1. Blood, Lymph and Immunity

2. Learning Objectives List and describe the functions of blood.
Describe the composition of blood plasma and the characteristics of mature erythrocytes.
Describe the structure of the hemoglobin molecule. Explain the fate of hemoglobin following intravascular and extravascular hemolysis.
Give the origin of thrombocytes. Describe their characteristics and functions.
List the types of leukocytes and describe the functions of each.
Describe the formation of lymph fluid and its circulation through the lymphatic system.
List the functions of the lymphatic system. Describe the structure and function of the lymph nodes, spleen, thymus, tonsils, and GALT.
List the functions of the immune system. Differentiate between specific and nonspecific immune reactions; cell-mediated and humoral immunity.
List the components of cell-mediated immunity. Explain the role of each.
List and describe the classes of immunoglobulins. Differentiate between active and passive immunity.

3. Functions of Blood Transportation Regulation Defense System
Oxygen, nutrients ,waste products, hormones
Regulation
Body temperature, tissue fluid content, blood pH
Defense System
White blood cell phagocytosis, platelets, clotting factors

4. Composition of Blood Liquid portion: Plasma Cellular portion:
Red blood cells (erythrocytes)
White blood cells (leukocytes)
Platelets (thrombocytes)

5. Plasma
45% to 78% of a blood sample volume, depending on the species of the animal and the size of its red blood cells
93% water
Substances dissolved or suspended in plasma
Albumin, globulins, and fibrinogen
Oxygen, carbon dioxide, and nitrogen
Lipids, amino acids, metabolic wastes, and electrolytes

6. Cellular Components Erythrocytes - carry oxygen
Thrombocytes - help prevent leaks from damaged blood vessels
Leukocytes
Granulocytic or agranulocytic
Multiple functions

7. Hematopoiesis Production of all blood cells
Occurs primarily in red bone marrow
Fetal hematopoiesis occurs in the liver and spleen
Neonatal hematopoiesis occurs in red bone marrow

8. Hematopoiesis Older animals
Some red bone marrow is converted to inactive, yellow bone marrow
Composed of fat cells that have replaced some of the active red marrow
In a mature animal most red bone marrow is found at the ends of long bones and in flat bones
Liver and spleen have a limited capacity to participate in hematopoiesis

9. Hematopoiesis Hematopoietic stem cell Pluripotent
Can develop into any one of the blood cells
Depends on chemical or physiological stimuli
Involves numerous cell divisions

10. Erythropoiesis Production of red blood cells
Erythropoietin: hormone released from cells in kidney in response to hypoxia
Triggers stem cell to divide and differentiate
Multiple maturation steps

11. Red Blood Cells Characteristics ~65% water and 35% solids
Primary solid - hemoglobin
Round, anuclear biconcave disks (most mammals)
Variable sizes
Uses plasma glucose for energy

12. Hemoglobin Composed of heme and globin
Heme - pigment portion; produced in the mitochondria; contains iron atoms (Fe++)
Every heme group can carry one molecule of oxygen.
Four heme groups attach to each globin molecule.
Globin - protein portion; produced by ribosomes

13. Normal Hemoglobin Types
Embryonic hemoglobin (HbE) - found in developing fetuses.
Fetal hemoglobin (HbF) - found in fetal blood during mid- to late gestation and up to a couple months after birth
Adult hemoglobin - found in the red blood cells of all animals beginning a couple of weeks to a couple of months after birth

14. Hemoglobin Oxyhemoglobin - hemoglobin that is carrying oxygen.
One oxygen molecule is associated with each iron (Fe++) molecule
Deoxyhemoglobin - hemoglobin that has released its oxygen

15. Carbon Dioxide Transport
CO2 diffuses into red blood cells and is transformed into carbonic acid
Ionizes into hydrogen ions and bicarbonate ions
H2O + CO2 = H2CO3 = H+ + HCO3–
Deoxyhemoglobin accepts the hydrogen ion.
Bicarbonate diffuses back into the plasma.

