Cellular response to nanoscale elastin-like polypeptide polyelectrolyte multilayers.

Ionic elastin-like polypeptide (ELP) conjugates are a new class of biocompatible, self-assembling biomaterials. ELPs composed of the repeat unit (GVGVP)(n) are derived from the primary sequence of mammalian elastin and produced in Escherichia coli. These biopolymers exhibit an inverse transition temperature that renders them extremely useful for applications in cell-sheet engineering. Cationic and anionic conjugates were synthesized by the chemical coupling of ELP to polyethyleneimine (PEI) and polyacrylic acid (PAA). The self-assembly of ELP-PEI and ELP-PAA using the layer-by-layer deposition of alternately charged polyelectrolytes is a simple, versatile technique to generate bioactive and biomimetic surfaces with the ability to modulate cell-substratum interactions. Our studies are focused on cellular response to self-assembled multilayers of ionic (GVGVP)(40) incorporated within the polymeric sequence H(2)N-MVSACRGPG-(GVGVP)(40)-WP-COOH. Angle-dependent XPS studies indicated a difference in the chemical composition at the surface ( approximately 10A below the surface) and subsurface regions. These studies provided additional insight into the growth of the nanoscale multilayer assembly as well as the chemical environment that the cells can sense. Overall, cellular response was enhanced on glass substrata coated with ELP conjugates compared with uncoated surfaces. We report significant differences in cell proliferation, focal adhesions and cytoskeletal organization as a function of the number of bilayers in each assembly. These multilayer assemblies have the potential to be successfully utilized in the rational design of coatings on biomaterials to elicit a desired cellular response.

Characterizing the dielectric properties of human mesenchymal stem cells and the effects of charged elastin-like polypeptide copolymer treatment.

HUMAN MESENCHYMAL STEM CELLS (HMSCS) HAVE THREE KEY PROPERTIES THAT MAKE THEM DESIRABLE FOR STEM CELL THERAPEUTICS: differentiation capacity, trophic activity, and ability to self-renew. However, current separation techniques are inefficient, time consuming, expensive, and, in some cases, alter hMSCs cellular function and viability. Dielectrophoresis (DEP) is a technique that uses alternating current electric fields to spatially separate biological cells based on the dielectric properties of their membrane and cytoplasm. This work implements the first steps toward the development of a continuous cell sorting microfluidic device by characterizing native hMSCs dielectric signatures and comparing them to hMSCs morphologically standardized with a polymer. A quadrapole Ti-Au electrode microdevice was used to observe hMSC DEP behaviors, and quantify frequency spectra and cross-over frequency of hMSCs from 0.010-35 MHz in dextrose buffer solutions (0.030 S/m and 0.10 S/m). This combined approach included a systematic parametric study to fit a core-shell model to the DEP spectra over the entire tested frequency range, adding robustness to the analysis technique. The membrane capacitance and permittivity were found to be 2.2 pF and 2.0 in 0.030 S/m and 4.5 pF and 4.1 in 0.10 S/m, respectively. Elastin-like polypeptide (ELP-) polyethyleneimine (PEI) copolymer was used to control hMSCs morphology to spheroidal cells and aggregates. Results demonstrated that ELP-PEI treatment controlled hMSCs morphology, increased experiment reproducibility, and concurrently increased hMSCs membrane permittivity to shift the cross-over frequency above 35 MHz. Therefore, ELP-PEI treatment may serve as a tool for the eventual determination of biosurface marker-dependent DEP signatures and hMSCs purification.