Apoptotic human cells inhibit migration of granulocytes via release of lactoferrin
Irini Bournazou, John D. Pound, Rodger Duffin, Stylianos Bournazos, Lynsey A. Melville, Simon B. Brown, Adriano G. Rossi, Christopher D. Gregory
Irini Bournazou, John D. Pound, Rodger Duffin, Stylianos Bournazos, Lynsey A. Melville, Simon B. Brown, Adriano G. Rossi, Christopher D. Gregory
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Research Article Immunology

Apoptotic human cells inhibit migration of granulocytes via release of lactoferrin

Abstract

Apoptosis is a noninflammatory, programmed form of cell death. One mechanism underlying the non-phlogistic nature of the apoptosis program is the swift phagocytosis of the dying cells. How apoptotic cells attract mononuclear phagocytes and not granulocytes, the professional phagocytes that accumulate at sites of inflammation, has not been determined. Here, we show that apoptotic human cell lines of diverse lineages synthesize and secrete lactoferrin, a pleiotropic glycoprotein with known antiinflammatory properties. We further demonstrated that lactoferrin selectively inhibited migration of granulocytes but not mononuclear phagocytes, both in vitro and in vivo. Finally, we were able to attribute this antiinflammatory function of lactoferrin to its effects on granulocyte signaling pathways that regulate cell adhesion and motility. Together, our results identify lactoferrin as an antiinflammatory component of the apoptosis milieu and define what we believe to be a novel antiinflammatory property of lactoferrin: the ability to function as a negative regulator of granulocyte migration.

Authors

Irini Bournazou, John D. Pound, Rodger Duffin, Stylianos Bournazos, Lynsey A. Melville, Simon B. Brown, Adriano G. Rossi, Christopher D. Gregory

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Figure 2

Biochemical characterization of the inhibitory factor.

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Biochemical characterization of the inhibitory factor. Conditioned media...
Conditioned media from BL2 cells cultured for 24 hours were size fractionated using filters with 50 kDa (A) molecular weight cutoff sizes. Unfiltered medium was included as control. *P < 0.001 compared with the corresponding positive control. Error bars indicate SEM. Ion exchange analysis included the use of Q Sepharose beads (positively charged) in order to distinguish positively and negatively charged molecules in the <100 kDa fraction (B) of the BL medium. Unbound molecules (Q1 fraction) were collected, whereas bound molecules were eluted from the beads (Q2 fraction). Neutrophil migration toward these fractions in the presence of fMLP (100 nM) was assessed. Q1 and Q2 fractions (unbound and eluant fraction) of serum-free medium (no BL) were included as control. †P < 0.05 compared with the corresponding control. Error bars indicate SEM. (C) Chemotaxis assay of neutrophils toward BL-conditioned medium that was heat inactivated (90°C for 10 minutes). (D) MALDI-TOF mass spectrum for the tryptic digest of the peptide band identified as lactoferrin.