Cellular and Molecular Physiology Commons^™

Caspases Protect Against Hyperglycemia Induced Necroptosis-Like Cell Death In Brain Endothelial Cells, Aleena S. Mathew

Theses and Dissertations

Acute diabetes characterized by high spikes in blood glucose levels in association with severe hyperglycemia can lead to an increased risk of ischemic stroke in the brain. To better understand high blood glucose levels, hyperglycemia, in the brain we explored its effects on endothelial cells. Endothelial cells line the inner walls of blood vessels and are directly impacted in hyperglycemia. Brain endothelial cells are a major component of the blood brain barrier (BBB) which tightly regulates the brain’s microenvironment. The type of cell death such as apoptosis, caspase dependent, or necroptosis, caspase independent, can exacerbate diabetes associated complications. There is …

Sphingosine Kinase 2 Depletion Dysregulates Hepatic Redox Metabolism And Enhances Inflammation In Early Metabolic Dysfunction-Associated Steatohepatitis (Mash), Kaitlyn Georgene Jackson

Theses and Dissertations

Metabolic dysfunction-associated steatotic liver disease (MASLD) currently affects one in three people worldwide; its prevalence is growing alongside other cardiometabolic disorders. Aberrations to redox and sphingolipid metabolism contribute to MASLD pathologic progression. Sphingosine-1-phosphate (S1P) is an active lipid, meaning that S1P participates in biochemical processes within the cell. S1P is generated by two distinct isoenzymes, sphingosine kinase (SphK)1 and SphK2. The localization of the SphK enzyme dictates S1P function. While cytosolic SphK1-generated S1P is well-characterized, nuclear and mitochondrial SphK2-generated S1P is less known. Moreover, SphK2 depletion effects on cellular redox processes and inflammation are not well-defined.

This study defines the …

Targeting Bcl-2 Family Proteins In Therapy Induced Senescent Cancer Cell Models, Wade Cook

Theses and Dissertations

Non-small Cell Lung Cancer (NSCLC) originates from numerous different cell types in the lungs and is among the deadliest of cancers. Head and Neck Squamous Cell Carcinomas (HNSCC) are derived from the mucosal membranes of the oral cavity, pharynx, and larynx. Both NSCLC and HNSCC are predominately caused by tobacco smoke inhalation and as such mutations in the tumor suppressor gene TP53 are common. Since similarities exist in the root cause of NSCLC and HNSCC, they may also share similarities in treatment methods. Cisplatin is a platinum-based DNA damaging agent that has been used as a cancer chemotherapy for decades. …

Role Of P53 Deficient Clonogenic Endothelial Cells In The Pathogenesis Of Pulmonary Hypertension, Aneel Roy Bhagwani

Theses and Dissertations

Pulmonary Artery Hypertension (PAH) is a progressive disorder of the small vessels of the pulmonary vasculature. It is characterized by occlusive vascular remodeling leading to right heart failure and ultimately death. PAH pathogenesis involves a multifactorial etiology leading to formation of occlusive lesions in small (preacinary) pulmonary arteries. These lesions contain hyper-proliferative, apoptosis resistant endothelial cells (ECs), smooth muscle cells and fibroblasts. ECs play a major role in the development of PAH. These ECs in vascular lesions are primitive and express progenitor cell markers, including CD117 (c-kit).

We have isolated clonally expanded, c-kit+ ECs from the lungs of Green Fluorescent …

A Novel Review Of Heat Shock Protein 110 Kda: A Basis For Research And Continued Experimentation Through Biochemical Analysis, Crist W. Cuffee

Theses and Dissertations

Heat shock protein 110 kDa, Hsp110, is a distinct cellular protector, different in form and function from Hsp70, a close relative of Hsp110. Functioning primarily as a holdase or in tandem with other molecular chaperones, a review of current accomplishments elucidates the uniqueness of this protein and the continued mysteries that surrounds it. Found only in eukaryotes, Hsp110 has been linked to many diseases, ranging from parasitic infection to neurodegenerative disorders. While still lacking, studies of this protein have provided much in the realm of speculation on the mechanisms behind Hsp110s’ contribution to different pathologies. This review will serve as …

