Week 9 - Part 2 - Platelets and Platelet Disorders

• The Megakaryocyte

Megakaryoblast:

• 20 - 50 microns

• Cytoplasm dark blue with blebs or pseudopods and non granular

  • Nucleus: round to oval centrally located, may contain nucleoli

Promegakaryocyte:

• Size 20 - 80 microns

• Undergoes mitotic divisions (endomitosis)

• Granulation starts to appear

• Nucleoli are usually visible

The Platelet:

• Small disc shaped cellular fragments

• Megakaryoblast develops into the megakaryocyte which

  gives rise to the thrombocyte

Life span of a platelet?:

• 10 days (9.5 days)

Platelet production:

• Thrombopoietin (TPO) hormone primarily produced by liver stimulates stem cells to differentiate into the megakaryocytic lineage

• Megakaryocytic cells undergo multiple mitotic divisions without cytoplasmic division (called endomitosis)

• Generates giant multinucleated cells

• Platelets are produced directly from the megakaryocyte cytoplasm

• As the megakaryocyte matures the granules in the

  cytoplasm begin to cluster into small groups.

Thrombopoiesis:

• The megakaryocyte membrane ruptures and the shedding of platelets occurs.

• Each megakaryocyte produces 2,000 to 4,000 platelets

• Platelet release from Mature Megakaryocyte

Platelet characteristics:

• Younger platelets are larger than old platelets

• Approximately 2/3 of platelets produced are in the peripheral blood and 1/3 are in the spleen

• The platelets in the spleen are interchangeable with the peripheral blood

• Platelet turnover is 35,000/mm per day

Platelet Estimations:

• In a field of 100 RBC’s 1 platelet is equal to 20,000 platelets

• Large platelets

  • Diameter greater than 4 microns

  • Seen during increase turn over

  • such as with immune thrombocytopenic purpura (ITP)

• Giant platelets

  • Diameter greater than 7 microns

  • Size of a RBC

Giant platelets and May Hegglin:

• Red arrow points to giant platelet

• Black arrow points to May-Hegglin body. Note

  the neutrophil with the blue inclusion

• If large quantity of giant platelets exist, the

  automated platelet count might be falsely decreased

• Size falls outside upper threshold of automated analyzers

Platelet Clumping:

• EDTA can cause platelets to surround the neutrophil – called satellitism

• EDTA dependent antibodies react with platelet glycoprotein IIb/IIIa

• Platelet clumping may also be caused by traumatic venipuncture

• Decrease PLT count on automation

• WBC count flagged on automation

Corrective Action of Platelet Clumping:

• redraw patient using sodium citrate tube (only time you would use this instead of EDTA)

• Correction PLT count x 1.1 (to compensate for sodium citrate)

• Platelet clumping

• Platelet Satellitism

Platelet structure:

• may be very small, but they are actually very complex and metabolically active

Four layers of Platelet structures:

• Peripheral Zone

• Structural Zone (Sol-gel Zone)

• Organelle Zone

• Platelet Membrane system

Peripheral Zone:

• Composes the surface coat

• Responsible for adhesion which is the attachment of the platelet to a foreign surface

• Responsible for aggregation which is the attachment of platelets to other platelets

• Important for coagulation

Sol Gel Zone:

• Sometimes referred to as structural zone

• Matrix of platelet cytoplasm

• Contains several fiber systems in various states of polymerization (cytoskeleton)

• Composed of microfilaments and microtubules

• Provides the cytoskeleton and contractile system

• Needed for platelet shape

  *Platelets flat biconcave disc until activated, once

  activated change shape “inside out” signaling

Organelle Zone:

• Storage and secretion of substances essential for platelet function

• Contains mitochondria, alpha granules, dense bodies and lysosomal granules

• Important for metabolism, hemostasis, and vessel repair

Platelet Membrane System:

• Important regulators of intracellular calcium

  concentration

• Calcium is important for platelet metabolism and

  activation of the coagulation process

• Production of prostaglandin synthesis, needed for platelet aggregation

• Platelet Ultrastructure and Functions

Platelet structure and physiology:

• Size: 2 to 3 micrometers in diameter

• Dense granules and alpha granules

• Platelets are biconvex in circulation

• When platelets are activated, they change shape and become tiny spheres, forming pseudopodia

Normal platelet count:

• 150,000 to 450,000

Where is one-third platelet volume?

