Transfusion Reactions and Complement PDF

Transfusion reactions and complement Video learning aims and objectives In these videos I aim to cover: 1) The real life causes and reasons for transfusion reactions 2) the interactions that occur between Antibody and Complement on the erythrocyte surface 3) the role of complement activat...

Transfusion reactions and complement Video learning aims and objectives In these videos I aim to cover: 1) The real life causes and reasons for transfusion reactions 2) the interactions that occur between Antibody and Complement on the erythrocyte surface 3) the role of complement activation in the process of immune haemolysis in vivo and in vitro By the end of these sessions, you should be able to explain: 1) Explain some of the major causes of transfusion reactions and what is used to try and prevent them. 2) describe the mechanisms by which antibody induced complement activation aids the immune haemolyis of red cells 3) discuss the relevance of antibody associated complement activation in vivo and in vitro Contents Preventing transfusion reactions Transfusion reactions; reasons for and investigations of Action of complement in transfusion reactions Complement cascade Complement activity in transfusion reactions Preventing transfusion reactions. Learning objectives The theory of audit trails and how blood products are linked between donors and recipients. Understand how product labelling is used to link donations to donors and donations, donors and patients to reactions. Blood pack information Taken from: https://www.bartshealth.nhs.u k/news/landmark-uk-study-lau nches-to-save-hundreds-more -lives-4592 08/11/2023 Mandatory and discretionary matching criteria Taken from: https://www.bartshealth.nhs.u k/news/landmark-uk-study-lau nches-to-save-hundreds-more -lives-4592 08/11/2023 Patient Identification Taken from: https://www.impressionsi d.com/vinyl-patient-id-ba nds/standard-colours/143 / 10/12/2023 Blood tracking Taken from: https://www.ehealthireland.ie/case- studies/electronic-blood-tracking/ 16/09/2024 Summary Methods in which blood products are traced from donor to recipient How this prevents Transfusion reactions; reasons for and investigations of. Learning objectives 1) Which blood group antigens are associated with haemolytic transfusion reactions Types of reaction Haemolytic Immediate: ABO mediated; IgM, MAC and complement activated. ABO antibodies, donor or recipient derived. Delayed: Protein or glycoprotein blood group antigens. IgG complement progression to C3b. Non-Haemolytic Febrile reactions; Increase in temperature but no symptoms of haemolysis Antibody reactions; Acute inflammatory response  Clinically significant antigens and statistics Frequency Frequency of Frequency of Among All Antigen System Antigen Antigen Potency* Detected (Whites) (Blacks) Alloantibodies E Rh 16–40% 30% 22% 4% K Kell 5–40% 9% 2% 9% D Rh 8–33% 85% 92% 70% c Rh 4–15% 80% 96% 4% Jk(a) Kidd 2–13% 77% 92% 0.14% Fy(a) Duffy 4–12% 66% 10% 0.46% C Rh 2–10% 68% 27% 0.22% e Rh 2–3% 98% 98% 1% Jk(b) Kidd 2% 74% 49% 0.06% S MNSs 1–2% 55% 31% 0.08% s MNSs < 1% 89% 94% 0.06% Taken from: https://emedicine.medscape.com/article/134958-overview?form=fpf#a6 on 22/11/23  Clinically significant antigens and reactions Frequency Among Frequency of Frequency of Antigen System All Detected Antigen Antigen Potency* Alloantibodies (Whites) (Blacks) E Rh 16–40% 30% 22% 4% K Kell 5–40% 9% 2% 9% D Rh 8–33% 85% 92% 70% c Rh 4–15% 80% 96% 4% Jk(a) Kidd 2–13% 77% 92% 0.14% Fy(a) Duffy 4–12% 66% 10% 0.46% C Rh 2–10% 68% 27% 0.22% e Rh 2–3% 98% 98% 1% Jk(b) Kidd 2% 74% 49% 0.06% S MNSs 1–2% 55% 31% 0.08% s MNSs < 1% 89% 94% 0.06% Taken from: https://emedicine.medscape.com/article/134958-overview?form=fpf#a6 on 22/11/23 Investigations Patient Antibody panel DAT, presence of antibody and complement Phenotype of antigens related to recently created antibodies (Landsteiners’s law) Donor / Donation Donation history / history of previous reactions Product recall Phenotyping for reaction specific antigen Non-specific testing for reasons for reaction Summary How antigens determine the class of antibody involved in the reaction. How antibody classes determine complement activity and patient symptoms. Investigations of both patients and donors / donations. Action of complement in transfusion reactions Learning objectives 1) The role of complement in immune reactions. 