The role of sodium(I)/hydrogen(I) exchanger-1 (NHE1) in mammary branching morphogenesis and maintenance of tissue architecture
Item
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Title
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The role of sodium(I)/hydrogen(I) exchanger-1 (NHE1) in mammary branching morphogenesis and maintenance of tissue architecture
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Identifier
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d_2009_2013:d7d58b26b794:11874
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identifier
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12514
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Creator
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Jenkins, Edmund C., Jr.,
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Contributor
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Jimmie E. Fata
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Date
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2013
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology | Developmental biology | Biochemistry | branching | mammary gland | NHE1
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Abstract
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Branching morphogenesis in vivo is a highly ordered process that necessitates spatially and temporally choreographed cues by growth factors and hormones, as well as mechanical and signal feedback from the extracellular matrix. Successful completion of this developmental process results in the architectural, and thereby the functional, basis for the lung, collecting ducts of the kidney, salivary, and mammary glands. The quest to understand the basic biological mechanisms underlying this developmental morphogenesis has lead to many seminal findings in the field of epithelial tube generation, as well as provided valuable insight into the pathogenesis of cancer. The primary focus of this thesis was elucidating the role of the Na+/H+ exchanger type 1 (NHE1) in branching morphogenesis of the mouse mammary gland. To accomplish this goal, we used three-dimensional (3D) primary tissue culture of mammary gland pieces (organoids) in a four day organotypic assay of growth factor induced branching morphogenesis. NHE1 is a ubiquitously expressed master regulator of intracellular pH (pHi). We found that blocking the function of this exchanger in the presence of growth factor stimulation led to altered kinase signaling, inhibition of growth factor induced alkalization, sustained proliferation after four days, ectopic expression of keratin 6 (K6), and a dramatic failure to undergo branching morphogenesis.;These findings led us to question the role of NHE1 in the maintenance of branched mammary tissue architecture. We, therefore, inhibited NHE1 function on fully branched structures in our assay and found that NHE1 inhibition led to rapid loss (within 24 hours) of branched architecture by a process of branch fusion, with complete loss of the branched morphology after four days. This was not accompanied by cell death or altered proliferation, however, we did record altered intracellular pH (pHi) in the end buds of branched structures that had NHE1 inhibited. NHE1 localization, F-actin organization, and myoepthelial cell location were altered in structures that had undergone a loss of architecture, indicating a loss of tissue organization. Finally, we found that NHE1 inhibition resulted in a decrease in mammary E-cadherin.;Having found that NHE1 function is vital for both branching morphogenesis and the maintenance of branched architecture, we considered the role that NHE1 could be playing in the pathology of breast cancer. Both intracellular and extracellular pH is deregulated in cancer. This could be attributed to over activity of NHE1. Additionally, NHE1 is over expressed in many cancers. We used the ER+ breast cancer cell line MCF7 to investigate the therapeutic potential of chemotherapy augmentation by NHE1 inhibition. We found that Cycylophosphamide, a DNA alkylating chemotherapeutic agent known to be more effective in an acidic environment, was roughly 5 times more effective when used along with NHE1 inhibition. These findings indicate that NHE1 is a critical regulator of branching morphogenesis and tissue stability, as well as suggests a potential therapeutic target for the treatment of breast cancer.
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Type
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dissertation
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Source
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2009_2013.csv
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degree
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Ph.D.
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Program
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Biology
