LEUCOCYTES

LEUCOCYTES

White blood cells are vitally important in the disposal of damaged and ageing tissue and in the immune responses, which protect us from infections and cancer cell proliferation. The total blood white cell count is normally in the range 4 x 104 to 10 x 104 /L (may increase markedly during infection or inflammation).

LEUCOCYTE TYPESBased on their histological appearances, two morphological groups of leucocyte may be identified:1. Polymorphonuclear granulocytes have irregular, multilobed nuclei and a high density of cytoplasmic granules. Neutrophils, eosinophils and basophils all belong in this group.2. Lymphocytes and monocytes lack granules and have large, regular nuclei, and so they are classified as mononuclear agranulocytes.

LEUCOCYTE TYPES

• NEUTROPHILS comprise 60-70% of circulating leucocytes. They are highly mobile and can engulf debris or foreign organisms through the process of phagocythosis, trapping the target in a vesicle, which then fuses with a lysosome.

• EOSINOPHILS make up 1-4% of circulating leucocytes. They are phagocytic and are particularly involved in the destruction of parasitic worms but may also contribute to allergic responses.

• BASOPHILS generally account for under 0.5% of leucocytes. These phagocytes release histamine and heparin and are involved in allergic responses.

• LYMPHOCYTES are the only nonphagocytic white cells and represent 25-30% of blood leucocytes. They are central to specific immune defences within the body and can be subdivided into B and T lymphocytes.

• MONOCYTES constitute 2-5% of leucocytes. They have the greatest phagocytic potential of all body cells. The monocyte/macrophage system forms the core of the reticuloendothelial system.

LEUCOCYTE PRODUCTION

Leucocytes originate from the pluripotential stem cells in the bone marrow, which divide and mature giving two separate leucocyte stem cell lines.

THE MYELOID LINE

This gives rise to the three types of granulocyte as well as monocytes and macrophages. These cells all have important phagocytic roles.

The myeloid stem cell line also produces large multinucleate megakaryocytes from which platelets are derived.

THE LYMPHOID LINE

• From this stem cell line the lymphocytes are produced.

• B cells mature in the marrow before being distributed to the lymphoid tissues of the body (the lymph nodes, spleen, thymus and Peyer's patches in the intestinal submucosa).

• T lymphocyte precursor cells are believed to migrate initially to the thymus, where they mature fully before being redistributed to other lymphoid sites.

• Lymphocytes can replicate and develop further within the lymphoid tissues of the body and there is a continuous recirculation of lymphocytes from blood to lymph and back again.

FUNCTIONS OF GRANULOSYTES:

NEUTROPHILS • The opsonised bacteria, i.e., bacteria coated with

antibodies, then attach to the receptors on the neutrophils, which are now activated. The activated neutrophils engulf the bacteria (phagocytosis) and release different enzymes into the phagocytic vesicle, which kill the bacteria and then digest it.

• The enzyme NADPH oxidase from the cell membrane leads to formation of O- (super oxide) and H2O2 is formed from O2 by the action of super oxide dismutase. Both the chemicals are highly toxic and are used for killing the bacteria.

FUNCTIONS OF GRANULOSYTES: EOSINOPHILS

• Eosinophils contain a major basic protein (MBP), which damages the larvae of parasites.

• There is one eosinophilic cation protein which neutralises heparin.

• The sulphatase-B present in eosinophil inactivates the slow reacting substance (SRS) released from mast cells and prevent anaphylaxis (anti-allergic action).

• Histaminase from eosinophils destroy the substances released from mast cells. Eosinophils also contain peroxidase.

• They are motile and phagocytic (take up antigen-antibody complexes). Chemotaxis is also shown by the eosinophils.

FUNCTIONS OF GRANULOSYTES: BASOPHILS

• Basophils like the mast cells have histamine in their granules and when the histamine releasing factors (e.g., IgE-antigen complex) bind to them, they release the histamine resulting in immediate hypersensitivity reaction like urticaria, rhinitis, anaphylactic reactions, etc.

• They contain heparin, proteases and some other mediators of inflammation.

• They are motile and phagocytic.

