The immune system

Chapter 2


The immune system




Chapter contents



EQUINE IMMUNOLOGY



HYPERSENSITIVITY REACTIONS



NEONATAL ISOERYTHROLYSIS


NEONATAL ALLOIMMUNE THROMBOCYTOPENIA


EQUINE PERINATAL IMMUNOLOGY



IMMUNODEFICIENCIES



IMMUNOMODULATORS



LEUKOPROLIFERATIVE DISEASES




EQUINE IMMUNOLOGY



INTRODUCTION


Advances in immunology and technology have contributed significantly to the understanding of fundamental concepts of disease, to the identification of preventive methods and to the evaluation of treatment responses. Nevertheless, many areas in the field of equine immunology await reagents and technical capabilities to expand our knowledge of the mechanisms involved in many infectious and non-infectious processes.


The development and characterization of monoclonal antibodies for equine molecules and the decoding of the horse genome are dependent on a cooperative effort among investigators concerned about the well being of the horse. When available, however, traditional and state of the art techniques can be used to reveal the role of immune cells in the pathogenesis and regulation of diseases. This chapter reviews the major conditions of the horse that directly involve the immune system and indicates the available diagnostic resources for the identification and monitoring of these processes.


A glossary of principal immunologic terminology is given in Box 2.1.



Box 2.1   A glossary of immunologic terms




Adjuvant: a substance that enhances the immune response to antigens.


Allele: any one of a number of alternative forms of the same gene occupying a given locus (position) on a chromosome.


Allergen: an antigen that induces IgE-mediated hypersensitivity reactions.


Alloantibody: antibody produced against a protein of an allele from another member of the same species and which can function as an antigen.


Anaphylatoxin: activated complement components (C3a, C5a) that induce mast cell degranulation.


Antibody: protein with specific binding properties produced by plasma cells in response to antigenic exposure.


Antigen: foreign molecule that can induce an immune response.


Antigen-presenting cell (APC): specialized type of cell (dendritic cells, macrophages) that can process and present antigens to lymphocytes.


Antigen processing: breakdown of molecules into peptides that can be presented to lymphocytes via major histocompatibility molecules.


Antigen receptor: the specific antigen-binding receptor on T (T cell receptor) or B (B cell receptor) lymphocytes.


B lymphocyte (B cell): lymphocytes that develop in the bone marrow, finish their maturation and differentiation in the periphery, and become antibody-secreting cells.


Blocking antibody: antibody molecule capable of blocking the interaction of antigen with other molecules.


Cell-mediated immunity: immune response mediated by T cells, often CD8+ T cell cytotoxic response.


Chemotaxis: directional migration of cells toward a concentration gradient of an attractant (chemotactic substance).


Classical pathway: the mechanism of complement activation initiated by antigen—antibody aggregates that induces the activation of C1, C4 and C2 components.


Cluster of differentiation (CD): cell surface molecules (markers) of leukocytes that can be identified by monoclonal antibodies to differentiate cell populations.


Complement: group of serum proteins involved in inflammation, phagocyte activity and lysis of cell membranes; the complement cascade is triggered classically by the interaction of antibody with specific antigen.


Complement receptor: a structure that binds C3 fragments found on phagocytes (neutrophils, monocytes and macrophages) and lymphocytes (B cells).


Coombs test: a test used to detect antibodies or complement on red blood cells using anti-immunoglobulin antibodies.


Cytokines: soluble substances secreted by cells, which have a variety of effects on other cells.


Cytotoxic T cell: lymphocyte (CD8+ T cell) that kills infected cells expressing processed antigen peptides via major histocompatibility class I.


Dendritic cells: cells present in tissues that capture and process antigens, migrate to regional lymph nodes and present processed antigen peptides to T lymphocytes.


Enzyme-linked immunosorbent assay (ELISA): assay that measures quantitatively the amount of a substance using enzyme-conjugated antibodies and a substrate that produces a colored end-product upon binding of the enzyme.


Epitope: part of the antigen that contacts the antibody or T cell receptor.


Fc receptor: receptor on a cell surface with specific binding affinity for the Fc portion of an antibody molecule.


