title: A combinatorial approach for directing the amount of fibronectin fibrils assembled by cells that uses surfaces derivatized with mixtures of fibronectin and cell binding domains.
abstract: Fibrillar fibronectin (FN) has the crucial role of attracting and attaching cells as well as molecules that mediate tissue repair during wound healing. A previous study demonstrated higher extracellular staining of FN fibrils in cells cultured on surfaces tethered with an equimolar mixture of a FN binding domain and FN's cell binding domain, III1-2 and III9-10 respectively, than on surfaces with III9-10 alone. The effect of varying surface amounts of III1-2 and III9-10 on the quantity of FN fibrils formed by NIH-3T3 fibroblasts was examined. GST tagged III1-2 and III9-10 were conjugated to polyurethane surfaces and ELISAs were used to identify the experimental design space or the range of concentrations of GST-III1-2 and GST-III9-10 that demarcated the limits of protein loading on the surface. When GST-III1-2 was fixed and GST-III9-10 varied within the design space, the amount of FN fibrils measured by immunoblotting detergent insoluble cell lysates was dependent on the ratio of III9-10 to III1-2. When the total protein concentration was fixed and the mixture composition of GST-III1-2 and GST-III9-10 varied such that it optimally covered the design space, a parabolic relationship between FN fibril amount and the ratio of III9-10 to III1-2 was obtained. This relationship had a maximum value when the surface was bonded to equal amounts of III1-2 and III9-10 (P<0.05). Thus the ratio of III9-10 to III1-2 can be utilized to direct the quantity of FN fibrils formed on surfaces.
authors.affiliation: Department of Chemical and Biological Engineering, Illinois Institute of Technology, Chicago, IL 60616, USA.
authors.forename: Pradnya P
authors.initials: PP
authors.lastname: Kshatriya
authors.forename: Stella W
authors.initials: SW
authors.lastname: Karuri
authors.forename: Chunyi
authors.initials: C
authors.lastname: Chiang
authors.forename: Nancy W
authors.initials: NW
authors.lastname: Karuri
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D020650:Combinatorial Chemistry Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D005353:Fibronectins / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011485:Protein Binding
chemical_list: D005353:Fibronectins; D005982:Glutathione Transferase