RESUMEN
Genetic polymorphisms of immune genes that associate with higher risk to develop Alzheimer's disease (AD) have led to an increased research interest on the involvement of the immune system in AD pathogenesis. A link between amyloid pathology and immune gene expression was suggested in a genome-wide gene expression study of transgenic amyloid mouse models. In this study, the gene expression of lysozyme, a major player in the innate immune system, was found to be increased in a comparable pattern as the amyloid pathology developed in transgenic mouse models of AD. A similar pattern was seen at protein levels of lysozyme in human AD brain and CSF, but this lysozyme pattern was not seen in a tau transgenic mouse model. Lysozyme was demonstrated to be beneficial for different Drosophila melanogaster models of AD. In flies that expressed Aß1-42 or AßPP together with BACE1 in the eyes, the rough eye phenotype indicative of toxicity was completely rescued by coexpression of lysozyme. In Drosophila flies bearing the Aß1-42 variant with the Arctic gene mutation, lysozyme increased the fly survival and decreased locomotor dysfunction dose dependently. An interaction between lysozyme and Aß1-42 in the Drosophila eye was discovered. We propose that the increased levels of lysozyme, seen in mouse models of AD and in human AD cases, were triggered by Aß1-42 and caused a beneficial effect by binding of lysozyme to toxic species of Aß1-42 , which prevented these from exerting their toxic effects. These results emphasize the possibility of lysozyme as biomarker and therapeutic target for AD.
Asunto(s)
Enfermedad de Alzheimer/enzimología , Muramidasa/metabolismo , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/genética , Péptidos beta-Amiloides/metabolismo , Animales , Encéfalo/enzimología , Encéfalo/patología , Modelos Animales de Enfermedad , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Drosophila melanogaster/ultraestructura , Ojo/metabolismo , Ojo/ultraestructura , Femenino , Humanos , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Muramidasa/genética , Mutación , Fragmentos de Péptidos/metabolismo , ARN Mensajero/metabolismoRESUMEN
Lysozyme amyloidosis is a hereditary disease in which mutations in the gene coding for lysozyme leads to misfolding and consequently accumulation of amyloid material. To improve understanding of the processes involved we expressed human wild type (WT) lysozyme and the disease-associated variant F57I in the central nervous system (CNS) of a Drosophila melanogaster model of lysozyme amyloidosis, with and without co-expression of serum amyloid p component (SAP). SAP is known to be a universal constituent of amyloid deposits and to associate with lysozyme fibrils. There are clear indications that SAP may play an important role in lysozyme amyloidosis, which requires further elucidation. We found that flies expressing the amyloidogenic variant F57I in the CNS have a shorter lifespan than flies expressing WT lysozyme. We also identified apoptotic cells in the brains of F57I flies demonstrating that the flies' neurological functions are impaired when F57I is expressed in the nerve cells. However, co-expression of SAP in the CNS prevented cell death and restored the F57I flies' lifespan. Thus, SAP has the apparent ability to protect nerve cells from damage caused by F57I. Furthermore, it was found that co-expression of SAP prevented accumulation of insoluble forms of lysozyme in both WT- and F57I-expressing flies. Our findings suggest that the F57I mutation affects the aggregation process of lysozyme resulting in the formation of cytotoxic species and that SAP is able to prevent cell death in the F57I flies by preventing accumulation of toxic F57I structures.