Flow cytometry: analysis of cell populations in solutions according to their size, granularity and fluorescence of cell markers detected by fluorescent monoclonal antibodies.


Fluorescent antibody: antibody conjugated with fluorescent dye often used in flow cytometric analysis and immunofluorescent assays.


Germinal centers: distinct areas in secondary lymphoid tissues (spleen, lymph node) in which B cells aggregate and go through differentiation and antibody class switching.


Helper T (Th) cells: class of T cells (CD4+ T cells) that help B cells to produce antibody and generate cytotoxic CD8 T cells; helper T cells recognize processed antigen peptide presented via major histocompatibility class II.


Humoral response: immune response mediated by antigen-specific antibodies present in serum and tissues.


Hypersensitivity: state of reactivity to an antigen that is far greater than the antigenic challenge presented.


Idiotype: the combined antigenic determinants (idiotopes) found in the variable region of antibodies.


Immune complex: antigen bound to antibody that may or may not contain complement components.


Immunoglobulins: proteins (antibodies) produced by plasma cells. There are five main types of antibodies: IgA, IgD, IgE, IgG and IgM.


Immunomodulators: substances that enhance or decrease non-specifically the immune response.


Immunosuppression: a mechanism (treatment) for producing a specific state of immunologic unresponsiveness.


Interferons: proteins with antiviral activity, which are involved in signaling between cells.


Interleukins: glycoproteins secreted by a variety of leukocytes, which are involved in signaling between cells.


Isotypes: classes of antibody that differ in the constant region of their heavy chain (Fc portion).


Isotype switching: the shift of a B cell from the secretion of antibody of one isotype (e.g. IgM) to a different isotype (different heavy-chain, e.g. IgG).


Leukotrienes: metabolites of arachidonic acid involved in inflammation.


Lymphokine-activated killer cells: cytotoxic cells generated ex vivo by stimulation with IL-2.


Major histocompatibility complex (MHC): cluster of genes encoding cell surface molecules that are polymorphic (multiple alleles at a particular genetic locus); this polymorphism codes for antigenic differences that can lead to graft rejection between members of a single species; the molecule is also involved in processed antigen peptide presentation to CD8+ T cells (MHC class I) and CD4+ T cells (MHC class II).


Mitogen: substance that stimulates the proliferation of many different clones of lymphocytes (e.g. phytohemagglutinin, PHA; concanavalin A, ConA; pokeweed mitogen, PWM; lipopolysaccharide, LPS).


Monoclonal antibody: antibody that is produced from a single A clone is the progeny of a single cell. In immunology, monoclonal generally describes a preparation of antibody that is monogenous, or cells of a single specificity.


Opsonin: antibody or complement component that coats a foreign particle (opsonization) to enhance phagocytosis by phagocytic cells.


Oxidative (respiratory) burst: increased oxidative metabolism within phagocytic cells stimulated by the phagocytosis of organisms; the reactive oxidative products that originate from this metabolism have killing properties.


Passive immunity: preformed antibody solution (e.g. colostrum, plasma) transferred to an individual.


Phagocytosis: the engulfment of a particle or a microorganism by phagocytes (neutrophils, macrophages); phagocytosis is facilitated by the binding of immunoglobulin- or complement-coated particles to the Fc receptors on the cell surface.


Primary immune response: cellular and humoral response to the first encounter with antigen; in general, this response has a long induction phase.


Protein A: component of the cell wall of certain staphylococci that binds the Fc portion of most IgG antibodies.


Reticuloendothelial system: network of phagocytes present in the liver and spleen, which remove abnormal circulating cells (e.g. antibody-bound erythrocytes).


Secondary response: cellular and humoral immune response that follows a subsequent encounter with a known antigen; often this response is faster and more specific than the primary response.


T cell: lymphocyte originating in the bone marrow that goes through differentiation and selection in the thymus.


Toxoid: nontoxic derivative of a toxin used as an antigen for the induction of antibodies.



