Autonomously vascularized cellular constructs in tissue engineering: opening a new perspective for biomedical science

Abstract

In tissue engineering cell cultures play a crucial role besides the matrix materials for the end of substituting lost tissue functions. The cell itself is situated at the cross-roads leading to different orders of scale, from molecule to organism and different levels of function, from biochemistry to macrophysiology. Extensive in vitro investigations have dissected a vast amount of cellular phenomena and the role of a number of bioactive substances has been elucidated in the past. Further, recombinant DNA technologies allow modulation of the expression profiles of virtually all kinds of cells. However, issues of vascularization in vivo limit transferability of these observations and restrict upscaling into clinical applications. Novel in vivo models of vascularization have evolved inspired from reconstructive microsurgical concepts and they encompass axial neovascularization by means of vascular induction. This work represents a brief description of latest developments and potential applications of neovascularization and angiogenesis in tissue engineering.

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The arteriovenous fistula in situ. The femoral vein (v) and artery (a) in the left medial thigh distal to the inguinal ligament (i) are dissected from the femoral nerve (n) and a vascular graft (g) from the contralateral femoral vein is interposed by means of micro-surgical anastomosis.

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Schematic representation of the isolation chamber. It is comprised by a base plate (B) (diameter: 15 mm), under a cylindrical shell (height 6 mm × diameter 12 mm) (C) and an upper lid (L) (height: 2 mm × diameter: 14 mm). At the sides there are perforations for fixation of the chamber on the fascia of the medial musculature of the medial thigh.

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Vascularization into the fibrin clot after 2 weeks. Prior to explantation the caudal circulatory system was perfused with an India ink gel for black coloration of the capillary network. Four poly-ethylene stabs serve for stabilization of the AV loop in the isolation chamber.

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Custom-made design of a biogenic processed bovine cancellous bone matrix. A scanning electron image with the discoid matrix displaying a circular notch for optimal accommodation of the AV loop is shown here. The matrix was rendered conductive prior to sputtering by means of minute copper wires. Eight canals for secondary injection of fibrin immobilized osteoblasts were included in the design.

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Cell survival study with CFDA marker.Transplantation of differentiated primary rat osteoblasts into the prevascularized matrices significantly enhanced initial cell survival (40 × magnification).

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Construction of the AV loop generates a dense neovascular network. Here scanning electron microscopical images of a corrosion cast two weeks after implantation of an AV loop. Note the uniform diameter of the relatively immature neocapillaries.

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Sprouting (*) and branching (+) in the neocapillary network. Incomplete filling of the capillaries (#) is characterized by a rounded tip. Sprouts are marked by spiked tips. (Scanning electron microscopy of corrosion casts.)

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Non-sprouting angiogenesis or intussusceptive angiogenesis.(Scanning electron microscopy of corrosion casts.)

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Micro-magnetic resonance angiography of the AV loop 1 weekafter implantation.

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The process of neovascularization as induced by the AV loop.

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Molecular regulation of angiogenesis. Vascular development proceeds through three different steps (sprouting, assembly and maturation). Each step is regulated by a distinct set of agonists acting via specific vascular receptor tyrosine kinases (red).