Hematology 1 MLS 113B LEC PDF

# HEMATOLOGY 1 MLS 113B LEC PRELIM - 2ND SEMESTER <3.d BLOOD Blood is a fluid connective tissue which transports substances from one part of the body to another. It provides nutrients and hormones to the tissues and removes their waste products. The image shows a schematic of blood composition as...

# HEMATOLOGY 1 MLS 113B LEC PRELIM - 2ND SEMESTER <3.d BLOOD Blood is a fluid connective tissue which transports substances from one part of the body to another. It provides nutrients and hormones to the tissues and removes their waste products. The image shows a schematic of blood composition as well as some blood cells: - Blood is composed of 55% plasma and 45% cells. - Plasma consists of 92% water, 7% proteins and 2% other solutes. - Proteins include: Albumin, globulin and coagulation factors - Globulins are divided into $\alpha$, $\beta$ an $\gamma$ globulins. - Cells include: RBCS, WBCS, PLT Cells derive from two types of stem cells: - Pluripotent stem cells: * Lymphoid stem cells which differentiate into B lymphocytes, T lymphocytes and natural killer cells. * Myeloid stem cells differentiate into: monoblasts, myeloblasts, proerythroblasts, and megakaryoblasts. - Myeloblasts differentiate into granulocytes, which are neutrophilic, eosinophilic and basophilic. - Monoblasts differentiate into monocytes. - Proerythroblasts differentiate into erythrocytes. - Megakaryoblasts differentiate into megakaryocytes which give rise to platelets. The image also shows a red blood and white blood cell, and they are aproximately 7.5 and 2-4 microns in size, respectively. The white is cell is 10-14 microns. | | | | :---- | :-------------------- | | HSC | Location: Bone marrow | | | Characteristics: | | :----------------- | :----------------------------------- | | | Multipotent and self-renewing. | | | Can differentiate into all blood cell lineages (myeloid and lymphoid) | | 2. CMP | HSCs differentiate into CMPs, which are progenitors for all myeloid cells including erythrocytes, granulocytes, monocytes, and megakaryocytes. | | 3. MEP | CMPs give rise to MEPs, which are committed to the erythroid and megakaryocytic linkages. | | 4. BFU-E | MEPs differentiate into BFU-Es under the influence of erythropoietin (EPO) stem cell factor (SCF), and other growth factors. | | | BFU-Es are the earlist erythroid-specific progenitors. | | 5. CFU-E | BFU-Es mature into CFU-Es, which are highly sensitive to erythropoietin. | | Pronormoblast | Rubriblast | Proerythroblast | First morphologically recognizable erythroid precursor | | :-------------------------------------------- | :------------------------------------------ | :-------------------------------------------- | :------------------------------------------------------- | | | | | Large nucleus with fine chromatin. | | | | | High RNA content (cytoplasm appears basophilic under a microscope). | | Basophilic Normoblast | Prorubricyte | Basophilic Erythroblast | Cytoplasm is deeply basophilic due to active ribosome production. | | | | | Hemoglobin synthesis begins. | | Polychromatophilic Normoblast | Rubricyte | Polychromatophilic Erythroblast | Cytoplasm becomes grayish-blue as hemoglobin accumulates, and RNA content decreases. | | Orthochromic Normoblast | Metarubricyte | Orthochromatic Erythroblast (Normoblast) | Cytoplasm becomes pink due to high hemoglobin concentration. | | | | | Nucleus condenses and is eventually expelled. | | Polychromatophilic Erythrocyte | Polychromatophilic Erythrocyte | Reticulocyte | Immature erythrocyte released into circulation. | | | | | Still contains remnants of RNA, which give it a reticular (mesh-like) appearance under special stains. | | | | | Continues maturing in the bloodstream or spleen. | | Erythrocyte | Normoblast | Mature Erythrocyte | Fully differentiated, biconcave cell with no nucleus or organelles. | | | | | Specialized for oxygen and carbon dioxide transport hemoglobin | Megakaryocyte and the Platelet System * Mega - "Large, huge, big" * Karyo - "Nucleus" * Cyte - "cell" Endomitosis: * It is