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DTSTART;TZID=America/New_York:20250721T140000
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SUMMARY:ChE PhD Dissertation Defense: William Doherty
DESCRIPTION:Name: William Doherty \nTitle: Stimulating Excitable Cells with Optosomes: Development of a Non-viral Cell Derived Vesicle Capable of Stimulating Excitable Cells in Response to Light Stimulus \nDate: 07/21/2025 \nTime: 2:00:00 PM \nCommittee Members:\nProf. Ryan Koppes (Advisor)\nProf. Abigail Koppes\nProf. Benjamin Woolston\nProf. Rebecca Shansky \nLocation: Shillman Hall 420 \nAbstract: \nFor years\, researchers have studied and developed neuromodulation techniques meant to stimulate and/or inhibit excitable cells both in research and clinical settings. A method to excite cells with light\, termed Optogenetics\, has been researched extensively since its discovery in the early 2000’s. A major constraint of Optogenetics is the expression of the necessary light-gated ion channels most often achieved using a viral vector. While this is not overly concerning in research settings\, clinical applications of optogenetics have been slow to develop as the use of viral vectors in humans presents challenges regarding safety. Additionally\, foreign opsin genes are believed to be a permanent addition to the transfected cells. \nThis dissertation aimed to develop Optosomes; a cell-derived vesicle containing excitatory opsin that couples with excitable cells via Gap-Junctions that conduct the stimulus current from the opsin into the cell. Initial production of Optosomes followed established protocols for producing Giant Plasma Membrane Vesicles (GPMVs) in which small volumes of cytoplasm are encapsulated in a piece of the cell’s plasma membrane. The number of GPMVs produced varied with pH\, cell confluency\, and base medium having a noticeable impact on the number of GPMVs generated. Optosome production required the creation of a stable cell line expressing Channelrhodopsin-2 (ChR2) and connexin-43 (Cx43) proteins required to form Gap-Junctions. Two separate transfections in the series generated a ChR2-Cx43 Hek293 cell line capable of producing Optosomes at a high concentration. Finally\, a mathematical model was built to simulate Optosome stimulation of excitable cells and how changes in the size of Optosomes and cells affect the strength of stimulus generated. The result of these simulations and attempts to stimulate neonatal Cardiomyocytes (CM) in vitro confirmed that the majority of Optosomes produced were too small to generate a stimulus capable of exciting CMs. Production of Optosomes with larger diameters or the use of a different strand of ChR2 is needed to increase the number of Optosomes able to stimulate CMs will be needed moving forward. \nThe results of this dissertation provide the foundation for developing Optosomes as an alternative approach to stimulating excitable cells with light. \n\nWilliam Doherty Northeastern University-Department of Chemical Engineering After spending nearly two years working on the development of a new automated Biomanufacturing system in the Love Lab\, Bill was accepted and enrolled in the PhD program for Chemical Engineering. After finding his home for the next 7 years in the Koppes Lab\, he got to work both on forming his thesis and integrating into the community at Northeastern. In pursuing his Ph. D\, he had started to appreciate how applying mathematical modeling techniques to biological systems offers a whole new perspective when trying to understand the complex innerworkings of the human body. It offered a nice juxtaposition to the time spent in lab running hands on experiments that are less about math and academic prowess and more about technique\, adaptability\, and problem solving in real time. Bill has sed the better part of his twenties working in Research and its why he was so eager to pursue a Ph D as he hopes to work his way into scientist positions overseeing research and development projects. Still residing in Boston\, he hopes to find a position in the New England Area after submitting his Dissertation; staying close to family and friends in the area.
