The present thesis studied the application of polyallyldiglycol carbonate (or PADC) polymer films for fabrication of two types of specially-designed cell culture substrates. The process exploited the special property of PADC films that pits were formed on them after alpha-particle irradiation and subsequent chemical etching. The first type of substrates were just PADC films with pits (used in Chapters 2, 3 and 5), and were referred to as (psuedo) three-dimensional (3D) substrates. The second type of substrates were polydimethylsiloxane (PDMS) films with micropillars fabricated as replicas of PADC films with pits (used in Chapter 6). The HeLa cell line was used for testing these culture substrates. The primary objective was to identify changes in behavior in the HeLa cells cultured on these specially-designed cell culture substrates, in terms of cell forming mono-/multiple- layers and vinculin expression (Chapter 2), microtubule (MT) growth rate (Chapter 3), cell sensitivity to ionizing radiation (Chapter 5), as well as the percentage of cells entering the S-phase, attached cell numbers and cell spreading areas (Chapter 6). An introduction to the characteristics of pit formation on PADC films was first given in Chapter 1. These characteristics were used for fabrication of 3D substrates (Chapters 2-5) and pillar substrates (Chapter 6) In Chapter 2, we explored the fabrication of 3D substrates by creating micrometer-size pits on PADC films through irradiation of the films by alpha particles and subsequent chemical etching. HeLa cells cultured on these 3D substrates were observed using scanning electron microscope. Multiple directions and multiple layers of HeLa cells were found to have grown in the pits, with normal nuclei and cell membranes as well as good cell spreading. For the cells cultured in 3D substrates with or without additional small pits, no significant differences were observed between their vinculin expression profiles, which was in contrast to the observation made for cells cultured on 2D substrates showing that small pits could enhance vinculin expression. The presence of the large pits and/or the enhanced biocompatibility of the substrate in the present experiments might be the reasons. The protrusions of cells were confined by the small pits, which was similar to the observation made for cells cultured on 2D substrates. Cellular behavior on 3D substrates was further studied in Chapter 3 in terms of MT growth speed. In Chapter 3, we used end-binding protein 3 - green fluorescent protein (EB3-GFP) as a marker of MT growth to show that cells having migrated into the pits with curved surfaces (3D PADC films) had significantly slower MT growth speeds than those having stayed on flat surfaces (2D PADC films) without the pits. We proposed that engagement of a substrate with curvature was perceived by the cells as equivalent to the engagement of a less compliant substrate, which induced different MT growth speeds. The 3D substrates with pits with defined curvature resembled in some ways to the structure of 3D scaffolds employed for tissue engineering purposes, in that the latter also consisted of pores with defined curvature as their basic structural element, and could thus provide useful information for designing new tissue engineering scaffolds. With the study of cellular behavior in low-dose radiation (Chapter 4), the usefulness of 3D substrates was further studied in Chapter 5 in terms of radiation sensitivity. Previous studies showed that 3D cell cultures could change the cell sensitivity to ionizing radiation, when compared to 2D cultures. In the present thesis, we would like to quantify the cell damages in terms of the numbers of p53 binding protein 1 (53BP1) foci. In dosimetry studies, 53BP1 foci have been widely used as a biomarker of DNA double-strand breaks (DSBs) in cells induced by ionizing radiations. 53BP1 was shown to relocalize into foci shortly after irradiation, with the number of foci closely paralleling the number of DNA DSBs. However, consensus on criteria in terms of the numbers of 53BP1 foci to define cells damaged by direct irradiation or by bystander signals has not been reached, which is partly due to the presence of 53BP1 also in normal cells. In Chapter 4, study of the changes in the distribution of cells with different numbers of 53BP1 foci in a HeLa cell population culture on 2D PADC films after low-dose ionizing irradiation (< 0.1 Gy) provided by alpha particles was conducted, with a view to propose feasible criteria for defining cells damaged by direct irradiation or by bystander signals. It was proposed that the change in the percentage of cells with 1 to 3 foci should be used for such purposes. Therefore, the usefulness of 3D substrates could be studied in Chapter 5 in terms of low-dose radiation sensitivity. In Chapter 5, we extended our research to the effects of low radiation doses (< 0.1 Gy) on HeLa cells cultured on pseudo 3D PADC films. Thin 3D PADC cell-culture substrates with a bulk thickness of 35 μm were successfully fabricated by creating micrometer-size pits on thin PADC films by alpha-particle irradiation and subsequent chemical etching. These thin substrates led to the capability of alpha particle irradiation from the bottom of the substrates for cell hitting. Our results showed there were no changes in the cell-phase distribution of cells cultured on 2D and 3D substrates. However, comparatively fewer 53BP1 foci in alpha-particle irradiated as well as bystander HeLa cells cultured on pseudo 3D PADC substrates than in those cultured on 2D substrates. These supported the feasibility of our pseudo 3D substrates in providing in vivo conditions to the cell cultures, which could give results very different from those 2D cell monolayers cultured on flat substrates. 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A polydimethylsiloxane (PDMS) replica of this PADC film gave a micropillar substrate with micron-scale pillars. The PDMS micropillar substrates were found to be capable of changing the response of HeLa cells in terms of the percentages of cells in the S-phase and the attached cell numbers after 3-day cell culture. This demonstrated that the proliferation of the HeLa cells could be changed through mechanosensing the substrate curvature.