• Spleen

• Rest is in circulation

• Vascular System

Primary hemostasis:

• The process of platelet plug formation

  • Adhesion

  • Shape change

  • Secretion: discharge of alpha and dense granules

  • Amplification

  • Aggregation

Platelet kinetics: Adhesion:

• Exposed collagen and subendothelial cells initiate the

  first step of the platelet plug formation.

• Platelets reach the surface and change shape from

  discs to spiny spheres

Platelet kinetics: Granule Release:

• The contents of the dense bodies and alpha granules are released to regulate clot formation

Platelet kinetics: Aggregation:

• Platelets stick to each other to form stronger platelet plug along with the coagulation proteins

What does platelet adhesion require

• Occurs when platelets attach to collagen in the exposed basement membrane

  • gpIa/IIa on the platelet surface

  • vWF

  • gpIb/IX binding site

Platelet amplification requires:

• Secreted substances, such as TXA2, recruits more platelets to the site of injury

Platelet aggregation requires:

• The use of fibrinogen as the mediator protein

• Fibrinogen attaches to platelets at the site of gpIIb/IIIa

• Platelet plug is formed to stop blood loss

Platelet Granules: Alpha:

• Clot-activating granules

  • Glycoproteins IIb and IIIa

  • Fibrinogen

  • von Willebrand Factor (vWF)

  • Factor V (5)

  • Factor VIII (8)

Platelet Granules: Dense:

• Platelet-activiating substances

  • ADP

  • Serotonin

  • Calcium

  • Thromboxane A2 (TXA2)

  • *****Blockage of TXA2 impairs platelet function

  • Aspirin (and specifically the acetylation of

    aspirin inactivates cyclogenese, which blocks

    TXA2 production

Von Willebrand Factor:

• can be found in megakaryocytes, it’s a component of alpha granules and made by megakaryocytes as they mature

• Platelet function in primary hemostasis

Types of Platelet Disorders:

• Quantitative

• Qualitative

Platelet disorders leading to bleeding disorders are a result of?:

• Quantitative abnormality of platelets

Quantitative Disorders:

• Thrombocytopenia — most common cause of abnormal bleeding

What is thrombocytopenia caused by?:

• A decrease or ineffective production of platelets

• An increase in platelet destruction or utilization

• An increase in platelet sequestration (removal) by the spleen

• Loss from the body

• Dilution factors

Congenital Hypoplasia:

• Decrease cell (platelet) production in the bone marrow

Fanconis Syndrome:

• a chromosomal defect where the bone marrow fails to produce RBC’s, WBC’s and platelets. (Pancytopenia, decrease in all cells). Found in children

Other reasons for decreased production:

• Infection

  • Newborns infected with virus, such as Rubella

• Intrauterine exposure to certain drugs that are capable of damaging production sites

Acquired Hypoplasia:

• a result of the action of chemical or toxic drugs to the bone marrow

• Radiation or chemotherapy: the platelets are the last

  cell line to return to normal and are first to be affected.

