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Complex-phase (CP) steels are employed in applications that require high-strength and good edge formability. These steels derive their strength from a fine-grained bainite-ferrite microstructure, and alloying to provide solid-solution and precipitation strengthening. CP steels are produced industrially through a process of controlled rolling and cooling to produce desirable microstructures. Hole-expansion tests are typically used as a measure of edge formability for applications such as stretch-flanges. It has been shown that CP microstructures are susceptible to large fluctuations in hole-expansion performance with little change in processing or resulting tensile properties. The steel’s characteristics of damage evolution are critical to the hole-expansion performance. This study investigates the role of microstructure in the development of damage in CP microstructural variants. Two variant pairs of different thicknesses were produced from the leading and trailing edge of industrially produced hot-rolled sheet. Each pair consisted of a variant with poor hole-expansion performance, and a variant with good hole-expansion performance. Each variant was tested via interrupted double-notched uniaxial tension testing to induce damage. Damage evolution in each variant was quantified by X-ray micro-computed tomography (XµCT), and supplementary optical micrography. The damage results were correlated with microstructural characteristics. It was shown that poor hole-expansion variants failed by intergranular fracture. In these variants, void damage induced by hard martensite and retained austenite was not critical in producing failure. Purely void-damaged microstructures failed by ductile fracture, whereas cracked microstructures failed in a mixed brittle-ductile failure initiated by planar cracks. Microstructural banding of large elongated ferrite grains correlated with the existence of intergranular planar fractures. / Thesis (Master, Mechanical and Materials Engineering) -- Queen's University, 2013-12-17 15:03:02.206

Genomic integrity is preserved by checkpoints, which are signal transduction pathways that serve to delay cell cycle progression in the presence of DNA damage or replication stress. The heterotrimeric Rad9-Rad1-Hus1 (9-1-1) complex is a proliferating cell nuclear antigen (PCNA)-like clamp that is loaded onto DNA at structures resulting from damage, and is important for initiating and maintaining checkpoint signaling. Rad9 possesses a C-terminal tail unrelated to PCNA that is phosphorylated constitutively and in response to cell cycle position and DNA damage. Previous studies have identified tousled-like kinase 1 (TLK1) as a kinase that may modify Rad9. This thesis establishes that Rad9 is indeed phosphorylated in a TLK-dependent manner in vitro and in vivo, and that T355 within the C-terminal tail is the primary targeted residue. Phosphorylation of Rad9 at T355 is quickly reduced upon exposure to ionizing radiation before returning to baseline later in the damage response. In addition, TLK1 and Rad9 were shown to interact constitutively, and this interaction is enhanced in chromatin-bound Rad9 at later stages of the damage response. Furthermore, this thesis demonstrates that TLK1 is required for progression through S-phase in normally cycling cells, and that depletion of TLK1 results in a prolonged G2/M arrest upon exposure to ionizing radiation, a phenotype that is mimicked by over-expression of a Rad9-T355A mutant. Given that TLK1 is transiently inactivated upon phosphorylation by Chk1 in response to DNA damage, this work proposes that TLK1 and Chk1 act in concert to modulate the phosphorylation status of Rad9, which in turn plays a role in regulating the DNA damage response. / Thesis (Ph.D, Biochemistry) -- Queen's University, 2013-12-24 10:31:57.987

The investigation of renal pathophysiology and toxicology has traditionally been advanced by the development of increasingly defined and refined in vitro preparations. This study has sought to develop and evaluate various methods of producing pure samples of renal proximal tubules (PTs) from the Fischer rat. The introduction summarised the most common in vitro preparations together with the parameters used to monitor viability - particularly with regard to toxic events. The most prevalent isolation methods have involved the use of collagenase to produce dissociation of the cortex. However, the present study has shown that even the mildest collagenase treatment caused significant structural damage which resulted in a longevity of only 3hr in suspension. An alternative mechanical isolation technique has been developed in this study that consists of perfusion loading the renal glomeruli with Fe304 followed by disruption of the cortex by homogenisation and sequential sieving. The glomeruli are removed magnetically and the PTs then harvested by a 64μM sieve. PTs isolated in this way showed a vastly superior structural preservation over their collagenase isolated counterparts; also oxygen consumption and enzyme leakage measurements showed a longevity in excess of 6hr when incubated in a very basic medium. Attempts were then made to measure the cytosolic calcium levels in both mechanical and collagenase isolated PTs using the fluorescent calcium indicator Fura. However results were inconclusive due to significant binding of the Fura to the external PT surfaces. In conclusion, PTs prepared by the present mechanical isolation technique exhibit superior preservation and longevity compared with even the mildest collagenase isolation technique and hence appear to offer potential advantages over collagenase isolation as an in vitro renal system.