Regenerative Hemolysis – Metabolic Disorders  Pyruvate kinase deficiency  ↓ ATP production leads to reduced RBC lifespan and hemolysis  Dogs: Basenji dogs, many other breeds  Markedly regenerative (up to 50% reticulocytes!!)  Myelofibrosis and osteosclerosis  Survival 3-5 years  Cats: Abyssininians, Somalis, DSH  Variable with regeneration  Normal lifespan Regenerative Anemias Hemolysis – Metabolic Disorders  Phosphofructokinase deficiency  English Springer spaniels  Decreased 2,3 DPG production – less O2 offloading, greater alkaline fragility  Acute, intravascular triggered by respiratory alkalosis (exercise, excitement)  Not anemic between hemolytic crises  Hereditary  Brown blood, decreased O2 capacity  Dogs, cats and horses  No anemia, anesthesia concern Regenerative Anemias Hemolysis – Micro-organsisms  Red cell parasites  Primarily via a secondary IMHA  Predominantly extravascular, but many have some intravascular component  Other microorganisms  Clostridium and Leptospira species – intravascular hemolytic anemia  E. coli hemolyic uremic syndrome  Equine infectious anemia  Acute stages of disease  Likely immune mediated  Occasionally see thrombocytopenia as well Regenerative Anemias Hemolysis – Miscellaneous  Severe hypophosphatemia (<1 mg/dl, canine RI 3-5.3 mg/dl) causes intravascular hemolysis  Post parturient hemoglobinuria, hypophosphatemia believed to play a role  DM (cats) loss through urine, intracellular movement with insulin administration  Hyperalimentation  Snake venom – IV hemolysis, echinocytes, spherocytes Non-Regenerative Anemias  The most common type of anemia  Anemia of inflammatory disease  Reduced erythropoiesis (hypocellular  Anemia of endocrine disease – marrow) mild to occ mod  Defective erythropoiesis (hypercellular  Hypoadrenocortisism (Addison’s marrow) disease), hypothyroidism,  Additional information hypopituitarism  Cell lines affected, only RBCs or WBCs  Myelophthesis (crowding out of and ? ) – severe!  Chemistry panel and endocrine testing  Chronic, severe, iron deficiency  Bone marrow findings  Microcytic, hypochromic, +/- ↑  History and PE platelets and ↓TP  Weeks to months to manifest – long  Certain infectious diseases RBC lifespan  Drugs and toxins Causes  Idiopathic  Chronic renal failure – severe!  ↑ BUN, Creat, Phos, isosthenuric USG Non-Regenerative Anemias Anemia of Inflammatory Disease (AID)

 Most common cause!  Inflammatory, infectious and neoplastic diseases  Typically is mild to moderate (PCV 20-30%)  Normocytic and normochromic; Rarely microcytic and/or hypochromic Parameter Iron deficiency AID Serum iron Decreased Decreased TIBC (transferrin) Normal to increased Normal to decreased Transferrin saturation Decreased Decreased Ferritin Decreased Normal to increased BM iron (dogs, not cats!) Decreased Normal to increased History, other findings Nutrition, blood loss Underlying inflammation, neoplasia Response to Resolves Does not respond supplementation Non-Regenerative Anemias Infectious Agents  AID – any infectious agent  Ehrlichia canis  Feline Leukemia Virus  Classic findings  Selective damage to erythroid  ↑ globulins, ↓plts, ↑ lymphocytes precursors (pure red cell aplasia)  Mild non-regenerative anemia  AID  Neoplasia or myelodysplastic  Immune-mediated syndrome resulting in  myelophthesis Chronic disease may result in pancytopenia   Immune-mediated  Anemia of inflammatory disease  Aplastic anemia  Macrocytic, normochromic,  Equine infectious anemia non-regenerative anemia  Bone marrow suppression  Bone marrow dysplasia Non-Regenerative Anemias Drugs and Toxins  Many implicated drugs and  Estrogen in dogs and ferrets chemicals  Can be idiosyncratic or overdose  May result in only anemia or in dogs aplastic anemia  Sertoli or granulosa cell tumors of dogs  Aplastic – all cell lines affected  Mismate shot, pseudopregancy  Can be idiopathic or predictable termination in dogs  Can be reversible or permanent  Hyperestrogenism