RESUMEN
It is thought that monocytes rapidly differentiate to macrophages or dendritic cells (DCs) upon leaving blood. Here we have shown that Ly-6C⺠monocytes constitutively trafficked into skin, lung, and lymph nodes (LNs). Entry was unaffected in gnotobiotic mice. Monocytes in resting lung and LN had similar gene expression profiles to blood monocytes but elevated transcripts of a limited number of genes including cyclo-oxygenase-2 (COX-2) and major histocompatibility complex class II (MHCII), induced by monocyte interaction with endothelium. Parabiosis, bromodoxyuridine (BrdU) pulse-chase analysis, and intranasal instillation of tracers indicated that instead of contributing to resident macrophages in the lung, recruited endogenous monocytes acquired antigen for carriage to draining LNs, a function redundant with DCs though differentiation to DCs did not occur. Thus, monocytes can enter steady-state nonlymphoid organs and recirculate to LNs without differentiation to macrophages or DCs, revising a long-held view that monocytes become tissue-resident macrophages by default.
Asunto(s)
Diferenciación Celular , Células Dendríticas/metabolismo , Ganglios Linfáticos/citología , Macrófagos/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Animales , Antígenos Ly/metabolismo , Movimiento Celular , Ciclooxigenasa 2/genética , Células Dendríticas/citología , Células Dendríticas/inmunología , Endotelio/metabolismo , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Pulmón/citología , Ganglios Linfáticos/inmunología , Macrófagos/citología , Macrófagos/inmunología , Ratones , Ratones Endogámicos C57BL , Piel/citologíaRESUMEN
Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease globally. miRNAs (miRs) regulate various cellular events that lead to NAFLD. In this study we tested the hypothesis that miR-155 is an important regulator of steatohepatitis and fibrosis pathways. Wild type (WT) or miR-155 deficient (KO) mice received a high fat-high cholesterol-high sugar-diet (HF-HC-HS) for 34 weeks and liver tissues were analyzed. In patients with nonalcoholic steatohepatitis and in the mouse model of HF-HC-HS diet we found increased miR-155 levels in the liver compared to normal livers. Upon HF-HC-HS diet feeding, miR-155 KO mice displayed less liver injury, decreased steatosis, and attenuation in fibrosis compared to WT mice. ALT, triglyceride levels, and genes involved in fatty acid metabolic pathway were increased in WT mice whereas miR-155 KO mice showed attenuation in these parameters. HF-HC-HS diet-induced significant increase in the expression of NLRP3 inflammasome components in the livers of WT mice compared to chow fed diet. Compared to WT mice, miR-155 KO showed attenuated induction in the NLRP3, ASC, and caspase1 inflammasome expression on HF-HC-HS diet. Fibrosis markers such as collagen content and deposition, αSMA, Zeb2, and vimentin were all increased in WT mice and miR-155 KO mice showed attenuated fibrosis marker expression. Overall, our findings highlight a role for miR-155 in HF-HC-HS diet-induced steatosis and liver fibrosis.
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Inflamasomas/metabolismo , MicroARNs/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Animales , Dieta Alta en Grasa , Azúcares de la Dieta , Fibrosis , Regulación de la Expresión Génica , Metabolismo de los Lípidos , Hígado/patología , Masculino , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/patologíaRESUMEN
Cellular homeostais, that is normally maintained through autophagy, is disrupted in alcoholic liver disease (ALD). Because autophagy and exosome biogenesis share common elements, we hypothesized that increased exosome production in ALD may be linked to disruption of autophagic function. We found impaired autophagy both in ALD and alcoholic hepatitis (AH) mouse models and human livers with ALD as indicated by increased hepatic p62 and LC3-II levels. Alcohol reduced autophagy flux in vivo in chloroquine-treated mice as well as in vitro in hepatocytes and macrophages treated with bafilomycin A. Our results revealed that alcohol targets multiple steps in the autophagy pathway. Alcohol-related decrease in mechanistic target of rapamycin (mTOR) and Ras homolog enriched in brain (Rheb), that initiate autophagy, correlated with increased Beclin1 and autophagy-related protein 7 (Atg7), proteins involved in phagophore-autophagosome formation, in ALD. We found that alcohol disrupted autophagy function at the lysosomal level through decreased lysosomal-associated membrane protein 1 (LAMP1) and lysosomal-associated membrane protein 2 (LAMP2) in livers with ALD. We identified that micro-RNA 155 (miR-155), that is increased by alcohol, targets mTOR, Rheb, LAMP1, and LAMP2 in the authophagy pathway. Consistent with this, miR-155-deficient mice were protected from alcohol-induced disruption of autophagy and showed attenuated exosome production. Mechanistically, down-regulation of LAMP1 or LAMP2 increased exosome release in hepatocytes and macrophages in the presence and absence of alcohol. These results suggested that the alcohol-induced increase in exosome production was linked to disruption of autophagy and impaired autophagosome and lysosome function. Conclusion: Alcohol affects multiple genes in the autophagy pathway and impairs autophagic flux at the lysosome level in ALD. Inhibition of LAMP1 and LAMP2 promotes exosome release in ALD. We identified miR-155 as a mediator of alcohol-related regulation of autophagy and exosome production in hepatocytes and macrophages.