Asunto(s)
Amiloidosis/genética , Sistema Nervioso Central/metabolismo , Muramidasa/genética , Placa Amiloide/genética , Agregación Patológica de Proteínas/genética , Componente Amiloide P Sérico/genética , Amiloidosis/metabolismo , Amiloidosis/patología , Animales , Animales Modificados Genéticamente , Apoptosis , Sistema Nervioso Central/patología , Modelos Animales de Enfermedad , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Expresión Génica , Humanos , Longevidad/genética , Muramidasa/metabolismo , Mutación , Neuronas/metabolismo , Neuronas/patología , Placa Amiloide/metabolismo , Placa Amiloide/patología , Placa Amiloide/prevención & control , Factores Protectores , Agregación Patológica de Proteínas/metabolismo , Agregación Patológica de Proteínas/patología , Agregación Patológica de Proteínas/prevención & control , Componente Amiloide P Sérico/metabolismo , TransgenesRESUMEN
Aberrant expression of miRNAs, cytokines and chemokines are involved in pathogenesis of colon cancer. However, the expression of p53 mediated miRNAs, cyto- and chemokines after radiation and SN38 treatment in colon cancer remains elusive. Here, human colon cancer cells, HCT116 with wild-type, heterozygous and a functionally null p53, were treated by radiation and SN38. The expression of 384 miRNAs was determined by using the TaqMan® miRNA array, and the expression of cyto- and chemokines was analyzed by Meso-Scale-Discovery instrument. Up- or down-regulations of miRNAs after radiation and SN38 treatments were largely dependent on p53 status of the cells. Cytokines, IL-6, TNF-α, IL-1ß, Il-4, IL-10, VEGF, and chemokines, IL-8, MIP-1α were increased, and IFN-γ expression was decreased after radiation, whereas, IL-6, IFN-γ, TNF-α, IL-1ß, Il-4, IL-10, IL-8 were decreased, and VEGF and MIP-1α were increased after SN38 treatment. Bioinformatic analysis pointed out that the highly up-regulated miRNAs, let-7f-5p, miR-455-3p, miR-98, miR-155-5p and the down-regulated miRNAs, miR-1, miR-127-5p, miR-142-5p, miR-202-5p were associated with colon cancer pathways and correlated with cyto- or chemokine expression. These miRNAs have the potential for use in colon cancer therapy as they are related to p53, pro- or anti-inflammatory cyto- or chemokines after the radiation and SN38 treatment.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Camptotecina/análogos & derivados , Quimiocinas/metabolismo , Quimioradioterapia , Neoplasias del Colon/terapia , Citocinas/metabolismo , MicroARNs/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Camptotecina/farmacología , Quimiocinas/genética , Neoplasias del Colon/genética , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Biología Computacional , Citocinas/genética , Bases de Datos Genéticas , Relación Dosis-Respuesta a Droga , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Heterocigoto , Humanos , Concentración 50 Inhibidora , Irinotecán , MicroARNs/genética , Mutación , Dosis de Radiación , Transducción de Señal , Factores de Tiempo , Transfección , Proteína p53 Supresora de Tumor/genéticaRESUMEN
The hallmarks of Alzheimer disease are amyloid-ß plaques and neurofibrillary tangles accompanied by signs of neuroinflammation. Lysozyme is a major player in the innate immune system and has recently been shown to prevent the aggregation of amyloid-ß1-40 in vitro. In this study we found that patients with Alzheimer disease have increased lysozyme levels in the cerebrospinal fluid and lysozyme co-localized with amyloid-ß in plaques. In Drosophila neuronal co-expression of lysozyme and amyloid-ß1-42 reduced the formation of soluble and insoluble amyloid-ß species, prolonged survival and improved the activity of amyloid-ß1-42 transgenic flies. This suggests that lysozyme levels rise in Alzheimer disease as a compensatory response to amyloid-ß increases and aggregation. In support of this, in vitro aggregation assays revealed that lysozyme associates with amyloid-ß1-42 and alters its aggregation pathway to counteract the formation of toxic amyloid-ß species. Overall, these studies establish a protective role for lysozyme against amyloid-ß associated toxicities and identify increased lysozyme in patients with Alzheimer disease. Therefore, lysozyme has potential as a new biomarker as well as a therapeutic target for Alzheimer disease.
Asunto(s)
Enfermedad de Alzheimer/enzimología , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Muramidasa/metabolismo , Fragmentos de Péptidos/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/ultraestructura , Animales , Encéfalo/patología , Muerte Celular , Drosophila melanogaster , Femenino , Humanos , Proteínas de Insectos/metabolismo , Locomoción , Masculino , Persona de Mediana Edad , Muramidasa/sangre , Muramidasa/líquido cefalorraquídeo , Muramidasa/farmacología , Fragmentos de Péptidos/ultraestructura , Placa Amiloide/metabolismo , Placa Amiloide/ultraestructura , Células Tumorales Cultivadas , Proteínas tau/metabolismoRESUMEN
We have created a Drosophila model of lysozyme amyloidosis to investigate the in vivo behavior of disease-associated variants. To achieve this objective, wild-type (WT) protein and the amyloidogenic variants F57I and D67H were expressed in Drosophila melanogaster using the UAS-gal4 system and both the ubiquitous and retinal expression drivers Act5C-gal4 and gmr-gal4. The nontransgenic w(1118) Drosophila line was used as a control throughout. We utilized ELISA experiments to probe lysozyme protein levels, scanning electron microscopy for eye phenotype classification, and immunohistochemistry to detect the unfolded protein response (UPR) activation. We observed that expressing the destabilized F57I and D67H lysozymes triggers UPR activation, resulting in degradation of these variants, whereas the WT lysozyme is secreted into the fly hemolymph. Indeed, the level of WT was up to 17 times more abundant than the variant proteins. In addition, the F57I variant gave rise to a significant disruption of the eye development, and this correlated to pronounced UPR activation. These results support the concept that the onset of familial amyloid disease is linked to an inability of the UPR to degrade completely the amyloidogenic lysozymes prior to secretion, resulting in secretion of these destabilized variants, thereby leading to deposition and associated organ damage.