Innate immune and inflammatory responses contribute to myocardial and cerebral ischemia/reperfusion (I/R) injury. Toll-like receptors (TLRs) play a critical role in the induction of innate immune and inflammatory responses via activation of nuclear factor kappa B (NF-κB). We have shown that activation of NF-κB contributes to myocardial and cerebral I/R injury. Indeed, inhibition of TLR4-mediated NF-κB activation significantly decreased myocardial and cerebral I/R injury via activation of PI3K/Akt signaling. PI3K/Akt signaling is an important pathway in regulating cellular survival and inflammatory responses. Therefore, an important question is how to differentially modulate PI3K/Akt signaling and TLR/NF-κB-mediated signaling pathway during I/R injury? We demonstrated that pretreatment of mice with Pam3CSK4, a specific TLR2 ligand, significantly decreased cerebral I/R injury and improved neuronal functional recovery. Importantly, therapeutic administration of Pam3CSK4 also markedly decreased cerebral I/R injury. The mechanisms involved suppression of NF-κB binding activity and activation of PI3K/Akt signaling. We also demonstrated that CpG-ODN, a specific TLR9 ligand, induced protection against cerebral I/R injury via activation of PI3K/Akt signaling. Our findings were consistent with our previous reports showing that administration of Pam3CSK4 or CpG-ODN protected against myocardial I/R injury via a PI3K/Akt-dependent mechanism. In addition, we demonstrated for the first time that TLR3 located in endosomes played a deleterious role in myocardial I/R injury via activation of NF-κB. To investigate how to activate PI3K/Akt signaling during I/R injury, we examined the role of microRNA (miRs) in regulating PI3K/Akt signaling during myocardial ischemic injury. We discovered that Pam3CSK4 or CpG-ODN treatment significantly increased the expression of miR-130a in the myocardium, while myocardial infarction markedly decreased the levels of miR-130a in the myocardium. The data indicated that miR-130a served a protective role in myocardial ischemic injury. Indeed, we demonstrated for the first time that increased expression of miR-130a significantly attenuated cardiac dysfunction and promoted angiogenesis after myocardial infarction. The mechanisms involved activation of PI3K/Akt signaling via targeting PTEN expression by microRNA-130a. This dissertation discovers novel mechanisms of cerebral and myocardial ischemic injury and provides solid basis for developing new approaches for the treatment and management of stroke and heart attack patients.

CTRP3 is a secreted plasma protein of the C1q family that helps regulate hepatic gluconeogenesis and is downregulated in a diet-induced obese state. However, the role of CTRP3 in regulating lipid metabolism has not been established. Here, we used a transgenic mouse model to address the potential function of CTRP3 in ameliorating high-fat diet-induced metabolic stress. Both transgenic and wild-type mice fed a high-fat diet showed similar body weight gain, food intake, and energy expenditure. Despite similar adiposity to wild-type mice upon diet-induced obesity (DIO), CTRP3 transgenic mice were strikingly resistant to the development of hepatic steatosis, had reduced serum TNF-α levels, and demonstrated a modest improvement in systemic insulin sensitivity. Additionally, reduced hepatic triglyceride levels were due to decreased expression of enzymes (GPAT, AGPAT, and DGAT) involved in triglyceride synthesis. Importantly, short-term daily administration of recombinant CTRP3 to DIO mice for 5 days was sufficient to improve the fatty liver phenotype, evident as reduced hepatic triglyceride content and expression of triglyceride synthesis genes. Consistent with a direct effect on liver cells, recombinant CTRP3 treatment reduced fatty acid synthesis and neutral lipid accumulation in cultured rat H4IIE hepatocytes. Together, these results establish a novel role for CTRP3 hormone in regulating hepatic lipid metabolism and highlight its protective function and therapeutic potential in attenuating hepatic steatosis.

