TRALI: It’s Not Just For Blood Bankers Anymore Norman D. Means, MD, FCAP Blood Bank of Alaska

TRALI What is it?

Transfusion-Related Acute Lung Injury First described by Popovsky, Abel and Moore in 1983 Am Rev Resp Dis 1983;128:185-9 – Acute Pulmonary Edema – Respiratory Distress – Hypoxemia – Hypotension – Fever – In the setting of a recent transfusion

History Case reports as long ago as 1951 – “Noncardiogenic edema” – “Allergic pulmonary edema” – “Hypersensitivity reaction” – “Leukoagglutinin transfusion reaction”

History TRALI is a difficult diagnosis to make, since there are often confounding underlying medical conditions that tend to obscure the diagnosis TRALI is one subset of Acute Lung Injury – Spectrum of disorders

Acute Lung Injury Spectrum of injury Acute Respiratory Distress Syndrome (ARDS) is most severe form Many potential causes, including many causes which are treated by transfusion – Up to 40% of acutely ill, actively bleeding patients will develop ALI Pulmonary edema may develop simultaneously

ALI/ARDS Other possible causes of ALI/ARDS Sepsis Trauma Aspiration Shock others

Acute Lung Injury ARDS Mortality 30 – 40% TRALI Mortality 6 – 12%

Consensus conferences North American-European Consensus Conference on ARDS (1994) NHLBI Working Group (2002) Canadian Consensus Conference (2004)

North American European CC Acute Lung Injury definition: Acute hypoxemia – PaO 2 /FiO 2 <300 mm Hg Pulmonary edema on frontal CXR Pulmonary artery occlusion pressure <18 mm HG or no evidence of left atrial hypertension

NHLBI WG /Canadian CC Defined TRALI Adopted NAECC definition of ALI TRALI if 6 hours since transfusion Risk factors for ALI – Canadian: “possible TRALI” – NHLBI: interpret time course

Why is TRALI suddenly so important?

Transfusion-associated fatalities reported to the FDA Source: Lee, JH Workshop on Bacterial Contamination of Platelets, 9/24/99; adapted from Menitove, JE Complications of Transfusion (p 1617), in Clinical Laboratory Medicine, 2 nd Ed. (2001), McClatchey, KD, ed. Cases% HTR16150 Bacterial Contamination4614 TRALI299 Non-bacterial infections237 Transfusion-associated GVHD186

Transfusion-associated fatalities reported to the FDA Cases 6 Source: Fatalities reported to the FDA following blood collection and transfusion ( ); from Cases% TRALI6451 HTR (non-ABO)2520 Microbial infection1512 HTR(ABO)97 TACO97

Differential Diagnosis of TRALI

TACO Transfusion-Associated Circulatory Overload

TACO The most common transfusion reaction 1-8% of post surgical patients requiring transfusion Age > 60 may develop with only a single unit of pRBC Dyspnea, orthopnea, elevated BP, pedal edema, crackles, tachycardia, infiltrates Elevated BNP

Anaphylactic Reactions Bronchospasm Wheezing Laryngeal edema Urticaria/Erythema Hypotension Rapid onset after start of transfusion IgA deficiency

Transfusion-Associated Sepsis Fever Hypotension Shock Rapid onset Bacteria present Platelets most at risk

Pathophysiology of TRALI Traditional model (antibody transfer) Transfer of biologic response modifiers

Traditional Model Passive transfer of antibody via transfusion HLA Class II, HNA, HLA Class I antibodies Sensitized donors – Multiparous women – Organ and tissue transplant recipients – Previously transfused blood donors

Traditional Model Bray, et al (2004) – 308 randomly selected units of plts, pRBC, FFP, cryoprecipitate – Overall 22% of units had HLA antibodies FFP 29% Cryo 24% Hum Immunol 2004;65:240-4

Traditional Model Why don’t we see more TRALI? – Antigen-antibody pairing – Threshold concentration of antibodies – Preexisting endothelial “priming” may potentiate response

Bioactive Response Modifier Model Accumulation of IL-6, IL-8, PAF, IFN-γ, TNF-α, NO, bioactive lipids Transfusion leads to activation of inflammatory cascade (NO→vasoconstriction) Damage to alveolar capillaries or increased hydrostatic pressure – Pulmonary Edema

Bioactive Response Modifier Model These bioactive agents have been shown to accumulate in stored blood products over time. Levels may rise high enough to “prime” neutrophils or endothelial cells Some evidence that these may rarely trigger TRALI directly.

