Introduction...of their multilobed nucleus. Lymphocytes and monocytes are called agranulocytes because their cytoplasm not contains prominent granules. They are found mainly in tissues

Medical Biology Blood Cells

Dr. Mohammed Hussein Assi MBChB – MSc – PhD – DCH (UK) – MRCPCH

Introduction

The blood is a mixture of cellular elements, fluid, proteins and metabolites

Blood has four major elements:1. Red blood cells (erythrocytes) transport oxygen from the lungs to the peripheral tissues2. White blood cells (leukocytes) have a defensive role.3. Platelets (thrombocytes) are important in haemostasis.4. Plasma is the proteinaceous solution in which the above-mentioned cells circulate, and

carries nutrients, metabolites, antibodies, hormones, proteins of the blood clotting system andother molecules throughout the body.

Bone Marrow Derived Stem Cells

The bone marrow contains at least two kinds of stem cells

One population, called ‘haematopoietic stem cells’ can form all of the types of blood cells in thebody. These are the cells useful clinically for bone marrow transplants.A second population is called ‘bone marrow stromal stem cells’ or ‘mesenchymal stem cells’.These nonhaematopoietic stem cells make up a small proportion of the stromal cell population inthe bone marrow, and can generate bone, cartilage, fat, cells that support the formation of bloodand fibrous connective tissue. These cells are under study as sources of different cell types forregenerative medicine.

Methods of Studying the Blood Cells

Blood is readily accessible by sampling with a needle and syringe. The name for thetwo main types of cells in blood derives from what is seen if blood is prevented fromclotting and left to stand in a tube. Blood settles into several layers: a thick layer ofclear plasma is seen at the top of the tube, beneath which is a very thin layer of whitematerial on top of a thick layer of red material. The cells in the white material arecalled ‘white cells’ and the cells in the red material ‘red cells’.The usual way to look at blood is to make a very thin smear on a glass slide. Themain staining method used (Romanovsky stain) involves applying several dyeswhich have an affinity for different cellular constituents. Under the microscope it ispossible to count the proportion of different cell types in blood and, as it has beenfound that this reflects disease processes, a blood count is a valuable diagnostic tool.In modern laboratory practice, routine counting of cells in blood is doneelectronically on preparations of cells in suspension. Smears of blood are stillexamined for morphological abnormalities of cells.



Red Blood CellsRed blood cells are highly deformable and are specialized for carrying oxygen

The red blood cells are responsible for oxygen transport. Red cells in peripheral blood smearsappear as rounded, bright pink-stained cells. They are 6.5–8.5 µm in diameter and have abiconcave shape, appearing paler in the centre and darker at the periphery. The biconcave shapemaximizes their surface area/volume ratio and thereby maximizes oxygen exchange. The brightpink colour (acidophilia) is due to the content of oxygen-carrying haemoglobin, which binds theacidic eosin dye used in staining. Red cells do not have a nucleus, as this is lost duringformation.The normal concentration of erythrocytes in blood is approximately 3.9–5.5 million permicroliter in women and 4.1–6 million per microliter in men.

Ultrastructurally, red cells have a cell membranewhich surrounds an electron-dense cytoplasmthat contains haemoglobin. There are nodiscernible organelles as these have been lostduring differentiation. Despite a lack oforganelles, red cells are metabolically active andderive energy by anaerobic metabolism ofglucose, and through ATP generation by thehexose monophosphate shunt.Functionally, red cells are highly deformable andare able to squeeze through small blood vesselsdown to 3–4 µm in diameter. The cell membraneis braced by an actin/spectrin-containingcytoskeletal meshwork, which is largelyresponsible for maintaining the distinctivebiconcave shape.



Red cells have a limited lifespan and are eventually destroyed in the spleen

Red cells have a lifespan of 100–120 days in the circulation. The mature red cell is unable tosynthesize new enzymes to replace those lost during normal metabolic processes. Diminishingefficiency of ion pumping mechanisms is probably the main factor in red cell ageing, the cellbecoming progressively less deformable until it is unable to negotiate the splenicmicrocirculation and is removed by phagocytosis.

Key Facts: RBCs• Biconcave shape for high surface area/volume ratio• Main function is oxygen and carbon dioxide transport• Contain haemoglobin• Have no cell organelles• Cell membrane is braced by an actin/spectrin-containing cytoskeleton which maintains shape.