URL:https://che.northeastern.edu/event/che-phd-dissertation-defense-william-doherty/
LOCATION:420 Shillman Hall\, 360 Huntington Ave\, Boston\, MA\, 02115\, United States
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SUMMARY:ChE PhD Dissertation Defense: Matthew Fernez
DESCRIPTION:Name:\nMatthew Fernez \nTitle:\nDevelopment of a Hydrogen Sulfide Biosensor and Interrogation the Role of Sulfide on Mucus Barriers \nDate:\n06/05/2024 \nTime:\n10:00:00 AM \nCommittee Members:\nProf. Benjamin Woolston (Advisor)\nProf. Rebecca Carrier (Co-Advisor)\nProf. Erel Levine\nDr. Jodie Ouahed \nLocation:\nShillman 420 \nAbstract:\nInflammatory bowel disease (IBD) is a gastrointestinal condition that is estimated to afflict more than 1% of adults\, with prevalence increasing over time. Hydrogen sulfide (H2S)\, produced by the gut microbiota and partially by the host epithelium\, has long been associated with IBD through metagenomic evidence of increased populations of sulfate producing bacteria in patients. However\, the effects of sulfide are disputed and the mechanisms underlying disease pathogenesis remain unresolved. Sulfide literature suggests both a therapeutic and restorative effect at low levels yet also implies higher levels may be toxic\, resulting in disruption of mucus barriers and promotion of an inflammatory cascade. The mucus barrier is comprised of a mucin network crosslinked by disulfide bonds\, along with myriad heterogenous components\, which provides protection for the cell lining from the microbiota and toxic byproducts. We hypothesize that in excess\, sulfide reduces disulfide bonds yielding a weaker\, more permeable barrier that is susceptible to microbial penetration. However\, studying sulfide in the gut microenvironment is technically challenging due to its high volatility and reactivity\, making accurate GI measurements challenging in vivo. Here\, we seek to address measurement limitations through the development of a sulfide biosensor and evaluate the impacts of sulfide on mucus barriers. \nThis thesis has three core elements: optimization of a functional sulfide biosensor\, exploration of implementation in a mesofluidic gut-on-chip\, and direct evaluation of sulfide on mucus barrier properties. To construct a transcriptional sulfide biosensor\, a polysulfide sensitive repressor\, SqrR\, from Rhodobacter capsulatus was codon harmonized and inserted into a plasmid with a fluorescent reporter module driven by the SqrR affiliated promoter pSqr. To convert sulfide into polysufides that de-activate SqrR\, we coupled the sensing plasmid with an enzymatic plasmid bearing codon harmonized Sqr from R. capsulatus. Initial construction failed to generate a response to sulfide\, but an 18-fold dynamic range up to 750 M in aerobic conditions was produced through pSqr engineering and SqrR tagging and titration. Sqr activity and products were characterized\, but failed to generate a response under anaerobic conditions likely due to electron transport chain dependent quinone incompatibility. Given that the gut microenvironment is largely anaerobic\, sensor response under anaerobic conditions was recovered through isolation of a novel Sqr homolog from Wolinella succinogenes\, an organism that couples sulfide oxidation to reduction of fumarate\, a compound abundant in the gut. We demonstrate sensor activity under anoxic conditions using this enzyme with both fumarate and nitrate\, a marker of the inflamed gut. The group has previously developed a gut on chip PDMS device with vertical orientation for cross sectional imaging of the mucus barrier of primary intestinal epithelium under static conditions. The gas permeability of PDMS encouraged us to attempt gel wall modifications to permit consistent delivery of sulfide via flow. While modifications of the gut chip to operate under continuous flow culture were unsuccessful\, a simple demonstration of a partial response to sulfide by the biosensor on chip is presented in the absence of epithelium\, illustrating promise for implementation as a diagnostic in the future. Finally\, the impacts of sulfide on mucus were elucidated in the third part of the thesis. Using harvested porcine intestinal mucus (PIM)\, we demonstrate that diffusivity of polystyrene nanoparticles in PIM increases 1.6 fold when treated with 1 mM sulfide. Sulfide also impacted microbial velocity and motion type\, yielding a statistically significant increase of average velocity and proportion of microbes exhibiting penetration like behavior. While structural changes were not observed through lectin staining of PIM\, the thesis culminates in a simple demonstration of deleterious effects of sulfide on a living mucus barrier in our gut on chip system. Mucus thickness was reduced and resulted in an increased proportion of nanoparticles that penetrated the mucus barrier. Taken together\, this work produced a novel sulfide biosensor that functions under aerobic and anaerobic conditions and characterized the deleterious effects of sulfide on barrier permeability suspected by the underlying hypothesis of reduction of disulfide bonds in mucus structure. \n\n \nMatt Fernez is currently a sixth year PhD student in Chemical Engineering at Northeastern University. He completed a Bachelor of Science in Chemical Engineering in May\, 2019 at University of Massachusetts Amherst where he also produced an honors thesis. In June of this year\, he will defend his PhD Thesis to complete the doctoral degree. His research is co-advised and focuses on synthetic biology and tissue engineering\, with the focus of studying mucus barrier properties utilizing genetically engineered bacteria. His dissertation is titled “Development of a Hydrogen Sulfide Biosensor and Interrogation the Role of Sulfide on Mucus Barriers”. He has contributed towards authorship on 4 publications\, including one review paper and three technical papers. His first author sulfide biosensor manuscript was recently published in ACS Synthetic Biology in Spring\, 2025. His research has been presented at 4 conferences\, culminating in an oral presentation at the American Institute of Chemical Engineering international conference in October\, 2024. He has been a mentor to several undergraduate students who have produced research posters at Northeastern’s undergraduate research conference. He also assisted in building the foundation of the now well-established Woolston Lab\, being one of the first two students and serving as the Lab Safety Officer. Outside of research\, Matt has many recreational hobbies including poker\, hiking\, travelling\, and fantasy sports. He is the president and founder of a local poker club in the greater Boston area. He also enjoys spending time with family and writing poetry.
URL:https://che.northeastern.edu/event/che-phd-dissertation-defense-matthew-fernez/
LOCATION:420 Shillman Hall\, 360 Huntington Ave\, Boston\, MA\, 02115\, United States
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