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Efferocytosis of apoptotic HPV positive cervical cancer cells by human primary fibroblasts
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International audience. Efficient clearance of dying cells is a fundamental physiological process for tissue development and homeostasis. The contribution of amateur phagocytes like fibroblasts to apoptotic cell efferocytosis has been established though the underlying mechanisms are not well understood. We recently demonstrated that horizontal DNA transfer can occur through the uptake of apoptotic HPV positive cancer cells by human primary fibroblasts (HPFs) leading to their transformation. In the present study, by using in vitro co-culture approaches, we confirm the role of HPFs as non-professional phagocytes in the context of apoptotic cervical cancer cell removal. Confocal microscopy imaging and flow cytometry analyses revealed that HPFs engulf more efficiently late apoptotic cells than early apoptotic targets, but their phagocytic ability remains limited compared to professional phagocytes as human monocytes-derived macrophages. Efferocytosis by HPFs occurs in a time-temperature- and calcium-dependent manner. Remodeling of actin-fibers contributes to the biogenesis of macroendocytic vacuoles. Both morphological analyses and pharmacological approaches confirmed the involvement of actin driven-phagocytosis and likely -macropinocytosis in apoptotic target internalization by HPFs. The uptake of apoptotic cell requires phosphatidylserine recognition. In HPFs, immunofluorescence and functional experiments using specific RNA interference assay suggested that this recognition is mainly mediated by Brain-specific Angiogenesis Inhibitor-1 (BAI1) and that this receptor contributes to subsequent uptake orchestration. Confocal microscopy analyses with organelle-specific markers revealed that internalized apoptotic material traffics into late phagolysosomes and specific-features of Microtubule-associated protein 1 Light Chain 3 alpha (LC3)-associated phagocytosis are observed. We conclude that efferocytosis by HPFs is mediated by phagocytosis/macropinocytosis involving phosphatidylserine recognition likely by the phosphatidylserine receptor BAI1.
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