Background: Vestibular evoked myogenic potentials (VEMPs) have been recorded from the sternocleidomastoid muscle (cervical VEMP or cVEMP) and more recently from the eye muscles (ocular VEMP or oVEMP) in response to air conduction and bone conduction stimuli. Both cVEMPs and oVEMPs are mediated by the otoliths and thereby provide diagnostic information that is complementary to videonystagmography and rotational chair tests. In contrast to the air conduction cVEMP, which originates from the saccule/inferior vestibular nerve, recent evidence suggests the possibility that the air conduction oVEMP may be mediated by the utricle/superior vestibular nerve. The oVEMP, therefore, may provide complementary diagnostic information relative to the cVEMP. There are relatively few studies, however, that have quantified the effects of stimulus and recording parameters on the air conduction oVEMP, and there is a paucity of normative data. Purpose: To evaluate the effects of several stimulus and recording parameters on the air conduction oVEMP and to establish normative data for clinical use. Research Design: A prospective repeated measures design was utilized. Study Sample: Forty-seven young adults with no history of neurologic disease, hearing loss, middle ear pathology, open or closed head injury, cervical injury, or audiovestibular disorder participated in the study. Data Collection and Analysis: The effects of stimulus frequency, stimulus level, gaze elevation, and recording electrode location on the amplitude and latency of the oVEMP for monaural air conduction stimuli were assessed using repeated measures analyses of variance in an initial group of 17 participants. The optimal stimulus and recording parameters obtained in the initial group were used subsequently to obtain oVEMPs from 30 additional participants. Results: The effects of stimulus frequency, stimulus level, gaze elevation, and electrode location on the response prevalence, amplitude, and latency of the oVEMP for monaural air conduction stimuli were significant. The maximum N1-P1 amplitude and response prevalence were obtained for contralateral oVEMPs using a 500 Hz tone burst presented at 125 dB peak SPL during upward gaze at an elevation of 30°. Conclusions: The optimal stimulus and recording parameters quantified in this study were used to establish normative data that may be useful for the clinical application of the air conduction oVEMP.

The otoliths are vestibular organs that act as gravito-inertial force sensors and contribute to the perception of spatial orientation (earth verticality). The subjective visual vertical (SVV) is a psychophysical measure of the angle between perceptual vertical and true (gravitational) vertical. The otoliths contribute to the estimation of the physical vertical orientation, and individuals with normal vestibular function align the SVV within 2 degrees of true vertical (0 degrees). Impaired SVV has been documented in patients with unilateral vestibular disorders. Most research has focused on measuring the static SVV (head upright and stationary); however, more recently, methods have been developed to measure the SVV during stimulation of the otolith organs using on-axis yaw rotation (bilateral centrifugation), off-axis eccentric rotation (unilateral centrifugation), or head tilt for tests of bilateral or unilateral otolith function. The SVV test may be a useful method to assess utricular function in patients complaining of dizziness and/or imbalance and identify stages of recovery for otolith involvement.

No description available.

Background: The otoliths act as gravito-inertial force sensors and contribute to the perception of spatial orientation. The perception of gravitational vertical can be assessed by asking a subject to adjust a light bar to the vertical. Prior to clinical use of the SVV (subjective visual vertical) test, normative data and test-retest reliability must be established. Purpose: To obtain normative data and d etermine the test-retest reliability for the SVV test performed in static and dynamic test conditions. Research Design: A descriptive design was used to obtain normative data. Study Sample: Twenty-four young adults with no history of neurological disease, middle-ear pathology, open or closed head injury, cervical injury, or audiovestibular disorder participated in the study. Data Collection and Analysis: The SVV angle was measured in the static position and in three dynamic conditions: (1) on-axis clockwise (CW) rotation, (2) off-axis CW rotation of right ear, and (3) off-axis CW rotation of left ear. Results: In young healthy individuals, the SVV was Conclusions: The normative data obtained in this study may be useful in identifying patients with chronic utricular dysfunction. We recommend the use of difference angles (on-axis SVV ‐ off-axis SVV) to remove baseline bias and decrease the variability of the SVV angles for the off-axis conditions.

