published_or_final_version / abstract / Obstetrics and Gynaecology / Master / Master of Philosophy

published_or_final_version / abstract / Pathology / Master / Master of Philosophy

published_or_final_version / Surgery / Master / Master of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

published_or_final_version / Biochemistry / Master / Master of Philosophy

published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

Exercise induces morphological and metabolic adaptations that are highly specific to the mode of exercise training. These specific phenotypical changes are due to an equally specific molecular response that may depend on the activation and coordination intramuscular signaling pathways. Just as metabolic and morphological changes are influenced by the mode of exercise training, the signaling pathways that mediate exercise adaptation may also be directly related to the training status of skeletal muscle. For example, pre-conditioned skeletal muscle may exhibit a specific intracellular signaling response to an acute bout of exercise that is dependent on past training history. Both Akt (protein kinase B) and extra-cellular signal-related kinase (ERK 1 /2) have been shown to be phosphorylated in response to an acute bout of resistance exercise in human skeletal muscle and have been suggested to mediate the adaptive response to exercise. The purpose of this investigation was to examine the response of Akt and ERKI/2 to an acute bout of resistance exercise in three groups with distinctly different exercise training backgrounds. Twenty one subjects performed 3 sets of 10 repetitions of knee extension exercise at 70% 1-RM. The subjects consisted of a resistance-trained group (RE) (n=7), endurance trained group (END) (n=7) and a sedentary group (SED) (n=7). Muscle biopsies were taken from the vastus lateralis muscle before, immediately after, and 10 min post-exercise and were analyzed for phosphorylation of Akt and ERK1/2. ERK1/2 phosphorylation increased 47%, and 54% from pre-exercise to immediately post-exercise in the SED and RE groups respectively (p < 0.05). ERK1/2 phosphorylation increased 95%, 196%, and 47% from pre-exercise to 10 min post-exercise in the SED, RE, and END groups, respectively. (p < 0.05). The magnitude of ERK1/2 phosphorylation 10 min post-exercise was different between each group and may be linked to the group's training status. (p < 0.05) Akt phosphorylation decreased 42% and 37% from pre-exercise to immediately post-exercise in the SED and END group, respectively (p < 0.05). There was a 40 % increase in Akt phosphorylation from immediate post-exercise to 10 min post-exercise in the END group. In conclusion, training status appears to influence the magnitude and time course of activation of both Akt and ERK1/2 in response to an acute bout of resistance exercise. The immediate response of both ERK1/2 and Akt may play a key role in the adaptive response of skeletal muscle ultimately resulting in metabolic and morphological changes that are dependent on the past training history of the individual. / School of Physical Education, Sport, and Exercise Science

Isotope partitioning experiments were carried out with the adenosine 3',5'-monophosphate-dependent protein kinase catalytic subunit (cAPK) from bovine hearts to obtain information on the order of addition of reactants and the relative rates of reactant release from enzyme compared to the catalytic step(s). A value of 100% trapping for both ErMgATP-[γ-32P] and E:3H-Serpeptide at low Mgf indicates that MgATP and Serpeptide dissociate slowly from the enzyme compared to the catalytic step(s). The K_Serpeptide for MgATP trapping is 17 μM, while the K_MgATP for Serpeptide trapping is 0.58 mM. The latter data indicate that the off-rate for MgATP from the E:MgATP complex is 14 s^-1 while that for Serpeptide from the E: Serpeptide complex is 64 s^-1. At high Mg^, 100% trapping is obtained for the E:MgATP-[γ-32P] complex but only 40% is obtained for the E:Serpeptide complex. Thus, the off-rate for Serpeptide from the E:MgATP:Serpeptide complex becomes significant at high Mg_f. Data suggest a random mechanism in which MgATP is sticky. The V for the cAPK reaction increases 1.5-1.7 fold in the presence of the R_II in the presence of saturating cAMP at a stoichiometry of R:C of 1:1. No change is obtained with the type-I complex under these conditions. At higher ratio of R:C (up to 100) no further change is observed with the type-II complex but inhibition by the type-I R_2(cAMP)_4 complex competitive vs. Serpeptide is observed. The activiation observed in the presence type-II R_2(cAMP)_4 effects neither the K_m for Serpeptide nor the K_m for MgATP. Both the activating affect of the type-II complex and the inhibitory effect of the type-I complex are dependent on the Mg_f with more type-II activation obtained the higher the Mg_f and more type-I complex required for inhibition the higher the Mg_f. The activation and inhibition are discussed in terms of the mechanism of the protein kinase.