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Autofagia/fisiología , Exosomas/fisiología , Hepatopatías Alcohólicas/fisiopatología , Lisosomas/fisiología , MicroARNs/fisiología , Animales , Femenino , Hepatitis Alcohólica/genética , Hepatitis Alcohólica/fisiopatología , Hepatocitos/fisiología , Humanos , Hepatopatías Alcohólicas/genética , Proteína 1 de la Membrana Asociada a los Lisosomas/fisiología , Proteína 2 de la Membrana Asociada a los Lisosomas/fisiología , Macrófagos/fisiología , Ratones , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas de Unión al ARN/metabolismo , Serina-Treonina Quinasas TOR/fisiologíaRESUMEN
BACKGROUND & AIMS: Chronic, excessive alcohol consumption leads to alcoholic liver disease (ALD) characterized by steatosis, inflammation, and eventually cirrhosis. The hepatocyte specific microRNA 122 (MIR122) regulates hepatocyte differentiation and metabolism. We investigated whether an alcohol-induced decrease in level of MIR122 contributes to development of ALD. METHODS: We obtained liver samples from 12 patients with ALD and cirrhosis and 9 healthy individuals (controls) and analyzed them by histology and immunohistochemistry. C57Bl/6 mice were placed on a Lieber-DeCarli liquid diet, in which they were fed ethanol for 8 weeks, as a model of ALD, or a control diet. These mice were also given injections of CCl4, to increase liver fibrosis, for 8 weeks. On day 28, mice with ethanol-induced liver disease and advanced fibrosis, and controls, were given injections of recombinant adeno-associated virus 8 vector that expressed the primary miR-122 transcript (pri-MIR122, to overexpress MIR122 in hepatocytes) or vector (control). Two weeks before ethanol feeding, some mice were given injections of a vector that expressed an anti-MIR122, to knock down its expression. Serum and liver tissues were collected; hepatocytes and liver mononuclear cells were analyzed by histology, immunoblots, and confocal microscopy. We performed in silico analyses to identify targets of MIR122 and chromatin immunoprecipitation quantitative polymerase chain reaction analyses in Huh-7 cells. RESULTS: Levels of MIR122 were decreased in liver samples from patients with ALD and mice on the Lieber-DeCarli diet, compared with controls. Transgenic expression of MIR122 in hepatocytes of mice with ethanol-induced liver disease and advanced fibrosis significantly reduced serum levels of alanine aminotransferase (ALT) and liver steatosis and fibrosis, compared with mice given injections of the control vector. Ethanol feeding reduced expression of pri-MIR122 by increasing expression of the spliced form of the transcription factor grainyhead like transcription factor 2 (GRHL2) in liver tissues from mice. Levels of GRHL2 also were increased in liver tissues from patients with ALD, compared with controls; increases correlated with decreases in levels of MIR122 in human liver. Mice given injections of the anti-MIR122 before ethanol feeding had increased steatosis, inflammation, and serum levels of alanine aminotransferase compared with mice given a control vector. Levels of hypoxia-inducible factor 1 alpha (HIF1α) mRNA, a target of MIR122, were increased in liver tissues from patients and mice with ALD, compared with controls. Mice with hepatocyte-specific disruption of Hif1α developed less-severe liver injury following administration of ethanol, injection of anti-MIR122, or both. CONCLUSIONS: Levels of MIR122 decrease in livers from patients with ALD and mice with ethanol-induced liver disease, compared with controls. Transcription of MIR122 is inhibited by GRHL2, which is increased in livers of mice and patients with ALD. Expression of an anti-MIR122 worsened the severity of liver damage following ethanol feeding in mice. MIR122 appears to protect the liver from ethanol-induced damage by reducing levels of HIF1α. These processes might be manipulated to reduce the severity of ALD in patients.
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Proteínas de Unión al ADN/metabolismo , Hepatopatías Alcohólicas/metabolismo , Hepatopatías Alcohólicas/prevención & control , MicroARNs/metabolismo , Factores de Transcripción/metabolismo , Adulto , Anciano , Animales , Estudios de Casos y Controles , Modelos Animales de Enfermedad , Femenino , Hepatocitos/metabolismo , Humanos , Hepatopatías Alcohólicas/patología , Masculino , Ratones , Persona de Mediana EdadRESUMEN
Early pathological descriptions of Crohn disease (CD) argued for a potential defect in lymph transport; however, this concept has not been thoroughly investigated. In mice, poor healing in response to infection-induced tissue damage can cause hyperpermeable lymphatic collecting vessels in mesenteric adipose tissue that impair antigen and immune cell access to mesenteric lymph nodes (LNs), which normally sustain appropriate immunity. To investigate whether analogous changes might occur in human intestinal disease, we established a three-dimensional imaging approach to characterize the lymphatic vasculature in mesenteric tissue from controls or patients with CD. In CD specimens, B-cell-rich aggregates resembling tertiary lymphoid organs (TLOs) impinged on lymphatic collecting vessels that enter and exit LNs. In areas of creeping fat, which characterizes inflammation-affected areas of the bowel in CD, we observed B cells and apparent innate lymphoid cells that had invaded the lymphatic vessel wall, suggesting these cells may be mediators of lymphatic remodeling. Although TLOs have been described in many chronic inflammatory states, their anatomical relationship to preestablished LNs has never been revealed. Our data indicate that, at least in the CD-affected mesentery, TLOs are positioned along collecting lymphatic vessels in a manner expected to affect delivery of lymph to LNs.