Muscarinic Excitation Of Dopamine Neurons In The Ventral Tegmental Area Via Activation Of A Trpc-Like Cation Conductance, Yu Tzu Chen

Theses and Dissertations

Dopaminergic (DA) neurons in the ventral tegmental area (VTA) play a crucial role in reward and motivational behaviors, including the development of drug addictions. VTA DA neurons receive excitatory cholinergic inputs from the mesopontine tegmentum. Blockage of the M5 muscarinic receptor in DA neurons has been shown to attenuate drug-induced DA release and abuse-related behaviors, but the molecular mechanism is unknown. In this study, experiments were designed to identify the electrophysiological effects of muscarinic agonism in the modulation of action potential kinetics and firing patterns in VTA DA neurons of mice. Pharmacology of the muscarinic receptor-evoked current was also characterized. …

Genetic And Cellular Models For The Study Of Ptk2b And Adam10 In Adaptive Thermogenesis And Metabolism, Jared S. Farrar

Theses and Dissertations

In a cellular model, we demonstrate that the non-receptor protein tyrosine kinase 2 beta (PTK2B) plays a critical role in mouse cultured beige adipocyte differentiation. CRISPR/Cas9-mediated knock-out of Ptk2b results in non-differentiating white adipocytes and differentiated beige adipocytes with significantly reduced thermogenic gene and protein expression, enlarged lipid droplet size, and altered mitochondrial respiration. Together, these data in a cell culture system provide evidence for a role of PTK2B in the differentiation of mouse beige adipocytes.

In the process of developing a new mouse model utilizing the adipocyte selective Adipoq-Cre transgenic mouse, strong genetic linkage between a gene …

Focal Augmentation Of Somatostatin Interneuron Function And Subsequent Circuit Effects In Developmentally Malformed, Epileptogenic Cortex, Nicole Ekanem

Theses and Dissertations

Drug-resistant epilepsy (DRE) is a common clinical sequela of developmental cortical malformations such as polymicrogyria. Unfortunately, much remains unknown about the aberrant GABA-mediated circuit alterations that underlie DRE's onset and persistence in this context. To address this knowledge gap, we utilized the transcranial freeze lesion model in optogenetic mice lines (Somatostatin (SST)-Cre or Parvalbumin (PV)-Cre x floxed channelrhodopsin-2) to dissect features of the SST, PV, and pyramidal neuron microcircuit that are potentially associated with DRE. Investigations took place within developmental microgyria’s known pathological substrate, the adjoined and epileptogenic paramicrogyral region (PMR). As well, microcircuit relationships within the previously unexplored range …

Novel Duel Mtor Inhibitors/Ampk Activators Improve Therapeutic Efficacy Of Doxorubicin And Ameliorate Its Associated Cardiotoxicity In Mice, Sahak Eric Hovsepian

Theses and Dissertations

Triple-negative breast cancer (TNBC) is a particularly aggressive subset of breast cancer that has a molecular expression profile which lacks the estrogen receptor (ER), progesterone receptor (ER), and human epidermal growth factor receptor type 2 (HER2). No targeted treatment options currently exist for TNBC, unlike other types of breast cancer, and therefore survival rates are far lower for these patients. The current treatment for TNBC is systemic anthracycline chemotherapy, typically doxorubicin (DOX), which has great clinical efficacy in increasing survival. However, DOX induces a dose-dependent progressive cardiomyopathy, which can present years and up to decades after the last treatment. This …

An Identification Of Tgf-B'S Role In Ischemic Cholangiopathy, Courtney Chmielewski

Theses and Dissertations

Organ transplants are a necessary intervention for many diseases that result in end stage organ failure. The donation pool cannot match the demands of the transplant list, so expanding the pool to include donation after cardiac death (DCD) is desired. However, there are increased odds of graft failure and ischemic cholangiopathy leading to inferior outcomes when DCD livers are used. Ischemic cholangiopathy consists of multiple diffuse strictures and fibrosis of the bile ducts leading to a loss of epithelialization and fibrosis. The cellular mechanism is proposed to be epithelial-to-mesenchymal transition (EMT). TGF-�� is seen as a key initiator of EMT, …