Hereditary spherocytosisHereditary spherocytosis is caused by an abnormal arrangement of the internal cytoskeleton of redcells. Normally the internal surface of the cell membrane is braced by cytoskeletal proteins viainteractions between ankyrin and spectri. In hereditary spherocytosis, a defect in spectrin or theankyrin binding of spectrin is the main underlying abnormality. As a result, the red cell membrane isnot braced and is easily deformed.In hereditary spherocytosis, red cells do not form their normal biconcave disc shape, but appearround and convex. They are abnormally brittle and less deformable than normal red cells, and so donot pass easily through the splenic microcirculation. They are trapped there and rapidly destroyed inlarge numbers; this excessive breakdown of red blood cells is called haemolysis.

Hypochromic, microcytic anaemiaThe most common cause of anemia is deficiency of iron, which is essential for the formation ofhemoglobin. Red cells are released into the circulation containing much less hemoglobin than normal,and are therefore pale staining (hypochromic) and small (microcytic)

Sickle Cell AnaemiaPoint mutations in the haemoglobin gene may cause abnormal red cells. Sickle cell anaemia is causedby such a mutation, leading to precipitation of haemoglobin in red cells subject to hypoxia, whichcauses a sickle shape instead of the biconcave disc. sickled cells become disrupted and can also blockblood vessels



White Blood CellsThere are five main types of white blood cells

White cells use the blood for transport from the bone marrow to their major sites of activity. Themajority of the functions of white blood cells take place when they leave the circulation to entertissues.The total number of WBCs in peripheral blood is normally 4.0–11.0 × 109/L, (4000-11000/µL)There are five types of WBCs, and their names and relative proportions in the circulation are asfollows:

• Neutrophils 50–70% (~ 60%)• Lymphocytes 20–40% (~ 30%)• Monocytes 2–8% (~ 6%)• Eosinophils 1-4% (~ 3%)• Basophils 0-2% (~ 1%)

Neutrophils, eosinophils and basophils are known as granulocytes because their cytoplasmcontains prominent granules, and may also be referred to as myeloid cells because of their originfrom bone marrow.Neutrophils are also commonly called polymorphonuclear leukocytes (or polymorphs) becauseof their multilobed nucleus.Lymphocytes and monocytes are called agranulocytes because their cytoplasm not containsprominent granules. They are found mainly in tissues such as lymph nodes and spleen. In thetissues, monocytes transform into macrophages.



NeutrophilsNeutrophils are the most common type of white blood cell

Neutrophils are the most abundant of the circulating white cells. They circulate in a resting statebut, with appropriate activation, leave the blood and enter tissues, where they become highlymotile, phagocytic cells. Their primary function is to ingest and destroy invadingmicroorganisms in tissues.They play a central role in the early stages of the acute inflammatory response to tissue injuryand are the major constituent of pus.There are normally 1.5–10 × 109 /L neutrophils in peripheral blood; a rise to above 10 × 109/L iscalled neutrophilia and is usually an indication of bacterial infection or tissue necrosis (e.g.myocardial infarction).A reduction in numbers of circulating neutrophils below 1.5 × 109 /L is called neutropenia oragranulocytosis; this reduction in numbers can be due to decreased production in the bonemarrow or to increased destruction in the tissues. The danger of persistent neutropenia is that thepatient becomes very vulnerable to severe bacterial infections.

Neutrophils contain three types of granules

Neutrophil cytoplasm contains three types (i.e. primary, secondary and tertiary) of membrane-bound vesicles (granules).o Primary granules are similar to

lysosomes in other cells. They are thefirst granules to appear duringneutrophil formation, but as the cellmatures, their number falls withrespect to secondary granules, makingthem difficult to see with lightmicroscopy. Primary granules containacid hydrolases, and antibacterial anddigestive substances, most notablymyeloperoxidase, which can bedetected by the peroxidase stain.Myeloperoxidase is therefore a usefullight microscopic marker not only forthese granules, but also in establishing cell lineage in the diagnosis of leukaemias.

o Secondary granules are specific to neutrophils and twice as numerous as primary granules,and smaller than them, they are barely visible by light microscopy. They contain substancesinvolved in the mobilization of inflammatory mediators and complement activation. Thesesubstances are secreted into the extracellular environment.

o Tertiary granules have only recently been described and contain enzymes (e.g. gelatinase)secreted into the extracellular environment. They also insert some glycoproteins into cellmembranes, and this may promote cellular adhesion and hence may be involved in thephagocytic process.



The neutrophil nucleus has several lobes

The characteristic neutrophil nucleus is composed of 2–5 distinct lobes, joined to one another byfine strands of nuclear material, the lobulation developing with cellular maturity. The chromatinis highly condensed, reflecting a low degree of protein synthesis.

