AVIAN HEMATOLOGY Do Dr Yasemin SALGIRLI DEMRBA Resident

AVIAN HEMATOLOGY Doç. Dr. Yasemin SALGIRLI DEMİRBAŞ Resident ECAWBM (BM)

BLOOD • Blood; ütransports O 2, nutrients and hormones to body cells/ carries CO 2, water and other wastes away from cells üis composed of plasma (contains protein, salts, sugar) and blood cells üAvian blood plasma has a higher sugar and fat content – nitrogenous waste product • Blood cells float in the plasma • The blood of every bird contains erythrocytes or Red Blood Cells (RBCs), leukocytes or White Blood Cells (WBCs) and Thrombocytes, the avian equivalent to mammal platelets.

ERYTHROCYTES üThe mature avian erythrocte is oval with a centrally positioned oval nucleus. üEx-Theory: The anucleated erythrocyte, as it is seen in mammals, is considered more evolutionarily ‘‘advanced’’ üMammals have smaller end-blood-vessels (capillaries of about 3 lm in diameter) than birds - The presence of a nucleus may prevent big nucleated RBC to squeeze through small capillaries. üDuring the evolutionary development, nature has found that it was better to extrude the nucleus and also other cell organelles, such as endoplasmic reticulum for protein synthesis, which were not needed for their actual function as oxygen carrier. üHowever, nucleus in the erythroblast is about 2 lm in diameter. And, if it was distributed at peripherally inflated region of the erythrocyte, it would neither hinder erythrocyte deformation nor its entrance into end-blood-vessels.

ERYTHROCYTES ü New Theory: Enucleation may facilitate erythrocyte deformation into the biconcave shape and becoming soft and elastic. ü Nuclear and mitochondrial extrusion may help mammal erythrocytes to better adapt to high-sugar and high-heme conditions, where they live. ü Avian erythrocytes are much larger than most mammalian erythrocytes. ü The size varies among species (7, 5 µm- 13, 5 µm). ü A small degree of anisocytosis* is usually seen in clinically healthy birds ü Besides typical mature RBCs, lesser numbers of other cells representing different stages of RBC development can be found on the blood of healthy birds - The presence of small numbers (between 1 to 5% of total circulating RBCs) of these immature cells in peripheral blood is normal ü elevated counts are described as polychromasia or polychromatophylia and indicative of increased erythropoiesis as seen in regenerative anemias • *Anisocytosis: RBCs are of unequal size

ERYTHROCYTES üTotal RBC count in birds is usually estimated by manual methods but more recent flow cytometric analysers properly adjusted can also be employed. üTotal RBC count is lower in birds (1, 5 to 4, 5 x 106 cells/µL) than in mammals. üAvian RBCs have a shorter half-life (25 to. 45 days) than mammal cells

ERYTHROCYTES üRbcs number depends upon : 1. Age: is higher in young animals 2. Sex: Male is higher than females 3. Hormones: üAnabolic hormones as thyroid hormones increase number of Rbcs. üEstrogen interfere iron absorption so decrease Rbcs count. 4. Seasons: Winter stimulate thyroid activity so increase Rbcs count. 5. Altitude: High altitude increase Rbcs due to increase 6. Erythropoitin hormones.

ERYTHROPOESIS: process of RBC formation ü Normal erythropoiesis takes place in the bone marrow although ectopic erythropoiesis can be occasionally found on the spleen and liver ü This process comprises eight sequential stages of cell development: 1. Rubryblasts (or erythroblasts) are the first stage of RBC development. 2. Prorubrycites are the second stage of RBC development-the lack of nucleoli and mitochondrial spaces in the cytoplasm 3. Basophilic rubrycites are third stage of RBC development and have homogenous basophilic cytoplasm and round nuclei 4. Initial polychromatic rubrycites - beginning of hemoglobin synthesis. 5. Polychromatic RBCs (Rubricyst) - There is a spectrum of blue color due to synthesis of hemoglobin 6. Metarubrucyte - This is the last nucleated erythrocyte stage 7. Reticulocyte - The reticulocyte is slightly larger than the mature erythrocyte 8. RBC

ERYTHROPOESIS • Erythropoietin (EPO): • kidney hormone that controls erythropoiesis - Glycoprotein hormone • In case of low O 2 levels –– EPO is secreted - stimulates red blood cell formation – (Humans - Anemia Treatment) – EPO

ERYTHROCYTES üAvian RBCs have a shorter half-life (25 to 45 days) than mammal cells - this may be associated with a higher body temperature in birds and rapid metabolic rate of avian rbcs, which consume higher rates of oxygen and nutrients than their mammal counterparts üBlood volume in birds is estimated between 4, 4 and 8, 3 % of body weight (converting grams to mililiters) with younger birds having a higher blood volume than adults. üCompared to mammals, avian blood is more viscous because RBCs are larger and less deformable. üBlood density is mostly influenced by the concentration of plasma proteins üBirds have lower blood albumin concentration and lower oncotic pressure than mammals

