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    Please use this identifier to cite or link to this item: https://ir.fy.edu.tw:8080/ir/handle/987654321/16496


    Title: Vibrio vulnificus RtxA1 modulated calcium flux contributes reduced internalization in phagocytes
    Authors: Soong-Yu Kuo;Miao-Chen Chou;Shin-Luen Lee;Yu Wang;Chun-Liang Chen;Pei-Ting Lin;Horng-Ren Lo
    Contributors: 輔英科技大學 醫學檢驗生物技術系
    Keywords: Vibrio vulnificus;RtxA1;Internalization;Cytotoxicity;Calcium flux
    Date: 2015-04-01
    Issue Date: 2015-08-24 20:32:42 (UTC+8)
    Abstract: Aims
    Vibrio vulnificusis an opportunistic pathogen that causes primary septicemia and wound infection with high mortality rate. This pathogen produces an RTX toxin (RtxA1) which can cause host cell rounding, cell death and interference with internalization by host phagocytes. However, the mechanism of RtxA1-induced phagocyte paralysis is not clear.
    Main methods
    Using the murine macrophage cell line RAW264.7, we measured cytotoxicity and phagocytosis ofV. vulnificusin normal and calcium-depleted media. To deplete extracellular and cytosolic Ca2 +, cells were exposed to the calcium chelators ethylene glycol tetraacetic acid (EGTA) and 1,2-bis-(o-aminophenoxy)-ethane-N,N,N′,N′-tetraacetic acid, tetraacetoxymethyl esteris (BAPTA-AM), respectively. The cytotoxicity was examined by measuring the activity of lactate dehydrogenase (LDH) released from the damaged cells. The gentamicin protection assay was conducted to determine the number of internalized bacteria, while acridine orange staining was applied to visualize the intracellular bacteria. The fluorescent indicator fura-2-acetoxymethyl ester (fura 2-AM) was used to measure the Ca2 +signal post-infection.
    Key findings
    We revealed that extracellular Ca2 +was essential for phagocytes to internalizeV. vulnificus. Meanwhile, cytosolic Ca2 +flux in RAW264.7 cells induced by an RtxA1 isogenic mutant was repressed by the parent strain. Furthermore, depletion of extracellular Ca2 +level by EGTA significantly reduced the cytotoxicity but did not affect the antiphagocytic activity of RtxA1 toxin.
    Significance
    Our results indicated that RtxA1 may interfere with cytosolic Ca2 +flux of phagocyte to promote bacteria colonization.
    Relation: Life Sciences 0(132),55-60
    Appears in Collections:[醫學檢驗生物技術系] 期刊論文

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