a specialized form of cell division in which the cell's nucleus undergoes multiple rounds of DNA replication without the usual division of the cell (cytokinesis). * As a result, the cell becomes polyploid, meaning it contains multiple copies of its genetic material in a single, enlarged nucleus. * Division of chromosomes not followed by nuclear division that results in an increased number of chromosomes in the cell. * Ploidy number of chromosomes occuring in the nucleus of a cell. The image shows the differentiation of hematopoietic stem cells into megakaryocytes and then platelet formation. The process starts with Hematopoietic stem cells differentiating to myeloid stem cells. Then, a megakaryoblast, a 2N cell differentiates into 4N, 8N, 16N and 32N cells. Eventually, these cells become megakaryocytes which form platelets that enter into the blood. Features of Endomitosis: 1. DNA Replication: * The DNA duplicates during the S phase, similar to regular mitosis. 2. No Cytokinesis: * Unlike typical mitosis, there is no separation of the cytoplasm or formation of two daughter cells. 3. Nuclear Division: * The nuclear envelope remains intact or reforms after replication, preventing full nuclear division. Megakaryocytes Definition: Megakaryocytes are large bone marrow cells responsible for the production of platelets. Development: * Derived from hematopoietic stem cells through the myeloid lineage. * Undergo a process called endomitosis, where the nucleus replicates without cytoplasmic division, leading to a polyploid nucleus. Structure: * Extremely large cells (50-100 um in diameter). * Contain a lobulated nucleus and abundant cytoplasm with demarcation membranes. Function: * Produce platelets by cytoplasmic fragmentation along proplatelet extensions. * Each megakaryocyte can release 1,000-3,000 platelets into the bloodstream. Platelets (Thrombocytes) Definition: Small, anucleate cell fragments derived from megakaryocytes. Structure: * Size: 2-3 um in diameter. * Lifespan: Approximately 7-10 days in circulation. * Contain alpha granules (e.g., fibrinogen, von Willebrand factor and dense granules (e.g., ADP, calcium): * Surface glycoproteins (e.g., GPIb-IX-V, GPllb/lla) mediate adhesion and aggregation. Function: * Hemostasis * Adhere to damaged blood vessels via von Willebrand factor and collagen. * Aggregate to form a primary platelet plug. * Secrete granule contents to recruit more platelets and stabilize the plug. Clot Formation: Provide a phospholipid surface for the assembly of clotting factors, accelerating the coagulation cascade. Tissue Repair: * Release growth factors (e.g, platelet derived growth factor [PDGF]) to promote healing. Regulation 1. Thrombopoiesis: * a. Regulated by thrombopoietin (TPO), primarily produced in the liver and kidney. * b. TPO binds to c-Mpl receptors on megakaryocytes and platelets. 2. Platelet Clearance: Senescent platelets are removed by the spleen and liver. 3. Feedback Mechanism: High platelet levels suppress TPO production, while low levels stimulate it. Clinical Relevance 1. Thrombocytopenia: a. Low platelet count (<150,000/uL). b. Causes: Decreased production (e.g., aplastic anemia), increased destruction (e.g., immune thrombocytopenic purpura), or sequestration (e.g., splenomegaly). c. Symptoms: Petechiae, purpura, mucosal bleeding. 2. Thrombocytosis: a. Elevated platelet count (>450,000/uL). b. Causes: Reactive (e.g., infection, inflammation) or clonal (e.g., essential thrombocythemia). c. Risk: Thrombosis or bleeding. 