• Replacement of the bone marrow by neoplastic diseases (like leukemia)

• Decrease in proliferation of megakaryocytes caused by marrow replacement (aplastic anemia)

Ineffective Thrombopoiesis:

• the bone marrow contains normal or increased numbers of megakaryocytes but the number of platelets in peripheral blood are decreased

What is ineffective thrombopoiesis seen with?:

• Megaloblastic anemia

• Ethanol abuse

• Pre-leukemia

Hereditary Thrombocytopenia:

• Inherited disorders where the platelet number is decreased

Increased Destruction:

• Immune related factors

• Idiopathic Thrombocytopenia Purpura (ITP)

Idiopathic Thrombocytopenia Purpura (ITP)

• Unknown origin

• Patients often have very big bruises

• Acute condition (resolves in a few weeks)

• Found predominately in children (2-6 years)

• Majority of cases develop after recovery from viral infection

• 66% of cases are antibody directed

• Usually self limiting, spontaneous remission in about 80% of cases

• platelet destruction is increased

Idiopathic Thrombocytopenia Purpura (ITP) causes:

• Causes excessive bleeding

• Unusually low level of platelets

• Sometimes referred to as immune thrombocytopenia

• After infection such as measles or chicken pox, immune system produces IgG antibodies which coat platelets.

• Leads to filtering out by spleen

• Can treat with steroids if doesn’t resolve on its own

• Steroids “turn off” or slow down immune activity

Idiopathic Thrombocytopenia Purpura (ITP): Laboratory Findings:

• Platelet count <20,000

• Sudden onset of petechiae and bruising

• Image shows petechiae

• Frequent nose bleeds

Idiopathic Thrombocytopenia Purpura (ITP): Treatment:

• Usually last 3 weeks – 6 months

• Immunosuppressants are sometimes given

Chronic ITP:

• Can be seen at any age but more often found in women between ages 20-50 years old

• Platelets are sensitized by platelet antibodies (usually IgG type antibodies)

• The clinical course of CITP is some patients consists of alternate periods of remission and relapse (periods where platelet count is almost normal and then drops again)

Chronic ITP: Laboratory and Clinical Findings:

• Platelet count 30,000 – 60,000

• Platelets are large and abnormal in appearance

• Petechiae and bruising present

Chronic ITP: Treatment:

• Keep platelets above 40,000

• Steroids to reduce spleen removal

  • prednisone

• Splenectomy may be advised

Transplacental or Neonatal disorders:

• A self limiting form of thrombocytopenia

• Caused by maternal IgG platelet antibody crossing placenta

• IgG only antibody subtype that crosses placenta

• Normally good for IgG to pass placenta as it gives the

  newborn circulating protecting IgG while newborn forms own immune system

• Causes destruction of the newborn’s platelets

• Greatest danger for the fetus is during delivery and immediate post partum

• Danger of central nervous hemorrhage

• Can have intercranial bleeding

Neonatal Increased Destruction Laboratory Findings:

• Platelet count less than 30,000

• In then decreases further during the first few hours of

  life

Neonatal Increased Destruction: Treatment:

• Usually last an average of 3 – 4 weeks

• Recovery follows clearance of antibody from circulation

• Must evaluate severity

• May require platelet transfusion

• Self-limiting once antibody is gone

Drug Induced:

• An immune reaction with destruction of platelets results after exposure to certain drugs

  • Most common quinidine, heparin (HIT) and some antibiotics

  • Caused by drug-dependent antibodies

  • Usually seen 1-2 weeks after patient is on drug treatment

  • Treatment includes removing the offending drug

Post transfusion Purpura:

• Occurs one week after blood transfusion

• Antibodies are stimulated from foreign antigen and cross react with platelets

  • Disappears 2-5 weeks without treatment

  • Treat with intravenous immunoglobulin (IVIG)

• (uncommon problem, rare, but serious)

Other causes of decreased platelet counts:

• HIV

• Hepatisis C Virus

• Helicobacter pylori

• Some infections may lead to decreased platelet count as disease progresses

• Though to occur as a result of immune reaction

Non-Immune: Thrombotic Thrombocytopenia Purpura (TTP):

• Intravascular platelet aggregation

• A rare disorder which the exact cause is unknown

• Characterized by low platelet count and hemolytic anemia (red blood cells are damaged)

