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Electrochemical Evaluation of Plasma Membrane Cholesterol in Live Cells and Mouse Tissues

Jiang, Dechen
http://rave.ohiolink.edu/etdc/view?acc_num=case1212761216

Abstract Details

2008, Doctor of Philosophy, Case Western Reserve University, Chemistry.
In this work, microelectrodes covalently modified with cholesterol oxidase were implemented for evaluation of the cholesterol content in the plasma membrane of single cells at physiological temperature. Cholesterol is a tightly regulated structural component of the cell plasma membrane. Electrochemical data indicated that transport of intracellular cholesterol to the plasma membrane was active in an atherosclerotic macrophage model and was inhibited in a human fibroblast model of Niemann-Pick disease. Electrochemical detection of plasma membrane cholesterol at the surface of excised mouse trachea tissue was reported. Experiments involving direct contact between the cholesterol oxidase-modified electrode and the surface of excised trachea tissue at 37 °C indicated steady state responses that were largely independent of the contact position on the tissue surface and the contact force. Trachea tissue excised from a mouse model of cystic fibrosis showed an electrode response that was about 40% larger than the response observed for wild-type mouse trachea tissue. Electrochemical studies of cholesterol chemical activity in a 1, 2-dipalmitoyl-sn glycero-3-phosphocholine (DPPC) lipid monolayer formed at the air-water interface were also performed. The current responses showed an increase when the molecular ratio of cholesterol to lipid was above 0.6, suggesting an existence of the stoichiometric condensed cholesterol-phospholipid complex in lipid monolayers. Further study in cystic fibrosis showed excess cholesterol stored in lysosomes and elevated cholesterol content in the plasma membrane. This suggested a possibility of increased Niemann-Pick disease, type C1 (Npc1) - driven membrane cholesterol transport in the absence of CFTR function. An application of an oxidase-modified electrode in the research of germ cell migration led to the observation of an elevated cholesterol content at the surface of genital ridge. The data supported a model where cholesterol was required for primordial germ cells development, but can be supplied either via uptake or synthesis. Scanning force microscopic images on mica exhibited features assigned to Streptomyces Sp. Cholesterol oxidase (class I) of 1-2 nm in height immobilized on membrane plateaus and 3-4 nm in height in Pseudomonas Sp. Cholesterol oxidase (class II). This phenomenon could be reasonably explained for the larger ternary crystal structure of Class II cholesterol oxidase.
James Burgess (Advisor)
Daniel Scherson (Committee Chair)
Robert Salomon (Committee Member)
Carlos Crespo (Committee Member)
Thomas Kelley (Committee Member)
133 p.
Chemistry
oxidase modified microelectrode; intracellular cholesterol trafficking; cystic fibrosis

Recommended Citations

Citations

  • Jiang, D. (2008). Electrochemical Evaluation of Plasma Membrane Cholesterol in Live Cells and Mouse Tissues [Doctoral dissertation, Case Western Reserve University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=case1212761216

    APA Style (7th edition)

  • Jiang, Dechen. Electrochemical Evaluation of Plasma Membrane Cholesterol in Live Cells and Mouse Tissues. 2008. Case Western Reserve University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=case1212761216.

    MLA Style (8th edition)

  • Jiang, Dechen. "Electrochemical Evaluation of Plasma Membrane Cholesterol in Live Cells and Mouse Tissues." Doctoral dissertation, Case Western Reserve University, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=case1212761216

    Chicago Manual of Style (17th edition)

case1212761216
625
© 2008, all rights reserved.
This open access ETD is published by Case Western Reserve University School of Graduate Studies and OhioLINK.