16. Red Blood Cell Life Span
Varies with the species
Dogs ~ 110 days
Cats ~ 68 days
Horse and sheep ~ 150 days
Cow ~ 160 days
Mice ~ days
Senescence - process of aging

17. Red Blood Cell Senescence
Enzyme activity decreases
Cell loses its deformability
Becomes rounder; volume decreases
Extravascular hemolysis
Macrophages (especially in the spleen) remove senescent RBCs from circulation
RBCs broken down into components that can be recycled in the body or eliminated as waste material

18. Red Blood Cell Senescence
Extravascular hemolysis
RBC membrane is destroyed
Iron is transported to the red bone marrow
Amino acids from globin molecules are transported to the liver for re-use

19. Red Blood Cell Senescence
Heme is converted to bilirubin, bound to albumin, and transported to liver (unconjugated bilirubin)
Bilirubin is conjugated to glucuronic acid.
Conjugated bilirubin excreted as a bile pigment into intestines
Converted into urobilinogen by bacteria
Some is reabsorbed and eliminated in urine as urobilin; some is converted to stercobilinogen and excreted in stool as stercobilin.

20. Red Blood Cell Senescence
Intravascular hemolysis
RBCs in circulation subjected to stresses
Can result in RBC fragmentation and/or destruction
Hemoglobin released directly into the blood

21. Red Blood Cell Senescence
Intravascular hemolysis
Unconjugated hemoglobin attached to haptoglobin
Transported to macrophages in the liver
Processed as with extravascular hemolysis
Excess unconjugated hemoglobin in the plasma (hemoglobinemia) is eliminated in urine (hemoglobinuria)

22. Anemia Results in decreased O2 carrying capacity of the blood
Caused by:
Low number of circulating mature red blood cells (blood loss, increased RBC destruction, decreased RBC production)
Insufficient hemoglobin production (e.g., iron deficiency)

23. Polycythemia Increase in number of RBCs Three types:
Relative polycythemia - hemoconcentration due to fluid loss (e.g., vomiting, diarrhea)
Compensatory polycythemia - result of hypoxia (e.g., high altitudes, congestive heart failure)
Polycythemia rubra vera - rare bone marrow disorder; cause unknown

24. Platelets (Thrombocytes)
Cytoplasmic fragments of bone marrow megakaryocytes
Thrombopoiesis - production of platelets
Megakaryocyte undergoes incomplete mitosis during maturation
Nuclei divide, cytoplasm doesn’t
Results in multinucleated cell with abundant cytoplasm

25. Platelet Characteristics
Circulating platelets are round with numerous small, purple granules
Granules contain some of the clotting factors and calcium
Platelets remain in peripheral blood until they are removed by tissue macrophages because of old age or damage.

26. Platelet Functions Maintain vascular integrity
Release endothelial growth factor into blood vessel endothelial cells
Formation of platelet plug
Attracted to exposed connective tissue of damaged blood vessel
Adhere to exposed connective tissue and each other
Stabilize the hemostatic plug
Fibrin strands form a netlike mesh around and through the platelets.

27. White Blood Cells (Leukocytes)
Classifications:
Functions
Presence or absence of granules
Nuclear shape

28. Don’t stain (usually invisible) Polymorphonuclear Phagocytosis
Table 9-2. White Blood Cells
NAME
CYTOPLASMIC
GRANULES
NUCLEAR
SHAPE
FUNCTION
SITE OF
ACTION
Neutrophil
Don’t stain (usually invisible)
Polymorphonuclear
Phagocytosis
Body tissues
Eosinophil
Stain red
Allergic reactions, anaphylaxis, phagocytosis
Basophil
Stain blue
Initiation of immune and allergic reactions
Monocyte (Macrophage)
None
Pleomorphic
Phagocytosis and process antigens
Body tissues or blood
B cell
Mononuclear
Antibody production and humoral immunity
Lymphoid tissue
T cell
Cytokine production and cell-mediated immunity
Lymphoid tissue and other body tissues

