RESUMEN
Glomerular scarring, known as glomerulosclerosis, occurs in many chronic kidney diseases and involves interaction between glomerular endothelial cells (GECs), podocytes, and mesangial cells (MCs), leading to signals that promote extracellular matrix deposition and endothelial cell dysfunction and loss. We describe a 3D tri-culture system to model human glomerulosclerosis. In 3D monoculture, each cell type alters its phenotype in response to TGFß, which has been implicated as an important mediator of glomerulosclerosis. GECs form a lumenized vascular network, which regresses in response to TGFß. MCs respond to TGFß by forming glomerulosclerotic-like nodules with matrix deposition. TGFß treatment of podocytes does not alter cell morphology but increases connective tissue growth factor (CTGF) expression. BMP7 prevents TGFß-induced GEC network regression, whereas TGFß-induced MC nodule formation is prevented by SMAD3 siRNA knockdown or ALK5 inhibitors but not BMP7, and increased phospho-SMAD3 was observed in human glomerulosclerosis. In 3D tri-culture, GECs, podocytes, and MCs form a vascular network in which GECs and podocytes interact intimately within a matrix containing MCs. TGFß treatment induces formation of nodules, but combined inhibition of ALK5 and CTGF is required to prevent TGFß-induced nodule formation in tri-cellular cultures. Identification of therapeutic targets for glomerulosclerosis depends on the 3D culture of all three glomerular cells. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Asunto(s)
Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Nefropatías Diabéticas/metabolismo , Nefropatías Diabéticas/patología , Glomérulos Renales/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Matriz Extracelular/metabolismo , Humanos , Enfermedades Renales/patología , Glomérulos Renales/metabolismo , Células Mesangiales/citología , Receptor Tipo I de Factor de Crecimiento Transformador betaRESUMEN
The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a variety of vascular beds. The role of developmental programming versus the tissue contextual environment for this specialization is not well understood. Here we describe a hierarchy of expression of HOX genes associated with endothelial cell origin and location. In initial microarray studies, differential gene expression was examined in two endothelial cell lines: blood derived outgrowth endothelial cells (BOECs) and pulmonary artery endothelial cells. This suggested shared and differential patterns of HOX gene expression between the two endothelial lines. For example, this included a cluster on chromosome 2 of HOXD1, HOXD3, HOXD4, HOXD8 and HOXD9 that was expressed at a higher level in BOECs. Quantative PCR confirmed the higher expression of these HOXs in BOECs, a pattern that was shared by a variety of microvascular endothelial cell lines. Subsequently, we analysed publically available microarrays from a variety of adult cell and tissue types using the whole "HOX transcriptome" of all 39 HOX genes. Using hierarchical clustering analysis the HOX transcriptome was able to discriminate endothelial cells from 61 diverse human cell lines of various origins. In a separate publically available microarray dataset of 53 human endothelial cell lines, the HOX transcriptome additionally organized endothelial cells related to their organ or tissue of origin. Human tissue staining for HOXD8 and HOXD9 confirmed endothelial expression and also supported increased microvascular expression of these HOXs. Together these observations suggest a significant involvement of HOX genes in endothelial cell positional identity.
Asunto(s)
Células Endoteliales/metabolismo , Perfilación de la Expresión Génica , Genes Homeobox , Proteínas de Homeodominio/genética , Adulto , Animales , Diferenciación Celular/genética , Línea Celular , Análisis por Conglomerados , Células Madre Embrionarias/metabolismo , Células Endoteliales/citología , Células Endoteliales/ultraestructura , Feto/embriología , Feto/metabolismo , Ontología de Genes , Proteínas de Homeodominio/metabolismo , Humanos , Pulmón/citología , Pulmón/embriología , Ratones , Neovascularización Fisiológica/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Especificidad de Órganos/genética , Fenotipo , Arteria Pulmonar/citología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los ResultadosRESUMEN
TNF signals through two distinct receptors, designated TNFR1 and TNFR2, which initiate diverse cellular effects that include cell survival, activation, differentiation, and proliferation and cell death. These cellular responses can promote immunological and inflammatory responses that eradicate infectious agents, but can also lead to local tissue injury at sites of infection and harmful systemic effects. Defining the molecular mechanisms involved in TNF responses, the effects of natural and experimental genetic diversity in TNF signalling and the effects of therapeutic blockade of TNF has increased our understanding of the key role that TNF plays in infectious disease.
Asunto(s)
Enfermedades Transmisibles/metabolismo , Receptores Tipo II del Factor de Necrosis Tumoral/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Muerte Celular , Enfermedades Transmisibles/genética , Regulación de la Expresión Génica , Humanos , Receptores Tipo I de Factores de Necrosis Tumoral/genética , Receptores Tipo II del Factor de Necrosis Tumoral/genética , Transducción de Señal , Factor de Necrosis Tumoral alfa/antagonistas & inhibidoresRESUMEN
Tumour necrosis factor (TNF) was originally described as a circulating factor that can induce haemorrhagic necrosis of tumours. It is now clear that TNF has many different functions in cancer biology. In addition to causing the death of cancer cells, TNF can activate cancer cell survival and proliferation pathways, trigger inflammatory cell infiltration of tumours and promote angiogenesis and tumour cell migration and invasion. These effects can be explained by the diverse cellular responses TNF can initiate through distinct signal transduction pathways, opening the way for more selective targeting of TNF signalling in cancer therapy.