related to prolonged estrus in ferrets  Idiosyncratic – many reported causes  Bracken fern toxicity in cattle Classic causes of aplastic anemia  Chloramphenicol  Chloramphenicol  Griseofulvin in cats  Anti-neoplastic and  Trimethoprim-sulfa, chemotheraputic agents cephalosporin in dogs Non-Regenerative Anemia Drugs and Toxins - Lead  Species sensitivity varies  Highly sensitive: Dogs  Intermediate: Cattle, horses, sheep  Resistant: Pigs  Classic blood smear findings  Most likely seen in dogs  Anemia with inadequate regeneration  Basophilic stippling  Inappropriate nRBCs (aka Erythrocytosis)  Some breeds normally have higher Hcts: Racing horses, warm- blooded breeds, Greyhounds, sight hounds, some Dachshunds Polycythemia

Relative Absolute a) Dehydration True increase in RBC ↑TP, USG, azotemia, PE production findings, PP:Fib > 15 b) Splenic contraction Primary Secondary Excitable animal (young cats, (Epo independent) (Epo dependent) foals), epinephrine response , dx of ↑ Epo exclusion, N or ↓ epo levels

Appropriate Inappropriate Hypoxia induced; ↓arterial PAO2, No hypoxia, N arterial PAO2, CHF, R→L shunting, pulmonary Renal tumors or cysts, tumors dz, high altitude secreting epo WBCs – Major Patterns of Response Neuts Left Toxic Lymphs Monos Eos shift change Classic inflammation ↑ + * +/- N, ↓, ↑ ↑ N to ↓

Inflammation, ↓ + + ↓ N to ↑ ↓ overwhelming tissue demand** Stress ↑ - - ↓ ↑ ↓ (corticosteroid resp.) Inflammation with ↑ +/- +/- ↓ ↑ ↓ stress Epinephrine ↑ - - ↑ N N to ↓

*Could be normal with chronic or mild inflammation **More likely in cattle d/t small storage pool Inflammation  Degenerative left shift  Species with less responsive BM:  Immature forms > mature forms OR ruminants, +/- horses significant left shift with normal or low  Plasma protein / fibrinogen ratio neuts  <10 inflammation, >15 normal or  Regenerative left shift dehydration   Mature forms > immature forms in Ruminants, inverted N:L ratio (may patients with a normal or high not be true, but everyone says it) neutrophil count , other causes  Toxic change – moderate to marked,  Chemotheraputic agents, causes of think infection! aplastic anemia/pancytopenia,  Döhle bodes, cytoplasmic , immune mediated, cyclic vacuolization, toxic granulation** hematopoiesis of grey collies and  A rare Döhle body in cats is OK FeLV+ cats, familial neutropenia of  **Rare in LAs, nearly unheard of in SAs, Belgian Tervuren dogs, idiopathic common in avian, reptile and amphibian species  Toxic change ≠ degenerative changes! Neutrophils, Hereditary Disorders  Leukocyte adhesion defect – neuts can’t leave blood  Holstein cattle, Irish setters  Marked to extreme  Recurrent infections with failure to form puss  Pelger-Hüet anomaly: failure of nuclei to segment, no function defect, don’t mistake for LS!  Lysosomal storage disorders; group of disorders  May see dwarfism, skeletal abnormalities, hepatomegaly, neurologic signs  Blood smear; neutrophils with pink-purple granules, lymphocytes with vacuoles or lymphocytes with granules  Birman cat neutrophil granulation anomaly  Granulated neutrophils, no defect. Also seen in some Orientals Neutrophil Inclusions  Granulocytic Ehrlichiosis and Anaplasmosis  E. ewingii: Can also find in joint fluid in acute disease with polyarthritis  A. phagocytophila (formerly E. equi)  Primarily dogs, rarely cats  Thrombocytopenia and anemia  Morulae  Pale blue to lavender, ‘raspberry’ or ‘asterisk’ like  Distemper inclusions Extreme (Leukamoid Response)  Think localized, purulent responses, ‘puss pockets’: Pyometra (esp. closed), walled off abscesses, pyelonephritis, pyothorax  Hepatozoon infection in dogs: tick transmitted protozoal infection of canids  H. americanum , SW and SE United States  Pyogranulomatous myocytis and periosteal bone reactions  Gamonts rarely found in peripheral blood neutrophils and  Very pale staining elongated structure, ‘Tick-tacs’  Immune mediated hemolytic anemia  