Cells of the retinal pigment epithelium (RPE) are generated prenatally and generally survive the lifetime of the individual without undergoing proliferation or replacement. Therefore, the mechanisms promoting individual RPE cell survival and longevity in vivo may be distinct from, or a limited subset of, the mechanisms known to promote survival in proliferative cells in culture. To identify specific factors that sustain cell viability independent of effects on cell division, we studied RPE cells in low-density suspension culture, in which cell proliferation is inhibited. Single cells from Xenopus laevis eyes were plated onto a non-adhesive surface in protein-free medium, then assayed for survival using the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cell viability in these cultures was essentially undiminished over the initial 2 days. However, by approximately 1 week in culture, only an average of 53% of the cells remained alive. Plating cells on a fibronectin-coated substratum significantly enhanced survival, such that the number of cells alive at 1 week was 80-90% of the initial level. Essentially identical results were obtained with laminin- or collagen IV-coated substrata, or with insulin (5μg ml-1) in the medium. The absence of cell division in these cultures was confirmed by cell counting and BrdU incorporation experiments. Interestingly, in suspension cultures derived from monolayers previously established on microporous membrane filters, cells lost viability much faster (average of 80% dead at 3 days), and showed a relatively greater response to extracellular matrix proteins (five-fold increase in cell survival at 3 days). Enhanced RPE survival in response to fibronectin required spreading of the cell on a substratum, rather than mere adherence, as there was a high correlation between the percentage of spread cells and the percentage that were MTT-positive (r = 0·940). Cell spreading apparently enhanced survival by preventing the initiation of programmed cell death: unattached non-viable cells in culture exhibited morphological features expected of apoptosis, as well as positive staining by the TUNEL reaction. These studies demonstrate that, of several factors shown to maintain or increase cell number in proliferating cultures, some have their effect, at least in part, by promoting the survival of individual cells. The increased susceptibility of subcultured RPE to cell death has implications for clinical transplantation applications that may require manipulation of RPE in vitro.

The presented doctoral research was aimed at preparation and characterization of ultra thin polymer films on solid substrates using different physical techniques. Each of these physical techniques probes selectively different characteristics of the films. While some of the techniques are strong in the predetermination of some unique properties of the layers, they might be limited and give no specific/conclusive information about some other important characteristics. Therefore, only the combination of the techniques provides a profound picture of the thickness, architecture, composition and functionality of the films/layers. This combined characterization approach elucidates in details the physical characteristics and the mechanisms responsible for the unique behavior of different polymer films/layers in the application that they are intended for. In the thesis, of particular interest were films of high biomedical, biotechnological and tissue engineering importance, such as: 1. poly(lactide) films formed by grafting "from-" a silanized alacrite thin films (L605 Co-based super alloy), 2. polydopamine (PDA) films that could serve as substrate independent mod- ification platform for further surface modification steps, 3. poly(ethylene oxide)films formed by "grafting to-" PDA modified surfaces, 4....