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    Triptolide Induces Increases in Migration via Mitogen-activated Protein Kinase Phosphatase-1 control of P38 and JNK MAPK Activation

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    Author
    Parekh, Nili M.
    Keyword
    triptolide
    p38 Mitogen-Activated Protein Kinases
    JNK Mitogen-Activated Protein Kinases
    Date Published
    2010-07-16
    
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    URI
    http://hdl.handle.net/20.500.12648/1152
    Abstract
    Purpose Triptolide is a Chinese herbal extract known for its anti-inflammatory and immunosuppressive effects in treating chronic inflammatory diseases and tumors. As these attributes promote wound healing, we determined if triptolide enhances corneal wound healing by stimulating human corneal epithelial cell (HCEC) migration through changes in negative feedback regulation by a dual specificity protein Phosphatase (DUSP1) of MAPK signaling mediated effect. Methods SV40-adenovirus-immortalized HCEC were maintained in DMEM/F12. Specific shRNA for MKP-1(DUSP1) and c-jun NH2- terminal kinase JNK-1 were transduced to establish stable cell lines deficient in their respective gene expression. Scratch wound assay was employed to assess cell migration rates by taking time-dependent serial photographs of cells following wound creation. Hydroxyurea (2.5 mM) was also added to the medium to inhibit cell proliferation during the experiment. Cell Titer-Glo® luminescent cell viability assay was used to evaluate cell viability by measuring ATP production by HCEC. Results Triptolide did not affect cell viability up to 10nM and stimulated wound closure through increases in migration. Maximal responses occurred at 1nM. These increases in migration were suppressed below that in the untreated control when p38 or JNK MAPK activation was inhibited. In the MKP-1 knockdown cells, migration was stimulated relative to the control and triptolide failed to augment this response. In JNK-1 knockdown cells, migration is comparable to SV40 wild type cells. In JNK-1 knockdown cells, triptolide mediated increases are diminished completely in the presence of p38 inhibition. Conclusions Triptolide at concentrations up to 10 nM promotes cell migration without compromising cell survival. Such promotion is mediated by loss of MKP-1 negative feedback control of p38 and JNK activation. Therefore, triptolide stimulates cell migration through inhibition of MKP-1 (DUSP1) stabilization induced by kinase mediated phosphorylation.
    Description
    M.S. Research Paper in partial satisfaction of the requirements for the degree of Master of Science In Vision Science
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    SUNY Optometry Masters Thesis Collection

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