Atypical response to parasitic infections - HW disease  Leukocyte adhesion deficiency  Paraneoplastic; production is cause by neoplasia  Chronic myelogenous leukemia – RARE! Eosinophils and Basophils Eosinopenia  Paraneoplastic: Mast cell tumor (dogs,  Corticosteroid response horses), T-cell lymphoma, various carcinomas  Promoted by epinephrine, not a consistent feature  Hypereosinophilic syndrome  : “Wheezes, sneezes and weird Rottweiler dogs, other dog breeds diseases”  Infiltration of multiple tissues with eosinophils  Parasitic infections, especially those with tissue  Presumed to be dysregulation stages of eosinophils  Heartworm disease is often NOT associated with significant eosinophilia  Feline hypereosinophilic granuloma complex  Hypersensitivity reactions: Asthma, dermatitis, flea bite allergy  Hypoadrenocortism: 20% of patients  Localized eosinophilic responses  Hyperthyroidism in cats  May NOT see peripheral eosinophilia  Eosinophilic leukemia – rare!  Eosinophilic keratitis, eosinophilic granuloma, Basophilia without eosinophilia canine panosteitis  Uncommon likely similar Ddx as  Infections: Bacteria (staph and strep), fungal eosinophilia; horses, canine HW disease, infections – Cryptococcosis, pythiosis myeloproliferative disorders Lymphocytes  Predominant in adult ruminants, many birds, Lymphocytosis pigs, mice and rats  Epinephrine response: young cats, foals  Ruminants can have large lymphocytes  Chronic antigenic stimulation Lymphopenia  Chronic canine Ehrlichiosis – not  Corticosteroid response uncommon  Early acute infections  Counts of up to 30,000/µl  Often many granulated lymphocytes  Destruction of lymphocytes  Trypanosomiasis in cattle  Some viral infections; distemper, parvovirus,  Leishmaniasis in dogs panleukopenia, FIV  Brucellosis  Chemotherapy, radiation  Mild increases with vaccination  Chylothorax / abdomen, PLE, lymphangectasia  Hypoadrenocorticism  therapy  Lymphoid neoplasia   Destruction of lymph node architecture: ALL – high number of blasts lymphoma, granulomatous disease  CLL – high numbers of small lymphs   Severe, combined (SCID) Leukemic phase of lymphoma  Arabian foals, rarely in dogs  Persistent lymphocytosis of cattle  Lymphocyte count of <1000/µl  BLV causes non-neoplastic proliferation  <5% develop lymphoma Mastocythemia  Mast cells are bone marrow derived, but not typically found in blood in health  Increased numbers  GI disease, especially parvovirus infection  Other inflammatory diseases  Mast cell tumor  Systemic mastocytosis of cats  Mast cell tumor with high circulating numbers of mast cells  Often involves the spleen  True mast cell leukemia is rare Hematopoietic Neoplasia  Acute: blast cells present  Lymphoid  Acute lymphocytic leukemia – high blast count, no tissue infiltration, anemic  DDx: leukemic phase of lymphoma – lower blast count, enlarged lymphoid tissues +/- other tissues, not anemic  Myeloid, monocytic, myelomonocytic, others, less common  Chronic: cells are differentiated  Lymphoid  Chronic lymphocytic leukemia  DDx: epinephrine response, chronic antigenic stimulation (Ehrlichia)  Myeloid: really rare, rule out other causes of neutrophilia, eosinophilia first!! Tests of Coagulation Test Sample Evaluates count EDTA whole blood Platelet numbers Citrated whole blood PT Citrated plasma Extrinsic and common pathways aPTT Citrated plasma Intrinsic and common pathways ACT** Gray top tube, whole Intrinsic and common pathways blood FDPs and d-dimers Usually citrated plasma, Fibrinolysis (d-dimers specific for varies by kit clot lysis) BMBT N/A Platelet function Fibrinogen Amount of fibrinogen Heat precipitation EDTA whole blood Thrombin time Citrated plasma **Very insensitive 95% reduction needed!! Affected by significant thrombocytopenia Patterns of Common Diseases Disorder PT aPTT FDP / Fibrino Platelets BMBT** d-dimers gen vWD N N N N N ↑ Thrombopathy N N N N N ↑ Thrombocytopenia N N N N ↓ N -↑ Hemophilia A (VIII), B N ↑ N N N N (IX), fXII deficiency Early Vit K deficiency / ↑ N N N N N antagonism Classic Vit K def/antag ↑ ↑ N N N N Early or compensated N or ↑ N or ↑ N or ↑ N ↓ N DIC Fulminate DIC ↑ ↑ ↑ N or ↓ ↓ ↑ **Prolonged by anemia Thrombocytopenia / Thrombocytosis Thrombocytopenia Thrombocytosis  Spurious  Physiologic; exercise or epinephrine, excitable  Platelet clumping animals  Breeds: Cavalier King Charles spaniels (75-100K),  Transient mild to moderate increase (few Greyhounds and Shibas, mildly decreased hours)   Possible concurrent lymphocytosis and Decreased production neutrophilia  Usually pan or bicytopenia, only plts decreased is RARE!  Reactive  Look for causes of BM suppression  Post regenerative response, mild increased   Increased consumption/loss Chronic hypoxia  Iron-lack anemia  DIC, acute massive hemorrhage  Corticosteroids, mild increase  Destruction  Inflammation  1º or 2º IMT, non-immune mediated destruction  Splenectomy  Abnormal distribution  Myeloproliferative  Splenic congestion or disease, hemangiosarcoma  Rule out other causes first!!  Essential thrombocythemia, high numbers of platelets (1000K or >)  False ↑ K, released with clotting  Megakaryoblastic leukemia, may see thrombocytopenia Thrombocytopenia – IMT and non-IM  1º IMT  2º IMT, non-IM destruction  Platelet count usually <  Overlapping causes, often 50K/µl, often <10K/ µl both mechanisms are present  +/- regeneration (↑ MPV)  Systemic autoimmune  Often with IMHA (Evans’ diseases syndrome)  Neoplasia – 2º and non-IM  Diagnosis of exclusion: response  Infectious agents - May have to therapy, immunosuppression, both 2º and non-IM must r/o infectious causes!!  Rickettsial agents – common!  Supportive tests (can’t  Viral agents distinguish 1º from 2º )  Vaccination with modified live  Platelet surface-associated viruses - not uncommon immunoglobulin (PSAIg)  Requires special sample handling Thrombopathias – decreased function Acquired causes – Common! Hereditary dysfunction, Rare!  Uremia - common  Rule out other common causes of  Drug therapy - common platelet dysfunction first!  Cyclooxygenase inhibitors (aspirin,  Suspect if : NSAIDs)  BMBT is ↑, plt # are normal, no  Numerous others anemia, normal vWF:Ag, acquired causes are r/o  Marked hypergammaglobulinemia (Multiple myeloma, some infections)  Specialized equipment/assays required  Coat platelets and decrease ability to to determine underlying defect. aggregate Disease include (don’t bother  Anti-platelet antibodies directed against memorizing!): surface receptors  Basset hound thrombopathia, Chediak Higashi syndrome (Persian cats;  Snake venom Herford, Brangus, Japanese black  Neoplasia cattle), cyclic hematopoiesis of grey  Elevated FDPs with DIC – interferes with collies, Glanzmann’s thrombasthenia, aggregation Simmental hereditary thrombopathia, Spitz thrombopathia, Thrombasthenic thrombopathia of otterhounds Von Willebrands Disease Decreased vWF means poor platelet binding  Type II disease: least common. Mod to Screening test abnormalities severe bleeding  German shorthaired pointer and the  Mild to moderate disease, ↑ BMBT German wirehaired pointer  Severe disease, possible ↑ aPTT  Type III disease: uncommon. Severe Testing bleeding  Measurement of plasma vWF levels  Most animals do not survive to adulthood  <50% positive, 50-70% gray zone, >70%  Chesapeake retriever, Dutch kooiler, negative Scottish terrier, Shetland sheepdog  Doesn’t distinguish molecule size - need Treatment protein electrophoresis  Fresh, whole blood, fresh plasma, fresh- DNA testing – defect varies with breeds and type frozen plasma of disease, separate test for each  Stored products contain minimal vWF Disease types  Plasma crypoprecipitate; high levels of  Type I disease: most common! Mild to vWF in a small volume moderate bleeding  Airedale, Akita, dachshund, Doberman pinscher, German shepherd, golden retriever, greyhound, Irish wolfhound, Manchester