LC3-II and the expression of p62. Activated autophagy in Aß-treated ARPE-19 cells was detected by three methodologies: 1) generation of autophagic compartments by transmission electron microscopy, 2) altered expression pattern of LC3 by immunofluorescence, and 3) elevated light-chain-3 II (LC3-II) and decreased p62 expression by western blot. These results suggest that Aß could induce autophagy in RPE cells, which provided a potential protective mechanism for the retina cells that encountered Aß deposition.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Células Epiteliais/metabolismo , Degeneração Macular/metabolismo , Fragmentos de Peptídeos/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Peptídeos beta-Amiloides/genética , Linhagem Celular , Células Epiteliais/ultraestrutura , Humanos , Degeneração Macular/genética , Degeneração Macular/patologia , Degeneração Macular/prevenção & controle , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Fragmentos de Peptídeos/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Epitélio Pigmentado da Retina/ultraestrutura
17.
Microsc Res Tech ; 82(8): 1339-1344, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31070847

RESUMO

Inflammatory bowel disease (IBD) is a global, chronic intractable disease. The functions of drugs and food components have been evaluated in models of IBD induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS). Here, we used transmission (TEM) and osmium-maceration scanning (SEM) electron microscopy to evaluate the ultrastructure of colonic epithelial cells in rat models of IBD induced by TNBS. Histological evaluation revealed that the intestinal crypts in the most regions of the IBD-model colons were deformed and we classified them as having high cell migration rates (HMIG). The remaining regions in the intestinal crypts retained a relatively normal structure and we classified them as having low cell migration rates (LMIG). Osmium-maceration SEM revealed the mucosal fluid flowing in spaces without secretory granules in crypt goblet cells of both HMIG and LMIG regions, indicating the depletion of goblet cell mucin that is found in patients with IBD. The Golgi apparatus in absorptive cells was stacked and curled in both regions. Osmium-maceration SEM showed membrane network structures resembling endoplasmic reticulum that were large and expanded in absorptive cells with HMIG rather than with LMIG regions in IBD-model colons. These findings indicated that endoplasmic reticulum stress is associated with susceptibility to IBD and that the effects of various agents can be evaluated according to endoplasmic reticulum stress revealed by using electron microscopy in models of IBD induced by TNBS.


Assuntos
Colo/citologia , Células Epiteliais/ultraestrutura , Doenças Inflamatórias Intestinais/patologia , Animais , Colo/patologia , Modelos Animais de Doenças , Retículo Endoplasmático/patologia , Retículo Endoplasmático/ultraestrutura , Células Epiteliais/patologia , Células Caliciformes/ultraestrutura , Complexo de Golgi/patologia , Complexo de Golgi/ultraestrutura , Doenças Inflamatórias Intestinais/induzido quimicamente , Masculino , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Mucinas , Ratos , Ácido Trinitrobenzenossulfônico/administração & dosagem
18.
Int J Mol Sci ; 20(7)2019 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-30959855

RESUMO

Cathepsin D is one of the major lysosomal aspartic proteases that is essential for the normal functioning of the autophagy-lysosomal system. In the kidney, cathepsin D is enriched in renal proximal tubular epithelial cells, and its levels increase during acute kidney injury. To investigate how cathepsin D-deficiency impacts renal proximal tubular cells, we employed a conditional knockout CtsDflox/-; Spink3Cre mouse. Immunohistochemical analyses using anti-cathepsin D antibody revealed that cathepsin D was significantly decreased in tubular epithelial cells of the cortico-medullary region, mainly in renal proximal tubular cells of this mouse. Cathepsin D-deficient renal proximal tubular cells showed an increase of microtubule-associated protein light chain 3 (LC3; a marker for autophagosome/autolysosome)-signals and an accumulation of abnormal autophagic structures. Renal ischemia/reperfusion injury resulted in an increase of early kidney injury marker, Kidney injury molecule 1 (Kim-1), in the cathepsin D-deficient renal tubular epithelial cells of the CtsDflox/-; Spink3Cre mouse. Inflammation marker was also increased in the cortico-medullary region of the CtsDflox/-; Spink3Cre mouse. Our results indicated that lack of cathepsin D in the renal tubular epithelial cells led to an increase of sensitivity against ischemia/reperfusion injury.


