A. James Clark School of Engineering

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The collections in this community comprise faculty research works, as well as graduate theses and dissertations.

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Subject: search.filters.subject.Blood-brain barrier

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    Metabolic Profiling of Brain Microvascular Endothelial Cells: Investigating the Role of Sex, Stress, APOE Genotype, and Exercise in Alzheimer's Disease Risk
    (2024) Weber, Callie; Clyne, Alisa M; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Alzheimer’s disease (AD) is the 7th leading cause of death in the United States, yet there are still no effective treatments to prevent or slow the progression of the disease. AD develops from a combination of genetic and lifestyle risk factors including female sex, elevated stress hormone exposure, the apolipoprotein (APOE) ε4 genotype, and a sedentary lifestyle. In order to better identify the manifestations of AD, it is vital to understand how each of these risk factors impact brain health and lead to neurological dysfunction associated with AD. Brain microvascular endothelial cells (BMEC) line the blood vessels of the brain and have specialized tight junctions designed to strictly regulate nutrient and waste transfer between the blood and the brain. Two of the early indicators of AD development are breakdown of the tight junctions and whole brain glucose hypometabolism. Since BMEC form the first line of defense for the brain against neurotoxic compounds in the blood and are responsible for glucose transport to the rest of the brain, the overarching goal of this thesis is to understand how female sex, elevates stress hormone exposure, the APOE ε4 genotype, and a sedentary lifestyle induce breakdown of tight junction proteins and glucose hypometabolism in BMEC. I first demonstrate that female sex exacerbates endothelial dysfunction in response to high levels of a stress hormone, Angiotensin II (AngII). Specifically, I show that in response to AngII, female endothelial cells increase oxidative stress and inflammatory responses while male endothelial cells do not. Next, I used CRISPR/Cas9 to generate a set of induced pluripotent stem cells (iPSC) homozygous for the APOE ε3 and ε4 genotype and differentiated them into BMEC (hiBMEC). Using the hiBMEC I showed the APOE ε4 genotype induces barrier deficiencies that are partially mediated through reduced levels of protein deacetylase Sirtuin 1 (SIRT1), and that the APOE ε4 genotype causes glucose hypometabolism through decreased insulin signaling. Finally, by adding serum from sedentary and exercise trained individuals to genotype-matched hiBMEC, I show that APOE ε3 and ε4 hiBMEC have divergent responses to treatment with serum from sedentary and exercise trained individuals. Treatment with exercise trained serum increases SIRT1 and glycolytic enzymes compared to sedentary serum, while exercise trained serum decreases SIRT1 and glycolytic enzymes in APOE ε4 hiBMEC compared to sedentary serum. The work described in this thesis gives a fundamental, mechanistic understanding to the roles of female sex, stress hormone exposure, the APOE ε4 genotype, and a sedentary lifestyle in BMEC dysfunction and hypometabolism, giving insight into how these factors contribute to AD development and progression.
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    LIGHT ACTIVATABLE PURE PORPHYRIN NANOPARTICLES FOR THE PHOTODYNAMIC OPENING OF THE BLOOD-BRAIN BARRIER AND GLIOBLASTOMA TREATMENT
    (2022) Inglut, Collin Thomas; Huang, Huang Chiao; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Glioblastoma (GBM) consistently recurs due to infiltrating cancer cells that cannot be removed by surgery and chemotherapy. The diffusive nature of GBM makes complete surgical resection unsafe, and the intact blood-brain barrier (BBB) prevents the penetration and accumulation of nearly all chemotherapy in infiltrative GBM cells. Existing BBB opening strategies are often associated with increased risk of edema, hemorrhage, or neurotoxicity and thus have limited clinical success. Photodynamic therapy (PDT) is a photochemistry-based treatment modality that has shown promise in treating GBM and opening the BBB in the clinic. In fact, a single adjunctive dose of PDT has been shown to add as much as 18 months to patient survival. However, the full potential of PDT is limited by the light activation depth of the ‘gold standard’ pro-drug photosensitizer, 5-aminolevulinic acid (5-ALA). In addition, large doses of PDT can result in edema and neurotoxicity. To address these issues, our lab has developed a photodynamic priming (PDP) strategy using the verteporfin (VP) photosensitizer, which operates at low optical energy to enhance intratumoral drug accumulation without damaging the healthy brain tissues. Unfortunately, VP is hydrophobic and requires liposomal encapsulation for intravenous administration, which can alter the photosensitizers cellular pharmaceutics. Here, we develop and compare a novel carrier-free pure-photosensitizer nanoparticle to a clinically relevant liposomal formulation.This dissertation covers a complementary, four-pronged approach to enhance drug delivery to brain tumors and treat GBM: (1) Understand the photoactivation depth of clinically relevant photosensitizers in the rodent brain for the targeting of infiltrative GBM cells. (2) Explore the mechanisms of photochemistry-induced BBB opening. (3) Engineer light-activable nanotechnology that can open the BBB, improve drug delivery, and eradicate GBM cells. And (4) develop a high-throughput model to examine the BBB integrity and efflux transporter function. The central hypothesis of this dissertation is the delivery of photoactivatable pure-photosensitizer nanoparticles can eradicate GBM cells and enhance drug delivery to microscopic GBM tumors.