Ligands of the human monoamine transporters encompass a wide range of both illicit and therapeutic drugs that act upon neural circuitry related to reward, motivation, and the processing of salient stimuli. The present study utilizes two methods for analyzing transporter substrates and inhibitors in order to characterize activity and assess potency. The first measures transient changes in intracellular calcium as a surrogate for transporter activity by harnessing the electrical coupling of monoamine transporters and L-type calcium channels. This is used to analyze novel chimera of the strong hDAT inhibitors methylphenidate and 𝛼-PPP in order to assess the contribution of specific moieties to potency. The observed reduction in potency suggests that methylphenidate may bind to the transporter in a manner distinct from 𝛼-PPP, as chimera would otherwise be expected to show similar activity to parent compounds. These results highlight the importance of 𝛼-carbon substituents and the relatively small contribution of beta-carbon groups to inhibitor potency at hDAT, while the lack of activity at hSERT suggests potency is not strongly influenced by beta-carbon or N-alkyl substituents. In order to further characterize drug-transporter interaction, a method was developed to analyze the kinetics of binding and unbinding using both known and novel hNET ligands, including a series of N-alkyl derivatives of 4-methylamphetamine. The study emphasizes the importance of both association and dissociation kinetics to affinity and sets up a methodological framework with two ways for determining Kd, with notable advantages over current models. The results indicate that lengthening the N-alkyl chain of 4-methylamphetamine leads to a decrease in potency and a shift in activity from substrate to blocker, with the results of N-propyl 4-methylamphetamine in particular indicating the potential existence of multiple low-affinity binding sites, each with distinct on and off kinetics. The implications of these results help elucidate the mechanism of action of transporter ligands and set up a framework for future studies that can more specifically classify the interaction between transporters and inhibitors or releasing agents.

Medial tibial stress syndrome (MTSS) is a stress and overuse injury that presents as pain on the medial aspect of the lower two-thirds of the tibia. It is most often caused by repetitive actions on hard surfaces such as running, marching, and dancing. Individuals most affected by MTSS are runners, members of the military, dancers, and athletes that play soccer, volleyball and basketball. While MTSS has a relatively standard presentation of pain on the medial aspect of the tibia, it can occasionally be mistaken for other injuries such as stress fractures or compartment syndrome. If a diagnosis is unsure, methods such as x-ray, bone-scan, and MRI can be utilized to better obtain the correct diagnosis. A variety of treatments exist for MTSS including, ice, massage, muscle strengthening, and rest. A combination of these various techniques is most often what is employed. In this study, the effectiveness of a set of resistance ankle exercises in combination with ice and massage was tested and compared to that of ice and massage alone. The hypothesis was that athletes receiving the exercises as part of their treatment, in addition to the icing and massaging, would have a greater decrease in pain from MTSS than athletes just receiving ice and massage as treatment. The exercises would strengthen the muscles of the lower leg that, when weak, can contribute to the development of MTSS. Results indicated that the exercises yielded a more significant decrease in pain from MTSS than ice and massage alone.