FUNCTIONS OF AGRANULOSYTES: LYMPHOCYTES

• Lymphocytes are involved in the very important defence mechanism, called immunity.

• The B lymphocytes are responsible for humoral immunity and T lymphocytes are for cell mediated immunity.

• Lymphokines secreted by some T cells have functions of defence.

• Lymphocytes produce antibodies, interferons, lymphotoxin, and they also control the immune mechanism by the helper and suppressor T-cells.

FUNCTIONS OF AGRANULOSYTES: MONOCYTES

• Monocytes after a short stay in blood enter in the tissues to be converted into macrophages, where they take up specific positions and wait for the prey.

• In the tissues they are matured further, which enable them to process antigen for presentation to the lymphocytes: secretion of interleukin-1; phagocytosis of microorganism, dead tissue; formation of bilirubin.

• The monokines (interleukin-1) secreted by monocytes, stimulate T cells, take part in inflammation, act as pyrogen (fever producing).

• They phagocytose and kill like neturophils (their digesting ability is less than the neutrophils).

Type % Main targets Nucleus Granules Lifetime

Neutrophil 65bacteria fungi

multi-lobed

fine,faintly pink

10h in blooddays in spleenand other tissue.

Eosinophil 4parasites in allergic reactions

bi-lobedfull of pink-orange(stained)

Basophil 1 in allergic reactions bi- or tri-lobed

large blue

Lymphocyte 25

B cells: various pathogens T cells: CD4+ (helper): intracellular bacteria. CD8+ cytotoxic T cells: virus-infected and tumor cells. γδ T cells: Natural killer: virus-infected and tumor cells.

Deeplystaining,eccentric

only NK-cells weeks to years

Monocyte6 various

Kidneyshaped

none

Macrophage days

BLOOD WHITE CELL COUNT• Total count (TC): TC of WBC means the total

number of WBCs present per unit volume of whole blood. Normal value is 4 to 10 thousand/cmm (when this value is > 10000, it is called leucocytosis and a value < 4000 is called leucopenia).

• Differential count (DC): DC of WBC means the number of individual types out of one hundred leucocytes, e.g., DC of WBC—N-67, E-2, B-l, L-25, M-5 means, out of 100 WBC examined 67 are neutrophils, 2 are eosinophils, 1 is basophil, 25 are lymphocytes and 5 are monocytes (DC is the percentage of different WBCs of the total).

• The normal value is as follows : Neutrophil 60 to 70%, eosinophil 1 to 4%, basophil 0 to 1%, lymphocyte 25 to 30% and monocytes 5 to 10%.

BLOOD WHITE CELL COUNT

BLOOD WHITE CELL COUNT

There is an increase in the blood white cell count (a leucocytosis), most of the increase being the result of extra neutrophils (a neutrophilia). This reflects both the rapid mobilization of neutrophils already present in the bone marrow and an increased rate of production in the marrow.

LEUCOGRAMME EXAMPLE

TotalWBC

x 109

/L

Granulocytes(%)

Agranulocytes (%)

Neutrophils Eosino-phils

Baso-phils

Lympho- cytes

Mono-cytesMyelo

-cytesMeta-myelo-cytes

Bands Seg-men-ted

4.0 – 9.0

0 0-1 1-6 47-72 0.5-5 0-1 19-37 3-11

IMMUNE RESPONSES

• The immune defences of the body are classically subdivided into those, which are nonspecific and innate, and those, which are specific and acquired.

• There are many points of interaction between the two systems. For example, lymphocytes only produce specific antibodies against foreign molecules when these antigens are first processed by nonspecific phagocytic cells such as macrophages. At the same time, antibodies lead to antigen removal by amplifying preexisting, nonspecific responses. These two elements of immunity are, therefore, highly interdependent.

NONSPECIFIC OR INNATE IMMUNITY

• This depends on interrelated defence mechanisms, which act against any foreign or abnormal cell, i.e. they are nonspecific.

• They are also said to be innate since they do not depend on previous exposure to a particular organism.

• Nonspecific immune mechanisms include physical barriers to infection, inflammation, complement activation and natural killer cell activity.