Investigation Of Molecular Mechanisms Of Liver Preservation Injury: A Complication Preceding Organ Transplantation, Ria C. Fyffe-Freil

Theses and Dissertations

Of the over 108,000 American awaiting a life-saving organ transplant today, over 12,000 (11%) of those need a new liver (OPTN, 2020). Last year, only 35% of patients on the waiting list for an organ were transplanted. Improving the quality of marginal organs by preventing or reversing preservation injury could vastly increase the number of transplants performed. Washout of circulating blood during liver procurement with cold University of Wisconsin solution flushes out any endogenous pro-survival signaling molecules. We investigated lysophospholipid (LPL) surface receptors (G-protein coupled receptors for lysophosphatidic acid (LPA) and sphingosine- 1-phosphate (S1P)) and their role in protecting hepatocytes …

An Evaluation Of Induced Shear Stress On Endothelial Cellular Adhesion Molecules, Edward B. Crabb

Theses and Dissertations

The pathophysiology of atherosclerotic cardiovascular disease (CVD) is highlighted by vascular dysfunction and low-grade vascular inflammation. Furthermore, the site-specific distribution of atherosclerosis throughout the arterial vasculature is primarily determined by local hemodynamic force. Therefore, this dissertation outlines three experiments designed to investigate the role of acute mental and physical (i.e., aerobic exercise), and vascular wall shear stress (SS) on the inflammatory aspects of atherosclerosis. Chapter 2 examines the effect of acute laboratory-induced mental stress on intracellular pro-inflammatory signaling pathways in peripheral blood mononuclear cells. Chapter 3 investigates the impact of acute laboratory-induced mental stress and maximal aerobic exercise on the …

Embryonic Stem Cell-Derived Exosomes Increase The Antiproliferative Activity Of Doxorubicin In Breast Cancer, Alexander M. Hirsch

Theses and Dissertations

The field of cancer research has grown immensely in recent decades and has led to a better understanding of the causes of the disease, as well as greatly improved treatment for various types of cancers, especially breast cancer. One of the most effective treatments involves the chemotherapeutic drug doxorubicin (DOX). DOX is an effective tool against all types of breast cancer, especially against triple negative breast cancer. However, DOX causes adverse side effects that include damage to the heart and skeletal muscle, particularly above specific cumulative doses. Recent evidence suggests that embryonic stem cell-derived (ES) exosomes, nanoscale extracellular vesicles that …

Electrostatic Networks And Mechanisms Of Δph-Dependent Gating In The Human Voltage-Gated Proton Channel Hv1, Ashley L. Bennett

Theses and Dissertations

The structure of the voltage-gated proton (H⁺) channel Hv1 is homologous to the voltage sensor domain (VSD) of tetrameric voltage-gated Na⁺, K⁺ and Ca²⁺ channels (VGCs), but lacks a pore domain and instead forms a homodimer. Similar to other VSD proteins, Hv1 is gated by changes in membrane potential (V), but unlike VGCs, voltage-dependent gating in Hv1 is modulated by changes in the transmembrane pH gradient (DpH = pH_o - pH_i). In Hv1, pH_o or pH_i changes shift the open probability (P_OPEN)-V relation by ~40 mV per …

An Investigation On The Role Of Β-Arrestin 2, Protein Kinase C And Sex On The Mechanism Of Morphine Tolerance In The Mouse Ileum, Karan Hitesh Muchhala

Theses and Dissertations

Opioids such as morphine are frequently used in the clinic to treat pain. However, the perennial bane of chronic opioid use is the rapid development of tolerance to the analgesic effects but delayed development of tolerance to the respiratory depressant and constipating effects. As constipation is one of the most common opioid-related adverse effects in humans, it is important to delineate mechanisms that drive opioid tolerance in the ileum and lack of it in the colon. The overarching goal of this thesis was to investigate mechanisms of morphine tolerance in the ileum by comparing the mechanism of morphine tolerance in …

