Self-Assembling Membranes and Related Methods Thereof (27009)
A self-assembling liquid polymeric system for the synthesis of robust sacs and string membrane structures having properties suitable for cell and therapeutic delivery, diagnostic and tissue repair applications.
ADVANTAGES: Facile generation of biocompatible membrane structures with excellent mechanical, diffusion and stability properties to encapsulate and deliver a wide range of medicinal and biological materials.
SUMMARY: The organization of molecules at interfaces is of great importance in the preparation of chemically defined surfaces and ordered materials. This invention provides self-assembly of ordered materials well beyond the monolayer scale at an aqueous liquid-liquid interface by the direct combination of a peptide amphiphile (PA) and the polysaccharide hyaluronic acid (HA). PAs are small synthetic molecules containing a hydrophobic C16 alkyl segment and grafted to a peptide sequence such as V3A3K3. HA is a negatively charged high molecular weight disaccharide of N-acetylglucosamine and glucuronic acid. Combining equal volume solutions of PA and HA, immediately form a solid membrane localized at the interface between the two liquids. With the denser HA solution placed on top of the PA solution, the HA fluid sinks into the PA component causing renewal of the liquid-liquid interface and resulting in continuous growth of membrane until the entire volume of HA solution is engulfed, resulting in a polymer-filled sac. Alternatively, closed sacs can be made instantly by injecting one solution directly into the bulk of the other, creating either HA-filled or PA-filled sacs. Ordered structures of self-sealing sacs (Fig.1,B to D), films with tailorable size and shape (Fig.1E), as well as continuous strings (Fig.1F) are available with these systems. The PA-HA membranes exhibit significant tensile strength with elastic moduli in the dry and hydrated states of the membrane about 670 and 0.9 MPa, respectively. For comparison, the elastic modulus of PA-HA membranes was found to be about nine times higher than that of the high molecular weight chitosan-gellan membranes, also known to yield macroscopic capsules and strings. Hydrated PA-HA membranes also exhibit minimal dimensional change and remain stable over months in either water or phosphate-buffered saline, in contrast to the chitosan-gellan membranes.
Permeability of the PA-HA membrane was demonstrated by monitoring diffusion of transforming growth factor–β1 (TGF-β1, 25 kD) out of a PA gel-filled sac versus a control PA gel without the sac membrane. Results over a 2-week period revealed similar TGF-β1 release kinetics between the gels with and without the surrounding membrane (Fig. 2H), confirming the permeability of the HA-PA membrane to proteins.
PA-HA structures also support cell viability and differentiation. In vitro studies using human mesenchymal stem cells (hMSCs) incorporated within gel-filled sacs and cultured in growth media remained viable within the sacs up to 4 weeks in culture. Furthermore, results with real-time reverse transcription polymerase chain reaction after 2 weeks in culture revealed an increase in collagen type II gene expression when cells in sacs were stimulated with TGF or chondrogenic media, indicating that hMSCs were able to differentiate toward a chondrogenic phenotype within the sacs (Fig. 2J). The self-assembling sacs providing sufficient nutrient diffusion for cell survival and differentiation.
STATUS: A patent application has been filed and Northwestern University seeks to develop the technology. Reference Science 319, 1812-1816 (2008)
H) TGF-β1 release from PA gel-filled sacs and PA gels as a function of time, demonstrating a nearly identical protein-release profile. (J) Collagen type II gene expression of hMSCs cultured within the PA gel–filled sacs in growth medium (GM), growth medium supplemented with TGF-β1 (TGF-GM), chondrogenic medium (CM), and chondrogenic medium supplemented with TGF-β1 (TGF-CM).
Samuel Stupp, Helena Azevedo, Ramille Capito
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