Asunto(s)
Enfermedad de Crohn/diagnóstico por imagen , Ganglios Linfáticos/diagnóstico por imagen , Vasos Linfáticos/diagnóstico por imagen , Adulto , Animales , Linfocitos B/patología , Enfermedad de Crohn/patología , Enfermedad de Crohn/cirugía , Femenino , Humanos , Íleon/diagnóstico por imagen , Íleon/patología , Imagenología Tridimensional , Inflamación , Intestinos/patología , Intestinos/cirugía , Ganglios Linfáticos/patología , Ganglios Linfáticos/cirugía , Vasos Linfáticos/patología , Vasos Linfáticos/cirugía , Masculino , Mesenterio/diagnóstico por imagen , Mesenterio/patología , Mesenterio/cirugía , Ratones , Persona de Mediana Edad , Estructuras Linfoides Terciarias/diagnóstico por imagen , Estructuras Linfoides Terciarias/patologíaRESUMEN
BACKGROUND & AIMS: Inflammation and impaired hepatocyte regeneration contribute to liver failure in alcoholic hepatitis (AH). Interleukin (IL)-1 is a key inflammatory cytokine in the pathobiology of AH. The role of IL-1 in liver regeneration in the recovery phase of alcohol-induced liver injury is unknown. METHODS: In this study, we tested IL-1 receptor antagonist to block IL-1 signalling in a mouse model of acute-on-chronic liver injury on liver inflammation and hepatocyte regeneration in AH. RESULTS: We observed that inhibition of IL-1 signalling decreased liver inflammation and neutrophil infiltration, and resulted in enhanced regeneration of hepatocytes and increased rate of recovery from liver injury in AH. CONCLUSION: Our novel findings suggest that IL-1 drives sustained liver inflammation and impaired hepatocyte regeneration even after cessation of ethanol exposure.
Asunto(s)
Insuficiencia Hepática Crónica Agudizada/tratamiento farmacológico , Proliferación Celular/efectos de los fármacos , Hepatitis Alcohólica/tratamiento farmacológico , Hepatocitos/efectos de los fármacos , Proteína Antagonista del Receptor de Interleucina 1/farmacología , Interleucina-1/antagonistas & inhibidores , Regeneración Hepática/efectos de los fármacos , Hígado/efectos de los fármacos , Insuficiencia Hepática Crónica Agudizada/metabolismo , Insuficiencia Hepática Crónica Agudizada/patología , Insuficiencia Hepática Crónica Agudizada/fisiopatología , Animales , Modelos Animales de Enfermedad , Femenino , Hepatitis Alcohólica/metabolismo , Hepatitis Alcohólica/patología , Hepatitis Alcohólica/fisiopatología , Hepatocitos/metabolismo , Hepatocitos/patología , Interleucina-1/metabolismo , Hígado/metabolismo , Hígado/patología , Hígado/fisiopatología , Ratones Endogámicos C57BL , Infiltración Neutrófila/efectos de los fármacos , Recuperación de la Función , Transducción de Señal/efectos de los fármacos , Factores de TiempoRESUMEN
BACKGROUND & AIMS: Alcoholic liver disease (ALD) ranges from fatty liver to inflammation and cirrhosis. miRNA-155 is an important regulator of inflammation. In this study, we describe the in vivo role of miR-155 in ALD. METHODS: Wild-type (WT) (C57/BL6J) or miR-155 knockout (KO) and TLR4 KO mice received Lieber DeCarli diet for 5weeks. Some mice received corn oil or CCl4 for 2 or 9weeks. RESULTS: We found that miR-155 KO mice are protected from alcohol-induced steatosis and inflammation. The reduction in alcohol-induced fat accumulation in miR-155 KO mice was associated with increased peroxisome proliferator-activated receptor response element (PPRE) and peroxisome proliferator-activated receptors (PPAR)α (miR-155 target) binding and decreased MCP1 production. Treatment with a miR-155 inhibitor increased PPARγ expression in naïve and alcohol treated RAW macrophages. Alcohol increased lipid metabolism gene expression (FABP4, LXRα, ACC1 and LDLR) in WT mice and this was prevented in KO mice. Alcohol diet caused an increase in the number of CD163(+) CD206(+) infiltrating macrophages and neutrophils in WT mice, which was prevented in miR-155 KO mice. Kupffer cells isolated from miR-155 KO mice exhibited predominance of M2 phenotype when exposed to M1 polarized signals and this was due to increased C/EBPß. Pro-fibrotic genes were attenuated in miR-155 KO mice after alcohol diet or CCl4 treatment. Compared to WT mice, attenuation in CCl4 induced hydroxyproline and α-SMA was observed in KO mice. Finally, we show TLR4 signaling regulates miR-155 as TLR4 KO mice showed no induction of miR-155 after alcohol diet. CONCLUSIONS: Collectively our results demonstrated the role of miR-155 in alcohol-induced steatohepatitis and fibrosis in vivo.