Regulation Of Hcn Channel Function By Direct Camp Binding And Singlet Oxygen, Vinaykumar Idikuda

Theses and Dissertations

Hyperpolarization-activated, cyclic-nucleotide gated ion channels (HCN channels) are activated by membrane hyperpolarization and modulated by cyclic nucleotides. HCN channels are important to maintain the resting membrane potential and input resistance in neurons and have important physiological functions in the brain and heart. Four mammalian HCN isoforms, HCN1-4, and the isoform cloned from sea urchin, spHCN, have been extensively studied. Among these, only spHCN channel shows a voltage dependent inactivation. Previous studies have shown that the ligand binding in mHCN2 channel is activity dependent: cAMP binding increases along with channel opening or channels in the open state have higher binding affinity …

Utilizing Voltage-Gated Calcium Channels To Assess The Activity Of Cathinone Derivatives At Human Monoamine Transporters, Brian A. Ruiz

Theses and Dissertations

Cathinones are psychostimulant compounds heavily implicated as drugs of abuse. They exert their physiological actions at the monoamine transporters, which are responsible for maintaining synaptic neurotransmitter homeostasis. Monoamine transporters produce currents during transport and have been shown to depolarize cell membranes and activate voltage-gated calcium channels in mammalian expression systems. This phenomenon is harnessed in an assay which measures these induced calcium transients, allowing for quantification of pharmacodynamic effects of compounds at monoamine transporters. It is unknown if this electrical coupling occurs in neurons, but the implications if it does are significant. In the current work, fluorescent resonance energy transfer …

Role Of C121a In Mglur2 Homodimeric Expression And Function, Jong M. Shin

Theses and Dissertations

The group II metabotropic glutamate receptors are known for their involvement in various psychiatric disorders. The mGluR2 in particular is linked with etiology of schizophrenia especially in the context of crosstalk with 5-HT2A. Thus, the mGluR2 has attracted attentions for its potential therapeutic applications. Despite numerous physiological evidences on the actions of mGluR2, its mechanism is still unclear to this day. It is partially due to the lack of understanding in characteristics of mGluR2 homodimer which is its functionally active form. Therefore, the characterization of dimeric interaction serves as a foundation to advanced understanding of the role of mGluR2. On …

Impact Of Aerobic Exercise On Monocyte Subset Receptor Expression And Macrophage Polarization, Anson M. Blanks

Theses and Dissertations

Atherosclerotic cardiovascular disease (CVD) is hallmarked by inflammatory immune activation, particularly by the induction of a response by monocytes. Classical (CD14⁺⁺CD16^-) are anti-inflammatory mediators under homeostatic conditions, while intermediate (CD14⁺⁺CD16⁺) and non-classical (CD14^LowCD16⁺⁺) monocytes promote inflammation following activation. Monocyte activation and functionality is dependent upon receptor expression and ligand production by a variety of cells, including monocytes. Alterations in the expression of surface receptors often have a direct impact upon monocyte function, such as the increased pro-inflammatory cytokine production in response to activation that accompanies elevated CD14 expression …

Nicotinic Acetylcholine Receptor Dependent Effects Of Nicotine On Hek293t And Hbo Cells, James D. Larsen

Theses and Dissertations

T2R receptors are the classical bitter taste receptors which detect and transduce bitter taste in a subset of taste receptor cells (TRCs). The TRPM5-dependent T2Rs are G-protein coupled receptors (GPCRs) and are linked to G protein, gustducin to initiate an intracellular signaling cascade for the transduction of bitter tastants. Nicotine is bitter. However, at present the transduction mechanisms for the detection of nicotine in are poorly understood. Previous studies from our laboratory using TRPM5 knockout (KO) mice demonstrated that the T2R pathway is insufficient in explaining the taste perception of nicotine. TRPM5 KO mice elicited chorda tympani (CT) taste nerve …