terrier, schnauzer, poodle, Shetland sheepdog, many others Enhanced Platelet Function  Increased risk of thromboembolic disease, esp. pulmonary thromboembolism  Diabetes mellitus, Cushing’s disease, heartworm disease  Laboratory evaluation  Specific tests of enhanced function are available, require specialized testing and meticulous sample handling  Global tests of coagulation (thromboelastographyTEG) may be better at detecting Coagulopathies - Hemophilia  Hemophilia A (VIII) and B (IX)  Intrinsic, ↑ aPTT  Defect is X-linked and recessive  Factor type bleeding in young animals  Dogs, IX occasionally in cats  Factor XI deficiency (Hemophilia C)  Most common hereditary coagulopathy of cattle  Signs are typically mild, rarely see spontaneous hemorrhage post sx, procedures, parturition  Increased incidence of mastitis, pneumonia chronic dermatitis, metritis, rebreeding  Pink colostrum!  Factor XII deficiency  Most common hereditary deficiency in cats (still rare)  Markedly prolonged aPPT without bleeding Vitamin K Deficiency and Antagonism Vitamin K dependant factors: II (thrombin), Laboratory findings VII, IX, X, protein C and protein S  Classic: Markedly ↑ aPTT and PT Vitamin K antagonism, common!  Early; may see only ↑ PT (fVII short  Rodenticides : warfarin, brodifacoum t½) (common), moldy sweet clover  Additional diagnostics (dicoumerol)  PIVKA test: detects deficiencies of Vitamin K deficiency (uncommon) vitamin K dependant factors, highly  Usually does not cause clinical signs sensitive Pathogenesis  GI sterilization after antibiotic, severe GI disease (decreased absorption), EPI, obstructive cholestasis, dietary deficiency  Severe liver disease  70-80% loss of functional mass  Acute, massive necrosis  Terminal cirrhosis or other terminal chronic liver disease  Will see other lab evidence of liver disease DIC Triggers of DIC  Most difficult to diagnose, best time to  Release of activating factors intervene!  Tissue factor released from: Tumors (necrosis,  Clinical signs not evident – too early, endogenous production, tumor lysis potential for DIC recognized based on syndrome, AML), massive tissue trauma, underlying disorder burns, endotoxin mediated release,  Screening tests  Some snake venom  PT and/or APPT may be normal or  Amniotic fluid (obstetric accidents) shortened (not reliable)  Widespread endothelial injury  BMBT likely normal, may be prolonged -  Acute, necrotizing pancreatitis, heat stroke, FDPs infectious agents that cause vasculitis  FDPs may be normal or mildly increased (Rickettsial agents – RMSF, Gram negative  Mild thrombocytopenia likely bacteria, viral agents), severe metabolic acidosis or uremia, uncontrolled, systemic IM  Advanced tests of global coagulation disease, severe/widespread burns potential   Acute, severe, intravascular hemolysis Thromboelastography, others potentiates  Better at detecting hypercoagulable states  Not readily available outside referral Peractute: initial stage of DIC institutes and sample can’t be stored and  Hyper-coagulative state shipped DIC Low grade DIC  Moderate to severe  Signs often mild or absent thrombocytopenia   Possible mild ↑ in BMBT - elevated FDPs FDPs /d-dimers- moderate to markedly elevated  Mild thrombocytopenia  BMBT – normal to prolonged  Schistocytes  Additional tests  Common causes of low grade DIC  Blood smear evaluation – schistocytes  Cardiovascular disease  Fibrinogen levels N to ↓  Neoplasia  Anti-thrombin III levels ↓  Various causes of vasculitis  Inhibitor of activated thrombin Fulminant DIC  Also decreased with PLN, leads to TED  Signs associated with fulminate DIC  What are the most consistent findings?  End-organ failure/dysfunction  Schistocytes in peripheral blood  Coagulopathy; frank hemorrhage or may  Thrombocytopenia only detect via laboratory tests  Low AT III levels  Screening tests  Increased FDPs / D-dimers  PT and aPTT, likely one or both prolonged due to consumption of factors