Cell Adhesion and Motility on Biocompatible Polyelectrolyte Multilayers
Martinez, Jessica Susanne (author)
Keller, Thomas C. S. (professor directing dissertation)
Schlenoff, Joseph B. (university representative)
Keller, Laura R. (committee member)
Ma, Teng (committee member)
Lenhert, Steven, 1977- (committee member)
Florida State University (degree granting institution)
College of Arts and Sciences (degree granting college)
Department of Biological Science (degree granting department)
To improve the design of prostheses surfaces, our research group investigates how biocompatible polyelectrolyte multilayers (PEMUs) can be constructed to serve as coatings for biomedical implants, providing a versatile, inexpensive, and potentially efficient solution to create anti-bacterial, anti-inflammatory, and biologically selective surfaces. More specifically, this dissertation research investigates how individual cells and cell sheets adhere and migrate on PEMUs constructed to have uniform and gradients of modulus and how individual cells and gram negative bacteria, Escherichia coli, adhere to PEMUs constructed to have an anti-adhesive surface chemistry. In this investigation, PAH/PAA PEMUs are shown to be biocompatible compared to the soluble polycation PAH at concentrations above 0.1mM. Soluble PAH concentrations at 1 and 10mM cause irreversible damage to the plasma membrane of smooth muscle, A7r5, and bone, U2OS, cells. Additionally, adhesive and motile responses of cells are dependent on PEMU surface chemistry. Cells on PEMUs terminated with the polycation PAH relocalize their focal adhesions to their cell periphery and are highly motile compared to cells cultured on PAA terminated PEMUs and uncoated glass coverslips. To investigate effects of PEMU modulus on cell adhesion and motility, PEMUs were made with the polyanion PAA (poly(acrylic acid)) modified with a photosensitive 4-(2-Hydroxyethoxy) benzophenone (PAABp) and the polycation PAH (poly(allylamine hydrochloride)). UV irradiating PAH/ PAABp PEMUs forms covalent bonds between PE layers and consequently increases its Young's Elastic Modulus, while retaining innate surface chemistry. Individual cells and cell sheets detect differences in PEMU modulus and respond by varying morphology and behavior. These PAH/PAABp PEMUs modulate the adhesion, spreading, and migration of individual cells, specifically smooth muscle, bone, and fibroblast cells. PAABp containing PEMUs were constructed to have either a shallow (~5MPa mm-1) or a steep (~50MPa mm-1) modulus gradient. Only smooth muscle cells durotax along steep modulus gradients toward increasing modulus and orient toward increasing modulus on shallow modulus gradients. In contrast, bone cells discriminately adhere to the stiffest region of both steep and shallow modulus gradients and fibroblasts show no difference in behavior along any region of the gradients. Epithelial sheets, isolated as primary explants of fish epithelial tissue from the scales of fish Poecilia sphenops (Black Molly) and Carassius auratus (Comet Goldfish), orient toward increasing modulus on steep modulus gradient. Cell sheets collectively durotax near the ~90MPa region of the gradient toward increasing modulus. Surfaces with substantial zwitterionic functionality (possessing a net neutral surface charge due to equal contribution of both positive and negative charges in polymer side groups) have been shown to effectively prevent cell and protein attachment. PEMUs built with PAH (poly(allylamine hydrochloride)) and PAA (poly(acrylic acid)) containing the AEDAPS zwitterionic group 3-(2-(acrylamido)-ethyldimethyl ammonio) propane sulfonate (PAH/PAA-co-AEDAPS PEMUs) and a new benzophenone crosslinker to stiffen the thin film were shown to prevent rat aortic smooth muscle (A7r5) and mouse fibroblast (3T3) cells attachment, but failed to prevent irreversible attachment of biofilm-forming gram-negative bacteria Escherichia coli, strain ATCC-8739. AEDAPS containing PEMUs are hydrophilic and have increased nanoroughness of ~10nm. 'Super soaking' AEDAPS PEMUs incorporates more zwitterions into the PEMU and significantly maximizes the surface presentation of PAA-co-AEDAPS, which promotes early attachment of bacteria, but eventually, causes a gradual decrease in bacteria attachment with increasing incubation time. This investigation provides further insight into the possible application of PEMUs as bioselective thin film coatings, which may have potential for use in biomedical applications.
Biocompatibility, Cell Adhesion, Durotaxis, Epithelial Cell Sheet, Motility, Polyelectrolyte Multilayers
October 26, 2015.
A Dissertation submitted to the Department of Biological Science in partial fulfillment of the Doctor of Philosophy.
Includes bibliographical references.
Thomas C. S. Keller, III, Professor Directing Dissertation; Joseph B. Schlenoff, University Representative; Laura R. Keller, Committee Member; Teng Ma, Committee Member; Steven Lenhert, Committee Member.
Florida State University