MICRO-AND NANOENGINEERING APPROACHES TO DEVELOPING GRADIENT BIOMATERIALS SUITABLE FOR INTERFACE TISSUE ENGINEERING - Micro and Nanotechnologies in Engineering Stem Cells and Tissues - page 59

aligned and moved along the gradient. 5 In an interesting study, DeLong et al. cultured
vascular smooth muscle cells (SMCs) on a gradient of basic fibroblast growth factor
(bFGF) immobilized on a PEG substrate with RGD adhesion sites and showed cell
migration along the gradient. 10 Jiang et al. have proposed a general method for
immobilizing biomolecular gradients on a surface through the use of avidin-biotin
bonding. 43 These researchers first immobilized a gradient of avidin on a surface by
adsorption and then added a biotinylated molecule of interest. Knapp et al. used a
chamber filled with collagen or fibrin gel that was divided into two parts by a Teflon
barrier. 44 Human foreskin fibroblast (HFF) cells were encapsulated in one gel part;
the other part contained the soluble fibronectin peptide Gly-Arg-Gly-Asp-Ser-Pro
(GRGDSP), which is a fibroblast chemotactic factor. When the Teflon barrier was
removed, the peptide diffused into the second gel part, forming a gradient of
GRGDSP that induced the alignment and migration of the fibroblasts toward the
region of higher peptide concentration. Other examples of morphogen gradients
include those related to angiogenesis and axonal growth. PC12 neurites were promoted
and guided when cultured in a nerve growth factor (NGF) gradient immobilized on
poly(2-hydroxyethyl methacrylate) (p(HEMA)) or when cultured in a p(HEMA)/poly-
L -lysine (PLL) scaffold loaded with NGF and neurotrophin-3 (NT-3) concentration
gradients. 45,46 Similarly, primary fetal neural stem cells (NSCs) showed a rapid
induction of glial fibrillary acidic protein (GFAP) when cultured on a hydrogel
with a ciliary neurotrophic factor (CNTF) gradient. 47 In the same context, endothelial
cells migrated along a surface density gradient of VEGF and formed sprouting
elements when exposed to a VEGF gradient in a collagen gel. 31,48 Biological gradients
have also been used to study cell metastasis in cancer. 49 Another dynamic research area
involving morphogen gradients focuses on bone and cartilage engineering. He et al.
have shown that RGD and BMP peptides found in bone morphogenetic protein-2
(BMP-2) acted synergistically when grafted onto a hydrogel to induce bone marrow
stromal (BMS) cell osteogenesis and mineralization. 50 Cooper et al. printed a BMP-2
gradient on a DermaMatrix scaffold to demonstrate the spatial control of osteoblast
differentiation. 51 Dormer et al. used poly(lactic-co-glycolic acid) (PLGA) micro-
spheres loaded with BMP-2 or transforming growth factor beta-1 (TGF 1) to generate

a three-dimensional (3D) scaffold with cross-gradients of those biomolecules. 52 When
loaded with human umbilical cord mesenchymal stromal cells (hUCMSCs) or human
bone marrow stromal cells (hBMSCs), these gradient scaffolds exhibited spatial and
temporal control of the protein release with ECM formation, glycosaminoglycan
production, or alkaline phosphatase activity along the increasing concentration
gradient. These and other experimental studies have demonstrated that the influence
of biological gradients regulates cell behavior.

b

3.3 MICRO- AND NANOENGINEERING TECHNIQUES FOR
FABRICATING GRADIENT BIOMATERIALS

An ideal tissue engineering scaffold should mimic the structure and function of
native ECM, in which cells and tissue are organized into 3D architectures and are