Neutrophils migrate into areas of tissue damage, where they have a defensive role

Phagocytosis is the process whereby cells ingest extracellular particles for destruction.Neutrophils have a role in the phagocytosis of bacteria and dead cells. To reach an area ofinfection or tissue damage, neutrophils leave the circulation by adhering to endothelial cellsusing adhesion molecules expressed in response to local secretion of cytokines, and movethrough the endothelium and basement membrane. Once in the extravascular tissue, neutrophilsrespond to chemicals (chemotaxins), moving towards the highest concentration. Neutrophilstypically die soon after phagocytosis, as this highly energy-dependent process uses up theirglycogen reserve. When they die, their lysosomal enzymes are released into the extracellularspace, causing liquefaction of adjacent tissue. The collection of dead neutrophils, tissue fluid anddebris is termed ‘pus’.

Key Facts: Neutrophils One of the myeloid series of white blood cells One of the granulocyte type of white blood cells Main role in phagocytosis and bacterial killing Contains three types of granule in cytoplasm Cells marked by myeloperoxidase Increase in number in blood in bacterial infection and inflammation (‘neutrophilia’).



EosinophilsEosinophils have a bilobed nucleus and acidophilic granules

Eosinophils have a bilobed nucleus and contain strongly eosinophilic granules. They arephagocytic, with a particular affinity for antigen–antibody complexes, but have less microbicidalactivity than neutrophils. After production in the bone marrow, eosinophils are stored forapproximately 8 days before release into the circulation, where they remain for about 8 h beforepreferentially migrating to the skin, lungs and GIT where they reside for about 8 days. They mayenter lung and gut secretions via lymphatics or by direct migration.Circulating eosinophil numbers show a marked diurnal variation, being maximal in the morningand minimal in the afternoon. They increase greatly in many types of parasitic infestation, andprotection against parasitic disease appears to be one of their main functions.Tissue (and sometimes blood) eosinophil numbers are also increased in certain allergic states, forexample in the nasal and bronchial mucosae in hay fever and asthma and in adverse reactions todrugs. Eosinophils do not usually re-enter the circulation after tissue migration.

BasophilsBasophils and mast cells have common lineage and similar functions

Basophils are the least common white cell in the blood. They are characterized by large,intensely basophilic, cytoplasmic granules, with two- to three-lobed nucleus (usually obscuredby the cytoplasmic granules)They share a common lineage with tissue mast cells, with which they have many structural andfunctional similarities.The granules of basophils and mast cells contain the proteoglycans heparin and chondroitinsulfate, together with histamine and leukotriene 3.Both basophils and mast cells have highly specific membrane receptors for the IgE produced inresponse to allergens. Exposure to allergens results in rapid exocytosis of their granules, therebyreleasing histamine and other vasoactive mediators, and resulting in an immediatehypersensitivity (anaphylactoid) reaction. Such a reaction causes allergic rhinitis (hay fever),some forms of asthma, urticaria and anaphylaxis.Mast cells reside in support tissues, especially those beneath epithelia, around blood vessels andlining serous cavities. They are long-lived (weeks to months) and can proliferate in the tissues. Inmucosae, but not in other sites, proliferation appears to depend on interaction with Tlymphocytes.



MonocytesMonocytes are part of a cell network, the monocyte–macrophage system

Monocytes are the blood- and bone marrow-located precursors of the macrophages found intissues and lymphoid organs, and are members of a single functional unit, the monocyte–macrophage system (mononuclear phagocyte system). This system consists of:

1. The bone marrow precursors (monoblasts and promonocytes)2. Circulating monocytes3. Tissue macrophages, both free and fixed (histiocytes)4. Kupffer cells of the liver5. Sinus lining cells of the spleen and lymph nodes6. Pulmonary alveolar macrophages7. Free macrophages in synovial, pleural and peritoneal fluid8. Dendritic antigen-presenting cells

Monocytes are large motile, phagocytic cells. In blood films, they often have vacuolatedcytoplasm. Each cell has a large, elongated nucleus, often assuming kidney or horseshoe shapes.Ultrastructurally, monocyte cytoplasm contains numerous small lysosomal granules andcytoplasmic vacuoles.The granules are of two types. One type represents primary lysosomes and is analogous to theprimary granules of neutrophils. The content of the other group of granules is less certain.Numerous small pseudopodia extend from the monocyte, reflecting its phagocytic ability andamoeboid movement.Monocytes respond chemotactically to the presence of necrotic material, invadingmicroorganisms and inflammation, and leave the blood to enter the tissues, where they are calledmacrophages.