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    Investigation of Vesicular-Mediated Transport of Intercellular Adhesion Molecule-1-Targeted Carriers for Treatment of Lysosomal Storage Disorders
    (2017) Manthe, Rachel Lee; Muro, Silvia; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Numerous cellular processes and therapeutic interventions rely on vesicular-mediated endocytosis to gain entry into cells and sub-cellular compartments, as well as for transcellular transport across biological barriers such as found at the blood-brain interface. Yet, endocytic behavior can be altered in disease, representing an additional hurdle in the design of effective therapeutic strategies. Lysosomal storage disorders (LSDs), characterized by lysosomal accumulation of undigested substrates as a result of deficient enzymatic activity, illustrate this paradigm. Currently, intravenous infusion of recombinant lysosomal enzymes to replace those deficient is the standard clinical approach for these disorders. However, clathrin-mediated endocytosis utilized by replacement enzymes for cellular uptake and lysosomal trafficking is altered, thereby impacting treatment efficacy as recently demonstrated in acid sphingomyelinase-deficient type A Niemann-Pick disease (NPD). Therefore, alternative means to bypass defunct routes is warranted. Therapeutic delivery via polymer nanocarriers targeting intercellular adhesion molecule-1 (anti-ICAM NCs), a cell-surface molecule overexpressed in endothelial and subjacent tissue cells during inflammation, such as in LSDs, represents a viable option since it permits uptake, intra- and transcellular transport via a unique endocytic route called the cell adhesion molecule (CAM) pathway. In this dissertation, cell culture and animal models were used to examine the (1) endocytic activity of the CAM pathway and other clathrin-independent routes in type A NPD, (2) role of targeting valency (i.e., density of ICAM-1-targeting molecules on the NC surface) in regulating the CAM pathway, and (3) effects induced via engagement of ICAM-1 on cells by anti-ICAM NCs. The results herein demonstrate the CAM pathway is more active in diseased cells compared to other classical endocytic pathways, making it the most amenable route for therapeutic enzyme replacement. Further, modulating targeting valency of NCs optimized this strategy for enhanced enzyme delivery to the brain, a target organ for type A NPD. Lastly, anti-ICAM NCs attenuated endothelial release of soluble ICAM-1, an inflammatory regulator, representing a secondary benefit of this system. Overall, this work validates utility of anti-ICAM NCs for enzyme replacement to treat NPD and likely other LSDs, and provides insight into biological processes and design parameters that influence the therapeutic efficacy of targeted drug carriers.
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    Targeting Intercellular Adhesion Molecule-1 to Enhance Delivery of Therapeutic Enzymes for Treatment of Lysosomal Storage Diseases
    (2014) Hsu, Janet; Muro, Silvia; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Lysosomal storage diseases (LSDs) are a group of more than 40 genetically inherited diseases that result from dysfunction of specific proteins, often an enzyme, located in lysosomes within cells which leads to abnormal lysosomal accumulation of specific macromolecules. As a result, cell malfunction occurs and escalates into multi-tissue and multi-organ failures, often resulting in premature death. For several early onset LSDs, the central nervous system (CNS) is also affected and manifests fatal neuropathic and/or neurodegenerative symptoms. Within the last two decades, treatment for selective LSDs has become clinically available. Specifically, enzyme replacement therapy (ERT) by intravenous injection of recombinant enzymes holds relevant promise. Yet current ERT results in suboptimal enzyme biodistribution to many target organs, including the peripheral organs and also the CNS. Delivery to the CNS is particularly impeded due to the tight blood-brain barrier (BBB) that strictly regulates passage between the circulation system and the brain tissue. We explored the use of targeted drug delivery systems to address this issue. Specifically, we focused on targeting intercellular adhesion molecule-1 (ICAM-1), a cell surface glycoprotein that is upregulated under pathological conditions, including LSDs. In this dissertation, using in vitro, cell culture, and in vivo techniques, we examined whether ICAM-1-targeted polymer nanocarriers: (1) enhance binding, uptake, and lysosomal delivery of different enzymes in cells, (2) provide targeting and transport across endothelial and subendthelial cells of the BBB, and (3) improve accumulation of lysosomal enzymes to peripheral organs and the brain. Results suggest that after intravenous injection of enzyme coupled to ICAM-1-targeted nanocarriers, ICAM-1 targeting shift these enzymes from the circulation to tissues, enhancing enzyme accumulation over non-targeted counterparts both in peripheral organs and the brain. This could be modulated by varying parameters such as the density of targeting antibodies on the carrier coat or the carrier bulk concentration. Also, ICAM-1-targeted nanocarriers were transported across BBB models followed by uptake and lysosomal transport to neuron-like cells. ICAM-1-targeted nanocarriers preferentially bound to diseased cells and were internalized and trafficked to lysosomes, resulting in degradation of the accumulated substrate. Therefore, overall, ICAM-1-targeting shows promise in improving ERT for LSD treatment.