Asunto(s)
Hígado Graso Alcohólico/etiología , Cirrosis Hepática Experimental/etiología , MicroARNs/fisiología , Animales , Tetracloruro de Carbono , ADN/metabolismo , Femenino , Interferón gamma/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , PPAR alfa/metabolismo , Elementos de Respuesta , Receptor Toll-Like 4/fisiologíaRESUMEN
Antibodies targeting receptor-mediated entry of HCV into hepatocytes confer limited therapeutic benefits. Evidence suggests that exosomes can transfer genetic materials between cells; however, their role in HCV infection remains obscure. Here, we show that exosomes isolated from sera of chronic HCV infected patients or supernatants of J6/JFH1-HCV-infected Huh7.5 cells contained HCV RNA. These exosomes could mediate viral receptor-independent transmission of HCV to hepatocytes. Negative sense HCV RNA, indicative of replication competent viral RNA, was present in exosomes of all HCV infected treatment non-responders and some treatment-naïve individuals. Remarkably, HCV RNA was associated with Ago2, HSP90 and miR-122 in exosomes isolated from HCV-infected individuals or HCV-infected Huh7.5 cell supernatants. Exosome-loading with a miR-122 inhibitor, or inhibition of HSP90, vacuolar H+-ATPases, and proton pumps, significantly suppressed exosome-mediated HCV transmission to naïve cells. Our findings provide mechanistic evidence for HCV transmission by blood-derived exosomes and highlight potential therapeutic strategies.
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Exosomas/metabolismo , Hepacivirus/fisiología , Hepatocitos/virología , ARN Viral/genética , Proteínas Argonautas/metabolismo , Células Cultivadas , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , MicroARNs/metabolismo , Unión Proteica , Receptores Virales/aislamiento & purificación , Replicación Viral/fisiologíaRESUMEN
BACKGROUND & AIMS: miR-122 is the most abundant miRNA in the liver particularly in hepatocytes where it targets cholesterol metabolism. Steatosis, a key component of non-alcoholic fatty liver disease, is regulated by hypoxia-inducible factor-1α (HIF-1α). Here, we hypothesized that reduced miR-122 has a pathogenic role in steatohepatitis. METHODS: miR-122 and its target genes were evaluated in mouse livers and/or isolated hepatocytes after methionine-choline-deficient (MCD) or methionine-choline-supplemented (MCS) diet. RESULTS: Liver and hepatocyte miR-122 expression was significantly decreased in steatohepatitis. A maximum reduction in miR-122 occurred at the fibrosis stage (8 weeks of MCD diet). MAP3K3, a miR-122 target gene, was induced at all stages of non-alcoholic steatohepatitis (NASH; 3-8 weeks) only at the mRNA level. Increased NF-κB activation was found in MCD diet-fed mice and MAP3K3 regulated the NF-κB DNA binding in naive hepatocytes. HIF-1α mRNA and DNA binding and expression of the HIF-1α target gene, profibrotic lysyl oxidase, was increased in advanced steatohepatitis (8 weeks). In addition, increase in vimentin and Sirius red staining (liver fibrosis) was found at 8 weeks of MCD diet. Using miR-122 overexpression and inhibition approaches, we confirmed that HIF-1α, vimentin and MAP3K3 are novel miR-122 targets in hepatocytes. We report transcriptional repression of miR-122 in NASH. Decreased liver miR-122 was associated with elevated circulating miR-122 in both exosome-rich and protein-rich serum fractions. CONCLUSIONS: Our novel data suggest that decreased liver miR-122 contributes to upregulation of modulators of tissue remodelling (HIF-1α, vimentin and MAP3K3) and might play a role in NASH-induced liver fibrosis.