Chronic Clozapine Treatment Impairs Functional Activation Of Metabotropic Glutamate Receptor 2 Via An Hdac2-Depedent Mechanism, Travis M. Cuddy

Theses and Dissertations

Schizophrenia is a chronic mental disorder affecting millions worldwide. It has no known cure. Current pharmaceutical treatments have shown efficacy in only one of the three symptom clusters of schizophrenia, providing little or no benefit in the other two. Furthermore, the current standard-of-care drugs, known as atypical antipsychotics, carry risks of severe side effects affecting multiple body systems. Most patients opt to discontinue drug therapy within two years of initiation due to lack of efficacy and/or preponderance of adverse effects. Previous findings have shown that chronic usage of atypical antipsychotics causes a 5-HT_2A-dependent upregulation of histone deacetylase 2 …

Identification Of Expression And Function Of The Glucagon-Like Peptide-1 Receptor In Gastrointestinal Smooth Muscle, Alexander T. May

Theses and Dissertations

In response to ingestion of nutrients, enteroendocrine L cells secrete the incretin hormone, glucagon-like peptide-1 (GLP-1), to enhance glucose-dependent insulin release. Therapies related to GLP-1 are approved for type 2 diabetes. The GLP-1 receptor (GLP-1R) is expressed in cells of the gastrointestinal tract and elsewhere. In pancreatic beta cells, GLP-1R are coupled to the Gs/cAMP/PKA pathway. The expression and function of GLP-1R in gastrointestinal smooth muscle are not known. Aim. To test the hypothesis that GLP-1 regulates smooth muscle function by acting on GLP-1R expressed on smooth muscle. Methods. Smooth muscle cells (SMC) were isolated and cultured. Expression of GLP-1R …

N-Alkyl 4-Methylamphetamine Enantiomers And The Implication For Potential Modulation Of Abuse Liability And Enhancement Of Psychoactive Drug Targeting., Ramsey Sitta

Theses and Dissertations

Drugs of abuse have a long history in humanity. Currently however, a subject of great interest is the phenylalkylamine family of drugs. Not only is the abuse liability of interest but also the potential therapeutic expansion of the capabilities of this family of drugs by utilizing the unique stereospecific effects of the newly discovered hybrid compounds. Based upon prior data of N-Alkyl 4-MA the enantiomers of N-Methyl, N-Ethyl, and N-Propyl were analyzed in hDAT, hNET, and hSERT. It was found that there was a negative correlation between chain length and potency and dopaminergic component. In agreement with the currently established …

Hydrogen Sulfide Regulation Of Kir Channels, Junghoon Ha

Theses and Dissertations

Inwardly rectifying potassium (Kir) channels establish and regulate the resting membrane potential of excitable cells in the heart, brain and other peripheral tissues. Phosphatidylinositol- 4,5-bisphosphate (PIP2) is a key direct activator of ion channels, including Kir channels. Gasotransmitters, such as carbon monoxide (CO), have been reported to regulate the activity of Kir channels by altering channel-PIP2 interactions. We tested, in a model system, the effects and mechanism of action of another important gasotransmitter, hydrogen sulfide (H2S) thought to play a key role in cellular responses under ischemic conditions. Direct administration of sodium hydrogen sulfide (NaHS), as an exogenous H2S source, …

Po2 Dependence Of Oxygen Consumption In Skeletal Muscle Of Hypertensive And Normotensive Rats, Habiba Shah

Theses and Dissertations

Human essential hypertension affects over 75 million people in the United States, and can lead to death due to its several serious health complications such as hypertension-related cardiovascular disease. The purpose of this research was to understand how hypertension could cause physiological changes to the microcirculation, specifically the PO₂ dependence of oxygen consumption (VO₂) in skeletal muscle of normotensive and hypertensive rats. The Spontaneously Hypertensive Rat (SHR) strain was used as the diseased model, and Wistar-Kyoto (WKY) rats were used as controls to conduct this study. The SHR strain develops hypertension between 5-6 weeks after birth with …