2) How complement works under normal immunological conditions and how complement is employed in haemolytic transfusion reactions 3) How classical and alternate complement pathways are activated and amplified Classical complement Activation of complement is through classical (antibody dependent) pathway. Multiple antibodies Close proximity IgM versus IgG stimulating complement No of antibodies per complex No. of complementary epitopes Dispersal of epitopes / antigens Red Cell Antibodies and Immune Haemolysis red cell antibodies can mediate Immune Haemolysis part of normal Immune Response Antibody Mediated Red Cell Destruction action of phagocytic cells carry FcyR receptors on membrane macrophages carrying FcyR bind to cells coated with IgG antibody results in extravascular haemolyis activation of Complement Complement and Immune Haemolysis vital role in immune defence destruction of pathogens; generation of inflammatory mediators three pathways composed of triggered enzyme cascades with amplification inherent regulatory mechanisms maintain localised response at least 29 different proteins half are involved in regulation two pathways mediate Immune Haemolysis Classical mediated by Red Cell Antibody bound to Antigen Alternative independent of Antibody The Classical Pathway enzyme cascade with amplification nine main glycoproteins : C1 - C9 many are precursor enzymes initiated by Ag/Ab reaction on a cell surface causes destruction of antibody coated red cells by Haemolysis cascade completion results in ‘doughnut-like’ lesions in red cell membrane Haemolysis is Intravascular Immune Adherence (Opsonization) sequence stops at intermediate (C3b) stage red cells destroyed by phagocytic cells carrying complement and IgG receptors Haemolysis is Extravascular Alternate complement pathway Starts at C3b and can either amplify or lead to MAC Binding either separate from antibodies and hydrolytic, or Amplifying based on pre-bound C3b Factor B, important part of alternate pathway C5 convertase Factor D, Factor B convertase Factors H and I inactivate alternate complement DAF, dissociates C3 convertase ccomponents Properdin stabilises C3bBb Alternate complement pathway acts as a complex positive feedback loop any C3b produced can enter the amplification loop large amounts of C3b are now generated Summary The activation of complement through the alternate and classical pathways. Where the reaction ends determines the type of transfusion reaction e.g. intravascular or extravascular What different components of the complement cascade actually do. Complement cascade Learning objectives 1) Individual stages of the complement cascade and their action. 2) The role of serine proteases in the complement cascade. 3) Regulation and stabilisation of specific areas of complement to prevent non-specific actions of complement and to continue directed actions of complement. Classical Complement Pathway - Stage 1. Attachment/Complement Binding Stage Aim: binding of C1 complex C1 is composed of a C1q:C1r2:C1s2 complex Ca2+ dependent requires formation of Ab/Ag complex two Fc regions must lie in close proximity C1q can then bind to the adjacent Fc sections via receptors on globular head regions results in a conformational change C1r becomes an active enzyme Classical Complement Pathway - Stage 2: Activation Stage activated C1r then cleaves and activates C1s C1s enzyme then acts on the next two components C4 is first cleaved into the C4b (largest fragment) and C4a C4b binds covalently to the cell surface C2 then binds to the C4b C2 is then cleaved by C1s to produce C2b the resulting C4b2a complex is the C3 convertase of the Classical Pathway Classical Complement Pathway - Stage 3: Amplification Phase C4b2a cleaves C3 into C3a and C3b there is major amplification as the large fragment C3b binds covalently to the local cell surface unbound C3b is quickly inactivated in the plasma phase bound C3b has immune adherence properties the smaller C3a fragment is released into the fluid phase where it acts as an anaphylatoxin a small amount of C3b binds to form a C4b2aC3b complex this is C5 convertase Classical Complement Pathway - Stage 4: Membrane Attack Phase (1) C5 binds to the C3b component of the C5 convertase the serine protease activity of C2a then cleaves C5 into C5b and C5a the smaller C5a fragment is released into the fluid phase this has both anaphlyatoxic and chemotactic properties