title: Effects of IL-6 and IL-6 blockade on neutrophil function in vitro and in vivo.
abstract: OBJECTIVE
  Reports on the regulation of neutrophil function by IL-6 are often conflicting. Therapeutic inhibition of IL-6 in RA is associated with occasional neutropenia, but the mechanisms underlying this observation are poorly understood. This study investigated interactions between IL-6, the anti-IL-6 receptor agent tocilizumab (TCZ) and neutrophils in vitro and in vivo.
  
  
  METHODS
  Neutrophils were isolated from healthy controls and incubated in vitro with pharmacologically relevant concentrations of IL-6 or TCZ. Neutrophils were also isolated from RA patients, including a cohort following TCZ therapy. Apoptosis was measured by annexin V/propidium iodide (PI) flow cytometry; phagocytosis was measured by incubating apoptotic neutrophils with THP-1-derived macrophages; chemotaxis was measured using cell migration through hanging-cell inserts towards IL-8 and cell surface proteins, including adhesion molecules CD11b (αMβ2 integrin) and CD62L (L-selectin) were measured by flow cytometry.
  
  
  RESULTS
  IL-6 (10-100 ng/ml) did not affect the rate of neutrophil apoptosis, priming of the respiratory burst or adhesion molecule expression nor act as a neutrophil chemoattractant. However, IL-6 enhanced signal transducer and activator of transcription 3 (STAT3) activation and neutrophil migration towards IL-8. TCZ in vitro did not induce apoptosis or phagocytosis of neutrophils, nor did it have a significant effect upon apoptosis or cell surface molecule expression. Neutrophil functions in ex vivo neutrophils from RA patients receiving TCZ treatment were unaffected.
  
  
  CONCLUSIONS
  Therapeutic blockade of IL-6, while inducing a transient neutropenia, does not directly affect neutrophil functions associated with host defence. TCZ-associated neutropenia cannot be explained by direct induction of apoptosis by TCZ, induction of apoptosis following depletion of IL-6, nor increased phagocytosis of neutrophils.
authors.affiliation: Institute of Integrative Biology, University of Liverpool and Institute of Ageing and Chronic Disease, University Hospital Aintree, Liverpool, UK.
authors.forename: Helen L
authors.initials: HL
authors.lastname: Wright
authors.affiliation: Institute of Integrative Biology, University of Liverpool and Institute of Ageing and Chronic Disease, University Hospital Aintree, Liverpool, UK.
authors.forename: Andrew L
authors.initials: AL
authors.lastname: Cross
authors.affiliation: Institute of Integrative Biology, University of Liverpool and Institute of Ageing and Chronic Disease, University Hospital Aintree, Liverpool, UK.
authors.forename: Steven W
authors.initials: SW
authors.lastname: Edwards
authors.affiliation: Institute of Integrative Biology, University of Liverpool and Institute of Ageing and Chronic Disease, University Hospital Aintree, Liverpool, UK. rjmoots@liv.ac.uk.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Moots
mesh_terms: D000368:Aged; D000911:Antibodies, Monoclonal / Q000009:adverse effects / Q000494:pharmacology / Q000627:therapeutic use; D061067:Antibodies, Monoclonal, Humanized / Q000009:adverse effects / Q000494:pharmacology* / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001172:Arthritis, Rheumatoid / Q000188:drug therapy* / Q000473:pathology / Q000503:physiopathology; D016022:Case-Control Studies; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D002633:Chemotaxis / Q000187:drug effects / Q000502:physiology; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D015850:Interleukin-6 / Q000276:immunology / Q000494:pharmacology*; D008297:Male; D008875:Middle Aged; D009503:Neutropenia / Q000209:etiology; D009504:Neutrophils / Q000187:drug effects* / Q000473:pathology / Q000502:physiology; D010587:Phagocytosis / Q000187:drug effects / Q000502:physiology; D050796:STAT3 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D000093242:Interleukin-6 Inhibitors
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D015850:Interleukin-6; D050796:STAT3 Transcription Factor; D000093242:Interleukin-6 Inhibitors; C494086:STAT3 protein, human; C502936:tocilizumab
keywords: IL-6; neutropenia; neutrophil; rheumatoid arthritis; tocilizumab