| Basic Structure and Function of Bone Marrow Before learning to examine bone marrow microscopically, it is important to understand the basic structure and function of the bone marrow.
The bone marrow is one of the largest organs in the body. The normal adult marrow on a daily basis produces approximately 2.5 billion red cells, 2.5 billion platelets and 1.5 billion granulocytes per kilogram of body weight. The main function of this organ is the formation and development of blood cells. Hematopoiesis begins in the yolk sac in the first weeks of embryonic life; stem cells from the yolk sac travel first to the liver and then to the spleen. These organs are the only blood forming sites during the first three months of fetal life. At the beginning of the fourth month the bone marrow begins its life-long function of cell production. | View Page |
| Location of Cells within Cord Within the hematopoietic cords each cell line has a specific location for development.
Erythroid precursors are located near a venous sinusoid and cluster around a macrophage. This is referred to as an erythroblastic island. Developing red cells obtain iron needed for hemoglobin production from macrophages.
Megakaryocytes are also located close to a venous sinus. They extend their cytoplasm in fingerlike projections through the sinus wall in order to release their platelets directly into the blood in the sinus.
Immature granulocytes lie within the hematopoietic cords.
The metamyelocyte stage is the first stage of the granulocyte series that is motile and able to move toward the sinus area. Mature neutrophils, eosinophils and basophils enter the sinusoidal blood through the basement membrane. As maturing erythrocytes also move toward the sinus wall any remaining nuclei are lost as the red cells move through small openings in the cells lining the sinus wall. | View Page |
| Changes in Cell Distribution Changes in the distribution of cells in the marrow are most apparent in the first month of life. At birth, granulocyte cells predominate. The myeloid to erythroid (M:E) ratio is somewhat higher in newborns and during infancy than it is later on in childhood and in adults. | View Page |
| Normal M:E Ratio The normal M:E ratio in adults varies from 1.2:1 to 5:1 myeloid cells to nucleated erythroid cells. An increased M:E ratio (6:1) may be seen in infection, chronic myelogenous leukemia or erythroid hypoplasia. A decreased M:E ratio (<1.2-1) may mean a decrease in granulocytes or an increase in erythroid cells. M:E ratios are somewhat higher in newborns and infancy than in later childhood and in adults. It is important to note that lymphocytes, monocytes and plasma cells are not included in the M:E ratio. | View Page |
| Representative Counting Field The actual cell count is performed using the oil (100x) objective. This oil immersion field shows a representative counting field. Four granulocytes, a prorubricyte, and two rubricytes are completely visible here. 100 to 500 nucleated cells are generally counted,depending on the cellularity of the smear, and only cells completely visible in the field should be included in the count. | View Page |
| High Power Examination High power (40x objective) examination can be used to estimate the myeloid-to-erythroid ratio. The erythrocytes are nucleated, immature erythrocytes. Under high power, nucleated red cells appear to have a dark purple nucleus as opposed to the lighter staining nucleus of the myeloid or granulocyte series. Lymphocytes also have a dark staining nucleus and some may be erroneously included in the erythroid estimate. In the normal marrow these numbers are insignificant. | View Page |
| Granulocytes include neutrophils, basophils and platelets. | View Page |
| Leukocytes or White Blood Cells (WBC's) The second group of cells are the leukocytes or white blood cells (WBC's). The leukocytes can be divided into two groups: granulocytes and mononuclear cells. Leukocytes are involved in various ways with the body's defense mechanisms. The cell shown by the red arrow is a mononuclear cell, in this case a monocyte. The cell shown by the blue arrow is a granulocyte, in this case a neutrophil. These cells will be presented in much more detail later. | View Page |
| Glossary of Terms A through M. Antibody - A modified type of serum globulin synthesized by lymphoid tissue in response to antigenic stimulus. By virtue of specific combining sites each antibody reacts with only one antigen. Anucleate - Having no nucleus. Azurophilic granules - The well-defined large reddish granules (lysosomes) which may be present in large lymphocytes. They are called "azurophilic granules" because they stain blue with the azure stains which were originally used. Basophilic granules - Specific granules present in the cytoplasm of basophils. These granules are large and stain purple-black due to their strong affinity for basic stain. B-cell - Bone marrow derived lymphocytes which produce humoral antibodies. Biconcave - Having two concave surfaces. Cellular Immunity - The capacity of a small