The role oxygen plays in the developing ovarian follicle is of interest not only to the field of developmental biology but also to in-vitro fertilisation (IVF) technologists, as oxygenation of the oocyte is considered to be a potential determinant of oocyte competence. Oxygen transport through the developing ovarian follicle, and practical aspects of the analysis of oxygen in human follicular fluid were investigated in this work. Mathematical modelling of oxygen transport in the pre-antral, and antrallpreovulatory follicle revealed a number of interesting findings, Contrary to previous conclusions (Gosden & Byatt-Smith, 1986), oxygen can reach the oocyte in the small pre-antral follicle. Improved estimates of diffusion coefficients through the granulosa cell layer and the inclusion of fluid voidage in this layer showed that oxygen can also reach the oocyte in large pre-antral follicles. The amount of oxygen that reaches the oocyte in the pre-antral follicle is a function of its size and degree of vascularisation. Symmetrically distributed vascularisation is superior in achieving a well oxygenated follicle. However, the large pre-antral follicle will eventually reach a size beyond which it cannot grow without anoxic regions developing. The size at which this occurs is consistent with the size at which antrum formation is observed in human follicles. The model predicts that the follicle can avoid an anoxic state through antrum formation, and shows that the follicle develops in a way that is consistent with overcoming mass transport limitations. The oxygen status of the follicle during the antrallpre-ovulatory phase of growth requires that the volume of granulosa cells be balanced by the volume of follicular fluid. Further predictions suggest that oocyte respiration becomes sub-maximal at follicular fluid volumes below approximately 4m1, vascularisation levels below 38%, or fluid i dissolved oxygen levels below 5.1 ~01%. These values are consistent with observations in the literature. It was also shown that the measurement of follicular fluid dissolved oxygen levels could provide a simple measure of the respiratory status of the oocyte, and this may be superior to the measurement of follicular vascularisation which requires knowledge of more parameters. Methodology for the analysis of follicular fluid oxygen solubility and diffusivity was developed using a Clark oxygen electrode. Analysis of these parameters showed that they are similar to human plasma, and allowed the predictive uncertainty of the model to be reduced. Experimental studies into the effects of IVF aspiration on follicular fluid were carried out. Aspiration results in significant changes in the properties of follicular fluid. Dissolved oxygen levels rose 5 * 2 vol%, pH increased by 0.04 * 0.01 pH units, and temperature dropped by 7.7 * 1.3 "C. Mathematical modelling of blood contaminated follicular fluid also showed that contamination results in significant changes in the dissolved oxygen of the fluid. This suggests that if the composition of follicular fluid is to be determined (particularly dissolved oxygen), sampling andlor measurement of fluid must take place before the collection vial of the aspiration kit, and blood contamination must be eliminated. Based on this result, the design and testing of devices capable of reliable sampling andlor rneasurement of oxygen levels of follicular fluid was considered. This presents a continuing challenge, including the integration of routine follicular fluid oxygen measurement into clinical practice.

<p>Asthma is a disease characterized by transient airway smooth muscle contraction leading to episodes of reversible airway narrowing. It affects over 300 million people worldwide and is implicated in over 250 000 deaths annually. The primary clinical features of asthma include airway inflammation, hyperresponsiveness, and remodeling. Generally, asthmatic patients experience exacerbations between periods of diminished symptoms. Interestingly, in addition to these above mentioned hallmarks, asthmatics have also been shown to react differently to ventilatory mechanical strain. This is most evident when assessing the effect of a deep inspiration (DI), clinically measured as a breath taken from functional residual capacity to total lung capacity, in healthy individuals <em>versus</em> asthmatics. These deep inspiratory efforts have been shown to produce a bronchodilatory response in healthy individuals, whereas in asthmatics, DIs are less effective in producing bronchodilation, can cause more rapid airway re-narrowing, and even bronchoconstriction in moderate to severe asthmatics. The mechanism by which a DI is able to cause bronchoconstriction remains ambiguous. Previous theories suggest that this phenomenon is intrinsic to airway smooth muscle (ASM) itself. However, the airway inflammation present in asthmatic airways may also add to the increased ASM contractility following stretch, by the release of mediators that can prime the contractile apparatus to react excessively in the presence of stretch.</p> <p>Thus, collectively, the studies contained in this thesis are linked to the general theme of greater characterization of the signalling mechanisms that regulate airway stretch-activated contractions using a pharmacological approach in intact bovine bronchial segments, with the hope of providing novel insights into the mechanisms that regulate the DI-induced bronchoconstriction seen in asthmatics.</p> / Doctor of Philosophy (Medical Science)