29. Leukopoiesis Occurs in red bone marrow
Some lymphocytes develop further outside bone marrow
Same pluripotent stem cell that produces red blood cells and megakaryocytes
Each type of WBC has its own stimulus for production

30. Granulopoiesis
Production of the neutrophils, eosinophils, and basophils
Initially there are no cytoplasmic granules, then non-specific granules are formed
Specific granules produced during maturation
Granules contain different substances depending on the cell’s function
Example: neutrophil granules contain lysosomal enzymes used in phagocytosis

31. Neutrophil Characteristics
Polymorphonuclear cells; segs
Most numerous WBC in circulation in the dog, horse, and cat
Granules don’t stain with either the blue alkaline stain or the red acid stain (neutral staining).

32. Neutrophils
Mature neutrophils in circulation have two to five nuclear segments joined by a strand of chromatin.
Immature neutrophils have a horseshoe nucleus without any segmentation (band neutrophil).
Neutrophil function: phagocytosis
Granules contain lysosomes capable of destroying bacteria and viruses that have been engulfed

33. Neutrophil Characteristics
In peripheral circulation for about 10 hours
Diapedesis - process used by neutrophils to go from circulation into tissue spaces
Chemotaxis - process that attracts neutrophils to inflammatory chemicals at a site of infection

34. Neutrophil Action
Outer membrane flows around the microorganisms and encases them within a membrane-bound phagocytic vacuole
Cytoplasmic granules move to the edge of the vacuole and fuse with its membrane
Secrete lysosomes into the vacuole

35. Neutrophils
Hydrogen peroxide - produced by neutrophils during oxygen metabolism
Bactericidal
Myeloperoxidase - released from neutrophil granules
Enhances the bactericidal action of hydrogen peroxide
Capable of destroying the cell walls of microorganisms

36. Neutrophil Count Relatively stable range in peripheral blood
Controlled by
Release of mature neutrophils from the storage pool in bone marrow into the peripheral blood
Rate of escape from peripheral blood into tissue
Entrance of increased numbers of pluripotent stem cells into the neutrophil production line

37. Intravascular Pools of Neutrophils
Circulating pool - within lumen of blood vessels
Marginal pool - line the walls of small blood vessels mainly in the spleen, lungs, and abdominal organs

38. Eosinophil Characteristics
Red granules in the cytoplasm of mature cells
0-5% of the total white blood cell count
Produced in bone marrow from the same pluripotent stem cell that gives rise to all other blood cells
Segmented nucleus - usually two lobes

39. Eosinophil Characteristics
Granule shape varies
Dogs: round granules of varying sizes; pale staining
Cats: numerous small, rod-shaped granules
Horses: very large, round or oval-shaped granules; stain intensely
Cattle, sheep, and pigs: round, small granules; stain pink to red

40. Eosinophil Pools Bone marrow reserve Circulating pool Marginal pool
Migrate into tissue within a few hours of release from bone marrow

41. Eosinophil Functions
Anti-inflammatory: granules contain anti-inflammatory substances
Immunity: can ingest substances associated with the humoral immune response
Phagocytosis: minimal phagocytotic and bactericidal functions; usually large organisms such as protozoa and some parasitic worms

42. Eosinophilia Causes: Increased release of mature eosinophils
Migration of eosinophils from marginal pool to circulating pool
Increased production
Longer time in peripheral blood before entering tissue
Accompanying leukocytosis may not occur
Eosinopenia (decreased numbers) - difficult to detect because numbers are normally low.