proportion of lymphoid population to exhibit response to a specific antigen. Chromomere - The centrally located granular portion of the platelet. Clone - A population of cells descended from a single cell. Delayed Hypersensitivity - (part of cellular immunity) that develops slowly over a period of 24-72 hours after an antigenic stimulus. It consists of an accumulation of cells around small vessels and/or nerves. Example: Tuberculin skin test reaction. Digestive Enzyme - A substance that catalyzes or accelerates the process of digestion. Eosinophilic Granules - Specific granules present in the cytoplasm of eosinophils. These granules are large, refractile spheres which stain reddish-orange due to their strong affinity for acid stain. Erythrocyte (red blood cell, RBC) - One of the elements found in peripheral blood. Normally the mature form is a non-nucleated, circular, biconcave disk adapted to transport respiratory gases. Fixed Macrophage - A phagocyte that is non-motile. Free Macrophage - An ameboid phagocyte present at the site of inflammation. Graft Rejection - A transplanted tissue that is rejected by the body's antibodies. Graft vs. Host Reaction - A complication that occurs when an implanted piece of tissue, which contains antibodies, rejects the host's tissue. Granulocyte - A leukocyte which contains granules in its cytoplasm, i.e., neutrophilic, eosinophilic, or basophilic granules. Half-life - is the length of time it takes for half of the cells circulating at a given time to leave the blood for the tissues. Hemocyte - Any blood cell or formed element of the blood. Hemostasis - A mechanism of the vascular system to arrest an escape of blood. It involves an interaction between blood vessels, platelets, and coagulation. Heparin - A mucopolysaccharide acid which, when present in sufficient amounts, functions as an anticoagulant by inhibiting thrombin. Histamine - A powerful dilator of capillaries and a stimulator of gastric secretions. Humoral Immunity - Acquired immunity produced after response to an antigenic stimulus in which B cells produce circulating antibodies. Hyalomere - the clear, blue non-granular zone surrounding the chromomere of a platelet. Immune Response - The interaction of a cell and an antigen that results in a proliferation of the cell and a capacity to produce antibodies. Isotonic Fluid - A fluid whose elements have an equal osmotic pressure. Leukocyte (white blood cell, WBC) - One of the formed elements of the blood; involved primarily with the body's defense. Lysosome - A microscopic body within cell cytoplasm; contains various enzymes, mainly hydrolytic, which are released upon injury to the cell. Megakaryocyte - A giant cell of the bone marrow from which platelets are derived. Mononuclear - A cell having a single nucleus. | View Page |
| Glossary of Terms N through Z. N:C Ratio - Nuclear: cytoplasmic Ratio - The ratio of nuclear volume to cytoplasmic volume within any one cell.Neoplasm - Any new and abnormal growth, such as a tumor.Neutrophilic Granules - Specific granules present in the cytoplasm of neutrophils. These granules resemble pencil stippling and stain a lilac color due to their affinity for both basic and acid dyes.Phagocyte - Any cell that ingests microorganisms or other cells and foreign particles.Phagocytosis - The ingestion and destruction of microorganisms or other foreign particles.Plasma - The fluid portion of blood in which the various blood cells are suspended.PF3 (platelet Factor 3) - A lipoprotein component of the platelet membrane; functions as a surface catalyst during blood coagulation.Pseudopod - A temporary protrusion of the cytoplasm of a cell.Refractile - Capable of refracting or changing the direction of light.Senescence - The process or condition of growing old.Serotonin - A constituent of blood platelets and other cells and organs; induces constriction of the blood vessels.Specific Granules - Granules found in cells of the more mature stages of the granulocytic series. They have distinct staining reactions which differ with each type of granulocyte.T-cell - Thymus derived lymphocyte which mediates cellular immunity.Thrombocyte (Platelet) - A circular or oval disk found in the blood; concerned with hemostasis.Thymus - A ductless gland-like body situated in the anterior mediastinal cavity; reaches its maximum development during the early years of childhood.Vacuole - Any small space or cavity formed in the cytotoplasm of a cell. | View Page |
| Eosinophils Eosinophil is also known as eosinophilic granulocyte, or eo. Eosinophils are one of the easiest cells in the peripheral blood to recognize because of their large bright granules. The diameter of the eosinophil is 9-15 microns, and the N:C ratio is 1:3. Eosinophils are generally the largest granulocytes found in normal blood. | View Page |
| All of the following statements describe an eosinophil EXCEPT: | View Page |
| Segmented Neutrophil Segmented Neutrophil is also be referred to as seg, polymorphonuclear leukocyte, poly and PMN. Segmented neutrophils are the most mature neutrophilic granulocytes present in circulating blood. Their diameter is approximately 9-15 microns, and their N:C ratio is 3:1. | View Page |