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Factor 1 Inducible por Hipoxia/metabolismo , Cirrosis Hepática/patología , MicroARNs/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Vimentina/metabolismo , Animales , Western Blotting , Ensayo de Cambio de Movilidad Electroforética , Hepatocitos/metabolismo , Inmunohistoquímica , Cirrosis Hepática/etiología , MAP Quinasa Quinasa Quinasa 3/metabolismo , Ratones , FN-kappa B/metabolismo , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Reacción en Cadena en Tiempo Real de la Polimerasa , Estadísticas no ParamétricasRESUMEN
This study was undertaken to determine the influence of Aloe vera (AV) on changes induced in pulmonary tissue of cigarette smoke (CS) inhaling mice. CS inhalation for 4 weeks caused pulmonary damage as evident by histoarchitectural alterations and enhanced serum and tissue lactate dehydrogenase (LDH) activities. CS inhalation also led to increased mucin production as revealed by mucicarmine and Alcian Blue-Periodic Acid Schiff (AB-PAS) staining. Studies on bronchoalveolar lavage fluid (balf) of CS exposed animals revealed structural changes in phospholipids and increase in surface tension when compared with control counterparts. These changes were accompanied by enhanced nitric oxide (NO) levels, citrulline levels, peroxidative damage, and differential modulation of antioxidant defense system. AV administration (seven weeks, 500 mg/kg b.w. daily) to CS inhaling mice led to modulation of CS induced pulmonary changes as revealed by lesser degree of histoarchitectural alterations, lesser mucin production, decreased NO levels, citrulline levels, peroxidative damage, and serum LDH activity. AV treatment to CS inhaling mice was associated with varying response to antioxidant defense system, however balf of CS + AV treated animals did not exhibit appreciable changes when compared with that of CS exposed animals. These observations suggest that AV has the potential to modulate CS induced changes in the pulmonary tissue which could have implications in management of CS associated pulmonary diseases, however, further investigations are required to explore its complete mechanism of action.
Asunto(s)
Aloe/química , Pulmón/efectos de los fármacos , Humo/efectos adversos , Aloe/metabolismo , Animales , Antioxidantes/metabolismo , Líquido del Lavado Bronquioalveolar/química , Citrulina/metabolismo , Lactato Deshidrogenasas/sangre , Peroxidación de Lípido/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Mucinas/metabolismo , Óxido Nítrico/metabolismo , Oxidorreductasas/metabolismo , Extractos Vegetales/química , Extractos Vegetales/farmacologíaRESUMEN
Effective therapies are limited for patients with parenteral nutrition-dependent short bowel syndrome. We previously showed that intestinal expression of the transcriptional coregulator tetradecanoyl phorbol acetate-induced sequence 7 (tis7) is markedly increased during the adaptive response following massive small bowel resection and tis7 plays a role in normal gut lipid metabolism. Here, we further explore the functional implications of tis7 deletion in intestinal lipid metabolism and the adaptive response following small bowel resection. Intestinal tis7 transgenic (tis7(tg)), tis7(-/-), and wild-type (WT) littermates were subjected to 50% small bowel resection. Mice were fed a control or a high-saturated-fat (42% energy) diet for 21 days. Survival, body weight recovery, lipid absorption, mucosal lipid analysis, and the morphometric adaptive response were analyzed. Quantitative real-time PCR was performed to identify tis7 downstream gene targets. Postresection survival was markedly reduced in high-fat, but not control, diet-fed tis7(-/-) mice. Decreased survival was associated with anastomotic inflammation and intestinal obstruction postresection. High-fat, but not control, diet-fed tis7(-/-) mice had increased intestinal IL-6 expression. Intestinal lipid trafficking was altered in tis7(-/-) compared with WT mice postresection. In contrast, high-fat diet-fed tis7(tg) mice had improved survival postresection compared with WT littermates. High-fat diet feeding in the setting of tis7 deletion resulted in postresection anastomotic inflammation and small bowel obstruction. Tolerance of a calorie-rich, high-fat diet postresection may require tis7 and its target genes. The presence of luminal fat in the setting of tis7 deletion promotes an intestinal inflammatory response postresection.
