The ability to culture complex organs is currently an important goal in biomedical research. It is possible to grow organoids (3D organ-like structures) in vitro; however, a major limitation of... Show moreThe ability to culture complex organs is currently an important goal in biomedical research. It is possible to grow organoids (3D organ-like structures) in vitro; however, a major limitation of organoids, and other 3D culture systems, is the lack of a vascular network. Protocols developed for establishing in vitro vascular networks typically use human or rodent cells. A major technical challenge is the culture of functional (perfused) networks. In this rapidly advancing field, some microfluidic devices are now getting close to the goal of an artificially perfused vascular network. Another development is the emergence of the zebrafish as a complementary model to mammals. In this review, we discuss the culture of endothelial cells and vascular networks from mammalian cells, and examine the prospects for using zebrafish cells for this objective. We also look into the future and consider how vascular networks in vitro might be successfully perfused using microfluidic technology. Show less
One of the major limitations in culturing complex tissues or organs is the lack of vascularization in the cultured tissue. Development of a functional capillary bed could overcome this problem.... Show moreOne of the major limitations in culturing complex tissues or organs is the lack of vascularization in the cultured tissue. Development of a functional capillary bed could overcome this problem. The zebrafish is a promising model for in vitro vasculogenesis and angiogenesis studies, as a replacement for currently used mammalian models. However, the culture of endothelial cells from this species is not well characterized. Here, we test different culture strategies, medium supplementations and culture substrates for their effect on the generation of putative endothelial (fli:GFP+ and kdrl:GFP+) cells and vascular morphogenesis in zebrafish blastocyst cell derived embryoid body culture. we have also developed a perfused culture model, using microfluidic technology, to culture zebrafish vascular networks. This study is a step forward to the development of zebrafish vascular networks in vitro. Show less
A major limitation to culturing tissues and organs is the lack of a functional vascular network in vitro. The zebrafish possess many useful properties which makes it a promising model for such... Show moreA major limitation to culturing tissues and organs is the lack of a functional vascular network in vitro. The zebrafish possess many useful properties which makes it a promising model for such studies. Unfortunately, methods of culturing endothelial cells from this species are not well characterised. Here, we tried two methods (embryoid body culture and organ explants from transgenic zebrafish kdrl:GFP embryos) to develop in vitro vascular networks. In the kdrl:GFP line, endothelial cells expresses green fluorescent protein, which allows to track the vascular development in live cultures. We found that embryoid bodies showed significantly longer and wider branches of connected endothelial cells when grown in a microfluidic system than in static culture. Similarly, sprouting of kdrl:GFP+ cells from the tissue explants was observed in a 3D hydrogel matrix. This study is a step towards the development of zebrafish vascular networks in vitro. Show less
Ent, W. van der; Veneman, W.J.; Groenewoud, A.; Chen, L.P.; Tulotta, C.; Hogendoorn, P.C.W.; ... ; Langenau, D.M. 2016
Zebrafish embryos can be obtained for research purposes in large numbers at low cost and embryos develop externally in limited space, making them highly suitable for high-throughput cancer studies... Show moreZebrafish embryos can be obtained for research purposes in large numbers at low cost and embryos develop externally in limited space, making them highly suitable for high-throughput cancer studies and drug screens. Non-invasive live imaging of various processes within the larvae is possible due to their transparency during development, and a multitude of available fluorescent transgenic reporter lines. To perform high-throughput studies, handling large amounts of embryos and larvae is required. With such high number of individuals, even minute tasks may become time-consuming and arduous. In this chapter, an overview is given of the developments in the automation of various steps of large scale zebrafish cancer research for discovering important cancer pathways and drugs for the treatment of human disease. The focus lies on various tools developed for cancer cell implantation, embryo handling and sorting, microfluidic systems for imaging and drug treatment, and image acquisition and analysis. Examples will be given of employment of these technologies within the fields of toxicology research and cancer research. Show less