C5b then binds C6 and C7 Classical Complement Pathway - Membrane Attack Phase (2) the trimolecular complex of C5b67 then binds to the cell membrane C7 inserts into the lipid bilayer C8 and then 10-16 molecules of C9 then bind to create a membrane lesion this is the Membrane Attack Complex (MAC) pore is hydrophobic externally but hydrophillic internally allows influx of solutes and water across lipid bilayer result is cell lysis Complement activation by IgM >IgG3 > IgG1 > IgG2 : IgG 4 does not bind Membrane Attack Complex Regulation of Classical Pathway regulation occurs in Activation, Amplification and Membrane Attack phases of complement activation by inhibitors that act directly on activated complement components by rapid disassociation of newly formed complement complexes by transient binding sites of activated components Activation Phase action of C1 inhibitor; short 1/2 life of C2a; transient binding site of C4b Amplification Stage action of Factor I; short 1/2 life of C3b; cell surface inhibitors eg. Decay Accelerating Factor (DAF) prevents assembly and promotes dissociation of the C3 convertase components Membrane Attack Complex action of plasma and membrane bound inhibitors The Alternative Complement Pathway evolved as part of the innate immune system not dependant on antibody for activation activated by ‘biological factors’ feeds into the common complement pathway at the C3 convertase stage C3bBb is the C3 convertase of the alternative pathway C3BbC3b is the C5 convertase of the alternative pathway different plasma proteins involved (Factor B, D & P) Summary Where the different proteins in complement align in different portions of the complement response. The role of proteins such as C3b and C5-9 in a complement response. How complement is regulated to prevent non-specific reactions and enhance targeted reactions. Complement activity in transfusion reactions Learning Objectives 1) How specific elements of complement cause specific physiological effects. 2) What the physiological effects of complement mediated haemolysis is and how they present differently. Biological effects of complement activation generation of inflammatory mediators C3a, C5a (& C4a) are anaphylatoxins induce smooth muscle contraction cause release of cytokines from macrophages bind to basophils and mast cells cause release of histamine which acts as a vasodilator attract phagocytes to site C5a acts as chemotactic factor accumulation of neutrophils at site of infection results in patient’s symptoms of Inflammatory response - seen in a Haemolytic Transfusion Reaction Types of immune mediated red cell haemolysis antibody mediated intravascular haemolysis potent complement fixing alloantibodies already present in the patient’s circulation formation of MAC complex cause rupture and breakdown of the red cells in the bloodstream mostly caused by IgM antibodies that are still active at or near 37 oC Complement is optimally active at 31-33°C antibody mediated extravascular haemolysis involves the removal and destruction of antibody coated red cells by macrophages outside the vascular circulation mostly caused by clinically significant IgG antibodies complement activation enhances effect cells coated with C3b and large amounts of IgG are destroyed by phagocytic macrophages with FcR and C3b receptors in the liver cells coated with only small amounts of IgG are removed by phagocytic cells with FcR receptors in the spleen Role of Complement in Transfusion Science in vivo Haemolytic Transfusion Reactions enhances antibody mediated removal of transfused ‘foreign’ red cells significant effect on morbidity and mortality Autoimmune Haemolytic Anaemias (AIHA) enhances antibody mediated removal of ‘self’ red cells causes haemolytic episodes in ‘cold’ type AIHA amplifies anaemic state in ‘warm’ type AIHA causes some adverse effects of transfusion due to complement activation by white cell antibodies Role of Complement in Transfusion Science in vitro aids in the detection of clinically significant complement binding red cell antibodies (serum samples only) causes haemolysis in a test: a positive reaction ‘sensitises’ red cell surfaces with complement components Note: anticoagulants are anti-complementary Summary The role of complement in immune reactions and immune mediated haemolysis. How types of haemolysis from transfusion reactions are predictable based upon the antigen responsible.