In vitro culture of rat neuromicrovascular endothelial cells on polymeric scaffolds Maria Teresa Conconi, 1 Silvano Lora, 2 Silvia Baiguera, 1 Elisa Boscolo, 1 Marcella Folin, 3 Renato Scienza, 4 Piera Rebuffat, 5 Pier Paolo Parnigotto, 1 Gastone Giovanni Nussdorfer 5 1 Department of Pharmaceutical Sciences, University of Padua, Padua, Italy 2 Institute of Organic Synthesis and Photoreactivity, C.N.R., Legnaro, Italy 3 Department of Biology, University of Padua, Padua, Italy 4 Department of Neurosurgery, Padua Regional Hospital, Padua, Italy 5 Department of Human Anatomy and Physiology, Section of Anatomy, University of Padua, Via Gabelli 65, I-35121 Padua, Italy Received 8 March 2004; revised 30 July 2004; accepted 10 August 2004 Published online 21 October 2004 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.a.30198 Abstract: Polyphosphazenes are polymers possessing a skeleton composed of alternating phosphorous and nitrogen atoms, and two side-moieties linked to each phosphorous atom. Polyphosphazenes with amino acid esters as side- moieties are biocompatible and biodegradable polymers. Two polyphosphazenes, poly[bis(ethyl alanate) phosphazene] and poly[(ethyl phenylalanate) 0.8 (ethyl alanate) 0.8 (ethyl glyci- nate) 0.4 phosphazene] (PPAGP) were synthesized, and pro- cessed to form small fibers. Their ability to support rat neu- romicrovascular endothelial cell (EC) adhesion and growth has been studied, using poly(D,L-lactic acid) as reference com- pound. Scanning electron microscopy revealed that both poly- [bis(ethyl alanate) phosphazene] and PPAGP fibers were thin- ner than poly(D,L-lactic acid) fibers, and possessed a more irregular and porous surface. All polymers increased EC ad- hesion, compared with polystyrene, but only polyphospha- zenes were able to improve EC growth. The highest increase in EC proliferation was induced by PPAGP, which, as revealed by environmental scanning electron microscopy, was also able to induce ECs to arrange into tubular structures. The conclu- sion is drawn that PPAGP may provide the best scaffold for engineered blood vessels, because it promotes adhesion, growth, and organization of ECs into capillary-like structures. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 71A: 669 – 674, 2004 Key words: neuromicrovascular endothelial cells; cell growth; cell adhesion; biomaterials; polyphosphazenes INTRODUCTION The in vitro culture of neuromicrovascular endothe- lial cells (ECs) on polymeric scaffolds may represent an important tool to study in vitro the blood– brain barrier (BBB) physiology, and to improve neovascu- larization in tissue engineering. An in vitro BBB model has been proposed, in which rat ECs and astrocytes are cocultured on either poly(ethylene terephthalate)- or type IV collagen-coated polycarbonate filters. 1,2 To- day, constructs composed of synthetic or natural scaf- folds, supporting adhesion and proliferation of autol- ogous seeded cells, are often used in tissue repairing. 3 However, a major problem of in vivo implants is the low rate of cell survival because of delayed neovascu- larization, so that it has been proposed to coculture tissue-specific cells with ECs, which, being able to form capillary-like tubules, could accelerate the recon- stitution of the capillary bed of the recipient. 4,5 The most commonly used biocompatible and biode- gradable polymeric scaffold is poly(D,L-lactic acid) (PDLLA), which has been used to support the in vitro growth of ECs from rabbit cornea, human umbilical vein, and rat thoracic aorta. 4,6,7 Other potentially use- ful scaffolds are polyphosphazenes, high-molecular- weight polymers with the molecular structure [NPR 2 ] n (Fig. 1). All these macromolecules have a skeleton composed of alternating phosphorous and nitrogen atoms and two side-moieties (R) bound to each phos- phorous atom. The choice of the side groups allows modulation of mechanical properties, surface charac- teristics, and hydrolytic sensitivity of polyphospha- zenes. Polyphosphazenes with amino acid esters as side groups are biodegradable and their hydrolysis products (phosphate, ammonia, and amino acids) are well compatible with the growth of mammalian cells. Correspondence to: G. G. Nussdorfer; e-mail: gastone. nusdorfer@unipd.it © 2004 Wiley Periodicals, Inc.