RESUMO
The use of exogenous signals is gaining importance in renal regenerative therapies. We wanted to explore the role of extracellular matrix (ECM) constituents on renal structure formation during renal organogenesis. We used a recently established organ culture setup to expose embryonic kidney rudiments directly to a large set of surface-immobilized or dissolved ECM molecules and growth factors. Organ culture was also performed on immobilized adult kidney ECM extracts and on reactive polymer films without any biomolecular components. The applied conditions resulted in distinct differences of organ phenotypes, underlining the multifaceted role of exogenous signals during kidney development. Specific ECM components, including collagen I and laminin, supported nephronal and tubular structure formation of the developing organ. ECM biopolymers, e.g. hyaluronic acid, were found to determine the fate of developing explants in a concentration- and molecular weight-dependent manner. The organ culture system used was an effective and robust means to identify exogenous signals that direct kidney development. This system can provide valuable insight for future regenerative therapies of kidney diseases.
Assuntos
Matriz Extracelular/química , Rim/citologia , Rim/embriologia , Técnicas de Cultura de Órgãos/métodos , Engenharia Tecidual/métodos , Animais , Células Cultivadas , Colágeno/química , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Proteínas da Matriz Extracelular/química , Proteínas da Matriz Extracelular/metabolismo , Feminino , Imunofluorescência , Ácido Hialurônico/química , Ácido Hialurônico/metabolismo , Rim/metabolismo , Laminina/química , Laminina/metabolismo , Camundongos , Néfrons/citologia , Néfrons/metabolismo , Organogênese/fisiologia , GravidezRESUMO
Here, we present a novel method for culturing kidneys in low volumes of medium that offers more organotypic development compared to conventional methods. Organ culture is a powerful technique for studying renal development. It recapitulates many aspects of early development very well, but the established techniques have some disadvantages: in particular, they require relatively large volumes (1-3 mls) of culture medium, which can make high-throughput screens expensive, they require porous (filter) substrates which are difficult to modify chemically, and the organs produced do not achieve good cortico-medullary zonation. Here, we present a technique of growing kidney rudiments in very low volumes of medium-around 85 microliters-using silicone chambers. In this system, kidneys grow directly on glass, grow larger than in conventional culture and develop a clear anatomical cortico-medullary zonation with extended loops of Henle.
