Human neutrophil antigen-3a antibodies induce neutrophil stiffening and conformational activation of CD11b without shedding of L-selectin

Transfusion. 2015 Dec;55(12):2939-48. doi: 10.1111/trf.13299. Epub 2015 Sep 10.


Background: HNA-3a antibodies induce severe transfusion-related acute lung injury (TRALI) in which neutrophils play a major role. As neutrophil passage through the pulmonary microvasculature is a critical step in the pathogenesis of TRALI, we investigated the impact of HNA-3a antibodies on two important factors that could impair granulocyte passage through lung capillaries: the elasticity of neutrophils and the expression and activation of adhesion molecules.

Study design and methods: The impact of HNA-3a antibodies on the elasticity of neutrophils was investigated using atomic force microscopy (AFM). Neutrophils were settled on poly-2-hydroxyethyl-methacrylate-coated glass slides before treatment with anti-HNA-3a plasma samples, control plasma, or control plasma containing formyl-methionyl-leucyl-phenylalanine (fMLP). Elasticity measurements were carried out in a temperature-controlled perfusion chamber using an atomic force microscopy (AFM) device. The impact of HNA-3a antibodies on the surface expression of total CD11b, activation of CD11b, and L-selectin (CD62L) shedding was investigated by flow cytometry. The functional impact of HNA-3a antibodies on neutrophil adhesion was assessed using fibrinogen-coated plates.

Results: HNA-3a antibodies induced stiffening of neutrophils (+24%-40%; p < 0.05) to a similar extent as fMLP. This effect was blocked by treatment of neutrophils with cytochalasin D. While total surface expression of CD11b and L-selectin on neutrophils was largely unaffected, HNA-3a antibodies induced alloantigen-specific activation of CD11b (+72%-107%; p < 0.05) and increased adhesion of neutrophils to fibrinogen.

Conclusion: Accumulation of neutrophils in the pulmonary microvasculature during severe TRALI is likely mediated by increased rigidity and CD11b-mediated adhesion of neutrophils leading to retention of neutrophils.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acute Lung Injury / etiology
  • CD11b Antigen / chemistry
  • CD11b Antigen / physiology*
  • Cell Adhesion
  • Humans
  • Isoantibodies / physiology*
  • Isoantigens / immunology*
  • L-Selectin / physiology*
  • Microscopy, Atomic Force
  • Neutrophils / physiology*
  • Protein Conformation
  • Transfusion Reaction


  • CD11b Antigen
  • HNA-3a antigen, human
  • ITGAM protein, human
  • Isoantibodies
  • Isoantigens
  • L-Selectin