Asunto(s)
Dieta Alta en Grasa/efectos adversos , Enteritis/etiología , Proteínas Inmediatas-Precoces/deficiencia , Obstrucción Intestinal/etiología , Intestino Delgado/metabolismo , Proteínas de la Membrana/deficiencia , Síndrome del Intestino Corto/complicaciones , Anastomosis Quirúrgica , Animales , Modelos Animales de Enfermedad , Enteritis/genética , Enteritis/metabolismo , Regulación de la Expresión Génica , Proteínas Inmediatas-Precoces/genética , Interleucina-6/metabolismo , Absorción Intestinal , Obstrucción Intestinal/genética , Obstrucción Intestinal/metabolismo , Intestino Delgado/cirugía , Metabolismo de los Lípidos , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Síndrome del Intestino Corto/genética , Síndrome del Intestino Corto/metabolismo , Factores de TiempoRESUMEN
BACKGROUND: Chronic alcohol impairs gut barrier function and induces inflammatory cytokines. The effects of acute alcohol binge on the gut are partially understood. Micro-RNA-155 (miR-155), a modulator of cytokine and T-cell immune response in the gut, stabilizes tumor necrosis factor-α (TNFα) mRNA. Here, we investigated the role of the inflammation modulator miR-155 as well as the effects of acute binge and chronic alcohol feeding in the small bowel (SB) in mice. METHODS: For the acute alcohol binge, wild-type (WT) mice received 5 g/kg 50% alcohol/d or equal amount of water oral gavage for 3 days. WT and miR-155-deficient (miR-155-knockout [KO]) mice received ethanol containing Lieber-DeCarli or isocaloric control diet for 5 weeks. MiR-155, antimicrobial peptide, regenerating islet-derived 3-beta (Reg3b), inflammation markers, Src homology 2-containing inositol phosphatase-1 (SHIP1), TNFα, and nuclear factor-κB (NF-κB) were measured in proximal intestinal tissue. Endotoxin was measured in the serum. RESULTS: Acute alcohol binge enhanced, whereas chronic alcohol feeding decreased, Reg3b mRNA and protein levels in the SB. Both acute binge and chronic alcohol feeding increased serum endotoxin levels, intestinal NF-κB activation and TNFα mRNA levels. However, TNFα protein and miR-155 were increased only after chronic alcohol feeding in the SB. Furthermore, miR-155-KO mice were protected from chronic alcohol-induced increase in serum endotoxin, intestinal TNFα, and NF-κB activation. Also, alcohol-fed miR-155-KO mice had no decrease of Reg3b and SHIP1 levels. CONCLUSIONS: These results demonstrate that both acute binge and chronic ethanol administration result in increased serum-endotoxin levels. Our study identifies a novel role for miR-155 in chronic alcohol-induced intestinal inflammation and barrier dysfunction.
Asunto(s)
Endotoxinas/sangre , Etanol/efectos adversos , Enfermedades Inflamatorias del Intestino/prevención & control , MicroARNs/genética , Consumo de Bebidas Alcohólicas , Animales , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Enfermedades Inflamatorias del Intestino/inducido químicamente , Enfermedades Inflamatorias del Intestino/metabolismo , Inositol Polifosfato 5-Fosfatasas , Ratones , Ratones Noqueados , MicroARNs/metabolismo , FN-kappa B/metabolismo , Proteínas Asociadas a Pancreatitis , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatasas , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Exosomes, membranous nanovesicles, naturally carry bio-macromolecules and play pivotal roles in both physiological intercellular crosstalk and disease pathogenesis. Here, we showed that B cell-derived exosomes can function as vehicles to deliver exogenous miRNA-155 mimic or inhibitor into hepatocytes or macrophages, respectively. Stimulation of B cells significantly increased exosome production. Unlike in parental cells, baseline level of miRNA-155 was very low in exosomes derived from stimulated B cells. Exosomes loaded with a miRNA-155 mimic significantly increased miRNA-155 levels in primary mouse hepatocytes and the liver of miRNA-155 knockout mice. Treatment of RAW macrophages with miRNA-155 inhibitor loaded exosomes resulted in statistically significant reduction in LPS-induced TNFα production and partially prevented LPS-induced decrease in SOCS1 mRNA levels. Furthermore, exosome-mediated miRNA-155 inhibitor delivery resulted in functionally more efficient inhibition and less cellular toxicity compared to conventional transfection methods. Similar approaches could be useful in modification of target biomolecules in vitro and in vivo. From the clinical editor: In this study, exosome-based delivery of miRNA-155 mimicker or inhibitor was found to have significant biological response in hepatocytes and macrophages. Exosome-based approaches may be useful in the modification of other target biomolecules.
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Exosomas/metabolismo , Macrófagos/metabolismo , MicroARNs/antagonistas & inhibidores , Animales , Línea Celular , RatonesRESUMEN
Leishmania, a protozoan parasite, is responsible for the occurrence of leishmaniasis, a disease that is prevalent in tropical regions. Visceral Leishmaniasis (VL), also known as kala-azar in Asian countries, is one of the most significant forms of VL, along with Cutaneous Leishmaniasis (CL) and Mucocutaneous Leishmaniasis (ML). Management of this condition typically entails the use of chemotherapy as the sole therapeutic option. The current treatments for leishmaniasis present several drawbacks, including a multitude of side effects, prolonged treatment duration, disparate efficacy across different regions, and the emergence of resistance. To address this urgent need, it is imperative to identify alternative treatments that are both safer and more effective. The identification of appropriate pharmacological targets in conjunction with biological pathways constitutes the initial stage of drug discovery. In this review, we have addressed the key metabolic pathways that represent potential pharmacological targets as well as prominent treatment options for leishmaniasis.
