Medical Sciences Commons^™

Optimizing Rna Isolation And Histology Protocols For Characterization Of Preclinical Models Of Tendon Disease, Deirdre Finnigan, Melika Meshkat, Ali Naeij, Dawn Bryce, Matthew W. Grol

Undergraduate Student Research Internships Conference

Tendons join muscle to bone and are essential for posture and movement. Tenocytes are the resident tendon cells. Isolating RNA from small animal models such as mice can be difficult, especially from tendon, as much of the time, they are extremely small samples. Downstream analysis such as qPCR and especially RNA sequencing require greater quantity and quality of RNA than is often achievable. RNA degradation is extremely rapid following tendon isolation, with RNA quality decreasing as time increases between the collection of tendons and RNA isolation. Even tendons whose RNA was isolated immediately following tendon isolation had imperfect quality. The …

Bridging The Gap Between Muscle And Bone: Examining Tendon Identity And Development, Amel Sassi

Undergraduate Student Research Internships Conference

Tendons are fibrous connective tissues that connect skeletal muscle to bone. Functioning primarily to facilitate locomotion, these tissues are rich with collagen and other minor collagens and proteoglycans. Nevertheless, with over 4000 tendons in the human body, the molecular pathways that define different tendons remain largely unknown. As such, the clear distinction between tendon identity, particularly at the genetic level has yet to be fully established. Similarly, molecular pathways involved in tendon developments remain to be fully elucidated. Despite the important role of tendons in motor coordination and posture, a number of areas in tendon development and identity have yet …

Investigating Pathways Associated With Intervertebral Disc Degeneration And Back Pain., Geoffrey J. Kerr

Electronic Thesis and Dissertation Repository

Lower back pain (LBP) is one of the most common conditions worldwide, yet, current therapeutics are limited to symptomatic relief and do not directly treat the underlying cause of pain. This is largely due to an incomplete understanding of the biological pathways and tissues involved in LBP. While many tissues appear to be involved, intervertebral disc (IVD) degeneration is believed to be a major contributor.

The main aim of this thesis was to characterize the role of two environmental risk factors, mechanical loading and obesity, in the initiation of IVD degeneration and associated pain using the mouse as a preclinical …

Low Birth Weight And Post-Natal Diet Induced Alterations In Skeletal Muscle Oxygen Consumption And Fiber Type Composition, Megan Cedrone

Electronic Thesis and Dissertation Repository

Adverse in utero and postnatal conditions can increase susceptibility to metabolic syndrome (MS). Altered muscle respiration contributes to MS, but the effects of restricted oxygen and nutrients in utero on skeletal muscle mitochondria remain unknown. In this study guinea pig sows underwent uterine artery ablations mid-gestation, producing fetuses with low birth weight (LBW). Soleus muscle was collected near term or at four months of age, from LBW and control fetuses and offspring, where the offspring were fed either a Western Diet (WD) or a control diet (CD). Soleus muscles from LBW fetuses exhibit lower maximal respiration rates than normal birth …

Modulation Of Inflammation Driven Wound Healing After Glaucoma Surgery, James J. Armstrong

Electronic Thesis and Dissertation Repository

Dysregulated wound healing contributes to most currently unanswered ophthalmological morbidity. Opacification and structure altering contractures compromise the delicate ocular anatomy upon which ocular function and healthy vision are reliant. Glaucoma filtration surgery, corneal stromal injury, proliferative vitreoretinopathy and age-related macular degeneration are major contributors to ocular morbidity – all with myofibroblast transdifferentiation and pathognomonic scarring activity at their core.

This thesis aims to revaluate the means by which dysregulated ocular wound healing is combated with evidence describing a novel strategy to mitigate its effects. A translational approach was used. An initial retrospective analysis of over ten thousand glaucoma surgeries found …

Modulation Of Parathyroid Hormone 1 Receptor Signaling By Extracellular Nucleotides, Brandon Kim

Electronic Thesis and Dissertation Repository

Parathyroid hormone (PTH) activates the PTH/PTH-related peptide receptor (PTH1R) on osteoblasts and other target cells. Mechanical stimulation of cells, including osteoblasts, causes release of nucleotides such as ATP into the extracellular fluid. In addition to its role as an energy source, ATP serves as an agonist at P2 receptors and an allosteric regulator of many proteins. We investigated the effects of concentrations of extracellular ATP, comparable to those that activate low affinity P2X7 receptors, on PTH1R signaling. Cyclic AMP levels were monitored in real-time using a bioluminescence reporter and β-arrestin recruitment to PTH1R was followed using a complementation-based luminescence assay. …

Functional Anatomy Of Palmar Musculature, Colin W. Moore

Electronic Thesis and Dissertation Repository

The palmaris longus (PL) and palmaris brevis (PB) are upper limb muscles considered atavistic remnants of those found in animal species. Despite their use in surgical grafting and tendon transfer procedures, the functional role of the PL and PB have not been investigated comprehensively in vivo. Using a multi-modal experimental approach consisting of indwelling fine wire electromyography (EMG), ultrasonography and immunohistochemical muscle staining techniques, the function of the PL and PB in the hand was evaluated both in in vivo and in situ.

