Bacillus anthracis peptidoglycan (PGN) is a major component of the bacterial cell wall and a key pathogen-associated molecular pattern (PAMP) contributing to anthrax pathology, including organ dysfunction and coagulopathy. Increases in apoptotic lymphocytes are a late-stage feature of anthrax and sepsis, suggesting there is a defect in apoptotic clearance. Here, we tested the hypothesis that B. anthracis PGN inhibits the capacity of human monocyte-derived, tissue-like macrophages (Mφ) to efferocytose apoptotic cells. Exposure of CD206+CD163+ Mφ to PGN for 24h impaired efferocytosis in a manner dependent on human serum opsonins but independent of complement component C3. PGN treatment reduced cell surface expression of the pro-efferocytic signaling receptors MERTK, TYRO3, AXL, integrin αVβ5, CD36 and TIM-3, whereas TIM-1, αVβ5, CD300b, CD300f, STABILIN-1 and STABILIN-2 were unaffected. Soluble forms of MERTK, TYRO3, AXL, CD36, and TIM-3 were increased in PGN-treated supernatants, suggesting involvement of proteases. ADAM17 is a major membrane-bound protease implicated in mediating efferocytotic receptor cleavage. ADAM17 inhibitors TAPI-0 and Marimastat abolished TNF release, indicating effective protease inhibition, modestly increased cell-surface levels of MerTK and TIM-3 but only partially restored efferocytic capacity by PGN-treated Mφ. We conclude that human serum factors are required for optimal recognition of PGN by human Mφ and that B. anthracis PGN inhibits efferocytosis in part by reducing cell surface expression of efferocytic receptors.

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This study aimed to determine the impact of five proposed anti‐cancer agents (Berberine chloride, Azathioprine, Gossypol, Miltefosine, and Etoposide) on three female cancer cell lines: MES‐SA (uterine), C33A (cervical), and SKBR (breast). We hypothesized that treatment with these agents would decrease invasiveness based on our previous findings of their growth inhibitory properties. This was tested using the CultreCoat® 96 Well Medium BME Cell Invasion Assay. The cell lines were grown to 80‐90% confluency, then treated with the proposed anti‐cancer agents at 2µM for 24 hours. Cells were harvested and subsequently seeded in the invasion chamber for 24 hours. Cell migration, reflecting invasiveness, was monitored via a fluorimetric assay and quantified. Untreated cells served as controls. Invasiveness was influenced by the type of agent and cell line. Treatment with Gossypol increased invasiveness in all cell lines, the highest by 235% in MES‐SA. Azathioprine increased invasiveness in SKBR, and even more in C33A (141%). Berberine chloride reduced the invasiveness of SKBR by 33%, and C33A by 137%. The agent that showed the best reduction in invasiveness was Etoposide, with the biggest, 182% in C33A cells. Agents that showed an increase in invasiveness could have their potential as anti‐cancer therapy greatly reduced. Agents reducing invasiveness (particularly, Etoposide and Berberine Chloride) should be considered for further study, especially in C33A, the cell line most respondent to these agents. A dose‐dependent impact on invasiveness is currently being studied on a variety of cancer cells. Support for this project was provided by CSU Channel Island's Project Vista.