3. Platelet Function Disorders: a. Congenital: e.g., Glanzmann thrombasthenia (GPIIb/Illa deficiency), Bernard-Soulier syndrome (GPib deficiency). b. Acquired: e.g., aspirin use (inhibits cyclooxygenase, reducing thromboxane A2). Comparison of Reactive and Clonal Thrombocytosis | Feature | Reactive Thrombocytosis | Clonal Thrombocytosis | | :-------------- | :------------------------ | :---------------------- | | Cause | Secondary to | Primary bone | | another condition | marrow abnormally | | :---------------- | :---------------- | | Platelet Count | Usually <1,000 x10^9/L | Often >1,000 x10^9/L | | Platelet Function | Normal | May be dysfunctional | | Cytokine Role | IL-6 driven | Autonomous TPO production| | Genetic Mutation | Absent | JAK2, CALR, MP L mutations | | Bone Marrow | Normal megakaryocytes | Clonal proliferation of abnormal megakaryocytes | | Treatment | Treat underlying cause | Aspirin cytoreductive therapy | The image shows the process of stem cells differentiating into megakaryocytes, which then form platelets. Megakaryopoiesis proceeds initially through a phase characterized by mitotic division of a progenitor cell, followed by a wave of nuclear endoreduplication The origin of all blood cells, capable of differentiating into multiple lineages. A. Gives rise to the megakaryocyte erythroid lineage and is committed to producing megakaryocytes and erythrocytes. B. The earliest recognizable precursor of megakaryocytes. It has a large, round nucleus with scant cytoplasm C. A more developed form with a larger size and increased cytoplasm. The nucleus begins to show lobulation D. The final stage in the bone marrow before platelet release. The nucleus is highly lobulated, and the cytoplasm is abundant, ready to fragment into platelets The image includes a description that Megakaryopoiesis is a process wherein there will be megakaryocyte maturation and eventually the production of platelets, and that It is a part of hemopoiesis The image includes a summary of Key Steps in Megakaryopoiesis The process includes Differentiation, Maturation, endomitosis and platelet specific granule formation FACTORS THAT AFFECT MEGAKARYOPOIESIS 1. HORMONE: THROMBOPOIETIN (ΤΡΟ) * It is a glycoprotein hormone produced mainly by liver, smooth muscle cells, and kidney that regulates the production of platelets by the bone marrow. * Stimulates production and differentiation of meg. * It means that from a primitive cell line, ΤΡΟ affects the growth of a megakaryocyte until it becomes a mature one. * Concentration of TPO is Inversely proportional with megakaryocyte and platelet mass 2. CELLULAR DERIVED STIMULATORS * Cytokines are cellular secretions that gives signal for the cell to proliferate and differentiate. * Among which are: * IL3 acts synergistically with ΤΡΟ to induce early differentiation of stem cells. * IL6 with IL11 acts in the presence of TΡΟ to enhance the later phenomenon of the following processes: Endomitosis, Megakaryocyte maturation, and Platelet release/shedding. * Different cells produce different cytokines depending on what cell it is: Monocyte produce monokines Lymphocytes produce lymphokines. 3. GROWTH FACTORS * Growth factors are necessary for the growth, maturation and proliferation of megakaryocytes. Among which are: * Stem cell factors/Kit * Ligand/Mast cell growth factor * G-CSF * EPΡΟ - at least 20% of circulating EPO is needed for meg and platelet maturation 4. INHIBITORS OF MEGAKARYOPOIESIS * Inhibitors control and regulate production of platelet in the circulation because accumulation of platelet in the blood may lead to formation of thrombus that cuts the oxygen supply in the body causing tissue necrosis and eventually death. * Platelet factor 4 * B-thromboglobulin * NAP2 (Neutrophil activating peptide 2) Megakaryoblast * Earliest recognizable stage. * Contains mitochondria and a primitive endoplasmic reticulum. * This cell displays blunt protrusions from its cytoplasmic membrane and contains a multitude of polyribosomes and clear vacuoles with diameters as large as 0.2 um. * Normally found only in the bone marrow (1 to 4 per 1000 nucleated cells). * Single, centrally located nucleus or multiple round and oval nuclei containing several nucleoli and distinct but fine, delicate chromatin strand. * The cytoplasm stains a diffuse blue, indicating absence of specific granules. * Has a diameter of 15 to 50 um Promegakaryocyte * Cell size ranges from 20 to 80 um. * The cell membrane retains its characteristic blunt protrusions, and the cytoplasm is rich in polyribosomes. * The number of nuclear lobes begins to