The purpose of Study 1 was to determine whether the PL provides synergistic contributions to …

Pannexins 1 And 3 In Skeletal Health And Osteoarthritis, Paxton M. Moon

Electronic Thesis and Dissertation Repository

Osteoarthritis (OA) is a disease that gradually robs affected individuals of their mobility and places a substantial burden on health care systems and economies. OA results from pathological changes in bone, cartilage, synovium and supporting structures which are all essential to joint function. Cartilage degeneration, a hallmarking feature of OA, is an irreversible and untreatable consequence of these changes. Though the mechanisms underlying the initiation of cartilage breakdown remain unclear, joint injury and aging are risk factors. Pannexins (PANXs), a novel family of channel forming glycoproteins, have been suggested as important mediators of early OA. Therefore, my thesis tested the …

Markerless Radiostereogammetry Of The Shoulder Joint In Humans: Comparisons Of Scapulohumeral Kinematics Between Individuals With Healthy And Supraspinatus-Impaired Shoulders, Ashley N. Hannon

Electronic Thesis and Dissertation Repository

The purpose of this collection of studies was to further develop the knowledge of shoulder motion in order to better understand joint function through direct measurement of 3D scapulohumeral joint kinematics using a technique of high accuracy. Markerless, bi-planar fluoroscopic radiostereometric analysis using a generic shoulder model was developed in this thesis, reducing the amount of radiation exposure to subjects. The studies compared kinematic data of the scapulohumeral joint in six degrees of freedom with a precise, in-vivo measuring technique. Data were collected on young and older healthy individuals, individuals with a torn supraspinatus and post-surgical intervention.

Although this generic …

Angiogenic Therapy In A Fibrotic Murine Model Of Duchenne Muscular Dystrophy, Kelly M. Gutpell

Electronic Thesis and Dissertation Repository

Duchenne muscular dystrophy (DMD) is a devastating neuromuscular disorder that affects approximately 1 in 5000 males. Vascular-targeted therapy has been proposed as a treatment for DMD to reduce ischemia and enhance endogenous repair. Additionally, a more vascularized environment may enhance regenerative approaches currently under investigation. Vascular endothelial growth factor (VEGF) and angiopoietin-1 (ANG1) are two of the most studied pro-angiogenic factors for this approach. To date, little is known regarding the effect of these pro-angiogenic factors on muscle function and whether they may exacerbate fibrosis in a relevant murine model of DMD. The first aim of this thesis was to …

A Clinically Relevant Relevant Post-Traumatic Osteoarthritis Mouse Model, Chantel P. Arce

Electronic Thesis and Dissertation Repository

Osteoarthritis affects 13-20% of Canadians with the majority being under 65years of age. Post-traumatic osteoarthritis (PTOA) is of great concern in young athletes following knee injury. Current research attempts at modeling the disease fall short. This study aimed to incorporate two important aspects of injury, the nature of the injury and the post-injury standard of care in humans, to a model of PTOA in mice. The study validated a non-invasive protocol to elicit an anterior cruciate ligament (ACL) injury at varying loading speeds addressing the closed capsule nature of an ACL injury that occurs in humans. Secondly, we proposed a …

The Role Of Bone Sialoprotein In The Tendon-Bone Insertion, Ryan M. Marinovich

Electronic Thesis and Dissertation Repository

Tendons and ligaments insert into bone through a transitional tissue termed the enthesis which is susceptible to injury and difficult to repair. Entheses contain a region of calcified fibrocartilage (CFC), however mineral-associated proteins in this tissue remain poorly characterized. Bone sialoprotein (BSP) is a phosphoprotein associated with mineralizing tissues. In these studies BSP was identified in the CFC of entheses by immunohistochemistry. Analysis of the entheses of Bsp^-/- mice indicate abnormalities in the CFC. Compared to controls, the CFC of the quadriceps tendon enthesis is 28% and 41 % longer in 15 week and 14 month old Bsp^{-/- …}

The Ctcf Chromatin Organizer Is Required For Hindlimb Development, Katherine L. Rabicki

