Protein-Cell Assemblies as Tissue Mimics It is proposed is to directly assemble living cells and recombinant structural proteins into tissue mimics for subsequent tissue regeneration, therapeutic screening, and other biomedical applications. Electrospinning techniques have recently evolved into a powerful fiber formation tool for fabricating protein fibrous scaffolds of nano/micro-scale structures with excellent mechanical properties and enhanced cell-scaffold interaction. However, the overly small pores of an electrospun protein scaffold inhibit cells from efficiently migrating into the scaffold and regenerating a 3D tissue. A direct assembling of cells and proteins was recently achieved through a sophisticated, jet-based, 3D printing technology. Compared to an electrospun scaffold, the mechanical properties of a scaffold prepared by this method are far inferior. In this proposal, we plan to incorporate the cell printing concept into an electrospinning technique for the fabrication of protein-cell assemblies, which can resemble the structural and mechanical characteristics of native tissues. The development of tissue-mimetic protein-cell assemblies (TMPCA) may be an important step in engineering a functional tissue. In addition, TMPCA may provide researchers with a novel, in vitro cell- culture system for the study of biochemical pathways, disease mechanisms, and drug metabolisms, aiding in therapy development for many diseases including cancer and atherosclerosis.
Specific Aim 1. Coaxially electrospin recombinant silk-elastin-like proteins (SELPs) and crosslinking agents or coagulants into a robust fibrous scaffold without post-electrospinning treatments. The electrospinning parameters will be optimized to enhance the scaffold mechanical properties. Likewise, SELPs of varying polypeptide sequences and crosslinking sites will be synthesized, and the ability of these structural determinants to modulate the fiber formation and scaffold properties will be defined.
Specific Aim 2. Simultaneously integrate 3T3 fibroblasts (as a model system) into a SELP nanofibrous scaffold by concurrently electrospinning SELPs and electrospraying fibroblasts. Cell encapsulation by alginate and fast evaporation of crosslinking agents/coagulants will be pursued, in order to enhance cell viability. A mechanically robust tissue mimic with high cell densities will be fabricated.
Specific Aim 3. Examine the structural integrity, cell activities, and scaffold remodeling of the SELP- fibroblast assemblies during a post-fabrication period of up to one week. In particular, the mechanical properties will be accessed under both static and dynamic loading conditions. Likewise, cell viability, growth, and proliferation will be investigated during one week of post-fabrication follow-up. In this process, mathematical constitutive models will be used to aid in mechanical characterization.
Protein-Cell Assemblies as Tissue Mimics Native tissues are comprised of living cells and structural proteins in an organized way. We propose to assembly recombinant structural proteins and living cells into a robust, organized structure as a tissue mimic. Such a tissue mimic can be used to grow artificial tissues, and provide a mimetic tissue- environment to study disease mechanisms and drug metabolisms, aiding in the development of new therapy for various diseases.
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