increase but there is only barely detectable margination of the chromatin around the membrane. * Demarcating membrane system (DMS) forms by invagination of the plasma membrane. Megakaryocyte * Represents the maturation stage. * This cell is round and is expanded in volume, with multiple nuclei and even, peripheral margins. * The abundant cytoplasm contains numerous small, rather uniformly distributed granules with a reddish-blue hue. * Chromatin pattern is linear and coarse, with distinct spaces between the strands. * It begins to contain all of structural constituents of a megakaryocyte. * It is the known stage that does not ordinarily produce platelets however megakaryocytes with at least four nuclei can produce platelets. Metamegakaryocyte * The fourth stage of maturation. * Very large cell, many time the size of the mature granulocyte, with a decreased nuclear-cytoplasmic ratio compared with the immature stages of developments. * Nucleus is multilobed and ploidy. * There is aggregation of granular material in the cytoplasm. * Platelets may be seen adhering to the cell membrane as they begin to break away from the metamegakaryocyte. * Contain predominantly polyribosomes with occasional mitochondria. Platelets * Have an average diameter of 2 to 4 um, with younger platelets being larger than older ones. * Has no nucleus * Cytoplasm is light blue, with evenly dispersed, fine red-purple granules. Platelet functions In the image, Platelet functions shown sequentially include: Platelet adhesion, followed by aggregation and finally secretion. * Adhesion-platelets roll and cling to nonplatelet surfaces (reversible) Image shows platelets adhering to damaged endothelial cells through the attachment of collagen fivers * Aggregation Platelets adhere to each other (irreversible) Image shows activated platelets sticking together and platelets activating new platelets to form a mass, otherwise known as a platelet plug. The plug is reinforced by fibrin forming through the clotting process * Secretion - platelets discharge the contents of their granules A schematic shows regulatory molecules ADP, TXA that lead to platelet activation, and vasoconstriction STRUCTURE & COMPOSITION. * Platelets - small plate like * Thrombocytes. - thrombo - clot; cytes - cells. * Size - smallest blood cells; 2-4 um. * Shape spherical or oval discoid * Volume 5.8 um3 Colour colourless. * Leishmann's staining faint blue cytoplasm with reddish purple granules. * Nucleus - Absent Electron Microscope Picture * Cell membrane * Microtubules * Cytoplasm Cell Membrane 6 nm thick trilaminar membrane Consists of +Lipids - phospholipids, cholesterol & glycolipids. +Carbohydrates +Proteins - glycoproteins Phospholipids of membrane contains platelet factor - 3, imp for blood clotting process. Glycoproteins - forming surface coat of platelets Prevent sticking of platelets to endothelium. *Accelerate adherence to collagen & damaged endothelium.*Canalicular system - surface of platelets invaginate to form Canalicular system or surface connecting system.*Receptors - for combining with collagen & fibrinogen.*Precursors of substances like - Thromboxane A2, PG, LT & platelet factor 3,4 are also present. Microtubules *Made up of Polymerised proteins called Tubulins.*Forms compact bundle below membrane & encircle whole cytoplasm.*Responsible for discoid shape. Cytoplasm*Contains - Golgi apparatus, Endoplasmic reticulum, Mitochondria, microtubules, microvesicles, filament, granules, glycogen, Lysosomes, protein enzymes & hormonal substances. Golgi apparatus, Endoplasmic reticulum Synthesizes various enzymes. endoplasmic reticulum stores large amount of calcium Mitochondria powerhouse of the cell Produce ATP & ADP Contractile Proteins *Actin & Myosin - same as those contractile proteins of muscle. Responsible for platelet contraction & clot retraction. Thrombosthenin Other Proteins Fibrin stabilizing factor - Imp. in blood coagulation Platelet derived growth factor - for growth of vascular endothelial cells, vascular smooth cells & fibroblast. Involved