HEMOGLOBIN ü Normal values of Hb v. Chicken: 9. 3 gm% v. Ducks: 10. 3 gm% v. Turkey: 10. 3 gm% v 8 -12 g/dl in chicken blood ü In most vertebrates, hemoglobin is made up of four subunits, each one of which has its own binding site for oxygen. ü There are some significant differences between avian hemoglobin and hemoglobin found in other vertebrates: v. In adult birds there are two different types of hemoglobin, hemoglobin A and D, both of which vary from each other in their affinity for oxygen. v. Hemoglobin A is often the more prevalent form and has a lower affinity for oxygen compared to hemoglobin D. v. A lower affinity means that oxygen is more readily dissociated from hemoglobin as arterial blood

Hb v. In general, avian hemoglobin shows more cooperativity with oxygen than does hemoglobin in other vertebrates. v. Cooperativity is the phenomenon whereby the binding of one molecule of oxygen with hemoglobin facilitates the binding of the next molecule of oxygen and so on up to the binding of four oxygen molecules by a molecule of hemoglobin. v. This cooperativity accounts for the sigmoidal shape of the oxygen-hemoglobin binding curve. v. The advantage of this high cooperativity is that it increases the delivery of oxygen to tissues.

Hb v. Another feature of avian hemoglobin is its interaction with inositol pentaphosphate and inositol tetraphosphate which shifts the oxygen-hemoglobin dissociation curve to the right and decreases the affinity of oxygen for hemoglobin, enhancing the delivery of oxygen to tissues. v In mammals the principal organic phosphate is 2, 3 - biphosphoglycerate (2, 3 -BPG), formerly known as 2, 3 -diphosphoglycerate (2, 3 DPG), v. The presence of two hemoglobins with different oxygen affinities, means that the erythrocytes have a greater range of oxygen partial pressures over which oxygen can be bound and released, favoring the uptake of oxygen where there is little oxygen available in the environment v. Carbon dioxide also affects the binding of oxygen to hemoglobin. In mammals, the binding of carbon dioxide with hemoglobin is more. It appears that in birds the strong binding of the organic phosphates to hemoglobin prevents this carbon dioxide effect.

LEUKOCYTES üAs in mammals there are Granular and Agranular leukocytes. üNumber of leukocytes: o Chicken: 19 -30 thousands/mm 3 o Duck: 32 thousands/mm 3 üLeukocytes are either granular or Agranular. üGranular leukocytes 1 -Heterophils 2 -Eosinophiles 3 -Basophils üAgranular leukocytes 1 -Lymphocytes 2 -Monocytes

HETEROPHILS üIt is analogue of mammalian neutrophils. üIts diameter about 10 -15 µ. üThe nucleus is polymorphic with varying degree of lobulation. üThe cytoplasm contains a characteristics rod-like granules , pointed at their ends, acidophilic bodies. üThe percentage of heterophil in blood ranges 20 -30% (except in ostrich and pheasants 60 -70%) üHeterophils are phagocytic and use their enzyme-containing granules to lyse ingested materials. üHeterophils are motile and can leave blood vessels to engulf foreign materials

EOSINOPHILS üThe size of eosinophil is about the same size as the heterophil. üRounded cells , üThe cytoplasm contains large, spherical, dull red coloured granules. üThe cytoplasm is Faint blue colour. üThe nucleus is often bilobed and Darker than that of heterophils. üEosinophils make up about 2 to 3 % of the WBC population of healthy birds.

BASOPHILS üThe size of basophil is about the same size as the heterophil. üRounded cells , the cytoplasm is devoid of colour and contains basophilic granules which may mask the nucleus and protruded from the surface of the cells giving black berry appearance. üThe nucleus is weakly Basophilic , may be rounded or oval.

LYMPHOCYTES ü The lymphocytes constitute the majority of the leukocytes in the blood of the fowl. ü The cytoplasm is weakly basophilic , it may forms a narrow rim around the rounded nucleus in small lymphocyte. ü The cytoplasm may constitute the major portion of the cell, as in the large lymphocytes. ü The nucleus is usually round and has a fairly coarse pattern of chromatin ü The percentage of lymphocytes in blood ranges 60 -70% (except in ostrich and pheasants 20 -30%) ü Lymphocytes are either T-lymphocytes (formed in the thymus) or Blymphocytes (formed in the bursa of Fabricius). ü B-lymphocytes produce antibodies; T-lymphocytes attack infected or abnormal cells.

MONOCYTES üMonocytes are large cells with relatively more cytoplasm than the large lymphocytes. üThe cytoplasm of these cells has a blue-gray colour and vaculated. üThe nucleus is usually irregular in outline üMonocytes %=5 -10% üMonocytes are motile cells that can migrate using ameboid movements. üMonocytes are also phagocytic.

THROMBOCYTES üThey are rounded to oval in shape and usually present in clumps. üThey have rounded very darkely stained nucleus and a clear cytoplasm. üThrombocyte are active participants in blood coagulation. üIn addition to blood clotting, these cells may have the ability to phagocytize foreign materials (e. g. bacteria) and they are able to carry O 2 like RBCs if an extreme anemic condition exists. üAvian blood appears to clot more slowly than does mammalian blood. üFor instance, the whole blood clotting time is 38 min for Japanese quail compared 8 min for human blood. üAvian blood does not clot in responses to glass activation

BLOOD GROUPS • Blood transfusing is a process based on erythrocyte surface antigens. • Avian blood typing is very rudimentary. • Extensive investigation and description currently limited with chicken. • In chickens 28 blood groups have been described