A dominant ENU screen produced mouse line 7238 with large platelets. Sequence capture and Next Generation sequencing identified a mutation in Myh9 at Q1443L [1]. Mice were tested for aspects of MYH9-Related Disease (MYH9RD), a rare human condition caused by mutations within MYH9; macrothrombocytopenia and neutrophil inclusions are found in almost all cases, while deafness, cataracts and renal disease have variable penetrance and severity. Myh9Q1443L/+ and Myh9Q1443L/Q1443L animals have neutrophil inclusions [1] and increased cataracts at 2, 6 and 12 months; Myh9Q1443L/Q1443L animals at 12 months have changes in kidney output [2]. Immunofluoresence showed changes in protein expression in glomeruli at two months. This is the first ENU mouse model identified by a sequence capture mechanism, and the first mouse line to produce a point mutation within the Myh9 gene [1,2]. This mouse models MYH9RD, and is an invaluable tool to understand the role of this protein, and to determine mechanisms underlying this disease.

New neurons are continually produced throughout adult life in the dentate gyrus of the hippocampus, in a process termed adult neurogenesis. Although there is a significant effort in the literature to understand the functional significance of hippocampal neurogenesis, conflicting experimental reports have left the role of neurogenesis unclear. Recently, computational modelling studies have hypothesized that neurogenesis may play a role in allowing association between event and context to be formed in memory. By using a novel odour task and a raised plus maze task, our work demonstrates that the reduction of hippocampal neurogenesis using focal irradiation impairs the ability of animal subjects to utilize contextual information to learn interfering information. The result of this work provides experimental evidence of a unique role neurogenesis may play in learning and memory.

Cardiovascular disease is the most common cause of morbidity and mortality in type 2 diabetes. Short-term studies of glucagon-like peptide-1 (GLP-1)-targeted therapies suggest potential beneficial effects on cardiovascular outcomes. The mechanism behind this unexpectedly rapid effect is not known. In this study, full-length human GLP-1 receptor (GLP-1R) mRNA was cloned and sequenced from a human megakaryocyte cell line. Quantitative RT-PCR results showed that expression levels were comparable to other GLP-1R expressing tissues. Furthermore, incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in these cells. As megakaryocytes are the cellular precursors of platelets, the effect of GLP-1 and exenatide were studied in gel-filtered human platelet aggregation, where they were both shown to have an inhibitory effect on thrombin-stimulated platelet aggregation. Platelet inhibition by GLP-1 and GLP-1R agonists presents a potential mechanism for the reduced incidence of atherothrombotic events thought to be associated with GLP-1-targeted therapies.

The ability to respond rapidly with spatial precision is required in a number of facets of everyday life, whether catching a falling object, reacting to other drivers on a busy freeway or recovering one’s balance following an unexpected perturbation. The sophisticated central nervous system (CNS) control of these reactions is often overlooked until the speed of such reactions becomes delayed, either due to ageing or brain injury, wherein the individual becomes at risk of injury. Surprisingly, little is known regarding the control of these ‘temporally urgent’ movements. Therefore, the primary objectives of this dissertation were to develop an understanding of the control of these movements by exploring the factors that may be involved in the generation of temporally urgent movements in the healthy CNS, locating the areas within the CNS that such modulation occurs and identifying the relative weighted importance of those modulators based on the initial conditions of stimulus delivery. Specific characteristics of stimulus properties, such as intensity and modality were particularly influential in the latency of motor reactions and physiological electrodermal skin responses fluctuated in accordance with input stimulus parameters. Importantly, outcomes from this dissertation identified that rapid reactions likely utilize a CNS network that includes higher cortical regions such as somatosensory cortex and primary motor cortex, which may be modulated by physiological arousal, rather than the solitary involvement of subcortical structures. The findings from this dissertation have important implications for individuals with disordered speed of processing and indicate the potential modifiability of factors that influence reaction time.

The Apelin-APJ system is transiently up-regulated in murine models of cardiac dysfunction. We have previously shown that Apelin-deficient mice subjected to aortic constriction suffer from severe fibrosis. In turn, we hypothesized that Apelin deficiency will also exaggerate the fibrosis phenotype post experimental myocardial infarction, associated with changes in fibroblast cell activity. Apelin-deficient and wildtype mice were randomly subjected to sham operation or left coronary artery ligation. Apelin deficiency worsened cardiac functionality, enhanced fibrosis-related gene expression and morphology, and enhanced vimentin intermediate filament expression, which may be involved in increasing fibroblast proliferation. MicroRNA target prediction softwares predict that apelin and vimentin 3 ’UTRs are potential targets of microRNA-378 regulation, and were confirmed with Luciferase reporter assays and western blot. Apelin up-regulation may be a useful strategy for attenuating unfavorable fibrosis through down-regulating vimentin-mediated adverse fibroblast activity. MicroRNA-378 regulation may be partly responsible for changes in apelin and vimentin expression.