Stroke is one of the leading causes of human death in the United States. The debilitating effects of an ischemic stroke are due to the fact that mammalian neurons are highly susceptible to hypoxia and subsequent oxygen reperfusion. From studies in Drosophila melanogaster, cGMP-dependent Protein Kinase (PKG) enzyme is thought to affect anoxia tolerance by modifying the electrical current through potassium ion channels. In this research, two animal models were employed: Drosophila melanogaster and mammalian neurons exposed to stroke-like conditions. First, in vivo studies using Drosophila were performed to further our knowledge about the differences between the naturally occurring variants of the Drosophila foraging gene, which shows different protein levels of PKG. Mitochondrial density and metabolic activity between two fly genotypes exposed to anoxia and reoxygenation were compared. It was found that flies with less enzyme potentially showed mitochondrial biogenesis and higher metabolic rates upon reoxygenation. Next, in vivo studies where PKG enzyme was activated pharmacologically were performed; it was found that the activation of the cGMP/PKG pathway led to neuroprotection upon anoxia and reoxygenation. Furthermore, this model was translated into the in vitro model using Drosophila cells. Instead of anoxia and reoxygenation, hypoxia mimetics and hydrogen peroxide were used to induce cellular injury. After showing the cGMP/PKG pathway activation-induced cell protection, the potential downstream targets of the molecular signaling as well as underlying biochemical changes were assessed. It was found that mitochondrial potassium ion channels were involved in the protective signaling and the signaling modulated metabolic function. Furthermore, it was found that acidosis protected Drosophila cells from cell death, metabolic disruption, and oxidative stress. Finally, this research was translated to a mammalian in vitro model of neuronal damage upon stroke-like conditions; there, it was demonstrated that the cGMP/PKG pathway activation in rat primary cortical neurons and human cortical neurons was protective from low oxygen and acute oxidative stress. The results of this study lead to a better understanding of molecular mechanisms taking place during low oxygen and oxidative stresses. Consequently, this knowledge may be used to identify potential therapeutic targets and treatments that may prevent detrimental neurological effects of an ischemic stroke in humans. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2018. / FAU Electronic Theses and Dissertations Collection

Furthermore, ACS patients with a high N-terminal pro-B-type natriuretic peptide (NT-proBNP) and a high ROCK activity on admission had a five-fold risk to experience a cardiovascular event, when compared to those with low NT-proBNP and low ROCK activity. In addition, patients with high NT-proBNP and high ROCK activity were also more likely to die or experience a cardiovascular event at two years when comparing to those with high NT-proBNP and low ROCK activity. / In both ACS and CHF study cohorts, all the clinical parameters were recorded and analyzed. / In the first part of this thesis, 176 ACS patients and 51 control subjects were studied. All The patients were enrolled between December 2007 and May 2009 and followed up till 15th March 2010 (mean: 15.4+/-7.6 months, from 0.5 month to 27.5 months). The main outcome measures were all cause mortality, readmission with ACS or congestive heart failure (CHF) at 2 years from presentation. Altogether, there were 23 deaths (13.1%),33 readmissions with ACS (18.8%) and 13 admissions with CHF (7.4%) within 2 years. / Recent studies have shown that ROCK may playa pivotal role in cardiovascular diseases such as vasospastic angina, ischemic stroke, heart failure and metabolic syndrome via its involvement in regulation of vascular tone, endothelial dysfunction, inflammation, and remodeling. Indeed, inhibition of ROCK by statins or other selective inhibitors leads to upregulation and activation of endothelial nitric oxide synthase (eNOS) and reduction of vascular inflammation and atherosclerosis. In this thesis, we hypothesized that ROCK activity is increased in a selected population of patients with acute coronary syndrome (ACS) and congestive heart failure (CHF) and that ROCK activity is able to predict long-term clinical outcomes in these two populations. / Rho/rho-kinase (ROCK) is a serine-threonine protein kinase, which is one of the first immediate downstream targets of RhoA and expressed ubiquitously. ROCK is involved in many cellular functions, such as, cell growth, migration, apoptosis via actin cytoskeleton organization, and gene expression. They regulate cell contraction through serine-threonine phosphorylation of adducin, ezrin-radixin-moesin proteins, LIM kinase, myosin light chain phosphatase, and Sodium-Hydrogen ion (Na/H) exchanger. / The main findings are: ROCK activity was increased in ST elevation myocardial infarction (STEMI), non-STEMI (NSTEMI) and unstable angina (UA) groups when comparing with disease controls and healthy controls. On multivariate analysis, heart failure symptom on presentation, LDL-C level, and number of diseased coronary vessels were independent predictors of ROCK activity in ACS patients. / The ROCK activity in CHF patients was significantly higher than that of the disease control and normal control groups. New York Heart Association (NYHA) class, low left ventricular ejection fraction (LVEF) and high creatinine were independent predictors of the baseline ROCK activity in CHF. In terms of long-term heart failure mortality, ROCK activity was not an independent predictor. However, combining ROCK activity and NT-proBNP provided an incremental value in predicting long-term heart failure mortality over NT-proBNP alone. / Thus, increased ROCK activity is likely involved in cardiovascular diseases and further studies would be helpful to elucidate the potential role of ROCK activity inhibition in cardiovascular diseases. / We also recruited a group of 178 patients with CHF. All the patients were enrolled between December 2007 and January 2009 and followed up until 1st February 2010 (mean: 14.4+/-7.2 months, from 0.5 month to 26 months) or until the occurrence of cardiac death. Forty-five patients died (25.3%) within 2 years follow up. / Dong, Ming. / Adviser: Cheuk Man Yu. / Source: Dissertation Abstracts International, Volume: 73-04, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 140-164). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.