43. Basophil Characteristics
Blue granules in the cytoplasm of mature cells
Not always visible on stained smear
May completely fill the cytoplasm
Least often seen WBC in circulation
Dog has fewer granules than the other common domestic species
Nucleus usually has 2 to 3 lobes

44. Basophils and Tissue Mast Cells
Theories regarding the relationship between mast cells and basophils:
Two different cell types with similar characteristics; produced in different areas and don’t give rise to one another
Mast cells are tissue basophils

45. Basophil Functions Granules contain histamine and heparin
Histamine helps initiate inflammation and acute allergic reactions.
Heparin acts as a localized anticoagulant to keep blood flowing to an injured or damaged area.
Eosinophils are attracted to site of an allergic reaction by chemotactic factor released from basophil granules

46. Basophilia and Basopenia
Basophilia - can be associated with an allergic or hypersensitivity reaction in tissue
May occur in conjunction with eosinophilia
Basopenia - difficult to evaluate because numbers are normally low

47. Monocyte Characteristics
5-6% of all circulating WBCs in common domestic species
Largest white blood cells in circulation
Abundant cytoplasm stains gray-blue
May contain vacuoles of varying sizes
Cytoplasm may take on a fine granular appearance
Nucleus - pleomorphic, non-segmented

48. Monocyte Functions Important phagocytic cells
Known as tissue macrophages when in tissue spaces
Found in organs that remove or contain foreign invaders, damaged/old blood cells, and cellular debris (liver, spleen, lung, lymph nodes)
Tissue macrophages and monocytes are known as the mononuclear phagocyte system (MPS).

49. Monocyte Functions
Remove cellular debris that remains after an inflammation and/or infection clears up
Process certain antigens, making them more antigenic
Present antigens to lymphocytes as part of immune response
Ingest foreign substances

50. Monocyte Functions
Follow neutrophils into tissue (chemotaxis) in response to tissue damage caused by trauma or invading microorganisms
Remain at site of damage longer than neutrophils
Function in circulating blood to phagocytize damaged blood cells or microorganisms found in the blood (septicemia)

51. Monocytosis and Monocytopenia
Increased number of monocytes in peripheral blood
Often associated with a chronic inflammatory condition
Monocytopenia
Decreased number of monocytes in peripheral blood
Difficult to evaluate because numbers are normally low

52. Lymphocytes Primary circulating WBC in ruminants and pigs
No phagocytic capabilities
Most reside in lymphoid tissues and circulate between these tissues and blood
Thought to arise from the same pluripotent stem cell in the bone marrow as other blood cells
Some lymphocytes leave the bone marrow and mature in other central lymphoid organs

53. Types of Lymphocytes
Every lymphocyte has surface markers that differentiate subsets of each type of lymphocyte.
Markers are not visible under light microscopy.
Three type of Lymphocytes:
T-lymphocytes (T cells)
B-lymphocytes (B cells)
Natural Killer (NK) Cells

54. T-Lymphocytes
Processed in the thymus before going to peripheral lymphoid tissue
Pre-T cells in the thymus are thymocytes
Responsible for cell-mediated immunity and for activating B cells
Most of the lymphocytes in peripheral blood are T cells.

55. B-Lymphocytes
“Bursa equivalent” - refers to bone marrow and other lymphoid tissue thought to be the equivalent of a bird organ called the bursa of Fabricius
Inactive B cells travel through lymph nodes, the spleen, and other lymphoid structures
Rarely circulate in peripheral blood.

56. B-Lymphocytes Responsible for antibody production
Each B cell produces only one specific antibody type against one specific antigen (foreign protein).
Surface receptors are shaped to fit only one antigen shape (epitope).

57. B-Lymphocytes and Plasma Cells
Humoral immunity: B cells recognize an antigen and transform into plasma cells
Activated B cells multiply by mitosis (blastic transformation) to become plasma cells.
Plasma cells produce, store, and release antibodies (immunoglobins).