The mechanism and subsequent patterns of Heat shock protein (Hsp) expression in skeletal muscle specific to contraction type was determined. Rat tibialis anterior (TA) muscle was forcibly lengthened (LC) or shortened (SC) in 5 sets of 20 repetitions before being removed at 2, 8, 24, 48, 72, or 168 hours and analyzed for muscle damage and Hsp25 and Hsp72 expression. Isometric peak torque was reduced to 63% and 33% (P<0.001) at 3-minutes after SC and LC, respectively. Muscle fibre damage appeared at 8h and beyond following LCs, but no damage was observed after SCs. Hsp25 content in LC muscle increased by 3.1±0.53 fold (P<0.01) at 48h and remained elevated. Hsp72 content increased by 3.8±0.66 fold at 24h and remained elevated. Neither Hsp25 nor Hsp72 content was elevated following SCs. Muscle damage associated with LCs results in a greater Hsp accumulation than SCs and 100 SCs do not result in increased Hsp content.

Fetal and neonatal immune thrombocytopenia (FNIT) is a severe bleeding disorder which results from fetal platelet destruction by maternal antibodies against platelet antigens, including GPIIbIIIa (αIIbβ3 integrin) and GPIbα. β3 integrin is also expressed by angiogenic endothelial cells (ECs) and is required for angiogenesis. Therefore, we investigated whether anti-β3 antibodies in FNIT cross-react with blood vessels of the fetus/neonate and contribute to pathogenesis. Antibodies to GPIbα were used as controls. To mimic human FNIT, β3 integrin- or GPIbα-deficient female mice were immunized with wild-type platelets and bred with wild-type male mice. Pups in both groups had thrombocytopenia but intracranial hemorrhage was only observed in anti-β3-mediated FNIT. Anti-β3-mediated FNIT pups had increased apoptosis in the brain and impaired vascularization of the brain and retina. In addition, anti-β3 sera inhibited proliferation and vascular-like tube formation by ECs in vitro. Therefore, anti-β3 antibodies in FNIT likely impair angiogenesis in the developing fetus/neonate.

New neurons are continually produced throughout adult life in the dentate gyrus of the hippocampus, in a process termed adult neurogenesis. Although there is a significant effort in the literature to understand the functional significance of hippocampal neurogenesis, conflicting experimental reports have left the role of neurogenesis unclear. Recently, computational modelling studies have hypothesized that neurogenesis may play a role in allowing association between event and context to be formed in memory. By using a novel odour task and a raised plus maze task, our work demonstrates that the reduction of hippocampal neurogenesis using focal irradiation impairs the ability of animal subjects to utilize contextual information to learn interfering information. The result of this work provides experimental evidence of a unique role neurogenesis may play in learning and memory.

Fetal and neonatal immune thrombocytopenia (FNIT) is a severe bleeding disorder which results from fetal platelet destruction by maternal antibodies against platelet antigens, including GPIIbIIIa (αIIbβ3 integrin) and GPIbα. β3 integrin is also expressed by angiogenic endothelial cells (ECs) and is required for angiogenesis. Therefore, we investigated whether anti-β3 antibodies in FNIT cross-react with blood vessels of the fetus/neonate and contribute to pathogenesis. Antibodies to GPIbα were used as controls. To mimic human FNIT, β3 integrin- or GPIbα-deficient female mice were immunized with wild-type platelets and bred with wild-type male mice. Pups in both groups had thrombocytopenia but intracranial hemorrhage was only observed in anti-β3-mediated FNIT. Anti-β3-mediated FNIT pups had increased apoptosis in the brain and impaired vascularization of the brain and retina. In addition, anti-β3 sera inhibited proliferation and vascular-like tube formation by ECs in vitro. Therefore, anti-β3 antibodies in FNIT likely impair angiogenesis in the developing fetus/neonate.

Cardiovascular disease is the most common cause of morbidity and mortality in type 2 diabetes. Short-term studies of glucagon-like peptide-1 (GLP-1)-targeted therapies suggest potential beneficial effects on cardiovascular outcomes. The mechanism behind this unexpectedly rapid effect is not known. In this study, full-length human GLP-1 receptor (GLP-1R) mRNA was cloned and sequenced from a human megakaryocyte cell line. Quantitative RT-PCR results showed that expression levels were comparable to other GLP-1R expressing tissues. Furthermore, incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in these cells. As megakaryocytes are the cellular precursors of platelets, the effect of GLP-1 and exenatide were studied in gel-filtered human platelet aggregation, where they were both shown to have an inhibitory effect on thrombin-stimulated platelet aggregation. Platelet inhibition by GLP-1 and GLP-1R agonists presents a potential mechanism for the reduced incidence of atherothrombotic events thought to be associated with GLP-1-targeted therapies.