VACCINATION


Vaccination is an important component of disease prevention programs and it is a relevant example of modulation of the immune system. The development of an effective vaccine requires detailed knowledge of the organism, the host’s natural immune response to it, and risk factors of disease. Therefore, microbiologists, immunologists and epidemiologists work together in the design of vaccination programs. Great effort is dedicated to discovering vaccine elements that stimulate, similar to natural infection, a long-lived protective immunity with humoral and cellular components at the systemic and mucosal levels.


Serious side effects to vaccines, including anaphylactic or anaphylactoid reactions, swellings and abscesses at the injection site, laminitis and death, are rare. Fever and malaise are however expected, and they are indicators of response to the vaccine. In general, the small risk of vaccine side effects is greatly outweighed by the benefits.


All vaccine products must go through safety tests before commercialization. The same requirement is not always true for tests that evaluate efficacy. Rarely, there is scientific evidence of efficacy from experimental challenge trials, and the estimation of protection is often taken from clinical and epidemiologic observations. In addition, almost no data are available on the efficacy of multivalent vaccines. Although immunization with different antigens is practical, differences in the risk of exposure (e.g. time of the year, age and group of risk), the duration of immune protection to each antigen, and the quality of the desired immune response may require dissimilar frequencies of vaccine administration.



Types of vaccines


There are two main categories of vaccines:



1. Dead or inactivated vaccines: these vaccines contain the whole inactive pathogen or selective antigenic elements of the pathogen; they induce mainly humoral immunity, however DNA vaccines may induce both humoral and cell-mediated immunity.



2. Live vaccines: the attenuated organism is able to invade a cell and use its replication machinery. Therefore, the organism is processed by the endogenous pathway for MHC class I presentation to generate cytotoxic T cells (CTLs), and MHC class II presentation to generate long-lasting humoral responses.



(a) Modified live vaccines (MLV) are modified organisms that have attenuated pathogenicity but still replicate in the vaccinated animal; the organism may be attenuated in cell culture by growing it in abnormal conditions, by the creation of temperature-sensitive mutants (random mutants that may reacquire virulence), by using recombinant DNA technology for predicted deletion mutation that cannot be reversed, or by using variant forms of the organism that affect other species.


(b) Recombinant vector vaccines (RVV) are engineered bacteria or viruses (vaccinia virus) that become carriers (vectors) of selected antigenic peptides from other pathogens. The bacteria or virus infect cells in the host and carry a target peptide that is known to induce a protective response against the organism. A variant of this type of vaccine is a multivalent vaccine in which one modified-live virus of interest carries another virus peptide.



Vaccination programs


A vaccination program should be tailored according to the degree of exposure to pathogen, number of animals in the herd, age groups involved, type of activity, horse transit, degree of stress, geographic location, management and hygiene and ventilation of the facilities housing the horses.


Primary and secondary vaccinations should be timed to precede the period of greatest exposure to the organisms. In addition, all the animals of the herd should be vaccinated at the same time to minimize replication of the infectious organisms and increase the protection of those animals that respond poorly to the vaccine.


It is possible that maternally derived antibodies may interfere with antigenic response and endogenous antibody production in foals. Ideally, vaccination of the foals should be performed after antigen-specific serum maternal antibody levels decline. In general, colostral IgG half-life is 28–35 days; however, specific antibodies may be transferred to the foal in different concentrations, and their catabolism may depend on environmental challenge.


The protocol for the immunization of foals has been revised because of a better understanding of the foal’s immune response to distinct vaccines in early life. Nevertheless, vaccination of foals is initiated around 3–6 mo of age, followed by one to two boosters 3–4 wk apart. In addition, vaccination of mares in late gestation against agents that are responsible for disease in early life is recommended in order to confer passive transfer of organism-specific immunoglobulins, e.g. rotavirus (q.v.) and Clostridium botulinum (q.v.).



HYPERSENSITIVITY REACTIONS


Hypersensitivity reactions are immune responses to previously encountered antigens with subsequent detrimental effect to the host. The classification of hypersensitivity reactions is based on the type of cells and the immune mediators that promote tissue injury:



TYPES OF REACTION




1. Type I hypersensitivity reactions are mediated by antigen-specific IgE, mast cells, basophils and their mediators. Examples are urticaria (q.v.), insect-bite hypersensitivity (q.v.) and food allergy.


