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Connections Between Protein Misfolding and Monobenzone-Induced Vitiligo
Başlık:
Connections Between Protein Misfolding and Monobenzone-Induced Vitiligo
Yazar:
Carty, Senegal, author.
ISBN:
9780438007536
Yazar Ek Girişi:
Fiziksel Tanımlama:
1 electronic resource (81 pages)
Genel Not:
Source: Masters Abstracts International, Volume: 57-06M(E).
Advisors: Jillian Richmond; Noel D. Lazo Committee members: Denis Larochelle; Noel D. Lazo; Jillian Richmond; Luis Smith.
Özet:
Vitiligo, an autoimmune disease affecting melanocytes, causes areas of skin to completely lose pigment. These colorless lesions can spread, eventually leading to the complete absence of skin and hair pigment in some patients. Exposure to monobenzyl ether of hydroquinone (MBEH), or monobenzone, is an environmental trigger of this disfiguring disease. In this study, we aimed to elucidate the mechanism of this phenomenon by studying monobenzone's inhibition of tyrosinase, a melanocyte enzyme that plays a key role in melanin production. As tyrosinase is an oxidizing enzyme, we hypothesized that when inhibited by monobenzone, resulting elevated levels of reactive oxygen species (ROS) lead to tyrosinase misfolding and initiate the unfolded protein response, thus creating a link between a person's levels of active tyrosinase and their sensitivity to depigmentation by monobenzone. To test this hypothesis, studies of the effects of monobenzone on tyrosinase in simple solutions of tyrosinase and monobenzone, cell lines, primary cells and samples of human skin were conducted.
Structural studies of solutions of tyrosinase in the presence of monobenzone were conducted using circular dichroism (CD) spectroscopy. The CD results strongly suggest that the inhibitor's interaction with the enzyme does not cause any structural changes on a global scale. Chemiluminescence ROS assays in cell-free systems did not reveal any changes in ROS species concentration in solutions of tyrosinase in the presence of MBEH as opposed to L-tyrosine, its natural substrate. In light of these results, experiments using primary cells, cell lines and tissue samples were conducted to gain deeper insights into MBEH's effects within complex living systems.
We noted significant increases in ROS in B16 mouse melanoma cells treated with monobenzone, as well as evidence to suggest that this occurs in treated whole skin taken from donors. However, a similar response to monobenzone was not seen in the ROS assays performed on primary cells, possibly because the cells' adaptation to culture conditions reduced their sensitivity to the treatment. Immunohistochemistry and confocal microscopy were performed on cytospins to assess tyrosinase colocalization with the endoplasmic reticulum chaperonin calreticulin in cells treated with MBEH, as increased ER localization is a hallmark of aberrant protein folding. However, both MBEH-treated and vehicle-treated controls exhibited calreticulin localization at the cell membrane. This indicates that the vehicle alone caused a state of oxidative stress, possibly masking MBEH's effects.
As naturally occurring mutations in tyrosinase have been known to affect its folding stability, we hypothesized that this variation and the resulting reductions in cellular tyrosinase activity could lower monobenzone toxicity to exposed cells. In light of the potential of cultured cells expressing tyrosinase variants to shed some light on the variability in human sensitivity to this drug, HeLa cells were transfected with genes encoding the wild type and mutated forms of the enzyme. Certain cell populations show darkened coloration, a phenotypic sign of the presence of L-DOPA, suggesting tyrosinase activity.
Notlar:
School code: 0048
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Yer Numarası | Demirbaş Numarası | Shelf Location | Lokasyon / Statüsü / İade Tarihi |
---|---|---|---|
XX(694278.1) | 694278-1001 | Proquest E-Tez Koleksiyonu | Arıyor... |
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