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Leishmania donovani , Leishmania , Leishmaniasis Cutánea , Leishmaniasis Mucocutánea , Leishmaniasis Visceral , Leishmaniasis , Animales , Leishmaniasis/tratamiento farmacológico , Leishmaniasis Visceral/tratamiento farmacológico , Leishmaniasis Visceral/parasitología , AsiaRESUMEN
Green synthesis methods offer a cost-effective and environmentally friendly approach to producing nanoparticles (NPs), particularly metal-based oxides. This study explores the green synthesis of copper oxide nanoparticles using Aloe vera (Aloe barbadensis Miller) leaf extract. The characterization revealed a unique sago-shaped morphology revealed by field-emission scanning electron microscopy and X-ray diffraction analysis. Distinctive metal-oxygen bonds at 521 and 601 cm-1 were confirmed by Fourier-transform infrared (FT-IR) spectroscopy. Furthermore, UV-visible spectroscopy revealed absorbance at 248 nm, suggesting electron transitions across energy bands and varying surface conduction electrons. The band gap value indicated the presence of quantum confinement effects, which were probably caused by the distinctive morphology and surface structure of the biogenic NPs. Additionally, molecular docking studies were carried out against key proteins of Salmonella typhi and Listeria monocytogenes, namely, listeriolysin O (PDB ID: 4CDB), internalin (InlA) (PDB ID: 1O6T), Salmonella effector protein (SopB) (PDB ID: 4DID), and YfdX (PDB ID: 6A07) using AutoDock 4.2. The results revealed binding energies against S. typhi and L. monocytogenes proteins, indicating potential interactions establishing the foundation for further in-depth understanding of the molecular basis underlying the observed antibacterial effects in vitro against S. typhi, Klebsiella pneumoniae, Pseudomonas aeruginosa, and L. monocytogenes. Antibacterial activity evaluation yielded impressive results, with CuO NPs displaying significant activity against S. typhi and L. monocytogenes, exhibiting zones of inhibition values of 13 ± 0.02 and 15 ± 0.04 mm, respectively. Moreover, the CuO NPs demonstrated remarkable photocatalytic efficacy, resulting in the degradation of 77% of the methylene blue dye when exposed to UV irradiation. This study highlighted the potential of green-synthesized CuO NPs derived from A. vera with their unique morphology, interesting spectroscopic properties, and promising antibacterial and photocatalytic activities.
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Interactions between the epithelium and surrounding mesenchyme/stroma play an important role in normal gut morphogenesis, the epithelial response to injury, and epithelial carcinogenesis. The tumor microenvironment, composed of stromal cells including myofibroblasts and immune cells, regulates tumor growth and the cancer stem cell niche. Deletion of epimorphin (Epim), a syntaxin family member expressed in myofibroblasts and macrophages, results in partial protection from colitis and from inflammation-induced colon cancer in mice. We sought to determine whether epimorphin deletion protects from polyposis in the Apcmin/+ mouse model of intestinal carcinogenesis. Epim-/- mice were crossed to Apcmin/+ mice; Apcmin/+ and Apcmin/+/Epim-/- mice were killed at 3 mo of age. Polyp numbers and sizes were quantified in small intestine and colon, and gene expression analyses for pathways relevant to epithelial carcinogenesis were performed. Primary myofibroblast cultures were isolated, and expression and secretion of selected growth factors from Apcmin/+ and Apcmin/+/Epim-/- myofibroblasts were examined by ELISA. Small bowel polyposis was significantly inhibited in Apcmin/+/Epim-/- compared with Apcmin/+ mice. Apcmin/+/Epim-/- compared with Apcmin/+ polyps and adjacent uninvolved intestinal mucosa had increased transforming growth factor-ß (TGF-ß) expression and signaling with increased P-Smad2/3 expression. Myofibroblasts isolated from Apcmin/+/Epim-/- vs. Apcmin/+ mice had markedly decreased hepatocyte growth factor (HGF) expression and secretion. We concluded that Epim deletion inhibits polyposis in Apcmin/+ mice, associated with increased mucosal TGF-ß signaling and decreased myofibroblast HGF expression and secretion. Our data suggest that Epim deletion reduces tumorigenicity of the stromal microenvironment.