in repair of damaged blood vessel. Von Willebrand factor - for platelet adherence Granules Dense granules contains phospholipids, triglycerides, cholesterol, ATP, ADP, Serotonin (5HT which they obtain while passing through GIT) Alpha granules contains secreted proteins as clotting factors, & PDGF Enzymes Adenosine triphosphate & enzymes for synthesis of prostaglandins Prostaglandins- acts as a local hormone & have local vascular & tissue reaction Properties Adhesiveness - whenever comes in contact with any wet or rough surface gets activated & stick to surface. Factors responsible are collagen, thrombin, ADP, thromboxane A2, Ca ion The image shows the steps of Von Willebrand factors binding to collagen fibers at the site of injury. The platelets then adhere to the Von Willebrand factors via glycoproteins Aggregation - property to stick to each other Factors responsible are ADP & Thromboxane A2 Agglutination property of clumping together of platelet Functions Role in haemostasis Role in clot formation Role in clot retraction Role in repair of injured blood vessels Role in defense mechanism Transport & storage function Role in Haemostasis Haemostasis - spontaneous arrest of bleeding from injured blood vessel Vasoconstriction - by 5HT & other vasoconstrictors Temporary haemostatic plug - by platelets due to its property of adhesiveness & aggregation Definite haemostatic plug - also initiated by platelets Role in Clot Formation Play important role in formation of intrinsic prothrombin activator It is responsible for onset of blood clotting Role in Clot Retraction Contraction of contractile proteins i.e Actin, Myosin & Thrombosthenin Responsible for clot Retraction & wound healing Role in Repair of Injured Blood Vessels Platelet derived growth factor (PDGF) in cytoplasm of platelet important for Repair of Endothelium Role in Defense Mechanism Due to the property of agglutination, platelets are capable of Phagocytosis Mainly in Phagocytosis of carbon particles, viruses & immune complexes Transport & Storage Function Platelet when passes through GIT takes 5-HT against concentration gradient, stored & transported to the site of injury Normal Counts & Variations Normal count 150,000-400,000 per mm3 Average 2.5 L/mm3 Physiological Variations Age - less in infant, reach adult level by 3 months of age Sex no difference but during menstruation reduced in females Meal - increases after meal Muscular exercise - increases High Altitude- increases Thrombocytosis - increases in count more than 450,000/mm3 After splenectomy - after hemorrhage, severe injury, major operation & parturition Myeloproliferative disorders - CML, Polycythemia vera, Myelofibrosis Pathological Variations Thrombocytosis - increase in count more than 450,000/mm3 After Splenectomy - after hemorrhage, severe injury, major operation & parturition Myeloproliferative - Disorders - CML Polycythemia vera, Myelofibrosis Thrombocytopenia - decrease in count less than 150,000/mm3 Bone marrow depression Acute leukemia Injections Toxemia, septicemia & uremia Formation of Platelets Stages in platelets production (10 days) Megakaryoblast, Promegakaryocyte, Megakaryocyte Sites - Bone marrow Stem cells PHSC, CFU-Meg Megakaryoblast Earliest recognizable cells CFU-Meg differentiate to form Megakaryoblast Diameter - 20-30 mm Cytoplasm - small, blue, non-granular Nucleus - large, oval/kidney shaped Promegakaryocyte Megakaryoblast - Endoreduplication of nuclear chromatin Nuclear chromatin replicates in multiple of 2 without division of cell Large cell with 32 times diploid content of nuclear DNA formed Cytoplasm - Granular Megakaryocyte Diameter - large cell with 30-90 mm in diameter Nucleus - single multiloaded Cytoplasm- abundant with red purple granules Cell margin irregular with Pseudopodia which gets detached into blood & forms platelets *One Megakaryocyte - 4000 platelets* Control of Thrombopoiesis Megakaryocyte - colony stimulating activity (Meg-CSA) Life Span & Fate of Platelets Megakaryocyte: Average platelet count 150-400 x $10^9$/L,lifespan 8-9 days