A fluidic device to study directional angiogenesis in complex tissue and organ culture models

Abstract

Many signals that induce angiogenesis have been identified; however, it is still not clear how these signals interact to shape the vascular system. We have developed a fluidic device for generation of molecular gradients in 3-dimensional cultures of complex tissues and organs in order to create an assay for precise induction and guidance of growing blood vessels. The device features a centrally placed culture chamber, flanked by channels attached to a perfusion system used to generate gradients. A separate network of vacuum channels permits reversible attachment of the device to a flat surface. We show that the fluidic device can be used to create growth factor gradients that induce directional angiogenesis in embryonic mouse kidneys and in clusters of differentiating stem cells. These results demonstrate that the device can be used to accurately manipulate complex morphogenetic processes with a high degree of experimental control.

A disposable and multifunctional capsule for easy operation of microfluidic elastomer systems

“…This allows for reversible bonding of the elastomer insert to any surface of choice, if the users wish not to use the here described bottom piece together with the glass cover slip for closure of the system [10,12]. The central part of the capsule below the light window is open throughout the upper unit, importantly enabling conventional light microscopy of samples, as well as gas flux to centrally positioned microfluidic channels and cell culture compartments of the elastomer insert.…”

“…Gradients were readily formed in the cell culture chamber of the closed capsule system connected to a syringe pump (Harvard Apparatus) run at 0.5 µl min -1 . The cell culture chamber, flanked by a source and a sink channel was filled with a collagen I matrix that was allowed to polymerize before closing the capsule, as previously described [10]. Figure 4 shows how an approximately linear gradient (0-2 µg ml -1 ) of FITC-dextran (MW=20 kDa; Sigma Aldrich) was formed within 14 hours.…”

“…A commercial product based on the present design reaching end users would consist of an assembled upper unit, together with a bottom piece and a circular glass cover slip. The user would then load a biological sample of interest into the cell culture chamber of the insert [10], before assembling the capsule by snapping the bottom piece to the upper unit, thereby pressing the cover slip against the elastomer insert to seal the fluidic channels (figure 2).…”

“…In many of the microfluidic cell culture systems, the fluidic channels and the cavities for cell culture are created in polydimethylsiloxane (PDMS) sealed by bonding to a glass surface. These assays enable real-time studies of cell migration, proliferation and differentiation in response to concentration gradients of soluble signaling molecules (often proteins), in both two-and three-dimensional settings [5][6][7][8][9][10][11]. Conceivably, some of these methods based on microfluidic technology could become new global standards, and thereby in part replace techniques such as for example the Boyden chamber assay for the study of chemotaxis.…”

Microfluidic technology in vascular research : the endothelial response to shear stress

“…However, using the laminar flow properties discussed earlier in this review, hydrogels can also be formed in situ and even be patterned and confined to certain regions of the microfluidic device. 87 The great potential of these three-dimensional culturing techniques was recently underlined by Barkefors et al, 11 who cultured ex vivo kidney tissue and followed the formation of blood vessels in response to a VEGF 165 gradient. Because of the small scale of microfluidic devices, it is possible to advance this proof-of-concept study towards high-throughput assays in order to screen for compounds that affect blood vessel formation in such realistic models.…”