2. Type II hypersensitivity reactions involve autoantibodies IgM or IgG against specific (often self) cell surface or extracellular matrix antigens. Subsequently, there is opsonization and phagocytosis of these cells and complement- and/or Fc receptor-mediated cell destruction. Examples include autoimmune hemolytic anemia (q.v.), thrombocytopenia (q.v.), incompatible blood transfusions, pemphigus foliaceus (q.v.) and drug hypersensitivity (q.v.).


3. Type III hypersensitivity reactions are promoted by the random deposition of immunocomplexes of circulating antigens (self or foreign) and IgM or IgG antibodies in blood vessels, with subsequent complement- and Fc receptor-mediated recruitment and activation of leukocytes and vasculitis. Examples include serum sickness (antisera passive immunization) (q.v.), glomerulonephritis (q.v.) and purpura hemorrhagica (q.v.).


4. Type IV hypersensitivity reactions do not involve antibodies and are mediated by sensitized CD4+ T cells (Th1 response) and CD8+ T cells (direct cytotoxic effect), which induce infiltration of macrophages and inflammation mediated by cytokines. An example is contact dermatitis (q.v.).


Homeostasis is widely dependent on a protective and balanced response of the immune system, which involves complex and redundant regulatory mechanisms. Occasionally, the immune system responds to a non-self structure or a danger signal with extreme properties that may lead to disease. In these conditions, selective activation or suppression of specific responses without affecting peripheral immune function becomes necessary.



ANAPHYLAXIS AND ANAPHYLACTOID REACTIONS


Anaphylaxis is a severe form of immediate hypersensitivity that manifests within minutes following exposure to an allergen (e.g. insect bite/sting, drugs or vaccines, food). The hypersensitivity reaction involves a primary exposure to an antigen, CD4+ T cell cytokine-mediated activation of B cells for the production of IgE, and the binding of IgE to receptors on mast cells and basophils. Upon a successive exposure to the sensitizing antigen, its cross-linking with IgE triggers the release of mediators in cytoplasmic granules. These mediators (histamine, tryptase, leukotriene [LT] C4, prostaglandin [PG] D2, tissue necrosis factor [TNF]) cause early effects of vascular permeability and dilatation, and late effects of inflammation that last approximately 24 h.


The more rapid the onset of clinical signs, the more severe is the event. The release of allergic mediators may lead to mild effects including nasal swelling, urticaria, erythema and pruritus. However, anaphylactic responses may be moderate to severe and lead to vasculogenic shock characterized by hypotension (vasodilation, increased vascular permeability, reduced venous return, cardiac arrhythmias, myocardial ischemia), difficulty in breathing (increased bronchial smooth muscle tone, increased mucosal secretion, laryngeal edema, pulmonary edema), and death. In addition, there may be gastrointestinal clinical signs including colic and diarrhea.


Anaphylactoid reactions are anaphylactic-like reactions that do not involve IgE. The degranulation of mast cells and basophils is directly induced by the agent (e.g. drug) and activation of complement cascade components C3a and C5a. Therefore, this type of reaction may happen at the first time of exposure to the molecule. Nevertheless, the vasogenic effects, clinical signs and treatment do not differ from the anaphylactic reaction.


Confirmation of anaphylactic and anaphylactoid reactions may be pursued by measuring serum or plasma tryptase (peak concentration in 1 h of exposure) or methyl histamine in the urine (collect the second urine sample after exposure to antigen).


Anaphylaxis is an emergency condition that requires immediate diagnosis and treatment. Epinephrine (adrenaline) (0.01 mL/kg BW of a 1 mg/mL solution, IM) should be given by injection without delay. Epinephrine has positive inotropic and chronotropic effects, increases cardiac output, and promotes peripheral vasoconstriction and bronchodilation. IV fluid therapy may assist in fighting hypotension but should be used cautiously in the event of pulmonary edema; hence, colloids should be considered. Airway patency should be assessed and endotracheal intubation or tracheotomy considered in cases of severe angioedema.