Pathophysiology of TRALI Probably all of these models have some role to play in the causation of TRALI Other causes may emerge

Pathophysiology of TRALI End result is damage to the pulmonary capillary endothelium Neutrophil-induced damage – Activated neutrophils express HLA Class II and HNA antigens – Activated pulmonary endothelium express HLA Class II antigens

Pathophysiology of TRALI Activated neutrophils – Response to various priming agents PAF, TNF-α, IL-8, GM-CSF, IFN-γ, LPS, infectious agents – Anatomic and physiologic changes Stiffening (actin polymerization) Adhesivity (β 2 integrins, selectins) Clustering of surface receptors (FcγRIIa, β 2 integrins) Release of toxic granule enzymes Formation of NADPH oxidase → Reactive oxygen species

Pathophysiology of TRALI Activated neutrophils – Activation may be triggered by a number of events Infection Cardiovascular disease Leukemia Recent surgery Others (trauma?, hemorrhage?) – Mechanical sequestration and aggregation in pulmonary microvasculature

Pathophysiology of TRALI Activated endothelium – Increased adhesivity (selectins, ICAM-1) – Promotes priming of “captured” neutrophils – Interaction of activated endothelium and neutrophils leads to endothelial damage/TRALI Platelet activation Increased neutrophil aggregation Complement activation – Role unclear

Pathophysiology of TRALI Neutrophil specific antibodies HNA, HLA Class II, and HLA Class I – Some are strong enough to trigger TRALI alone Presence of leukoagglutinins is especially worrisome Leukoagglutinins may stimulate active neutrophil aggregation

Pathophysiology of TRALI Activated neutrophils are sequestered in the microvasculature of the lungs – Production of bioactive products ROS Enzymes Endothelial damage Exudation of fluid and neutrophils into alveoli  Respiratory compromise

Pathophysiology of TRALI Exudative (high protein) fluid in alveoli – TACO has transudative (low protein) protein in alveoli – Damage to the pulmonary capillary endothelium leads to leakage into alveoli – Due to activated neutrophils damaging endothelium

Pathophysiology of TRALI Presence of HLA-antibodies and HNA antibodies in a blood product does not necessarily mean that these antibodies will cause TRALI Cognate antigens in recipient may be necessary

HLA and HNA antibodies HLA antibodies identified in 70-75% of all TRALI cases – 85-90% antibody in transfused product – 10% in recipient – Rare inter-donor cases antibody in transfused product directed against antigen in other transfused product HNA antibodies 10-15% 10-15% ? ?

HLA and HNA antibodies # of cases of severe TRALI % HLA Class I410 HLA Class II1747 HLA Class I and II38 HNA (HNA-3a) 12 (10/12) 33 Total36 Reil A, et al. Vox Sanguinis 2008;95(4):

HLA and HNA antibodies HNA-3a antibodies in 6 of 10 fatal cases in this study Remainder were HLA Class II and HNA-2a

HLA testing All test rely on the AHG test Plasma or serum is reacted to test antigens Wash unbound antibody away Incubate with anti-IgG Wash unbound anti-IgG away Detection

HLA testing Complement Dependent Cytotoxicity – Live cells – Mix with complement – Count living cells (dye exclusion) – Very labor intensive – Least sensitive – Cellular test Not specific for HLA antibodies

HLA testing ELISA – Full range of antigens bound to test wells – Usually one well for Class I and one well for Class II – Less sensitive – Manual testing currently automation under development – Acellular (specific for HLA antibodies) – No indeterminant tests: cutoff value

HLA testing Flow Cytometry – Latex microparticle beads with antigen – Complex test that is labor intensive Indeterminate results possible – Acellular – Can distinguish between class I and class II due to differences in luminescence markers Single run tests for both

HLA testing Luminex – Similar to Flow cytometry Antigen coated microbeads Fluorescent-tagged antibodies – Simpler chemiluminescence test Instrument is smaller Less expertise to operate Both class I and class II can be simultaneously tested – Manual test, but automation looks easiest – No indeterminate tests…cutoff value

HNA testing Expensive ($ ), few labs doing this at the moment Generally tests are labor-intensive and require high level of expertise to perform

HNA testing Granulocyte agglutination testing – Serum incubated for 4-6 hours with fresh neutrophils – Presence of antibodies cause clumping of neutrophils – Least sensitive of tests

HNA testing Granulocyte immunofluorescence (GIF) – Similar to AHG test – Pretreatment with 1% paraformaldehyde to prevent antibody binding to Fc portion of neutrophil receptor – Microscopy: Can have high background fluorescence making it difficult to read weak results – Flow cytometer is most often used now

I got a TRALI case! What do I do now?

Tenth Law of the House of God IF YOU DON’T TAKE A TEMPERATURE, YOU CAN’T FIND A FEVER. -- Samuel Shem (1978)

Investigation of Suspected TRALI Increased numbers of reported TRALI fatalities in past few years is due in large part to increased awareness on part of the folks at the bedside Education of clinical personnel about TRALI recognition is critical. – CCC criteria – NHLBI WG criteria

Investigation of Suspected TRALI Clinician notifies transfusion service of possible TRALI case Initial prompt investigation of possible TRALI is performed by the transfusion service.