• Inflammation is a set of local cellular and vascular responses to tissue damage or infection, which accelerates the destruction and phagocytic removal of invading organisms and debris.

MACROPHAGES

RETICULOENDOTHELIAL SYSTEM

• This phagocytic system is almost synonymous with the tissue macrophage system. Mobile macrophages rove freely through connective tissues, but many more remain relatively fixed in a given region.

• When activated, these will also become mobile, being chemotactically attracted to local sites of infection or damage.

• There are dense aggregations of macrophage-type cells within the reticular tissue of the lymph nodes, spleen and bone marrow.

• Alveolar macrophages in the lung and the phagocytic Kupffer cells in the liver, along with microglial cells in nervous tissue, are also regarded as part of the reticuloendothelial system, as are the blood monocytes.

PHAGOCYTIC RESPONSESIN INFLAMMATION

• Tissue macrophages adjacent to a site of bacterial invasion become mobile and phagocytically active.

• Chemicals released from injured and infected cells (chemotaxins) act as attractants for these cells, directing their movements towards the damaged area (chemotaxis).

• Local macrophages are soon reinforced by the migration of blood neutrophils and monocytes into the region.

• These stick to the endothelium of capillaries in the affected area (leucocyte margination) and then invaginate themselves through the clefts between the endothelial cells using active amoeboid movements (diapedesis).

• The blood-borne phagocytes escape into the tissues where they are chemotactically attracted to assist in removal of infectious or toxic agents and tissue debris.

MECHANISM OF PHAGOCYTOSISSteps of a macrophage ingesting a pathogen:a. Ingestion through phagocytosis, a phagosome is formed.b. The fusion of lysosomes with the phagosome creates a phagolysosome; the pathogen is broken down by enzymes.c. Waste material is expelled or assimilated.

ACTIVATION OF THE COMPLEMENT SYSTEM

Foreign cells, especially certain bacteria, carry surface molecules, which activate the complement system of plasma proteins. The system is organized as a cascade so that each activated component activates the next in the sequence. Nonspecific activation is referred to as activation by the alternate pathway, distinguishing it from the classical pathway to activation, which requires antibody.

COMPLEMENT FUNCTION

• DIRECT CELL KILLING. The activated forms of the last five components in the system combine to form a protein called the membrane attack complex. This becomes inserted in the plasma membrane of the invading cell, where it forms a large pore. When the density of such pores is high, cell lysis results.

• PHAGOCYTOSIS. Complement activation produces a series of biologically active complement fragments, which increase the efficiency of phagocytosis.

• Phagocytosis involves several mechanisms:– OPSONIZATION makes target cells more susceptible to phagocytosis.

Complement fragments, which act as opsonins, bind to the surface of bacteria. Phagocytic cells, which carry receptors for these fragments, become attached to them and this increases the efficiency of bacterial phagocytosis.

– CHEMOTAXIS by complement fragments attracts more phagocytic cells to an infected or damaged region.

– VASODILATATION occurs and capillary permeability is increased following complement activation, thus amplifying the inflammatory response.

SPECIFIC OR ACQUIRED IMMUNITY

• Specific immunity refers to a number of mechanisms whereby susceptibility to infection by a particular organism is greatly reduced following initial exposure to it.

• This is clearly demonstrated with measles, mumps, rubella and other childhood infections, in which the immunity acquired following a first infection usually protects us, if we are exposed to the same organism later in life.

• Clinical infection is not necessary; vaccination uses weakened or killed pathogens to stimulate immunity without producing illness. In both infection and vaccination the protection acquired is highly specific.

ANTIBODY-MEDIATED RESPONSES

• B lymphocyte activation is the first step in antibody production and it occurs, when antigens are processed by macrophages and then presented to B lymphocytes. These cells carry surface immunoglobulins, or antibodies, and when these bind to an antigen the lymphocyte is activated.

• Each B cell will only bind with one type of antigen and this leads to production of circulating antibodies with the same specificity. Specific antibody only appears in body fluids once the relevant B cell has been activated by antigen binding, after which it multiplies and the daughter cells become transformed into plasma cells and memory cells.