After lifesaving measures and epinephrine administration, antihistamines (both H1 and H2 blockers) and corticosteroids (methylprednisolone 20–30 mg/kg BW IV slowly) may be given to further reduce clinical signs. Bronchodilators (β2 adrenergics such as albuterol, clenbuterol, salmeterol) (q.v.) and oxygen therapy may also be used in severe cases.


Anaphylactic reactions can be uniphasic, biphasic or protracted. In the uniphasic reaction, clinical signs resolve within hours. In the biphasic reaction, there is a recurrence of the anaphylactic signs any time between 1 and 30 h after the initial remission. In some cases, the severity of the biphasic reaction is comparable to and often involves the same clinical signs and body systems as the initial reaction. In human patients, delay in the administration of epinephrine is associated with an increase in biphasic reactions and anaphylactic death. It is not clear in human medicine whether steroid therapy prevents biphasic reactions although its anti-inflammatory properties could counter the delayed inflammatory response of anaphylaxis.


In the protracted reaction, profound hypotension may last beyond 24 h despite treatment. It is important to remember that, although rarely used in equine medicine, beta-blockers (e.g. propranolol) and angiotensin converting enzyme (ACE) inhibitors (e.g. enalapril) may increase the severity of anaphylaxis by decreasing blood pressure, promoting bronchoconstriction, and antagonizing the response to epinephrine. Therefore, in patients with refractory reactions, the dose of epinephrine may have to be repeated IM q 10–20 min, or epinephrine may be given IV (0.01 mL/kg BW of a 0. 1 mg/mL solution). Other beta-adrenergic drugs (e.g. dopamine 10 μg/kg BW/min IV, isoproterenol 0.05–0.2 μg/kg BW/min IV) may also be used in non-responsive cases.



ALLERGY OR ATOPY


Allergy or atopy is an immediate hypersensitivity response of skin and mucosal membranes mediated by IgE, mast cells and basophils in genetically susceptible individuals.


A primary exposure to the allergen is required, in which CD4+ T helper cells and Th2 cytokines induce B cell class switching and the production of high concentrations of allergen-specific IgE. Subsequently, the IgE binds to high-affinity Fc receptors specific for the image heavy chain on mast cells present in tissues or circulating basophils. Upon successive exposure, the allergen binds to the cell-associated IgE, and the cross-linking with other IgE molecules and Fc receptors triggers the release of mediators present in the cytoplasmic granules.


In addition to IgE-mediated allergy, the release of cytoplasmic granules by mast cells and basophils can be mediated by complement components C3a, C4a or C5a, which are known as anaphylatoxins. Elements that may trigger this hypersensitivity response include exercise, opiates, cyclo-oxygenase inhibitors and IV contrast materials.


Hypersensitivity diseases in horses may result from exposure to allergens via ingestion (food allergy), inhalation (heaves), injection (drug, insect bite) or percutaneous absorption (contact dermatitis). A familial or hereditary component seems to be involved, and it may be associated with major histocompatibility class I haplotype.


In urticaria (q.v.), cutaneous wheals of various sizes and shapes, pitting edema and pruritus develop within minutes to hours after exposure to the antigen. Urticaria may not involve an IgE-mediated hypersensitivity response, and angioedema secondary to systemic diseases, physical (dermatographism in areas of pressure), temperature-sensitive, cholinergic, and exercise-induced urticarias are possible. A complete history, physical examination and dermatologic testing may help identify the primary cause of the disease. Histopathologic findings in urticaria may include vascular dilation in the superficial and middle dermis and infiltration of inflammatory cells (mononuclear cells, neutrophils, mast cells and eosinophils) in the perivascular regions.


Sweet itch or insect-bite hypersensitivity (q.v.) is a seasonally recurrent hypersensitivity skin reaction to antigens in the saliva of Culicoides species characterized by widely distributed pruritic crusting dermatitis. Following antigen exposure, eosinophils are recruited to the affected skin in response to histamine, platelet-activating factor (PAF) and eotaxin. Eosinophils may contribute to the disease with the release of proteases and inflammatory mediators.