Deletion Of Cardiac Mir-17-92 Cluster Increases Ischemia/ Reperfusion Injury Via Pten Upregulation, Meeta B. Prakash

Theses and Dissertations

The miR-17- 92 cluster is necessary for cell proliferation and development of the cardiovascular system. Deletion of this cluster leads to death in neonatal mice. The role of this cluster still needs to be defined following ischemia and reperfusion. Methods and Results: Adult male mice were injected with Tamoxifen- was to induce inducible cardiac-specific miR-17- 92-deficient (miR-17- 92-def: MCM:TG:miR-17- 92 flox/flox ) and wild type (WT: MCM:NTG:miR-17-92 flox/flox ) mice were subjected to 30 minutes of myocardial ischemia via left anterior descending coronary artery ligation followed by reperfusion for 24 hours. Post I/R survival (48%) and ejection fraction were reduced, …

The Role Of Human Msc Derived Exosomes In The Treatment Of Periodontal Diseases, Sonia S. Talegaonkar

Theses and Dissertations

Periodontal disease affects 47% of Americans over 30. Characterized by microbial dysbiosis and unregulated inflammation, severe periodontitis causes degradation of bone and soft tissue around teeth. Current treatments have limited regenerative outcomes and frequent reinfection by harmful bacteria. Human mesenchymal stem cells (hMSCs) have been shown to promote wound healing and tissue regeneration. Many therapeutic benefits of hMSCs are due to their secretome products, like exosomes. Our long-term goal is to develop periodontal therapies with hMSC exosomes. The objectives of this study were to determine the effect of hMSC-derived exosomes on cellular activity of hMSCs and investigate whether hMSC exosome …

Hydrogen Peroxide And Pharmacological Agent Modulation Of Trpv2 Channel Gating, Tuoxin Cao

Theses and Dissertations

Transient receptor potential vanilloid 2 channel (TRPV2) is a Ca²⁺-permeable ion channel that is highly expressed in leukocytes but is also present in skeletal and cardiac muscle and endocrine cells. The TRPV2 function is implicated in a number of physiological processes, including bacterial phagocytosis, pro-inflammatory cytokine production, cardiac hypertrophy, and cancer development. TRPV2 knockout mice exhibit a high incidence of perinatal mortality, arguing that the channel plays essential roles in physiology. Despite the importance of TRPV2 for normal homeostasis, the mechanisms that control TRPV2 gating in response to pharmacological agonists, heating, membrane stretch, bioactive lipids and reactive oxygen …

Tissue Factor Expression, Regulation, And Signaling In Human Airway Cells, Michael D. Davis

Theses and Dissertations

Rationale: Tissue Factor (TF) is a transmembrane glycoprotein that canonically functions as the initiator of the coagulation cascade. Increased levels of TF have been associated with inflammatory airway diseases. Since lipopolysaccharide (LPS) is known to elicit and inflammatory response in airway epithelium, we hypothesized that airway epithelial cells release TF when exposed to LPS. Since TF aids in local wound healing, we also hypothesized that inhibition of TF would decrease NHBE growth. The specific aim of this work was to evaluate the effects of LPS exposure on TF production and release from airway epithelia and determine the signaling pathways involved. …

A Mechanistic Study Of An Ipsc Model For Leigh’S Disease Caused By Mtdna Mutataion (8993 T>G), John P. Galdun

Theses and Dissertations

Mitochondrial diseases encompass a broad range of devastating disorders that typically affect tissues with high-energy requirements. These disorders have been difficult to diagnose and research because of the complexity of mitochondrial genetics, and the large variability seen among patient populations. We have devised and carried out a mechanistic study to generate a cell based model for Leigh’s disease caused by mitochondrial DNA mutation 8993 T>G. Leigh’s disease is a multi-organ system disorder that depends heavily on the mutation burden seen within various tissues. Using new reprogramming and sequencing technologies, we were able to show that Leigh’s disease patient fibroblasts …