58. Natural Killer (NK) Cells
NK cells don’t have to be activated by a specific antigen.
Have the ability to kill some types of tumor cells and cells infected with various viruses
Must come in direct contact with these cells before they can destroy them

59. Lymphocyte Characteristics
Circulating lymphocytes are classified as either large or small lymphocytes.
No cytoplasmic granules
Nucleus - round or oval, non-segmented
Large lymphocytes - abundant sky-blue cytoplasm
Small lymphocytes -scant amount of cytoplasm

60. Memory Cells Both T cells and B cells can become memory cells.
Clones of an original lymphocyte
Reside in lymphoid tissue until second exposure to the same antigen encountered previously
Quicker and mounts a greater response than the initial immune response

61. Lymphocytosis
Lymphocytosis - increased number of lymphocytes in peripheral blood
Can result from leukemia, chronic infection, epinephrine release
Can be significant enough to cause a leukocytosis

62. Lymphopenia
Lymphopenia - decreased number of lymphocytes in peripheral blood
Can result from decreased production, corticosteroids, immune deficiency diseases, acute viral diseases
Can result in a leukocytopenia

63. Lymphatic system Series of vessels/ducts
Carry excess interstitial tissue fluid to blood vessels near the heart where fluid is put back into the bloodstream
Also includes lymph tissue scattered throughout the body (lymph nodes, spleen, thymus, tonsils and gut associated lymph tissue [GALT])

64. Lymph
Consists of:
Blood cells - mostly lymphocytes
Nutrients (proteins, fats, etc.)
Hormones
Some T cells circulate from blood to interstitial fluid to lymph and back to blood.
B cells are found primarily in lymph tissues and rarely recirculate.

65. Lymph Formation
Excess interstitial tissue picked up by small lymph capillaries that start blindly in the interstitial spaces of soft tissue
Fluid enters/leaves tissues spaces due to blood pressure and osmotic pressure

66. Lymph Circulation
Lymph capillaries join together to form larger and larger lymph vessels.
Many contain one-way valves that prevent lymph from flowing backwards.
Body movements propel lymph toward the heart.
Lymph vessels eventually join to form the thoracic duct that empties lymph into the vena cava just before it enters the heart.

67. Lymph Circulation
Lymph vessels pass through at least one lymph node and pick up lymphocytes.
Any microorganisms in the lymph are removed by macrophages found in the lymph nodes.

68. Lymph Characteristics
Transparent or translucent liquid containing varying numbers of cells, primarily lymphocytes
More water, sugar, and electrolytes than plasma
Fewer of the larger proteins found in plasma
Chyle - Lymph from the digestive system
Chylomicrons cause lymph to appear white or pale yellow and cloudy.

69. Lymphatic System Functions
Removal of excess tissue fluid
Waste material transport
Interstitial fluid contains some of the waste materials from the tissue cells
Filtration of lymph
Removal of microorganisms, cellular debris, and other foreign matter
Protein transport

70. Lymph nodes
Small kidney-bean-shaped structures located at various points along the lymph vessels
Connective tissue capsule sends branches (trabeculae) into the body of the lymph node
Afferent lymph vessels empty fluid just beneath the capsule.
Efferent vessels exit the lymph node in the indented hilus area.

71. Lymph nodes Cortex - location of resident lymphocytes
Lymph nodules: clusters of lymphocytes around periphery of the node
Medulla - contains tissue macrophages embedded in a coarse fibrous mesh

72. Spleen Tongue-shaped organ located on the left side of the abdomen
Near the stomach in simple-stomached animals
Near the rumen in ruminants
Largest lymphoid organ in the body

73. Spleen
Covered with a fibrous connective tissue capsule and smooth muscle
Capsule sends branches (trabeculae) into the soft tissue of the spleen
Trabeculae contain blood vessels, nerves, lymph vessels, and smooth muscle cells.
Trabeculae are very muscular in carnivores.
When the smooth muscle cells contract, they squeeze blood out of the spleen and back into circulation.