To date, clinical studies have indicated that allergic tests still lack levels of sensitivity and specificity when horses affected by sweet itch are compared to normal horses. Therefore, allergic testing alone should not be used to diagnose this hypersensitivity reaction, and results should be evaluated in combination with the history and clinical manifestation of the disease. Serologic testing (ELISA for IgE or radioactive [RAST] or fluorescent allergosorbent [FAST] tests) may be used to aid in the design of a treatment plan. A positive reaction indicates the presence of antigen-specific IgE but alone it does not indicate whether that specific antigen is the cause of the disease. Intradermal allergy testing is probably more clinically significant than serology because it tests not only the presence of antigen-specific IgE but also the level of sensitization upon exposure to the allergen.


Acute idiopathic hypersensitivity reactions may be responsive to antihistamines (e.g. hydroxyzine hydrochloride 0.5–1 mg/kg BW t.i.d. PO) and corticosteroids (dexamethasone 0.05–0.1 mg/kg BW s.i.d. IV or PO; prednisolone 1 mg/kg BW s.i.d. or b.i.d. PO).


Management of chronic allergic reactions is based on minimizing or removing exposure to the allergen, which is often not possible. Treatment of recurrent allergies may be frustrating and prevent horses from competing due to regulatory drug withdrawals.


Hyposensitization with a vaccine formulated with antigens selected on the basis of history and results of intradermal testing seems to be a long-term treatment option, and response rates vary from 50% to 85%. The chronic administration of low doses of allergen (1 mL of a 20 000 protein nitrogen unit [PNU] of allergen per milliliter of water SC or IM q 21 days) may stimulate the Th1 type response and/or production of IgG “blocking antibody” that anticipates the binding of IgE to the antigen. In addition, hyposensitization may decrease the number of B cells expressing IgE receptor and restore antigen-stimulated IL-4 production to the levels of non-affected individuals.



AUTOIMMUNITY



Autoimmune diseases


Autoimmunity is an immune response against self-molecules. The presence of autoimmunity is common to all individuals and may or may not have an adverse effect on the body. Autoimmune diseases are clinical manifestations that involve B lymphocytes (antibodies) or T lymphocytes (CD4+ helper or CD8+ cytotoxic cells) reactive to self-antigens. Auto-reactive B and T cells are controlled by mechanisms of central and peripheral tolerance. However, immune dysfunction involving antigen presentation and co-stimulation, genetic defects (often familial) and environmental factors may lead to failure of autoimmunity control and, consequently, to autoimmune disease.


The failure of the anti-idiotype control mechanism of antibody production may allow the production of autoantibodies. In addition, molecular mimicry of microbes and self-epitopes may result in immune responses that overcome immunologic tolerance and lead to tissue injury. Exposure of auto-antigens present in systems that are not normally visited by lymphocytes (e.g. a breakdown of the blood–brain barrier in the central nervous system) or the development of new epitopes on normal proteins may stimulate an immune response.


Viruses, particularly those that infect lymphoid tissues, may be capable of interfering with immunologic control mechanisms. Exotoxins produced by bacteria (e.g. Clostridium spp.) may promote damage of red cell membranes and the development of antigenic epitopes for the production of autoantibodies. Penicillin-induced immune-mediated hemolytic anemia is caused by the strong binding of the drug hapten to the surface of red cells. The complex induces the production and subsequent binding of antibodies. In some cases, autoimmunity is associated with the aging process and immune system malfunction, or the effect of sexual hormones. The mechanisms that induce tissue injury are those described in hypersensitivity reactions (q.v.).



Immune-mediated thrombocytopenia


Thrombocytopenia is a rare condition in horses, and most cases are of unknown cause. In addition to disseminated intravascular coagulopathy and decreased megakaryopoiesis, thrombocytopenia may be associated with autoimmunity. In the latter, it may occur in patients with infectious agents such as equine infectious anemia (q.v.) or Clostridium spp. (q.v.), after administration of drugs (e.g. penicillin, heparin, quinidine, thiazides, digoxin, sulfonamides, erythromycin) or in neoplasia (lymphosarcoma).