Electronic Thesis and Dissertation Repository

Mutations in chromatin organizer CTCF were identified in patients with intellectual disability and skeletal defects. Previous studies demonstrated that depletion of CTCF in murine limb mesenchyme results in apoptosis in the forelimb. The role of CTCF in the hindlimb, however, is unknown. My objective was to investigate effects of CTCF deletion on chondrogenesis and skeletal development in the hindlimb. In vitro wild-type micromass cultures demonstrate that chondrocyte gene expression is delayed in the hindlimb when compared to forelimbs. Embryonic Ctcf^Fl/Fl;Prx1Cre mice were investigated, and qRT-PCR and histology were performed on limb buds and long bones. Results show that …

Diabetic Bone Marrow & Stem Cell Dysfunction, Meghan A. Piccinin

Electronic Thesis and Dissertation Repository

ii Abstract Abstract Defects in the proliferation, differentiation, and activity of bone marrow (BM)-derived vasculogenic/vascular stem cells (VSCs) have been observed in diabetes and contribute to the development of vascular complications. Diabetes leads to enhanced bone marrow adipogenesis, altering the composition of the BM stem cell (SC) niche and potentially disrupting the normal functioning of resident VSCs. Here, I establish that adipocytes have a negative influence on SC survival in culture. I also show that adipocytes and osteoblasts are responsible for the creation of distinct extracellular microenvironments, with unique expression patterns of several pro- and anti-angiogenic factors with known effects …

Novel Techniques For Investigating The Regulation Of Skeletal Muscle Hemodynamics, Baraa K. Al-Khazraji

Electronic Thesis and Dissertation Repository

The effect of the sympathetic nervous system (SNS) on blood flow distribution within skeletal muscle microvasculature is conditional upon regional activation of SNS receptors. Due to a lack of appropriate experimental models and techniques, no study has systematically evaluated the effect of SNS receptor activation in continuously branching skeletal muscle arteriolar trees. In line with previous work, we hypothesize that there will be a spatially-dependent distribution of sympathetic receptor activation along the arteriolar tree. Specifically, we anticipate a progressive decrease of adrenergic activation and a progressive increase of peptidergic and purinergic activation with increasing arteriolar order. We developed a novel …

The Antisaccade Task: Visual Distractors Elicit A Location-Independent Planning 'Cost', Jesse C. Desimone

Electronic Thesis and Dissertation Repository

Prosaccades are rapid eye movements with direct stimulus and response relations and are designed to bring the fovea onto a target or area of interest. In contrast, antisaccades require the inhibition of a prosaccade and the evocation of a saccade to a target’s mirror-symmetrical location. Previous work has shown that a remote (i.e., midline, contralateral) – but not proximal (i.e., ipsilateral) – task-irrelevant distractor relative to a visual target delays prosaccade reaction times (RT) (i.e., remote distractor effect: RDE). To my knowledge, however, no work has examined whether antisaccade RTs are similarly influenced by a RDE. Accordingly, I sought to …

Roles Of P2x7 Receptors In Adipose And Skeletal Tissues Of Mice, Kim L. Beaucage

Electronic Thesis and Dissertation Repository

P2X7 is an ATP-gated ion channel in the plasma membrane of a number of cell types. Previous in vitro studies show that P2X7 plays a role in regulating the differentiation of osteoblasts and adipocytes from mesenchymal precursors, as well as cellular metabolism and energy homeostasis. Furthermore, P2X7 has been implicated in mediating the effects of mechanical stimuli in bone. Our objective was to investigate the roles of P2X7, mechanical vibration, and their possible interaction in regulating body composition in vivo. We developed a novel method to monitor body composition during growth and aging in mice using micro-computed tomography (Chapter …

Durand Et Al 2012 Supplemental Figures, M. Durand, Svetlana V. Komarova, Ajay Bhargava, Keying Li, Cara Fiorino, Osama Maria, Noushin Nabavi, Morris Manolson, Rene Harrison, S. Jeffrey Dixon, Stephen Sims, Marcin Mizianty, Lukasz Kurgan, Sonia Haroun, Gilles Boire, Maria Lucena-Fernandes, Artur De Brum-Fernandes

Physiology and Pharmacology Publications

Objective. Our objective was to compare the osteoclastogenic capacity of peripheral blood mononuclear cells (PBMCs) from patients with osteoarthritis (OA) to that of PBMCs from self-reported normal individuals.

Methods. PBMCs from 140 patients with OA and 45 healthy donors were assayed for CD14⁺ expression and induced to differentiate into osteoclasts (OCs) over 3 weeks in vitro. We assessed the number of the OCs, their resorptive activity, OC apoptosis, and expression of the following cytokine receptors: receptor activator of nuclear factor κB (RANK), interleukin-1 receptor type I (IL-1R1) and IL-1R2. A ridge logistic regression classifier was developed …