74. Spleen Interior of the spleen is divided into white pulp and red pulp
White pulp: localized areas of lymphoid tissue
Red pulp: blood vessels, tissue macrophages, and blood sinuses (storage)

75. Spleen Functions Blood storage in the red pulp
Removal of foreign material from circulation by the tissue macrophages in the red pulp
Removal of dead, dying and abnormal red blood cells by the tissue macrophages in the red pulp
Lymphocyte cloning in the white pulp during an immune response

76. Thymus
Lymphoid organ located in the caudal neck and cranial thoracic region on either side of the trachea.
Most prominent in young animals
Processes thymocyctes and T-cells

77. Tonsils Nodules of peripheral lymphoid tissue
Not covered with a capsule
Found close to mucosal surfaces all over the body
Pharynx, larynx, intestine, prepuce, and vagina
Tonsils in pharyngeal region prevent spread of infection into the respiratory or digestive systems
Located at the beginning of the lymph drainage system, not along the lymph vessels like lymph nodes

78. Gut Associated Lymph Tissue (GALT)
Lymphoid tissue found in the intestinal mucosa and submucosa
Largest lymphoid organ in the body
GALT is classified as both central and peripheral lymphoid tissue.

79. The Immune System Actions:
Phagocytosis and destruction of foreign cells
Lysis of foreign cell membranes
Inactivation of pathogenic organisms or chemical substances
Precipitation or agglutination of cells or molecules

80. Nonspecific Immunity Mechanical barriers - skin and mucous membranes
Chemical barriers (e.g., hydrochloric acid in the gastric mucosa)
Inflammatory response - tissue damage provokes release of chemical mediators (e.g., histamine) and other chemotactic factors
Phagocytosis by neutrophils, monocytes and tissue macrophages

81. Nonspecific Immunity
Natural Killer cells - come in direct contact with tumor cells and tissue cells that have been invaded by viruses and destroy them
Interferon – protein produced by a cell after it has been infected by a virus; inhibits further development and spread of the virus

82. Nonspecific Immunity
Complement - group of enzymes in plasma that can be activated by the attachment of an antibody to an antigen
Complement fixation - cascade of reactions that results in antigen lysis

83. Specific Immunity
B cells that produce antibodies or direct other cells to attack the antigen
T cells that attack more directly
Response is initiated as a reaction to the epitope on the invading cell’s wall
Lymphocytes primarily involved but may depend on the actions of other cells for activation

84. Table 9-3. Humoral Immune Response versus Cell Mediated Immune Response
Cell type involved
B cell that transforms into a plasma cell after antigenic stimulation
T lymphocyte that transforms into cytotoxic T cell, helper T cell or supressor T cell after antigenic stimulation
Substance produced
Immunoglobulins (antibodies)
Lymphokines
Cellular mobility
B cells and plasma cells stay in the lymphoid tissue. Antibodies are released into plasma.
T cells can enter circulation and travel to the site where an antigen entered the body
Memory cells produced?
Yes

85. Types of Immunoglobulins (Ig)
IgG - first Ig made during first exposure to an antigen
IgM - made when animal exposed to an antigen for a long time or when exposed to the antigen for the second time
IgA - can leave blood and enter tissue fluids; plays a role in protecting mucosal surfaces (e.g., intestinal tract and lungs)
IgE - associated with an allergic response
IgD - function is unknown

86. Passive Immunity Animal receives preformed antibodies
Antibodies produced by a mother that are passed to a fetus transplacentally
Ingestion of colostrum (antibody-rich first milk produced)
Antibodies produced by another animal and given to a sick animal (e.g., administration of tetanus antitoxin)
No memory cells produced

87. Active Immunity
Exposure to antigen that triggers animal’s own immune response
Memory T- or B cells are produced
Immunization: activate animal’s own immune systems
Vaccines contain epitope of the antigens
Killed or live-but-weakened (attenuated) antigens