The role of neutrophils in the pathogenesis of transfusion-related acute lung injury

Abstract

Transfusion-related acute lung injury (TRALI) is the major cause of transfusion related morbidity and mortality, world wide. Efforts to reduce or eliminate this serious complication of blood transfusion are hampered by an incomplete understanding of its pathogenesis. Currently, TRALI is thought to be mediated by donor alloantibodies directed against host leukocytes or the result of 2 distinct clinical events. For both proposed mechanisms, the neutrophil is the key effector cell. This article reviews TRALI pathophysiology, explores the role of the neutrophil, details practical information for appropriate diagnosis and promotes further studies into the pathogenesis of TRALI.

Fig. 1. Distribution of neutrophils

In a healthy normal adult, 36% of all PMNs reside in the circulating pool and 64% are in the non-circulating ‘reserve’ pool (1,2). Of the total number of PMNs 28% are in the pulmonary pool, and this consists of both circulating and non-circulating PMNs. Therefore, a substantial proportion of PMNs are normally located in the pulmonary system.

Fig. 2. PMNs deform as they pass through the pulmonary capillaries

PMNs are 6 – 8 µm in diameter, while the lumen of pulmonary capillaries range from 2 –15 µm (6). Therefore, PMNs are deformed and get squashed into an elliptical shape as they transit the pulmonary capillaries (9,10). This slows down the PMNs and allows them more time to “sense” the surrounding environment.

Fig. 3. Functional PMN phenotypes: quiescent, primed and activated

Activated Complement 5 (C5a); N-formyl-Met-Leu-Phe (fMLP); Granulocyte Monocyte Colony Stimulating Factor (GM-CSF); Interleukin (IL); Lipopolysaccharide (LPS); lysophosphatidylcholines (lyso-PC); Platelet Activating Factor (PAF); Reactive Oxygen Species (ROS); Tissue Necrotising Factor- alpha (TNFα) There are three distinct PMN phenotypes; quiescent, primed and activated (13). Quiescent PMNs become primed when they are exposed to priming agents such as PAF, TNFα, GM-CSF, IL-8, LPS or bacterial products. Primed PMNs become activated when stimulated with activating agents such fMLP or LTB₄, but they can also activated by other priming agents such as HNA-3a or MHC class I antibodies (56, 101). This culminates in the respiratory burst. Priming is reversible as PMNs can deprime if subsequent activation does not occur.

Fig. 4. Production of O₂ ⁻ and H₂O₂ following NADPH oxidase activation

Activated Complement 5 (C5a); N-formyl-Met-Leu-Phe (fMLP); Tissue Necrotising Factor- alpha (TNFα)

1. Activation of NADPH oxidase catalyses the reduction of oxygen (O₂) to superoxide (O₂ ⁻). This is a rapid reaction that occurs within 30 to 60 seconds of activation

2. O² ₋ alone is not effective in bacterial killing but is a good reducing agent. Dismutation of O₂ ⁻ produces oxygen and hydrogen peroxide (H₂O₂). H₂O₂ is cytotoxic and this ability is greatly enhanced by myeloperoxidase that is released from the azurophilic granules (14,24).

3. Myeloperoxidase uses the Cl⁻, Br⁻ or I⁻ as reducing substrates to produce hypohalous acid. The oxidation of Cl⁻ produces hypocholorous acid (HOCl).HOCL is a very potent cytotoxic agent for bacteria, virusus, fungi and mycoplasmas.

Fig. 5. Antibody mediated TRALI

Antibodies (Abs) in the transfused donation interacts with its cognate antigen (Ag) on recipient’s PMN, causing aggregation in the pulmonary microvasculature and activation of PMNs (27,28). Activated PMNs produce and release reactive oxygen species (ROS) and enzymes, which in the absence of a microbicidal target instead cause damage to the pulmonary endothelium. The damage to the pulmonary endothelium allows the leakage of fluids into the pulmonary space resulting in the clinical symptoms of the respiratory distress associated with TRALI.

Fig. 6. The Two Event Model

The patient’s clinical condition causes activation of the pulmonary endothelium, resulting in the adherence and sequestration of primed PMNs in the pulmonary capillary beds (49,50). Antibodies or biological response modifers (BRMs) in the transfused blood product activates the primed PMNs. Activation of primed PMNs produces an augmented release of the cytotoxic microbicidal arsenal resulting in damage to the pulmonary endothelium. Damage to the pulmonary endothelium allows the leakage of fluids into the pulmonary space that produces the respiratory distress associated with TRALI.

Fig. 7. Laboratory investigation of TRALI

Antigen (Ag); Antibody (Ab); Granulocyte Agglutination Test (GAT); Granulocyte Immunofluorescence Test (GIFT); Lymphocytotoxicity Test (LCT); Lymphocyte Immunofluorescence Test (LIFT) The first aim of the laboratory investigation of TRALI is to detect any leukocyte antibodies (Abs) (HNA, HLA class I and HLA class II) in either the patient (transfusion recipient) or associated donors/donations using well validated assays (27,57,86) . The second aim is to seek corroborative evidence that the detected Ab can react with an available corresponding antigen on the target cell. *In the majority of cases the Ab is of donor/donation origin and this is investigated either by confirming that the patient has a corresponding antigen or by a cross-match of donor sample against the recipient’s PMN. In the rare instance where the Ab is from the patient, a cognate antigen in the donors/donation is sought and the cross-match is between the patient serum/plasma against donor PMNs. If resources are available further investigation includes determining if the Abs detected are able to prime or activate for PMN respiratory burst using assays such as the superoxide dismutase-inhibitable reduction of cytochrome C assay.