• Formation of platelet clots in capillaries and arteriole

• See schistocytes (RBC fragments) in peripheral blood

• Due to red blood cell injury from damaged epithelium

• As blood flow through the RBC’s become damaged and an anemia results

  • Schistocytes

• TTP affects all ages, more commonly women in child bearing age

Non-Immune: Thrombotic Thrombocytopenia Purpura (TTP): characteristics & findings:

• Platelet counts average about 20,000

• PT and aPTT are normal

• not involving coagulations factors

• Platelets clot, not fibrin

• Increased lactate dehydrogenase (LDH) common for TTP

• TTP characterized by Macroangiopathic Hemolytic Anemia (MHA)

• Intravascular hemolysis

• Decrease haptoglobin

• Neurological complications

Etiology of Thrombotic Thrombocytopenia Purpura (TTP):

• Deficiency of ADAMTS-13

  • Molecular test looks at vWF attached to factor 8

  • The protein has a relation wit vWF (Von Willebrand Factor)

  • Function to break this large molecule into small pieces

  • Large vWF multimers tend to increase platelet adhesion

  • Supposed to be small but high in thrombocytopenic purpura (TTP)

  • vWF is important in adhesion of platelets for normal hemostasis

Non-Immune: Throbotic Thrombocytopenia Purpura (TTP): Treatment:

• Anti-platelet agents, plasma exchange transfusion, steroids

• Can be fatal due to formation of clots

Disseminated Intravascular Coagulation DIC:

• Results from a major tissue injury or infection

• Thrombotic occlusion of the microcirculation

  • Consumption of platelets and clotting factors

  • RBC fragments and hemolytic anemia

  • Decreased platelet count

  • Abnormal coagulation testing

  • DIC involves coagulation factors

Hemolytic Uremic Syndrome (HUS):

• Predominates in children age 6 months to 4 years (although any age can contract)

• Usually self limiting

• High occurrence with E. Coli 0157:H7 infection

• Can be caused by other infections

• Often follows an acute viral infection and is associated

  with vomiting and diarrhea

• Platelet consumption occurs in the kidneys

• Renal damage – abnormal kidney tests

• Signs are hemolytic anemia, (presence of schistocytes, thrombocytopenia and renal failure)

Mechanical Destruction of Platelets:

• Artificial heart valves

• ECMO: extracorporeal membrane oxygenation

  • Blood pumped outside the body to heart-lung machine that removes CO2 and sends oxygen back to blood

Increased splenic sequestration:

• When spleen becomes enlarged, the number of

  platelets sequestered is increased

  • Example: Hodgkin’s lymphoma, cirrhosis of liver

Dilution:

• Patients who experience massive hemorrhage

  requiring blood replacement

  • No viable platelets in transfusion units

Thrombocytosis:

• Overproduction of platelets

• Usually platelet count over 400,000 (or 450,000)

• Relative Thrombocytosis or Primary

  • Uncontrolled proliferation of platelets both in

    peripheral and bone marrow

• Example:

• Splenectomy, acute blood loss, acute and chronic

  inflammation, myeloproliferative disorders (CML), or

  increased hematopoiesis

Thrombocythemia:

• Platelet count >1-2 million

  • Platelets are clumped, bizarre shape and size

  • Function may be abnormal

  • Usually secondary to another disorder

Qualitative reasons for platelet disorder functioning:

• Glanzmanns

• Storage pool

• Bernard-Soulier

• Von Willebrand’s disorder

• Aspirin

Qualitative: Glanzmanns:

• Aggregation disorder

Qualitative: Storage Pool:

• Secretion and release action disrupted

• Granules not released

Qualitative: Bernard-Soulier:

• Adhesion disorder

  • Missing membrane receptors for vWF

Qualitative: Von Willebrand’s Disorder:

• Adhesion disorder

  • Receptor there but not functional

Qualitative: Aspirin:

• Loss of aggregations (prostaglandin synthesis)

Patients who present with bleeding disorders (due to platelet issue) will typically present with?:

• Mucosal Bleeding