Investigation of Suspected TRALI Acute hypoxemia – PaO 2 /FiO 2 <300 mm Hg NEW bilateral pulmonary infiltrates on frontal CXR Pulmonary artery occlusion pressure <18 mm HG or no evidence of left atrial hypertension Occurring within 6 hours of transfusion of blood products No evidence of pre-existing ALI prior to transfusion

Investigation of Suspected TRALI Easiest way to exclude TRALI ?? Onset of symptoms > 6 hours post transfusion

Investigation of Suspected TRALI The results of the initial investigation should be reviewed with the transfusion service medical director. Medical director of the Blood Bank of Alaska is available for telephone consultation with the transfusion service medical director, if needed. – (907) BBAK – (907) (after hours, urgent calls only) Close cooperation between transfusion service and donor center is essential.

Investigation of Suspected TRALI If TRALI is suspected after initial investigation is completed, then transfusion service should begin laboratory testing. If possible, have all implicated blood product units returned to the blood bank. – Even an “empty” bag may retain enough volume to be useful. Culture and gram stain of blood product(s) Blood cultures and gram stain of patient’s blood Obtain and preserve recipient blood specimen for possible future HLA/HNA antigen testing.

Investigation of Suspected TRALI If TRALI is suspected: Complete the TRALI/TTD investigation form and fax immediately to BB of AK for further investigation. – Include all pertinent clinical information and diagnoses, including time course of onset of symptoms – Include radiologic diagnoses of CXR – Include lab studies (ABG, CBC, GS, cultures, chemistries) – List attending physician and contact information – List contact information for transfusion service – Include complete list of all unit numbers transfused in the 12 hours prior to the reported onset of symptoms.

Investigation of Suspected TRALI BB of Alaska will notify Medical Director. Quarantine/recall involved units. Medical Director may contact transfusion service or clinician to obtain additional history or other information. Involved donors will be contacted to return for HLA antibody testing and possible HNA antibody testing. – HNA antibody and antigen testing is very expensive and difficult to obtain, limiting effectiveness

Investigation of Suspected TRALI Several weeks may be required to obtain testing on all donors. Involved donors will be reviewed to see if any donors have been previously “flagged” – “Flagged” means that the donor has been involved in previous TRALI case(s)

Investigation of Suspected TRALI Medical Director of BB of AK will obtain all necessary and available information and testing results and will make a determination: – TRALI – Possible TRALI – Not TRALI Written report of findings Fatalities will be reported to FDA

Investigation of Suspected TRALI Implicated donors will be deferred from future donations Multiply flagged donors will be evaluated for future eligibility

Donor Management TRALI prevention strategies are still a work in progress. Ongoing efforts to identify causes of TRALI Screening tests to reduce risk are not available. – Not yet cost effective to test for HLA/HNA antibodies on all donations

Donor Management AABB: “TRALI mitigation plan required”

Donor Management AABB standard – “Donors implicated in a TRALI event or associated with multiple events of TRALI shall be evaluated regarding their continuing eligibility to donate.” – “Implicated”: A donor who is the source of a blood product transfused into a recipient within 6 hours of symptom onset in a proven TRALI event that contains HLA and/or HNA antibodies against antigen(s) expressed by the recipient.

Donor Management UK National Blood Service 2003: male-only donors for plasma rich products (FFP, platelets) in 95% of FFP – Same day requirement – SHOT (Serious Hazards of Transfusion) surveillance system in UK

Donor Management UK NHS Blood and Transplant Service CE Chapman and LM Williamson Transfus Med Hemother 2008;35:93-96 TRALI cases reported to SHOT ( ) Incidence High plasma FFP/ cryo-poor plasma 31/2.3 million1:74,000 Platelets17/1.5 million1:88,000 Low Plasma Cryoprecipitate1/0.5 million1:500,000 pRBC28/15.6 million1:577,000

Donor Management Many US Blood centers have adopted similar policies Reduction of TRALI by 75%, and fatal TRALI by 90%

Donor Management Alternative is to use solvent-detergent (SD) plasma – Very low risk of TRALI, probably due to dilution effect – Reduction of infectious disease – Production of IVIG and albumin Very costly ($13.5 million/year in Canadian estimate)

Donor Management BB of AK – Male-only FFP donors – HLA antibody testing of established male and female apheresis platelet donors – New apheresis platelet donors male-only

What does the future hold?

SD plasma Clearer elucidation of the pathophysiologic mechanism of TRALI Specific antibody screening – Better technology – automation

Where have I seen this guy before? Words A Poppin’ 1975