• Plasma cells act as factories for the relevant antibody, which they secrete in large amounts. Memory cells are identical with the parent lymphocyte and they greatly expand the reserve of cells capable of being activated during subsequent antigen exposure.

PRIMARY AND SECONDARY RESPONSES

• On first exposure to an antigen, the immune response takes some weeks to develop fully. This is referred to as the primary response and, although it can limit the duration of an infection, it is generally too slow to prevent it altogether.

• Antibody levels fall again as time passes, but a second exposure to the same antigen produces a much more rapid antibody response which peaks at a higher concentration and declines more slowly. This secondary response depends on the memory cells formed following initial B lymphocyte activation.

• These provide a reservoir of lymphocytes, which can be rapidly activated during future exposure to the relevant antigen. This amplified antibody response may overwhelm a potential pathogen before it can cause the symptoms of infection, i.e. it provides acquired, antibody-mediated immunity.

ANTIBODY FUNCTION

• Antibodies bind to antigens, thus promoting their destruction through a range of different mechanisms.

• 1. Targeting and amplification of nonspecific immunity is the major strategy employed by antibody-mediated immunity to provide protection against specific pathogens. IgG and IgM class antibodies are particularly important in promoting bacterial lysis and phagocytosis in this way.

• Antibodies activate the complement system by the classical pathway. The Fc region of antigen-bound antibodies can bind and cleave the C1 complement component. This leads to activation of the entire complement cascade.

• Antibodies can act as opsonins, using receptors for the Fc region to bind phagocytes to antigens.

• Antibody-coated cells are more likely to be attacked by natural killer cells.• 2. Agglutination refers to the clumping together of bacteria or foreign cells into large-scale

lattices held together by antibody linkages. This is possible because all antibodies have more than one antigen-binding site and so can interconnect target cells, physically hindering the spread of infectious agents and increasing the likelihood of phagocytosis. IgM antibodies are the most effective because their pentameric structure has 10 binding sites per molecule. Agglutination reactions are used in blood group testing.

• 3. Neutralization of toxins and inactivation of some viruses may result from immunoglobulins cloaking biologically active sites. This is how antisera to snake and insect bites work.

CELL-MEDIATED IMMUNITY

This relies on T lymphocytes, which do not manufacture circulating antibodies. It is particularly important in combating viral and fungal infections as well as in immune responses against potential cancer cells.T lymphocytes are activated by exposure to a foreign antigen, which is identified and bound by specific surface receptors. This recognition step only occurs, however, if the antigen is closely associated with other cell surface antigens, which the T lymphocyte recognises as normal self-antigens (i.e. belonging to that individual).

T LYMPHOCYTE FUNCTION

There are three main types of T lymphocyte:– CYTOTOXIC T LYMPHOCYTES can lyse cells carrying the

antigen to which they are sensitive.– HELPER T LYMPHOCYTES (also called Th cells) are vital in

both antibody- and cell-mediated immunity, but they have no direct effects on foreign antigens or cells. Like B lymphocytes, they are activated by macrophage-processed antigens, rather than cell surface antigens as is the case for other T lymphocytes. Once stimulated by the appropriate antigen, they release a number of lymphokines, which stimulate other immune cells. This increases macrophage activity as well as promoting multiplication and activation of T and B lymphocytes.

– SUPPRESSOR T LYMPHOCYTES inhibit lymphocyte function. This may provide a mechanism for the control of the immune response, which reduces the risk of immune damage to normal body cells.

ACTIVE AND PASSIVE IMMUNITY

Activation of lymphocytes provides active defence against infection. If antibodies are injected or absorbed into the body these will also protect us from a given antigen. This is called passive immunity but, unlike active immunity, it only provides short-term protection (a few months) since there is no production of memory cells.A physiological example of passive immunity is the protection a newborn baby gains from the maternal immunoglobulins in its body (IgG crosses the placenta and IgA is present in breast milk). As these antibodies are degraded over the first 2-3 months of life, the level of passive protection wanes, but by this time the infant's immune system is mature enough to mount active responses.In adult life, injections of immunoglobulin may be used to give short-term passive protection, e.g. against some forms of viral hepatitis.