LymphocytesLymphocytes are responsible for generating specific immune responses

In adults and older children lymphocytes are the second most numerous white cell in the blood,their numbers increasing in response to viral infections; they are the most numerous white cell inyoung children.Most circulating lymphocytes are small, but about 3% are large. Their nuclei are ovoid, with thedense chromatin typical of cells with little biosynthetic activity. There are two main types oflymphocyte, termed B and T cells, which perform different but linked roles in the generation ofspecific immune responses.The small mature lymphocytes circulating in the blood emigrate into tissues and into specialorgans of the immune system. They are responsible for immune surveillance, constantlysampling their environment for foreign material. Lymphocytes then transform into active cells,mediating the immune responses, particularly in specialized lymphoid tissues.Large lymphocytes may be seen in the blood, representing such activated lymphocytes en routeto the tissues.



Plasma cellsPlasma cells are formed from B lymphocytes and secrete immunoglobulin

Plasma cells are a differentiated form of B lymphocyte and actively synthesize immunoglobulin.Plasma cells form a small population in normal marrow and are usually seen in support tissuesand specialized lymphoid organs.In health, they are not found in the blood.Plasma cells are large and have an eccentrically located, round or oval nucleus with thechromatin coarsely clumped in a characteristic cartwheel or clock-face pattern, reflecting activetranscription. Their cytoplasm is moderately basophilic owing to its large content of ribosomalRNA in abundant rough endoplasmic reticulum, required to manufacture the immunoglobulinprotein. A well-developed Golgi complex displaces the nucleus and is visible as a paranuclearhaloor pale zone (Golgi zone).

WHITE BLOOD CELL ABNORMALITIES Analysis of the peripheral blood is an important part of diagnosing disease in sick patients. Part of a

full blood count lists the numbers of white cells in the peripheral blood. The number of circulatingwhite blood cells is altered in many disease processes and changes in different cell types areassociated with different diseases.

Increased numbers of white cells appear in the peripheral blood in a variety of disorders and providea useful clue to underlying disease. For example:• An increase of circulating neutrophils in bacterial infections (neutrophilia)• An increase of circulating eosinophils in parasitic infestations and some allergies (eosinophilia)• An increase in circulating lymphocytes in certain viral infections (lymphocytosis)

Leukemias are classified according to the cell line involved (i.e. granulocytic, monocytic,lymphocytic) and also according to their degree of malignancy.

Golgi Zone



PlateletsPlatelets are small cell fragments derived from megakaryocytes and are important in haemostasis

Platelets (also called ‘thrombocytes’) are small, disc shaped anuclear structures that are formedby the cytoplasmic fragmentation of huge precursor cells (megakaryocytes) in the bone marrow.Platelets contain mitochondria, microtubules, glycogen granules, occasional Golgi elements andribosomes, as well as enzyme systems for aerobic and anaerobic respiration.Their most conspicuous organelles, however, are their granules, of which there are three types:1. α granules are variable in size and shape and contain: PF4 (platelet factor 4), vWF(von

Willebrand Factor and platelet derived growth factor, fibrinogen, fibronectin, and others.2. Dense granules (δ granules) are electron-dense and contain small molecules such as ADP,

serotonin and calcium. These components are critical for platelet activation andvasoconstriction

3. Lysosomes are membrane-bound vesicles containing lysosomal enzymes (e.g. cathepsins andhexosaminidase).

Platelets aggregate together and degranulate in haemostasis

Platelets are essential to normal haemostasis, undergoing aggregation in the process.Haemostasis is achieved by the following steps:After loss of the lining endothelium of blood vessels, platelets adhere to exposed collagen byinteracting with glycoprotein receptors for von Willebrand Factor attached to it.Platelet actin, myosin and microtubules cause reversible platelet moulding and adhesion along abroad surface. They then irreversibly release the contents of their granules through thecanalicular system, in a secretion reaction, and synthesize thromboxane.Thromboxane, ADP and Ca2+ ions mediate adhesion of other platelets.Platelet phospholipids (with Ca ions) activate the blood clotting cascade, leading to the formationof fibrin.

PLATELET DISORDERS• The normal platelet count is 150-400× 109/L• Severe reduction in the number of platelets well below 150 × 109/L in circulating blood is

called ‘thrombocytopenia’. It causes spontaneous bleeding because of the failure of plateletsto plug microscopic breaches in vessel walls, resulting from minor trauma.

• The presence of excessive numbers of circulating platelets is called ‘thrombocytosis’, andfrequently occurs as a transient phenomenon when a general burst of bone marrowhyperactivity follows acute blood loss. A more persistent thrombocytosis occurs as a part ofso-called ‘myeloproliferative disorder’, an uncontrolled clonal proliferation of the blood-forming cell colonies in the bone marrow.

• Thrombocytosis is an important predisposing factor in the development of pathologicalthrombosis.

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Introduction...of their multilobed nucleus. Lymphocytes and monocytes are called agranulocytes because their cytoplasm not contains prominent granules. They are found mainly in tissues