Assuntos
Catepsina D/deficiência , Túbulos Renais Proximais/enzimologia , Túbulos Renais Proximais/patologia , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/patologia , Animais , Autofagia , Catepsina D/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Integrases/metabolismo , Camundongos
19.
Microsc Res Tech ; 82(8): 1267-1276, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31002452

RESUMO

Amphibian skin secretions contain a variety of bioactive compounds that are involved in diverse roles such as communication, homeostasis, defence against predators, pathogens, and so on. Especially, the caecilian amphibians possess numerous cutaneous glands that produce the secretory material, which facilitate survival in their harsh subterranean environment. Inspite of the fact that India has a fairly abundant distribution of caecilian amphibians, there has hardly been any study on their skin and its secretion. Herein, we describe, using light microscopy and electron microscopy, two types of dermal glands, mucous and granular, in Gegeneophis ramaswamii. The mucous glands are filled with mucous materials. The mucous-producing cells are located near the periphery. The granular glands are surrounded by myoepithelial cells. A large number of granules of different sizes are present in the lumen of the granular gland. The granule-producing cells are present near the myoepithelial lining of the gland. There are small flat disk-like dermal scales in pockets in the transverse ridges of the posterior region of the body. Each pocket contains 1-4 scales of various sizes. Scanning electron microscopic (SEM) study of the skin surface showed numerous funnel-shaped glandular openings. The antibacterial activity of the skin secretions was revealed in the test against Escherichia coli, Klebsiella pneumoniae, and Aeromonas hydrophila, all gram-negative bacteria. SEM analyses confirm the membrane damage in bacterial cells on exposure to skin secretions of G. ramaswamii.


Assuntos
Anfíbios/anatomia & histologia , Escamas de Animais/ultraestrutura , Antibacterianos/farmacologia , Células Epiteliais/ultraestrutura , Glândulas Exócrinas/ultraestrutura , Pele/anatomia & histologia , Animais , Antibacterianos/química , Bactérias/efeitos dos fármacos , Transporte Biológico , Glândulas Exócrinas/química , Feminino , Masculino , Microscopia , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Pele/ultraestrutura
20.
Arthropod Struct Dev ; 50: 78-93, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31022533

RESUMO

Differentiation of transporting epithelial cells during development of animal organisms includes remodelling of apical and basal plasma membranes to increase the available surface for transport and formation of occluding junctions, which maintain a paracellular diffusion barrier. This study provides a detailed ultrastructural analysis of apical and basal plasma membrane remodelling and cell junction formation in hindgut cells during late embryonic and early postembryonic development of the crustacean Porcellio scaber. Hindgut cells in late-stage embryos are columnar with flat apical and basal plasma membranes. In early-stage marsupial mancae the hindgut cells begin to acquire their characteristic dome shape, the first apical membrane folding is evident and the septate junctions expand considerably, all changes being probably associated with the onset of active feeding. In postmarsupial mancae the apical labyrinth is further elaborated and the septate junctions are expanded. This coincides with the transition to an external environment and food sources. First basal infoldings appear in the anterior chamber of early-stage marsupial mancae, but in the papillate region they are mostly formed in postmarsupial mancae. In molting late-stage marsupial mancae, the plasma membrane acquires a topology characteristic of cuticle-producing arthropod epithelia and the septate junctions are considerably reduced.


Assuntos
Isópodes/crescimento & desenvolvimento , Isópodes/ultraestrutura , Animais , Diferenciação Celular , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Sistema Digestório/crescimento & desenvolvimento , Sistema Digestório/ultraestrutura , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Junções Intercelulares/metabolismo , Junções Intercelulares/ultraestrutura , Microscopia Eletrônica de Transmissão
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