Clinical signs include hemorrhagic diathesis characterized by epistaxis, prolonged bleeding from injection sites, mucosal petechiae, hematomas, occult blood in urine and feces, melena, hemoarthritis, and/or hyphema. These findings may be differentiated from vasculitis by the absence of heat, pain and swelling. In general, clinical signs are not detectable until platelet counts are <40 000/μL, and spontaneous bleeding may occur when counts are <20 000–10 000/μL.


A coagulation profile (q.v.) often reveals prolonged bleeding time, abnormal clot retraction, slightly prolonged activated coagulation, elevated fibrinogen degradation products (FDP), normal prothrombin time (PT) and normal activated partial thromboplastin time (APTT). Megakaryocytic hyperplasia is an expected response to the platelet destruction in the periphery and, when absent, may indicate immune-mediated destruction of megakaryocytes or myelophthisic disease.


Immune-mediated thrombocytopenia results either from the production and binding of immunoglobulins to platelets or megakaryocyte antigen surface (primary or idiopathic thrombocytopenia), or from the binding of immunocomplexes (antibodies against microorganisms, or a drug hapten) to the Fc receptors on platelets (secondary thrombocytopenia). In addition, platelet-bound IgM may fix complement. The antibody- or antibody– complement-coated platelets are non-specifically removed from the circulation by the reticuloendothelial system (macrophages in the spleen and liver that phagocytose the platelets via their Fc and complement receptors). The mechanism for autoantibody production may involve auto-reactive B cell clones that are stimulated during an immune response to infectious organisms, dysfunction in CD4+ T cell regulation of B cell response, antigenic mimicry and altered anti-idiotypic regulation of antibody production.


Thrombocytopenia should be confirmed by performing an automated platelet count (q.v.) using a blood sample preserved in citrate anticoagulant, and/or a platelet hand count to rule out pseudothrombocytopenia. Flow cytometric analysis of IgG and IgM coated platelets may confirm the diagnosis of an immune-mediated process. The immuno-injury assay that measures anti-platelet activity in plasma of affected horses often renders false negative results. The presence of young platelets in the circulation as an indicator of bone marrow response to thrombocytopenia can be assessed by the staining of blood platelets with thiazole orange and flow cytometric analysis. Physical examination and ancillary diagnostics should be used in an attempt to identify the presence of bacterial or viral infections and tumors. A Coggins test should be used to diagnose equine infectious anemia (q.v.). In confirmed cases, intermittent hemolysis occurs concomitantly with viremic states.


If drug-induced thrombocytopenia is suspected, discontinuation of the medication is imperative for remission of the autoimmune response.


Platelet-enriched plasma transfusion may partially replace the loss of platelets to levels that prevent spontaneous bleeding or diathesis. Platelet-enriched plasma is prepared by centrifugation at 200 g for 10 min at room temperature of aseptically collected blood in plastic bags containing 2.5% sodium citrate. This therapy may be more effective in horses of small size (foals). The shelf life of platelet-enriched plasma is just a few hours before platelet function inactivation.


Immunosuppressive therapy may be attempted with the use of corticosteroids (dexamethasone 0.1–0.2 mg/kg BW s.i.d. IV), which suppress phagocytic function and antibody production. After 4–5 days, an increase in platelet counts may allow progressive reduction of the dexamethasone dose (20% daily) and, potentially, its substitution with prednisolone (starting at 1 mg/kg BW b.i.d. PO, with subsequent reduction in dose). Low dose corticosteroid treatment or early discontinuation of therapy may result in persistence or recurrence of disease. Therefore, in many patients, immunosuppressive therapy may last for 30 days.


The use of other immunosuppressive therapy (vincristine sulfate 0.01 mg/kg BW IV one or two doses a week apart, in combination with dexamethasone; or azathioprine 3 mg/kg BW s.i.d. PO, and subsequent decreasing doses) is an option in cases refractory to corticosteroid therapy. Immunoglobulin transfer using IV plasma transfusion may assist by interfering with macrophage Fc-receptor clearance of immunoglobulin-coated platelets.

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Jul 8, 2016 | Posted by in EQUINE MEDICINE | Comments Off on The immune system

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