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Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Factor de Crecimiento de Hepatocito/metabolismo , Glicoproteínas de Membrana/metabolismo , Miofibroblastos/metabolismo , Poliposis Adenomatosa del Colon/genética , Poliposis Adenomatosa del Colon/metabolismo , Poliposis Adenomatosa del Colon/patología , Proteína de la Poliposis Adenomatosa del Colon/genética , Animales , Neoplasias del Colon/metabolismo , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica , Factor de Crecimiento de Hepatocito/genética , Mucosa Intestinal/fisiología , Masculino , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
UNLABELLED: MicroRNAs are fine tuners of diverse biological responses and are expressed in various cell types of the liver. Here we hypothesized that circulating microRNAs (miRNAs) may serve as biomarkers of liver damage and inflammation. We studied miRNA-122, which is abundant in hepatocytes, and miR-155, -146a, and -125b, which regulate inflammation in immune cells in mouse models of alcoholic liver disease (ALD), drug (acetaminophen, APAP)-induced liver injury (DILI), and Toll-like receptor (TLR) 9+4 ligand-induced inflammatory cell-mediated liver damage. We found that serum/plasma miR-122 correlated with alanine aminotransferase (ALT) increases in the liver damage caused by alcohol, APAP, and TLR9 (CpG)+4 (LPS) ligands. MiR-155, a regulator of inflammation, was increased in serum/plasma in alcoholic and inflammatory liver injury. Alcohol failed to increase serum miR-122 in TLR4-deficient and p47phox-deficient mice that were protected from ALD. We found the most robust increase in plasma miR-122 in DILI and it correlated with the highest ALT levels. Consistent with the massive inflammatory cell infiltration in the liver, plasma miR-155 and miR-146a were significantly elevated after CpG+LPS administration. We show for the first time that, depending on the type of liver injury, circulating miRNAs are associated either with the exosome-rich or protein-rich compartments. In ALD and in inflammatory liver injury, serum/plasma miR-122 and miR-155 were predominantly associated with the exosome-rich fraction, whereas in DILI/APAP injury these miRNAs were present in the protein-rich fraction. CONCLUSION: Our results suggest that circulating miRNAs may serve as biomarkers to differentiate between hepatocyte injury and inflammation and the exosome versus protein association of miRNAs may provide further specificity to mechanisms of liver pathology.
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Enfermedad Hepática Inducida por Sustancias y Drogas/sangre , Exosomas/metabolismo , Hepatitis/sangre , Hepatopatías Alcohólicas/sangre , MicroARNs/sangre , Acetaminofén/toxicidad , Alanina Transaminasa/sangre , Animales , Biomarcadores/sangre , Islas de CpG , Modelos Animales de Enfermedad , Femenino , Hepatocitos , Ligandos , Lipopolisacáridos , Hígado/lesiones , Ratones , Estadísticas no Paramétricas , Receptor Toll-Like 4/deficiencia , Receptor Toll-Like 9RESUMEN
A new pregnane, 12ß-O-acetyl-20(S)-O-N-methylanthranilyl-3ß, 8ß, 14ß, 17α-tetrol pregn-5-ene (12ß-O-acetyl-20(S)-O-N-methylanthranilyl-sarcostin) have been isolated from Marsdenia tenacissima (family: Asclepediaceae). The structure of new pregnane was elucidated by using spectroscopic techniques,1H,13C NMR, HMBC, HSQC, COSY and TOCSY and ESI-MS Mass spectrometry.
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Introduction: The use of information and communication technology (ICT) in education and clinical purposes is growing immensely. The usage skill, attitude of dental students, and difficulties faced by students are important concerns to be addressed. Objective: The study was designed to investigate the knowledge, attitude, and skills of ICT of undergraduate dental students via assessing the ease of computer and internet use among students, the level of computer skills and training of the students, and usage pattern of computer activities. Materials and methods: A cross-sectional survey was conducted among students of three dental colleges in Haryana using a self-designed questionnaire having sections related to demographics, devices, softwares and search engine usage, ICT skills, ICT attitude, and educational and social use. Results: The response rate was 81.96%. A total of 99.1% of the individuals responded affirmatively to the usage of mobile phones, 48.4 and 13.1% for laptops and tablets, respectively. The average duration of use of smartphones per day was 4.2121 ± 2.834 hours. Google was opted as the most popular search engine used and PubMed the least popular. Only 12.9% of participants showed a negative attitude toward the use of ICT in studies. More social use than academic use was observed. Attitude scores showed a significant correlation with the educational use of ICT (r = 0.89, p = 0.003). Conclusion: Most of the participants were well acquainted with the usage and had a positive attitude toward ICT for educational purposes. Sensitization and training in scientific literature search and basics of ICT and their practical utilization in dental education, research, and practice should be included in the curriculum. How to cite this article: Sikka N, Arya L, Bala S, et al. Knowledge, Attitude, and Perception of Undergraduate Dental Students on Information and Communication Technology and Computer-assisted Learning. Int J Clin Pediatr Dent 2023;16(5):711-715.
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Naturally occurring pentacyclic triterpenoids and their semisynthetic analogues have engrossed increasing attention for their anticancer potential and exhibiting promising role in discovery of new anticancer agents. Present study include the semi synthetic modifications of Lantadenes from the weed Lantana carama and their structures delineation by FT-IR, 1H-NMR, 13C-NMR & mass spectroscopy. All the compounds were scrutinized for in vitro cytotoxicity, ligand receptor interaction and in vivo anticancer studies. Most of the novel analogues displayed potent antiproliferative activity against A375 & A431 cancer cell lines and found superior to parent Lantadenes. In particular, 3ß-(4-Methoxybenzoyloxy)-22ß-senecioyloxy-olean-12-en-28-oic acid was found to be most suitable compound, with IC50 value of 3.027 µM aganist A375 cell line having least docking score (-69.40 kcal/mol). Promising anticancer potential of the lead was further indicated by significant reduction in tumor volume and burden in two stage carcinoma model. These findings suggests that the Lantadene derivatives may hold promising potential for the intervention of skin cancers.