| Which type of granulocyte has water soluble granules which may fade during the rinsing phase of the staining process? | View Page |
| Segmented and Band Nuclei The granulocytes found in normal peripheral blood are neutrophils, eosinophils and basophils.Most have segmented nuclei, and are therefore classified as being at the "segmented" stage of development. Some that are a little less mature have unsegmented nuclei. These are classified as "bands." Generally, we differentiate between the band and segmented forms of neutrophils, but since eosinophils and basophils are present in such low numbers, and since their nuclei are often obscured by cytoplasmic granules, we usually don't concern ourselves with designating the band forms.Since hematologists and textbooks use several different terms for these cells, synonyms for each term will be given and then may be used interchangeably throughout the course. | View Page |
| Basophil Basophil is also known as basophilic granulocyte and baso. Basophils are easily recognized because of their large, dark granules. The basophil's diameter is 9-15 microns, and its N:C ratio is approximately 1:3. | View Page |
| The Process of Phagocytosis Neutrophils have a relatively short life span.They are produced in the bone marrow, and when they reach the band or segmented stages are released into the peripheral blood.They remain there for approximately ten hours before randomly entering body tissues.Neutrophils in the blood stream can be divided into circulating granulocyte pool(CGP) and marginating granulocytic pool (MGP).The white blood cell count reflects the cells in the circulating pool.The cells in the marginating pool move quickly into the circulating pool when needed.During an infection the neutrophil concentration of the peripheral blood can increase almost immediately due to the shift of these cells from the marginating pool and release from the bone marrow storage pool, if needed.Neutrophils then migrate to areas of tissue damage or infection.Neutrophils do not reenter the blood stream from the tissues, thus end their life in the tissues either as a result of phagocytosis or senescence. | View Page |
| Mononuclear Cells: Lymphocytes and Monocytes. The mononuclear leukocytes consist of two cell types: lymphocytes and monocytes. In contrast to the granulocytes, these cells have rounded nuclei, some with indentations or folds. Granules are not prominent. | View Page |
| Dohle Bodies Dohle bodies are small blue irregular staining patches of cytoplasm seen near the edge of the cell in granulocytes under certain conditions. Electron microscopic examination reveals that they are composed of aggregates of rough endoplasmic reticulum remaining from an earlier stage of development. | View Page |
| Cytoplasmic Vacuolation Vacuoles are areas of the cytoplasm which do not stain with Wright's stain and appear as holes in the cytoplasm. Their composition may vary; some will contain remnants of bacterial digestion, autodigestion in an aging cell, while still others may contain fat. It is not possible to differentiate the various types of vacuoles on Wright's stained smears under light microscopy. They may be seen occasionally in an aging granulocyte, but are seen more frequently and are significant in cases of bacterial infection and septicemia. | View Page |
| More on May-Hegglin May-Hegglin bodies stain blue and appear to have a more definite shape than Dohle bodies. When examined under electron microscopy, they appear to be aggregates of thread-like structures in a crystal-like arrangement. May-Hegglin inclusions are RNA material believed to be derived from the endoplasmic reticulum.
May-Hegglin bodies can be seen in monocytes and platelets as well as in all mature granulocytes.
The platelets in May-Hegglin anomaly are very bizarre in appearance and thrombocytopenia is usually noted.
When examining a slide that has bizarre platelets and blue-appearing bodies in the cytoplasm, thought should be given to the possibility of the May-Hegglin anomaly. | View Page |
| Alder-Reilly Anomaly (Alder's Anomaly) Alder Reilly Anomaly is a rare autosomal recessive hereditary disorder in which the basic defect involves protein-carbohydrate complexes called mucopolysaccharides. The accumulation of partially degraded (broken down) protein-carbohydrate complexes within the lysosomes account for the larger than normal purple-staining granules seen in the granulocytes, monocytes and/or lymphocytes.
The granules may occur in clusters, rather than diffusely, throughout the cytoplasm as in toxic granulation.
These inclusions may be seen in the bone marrow more frequently than in peripheral blood. The physical characteristics associated with this disorder include gargoylism and dwarfism.
The function of the cells involved is not affected.
This morpholical change would be classified as pathological since the body is responding abnormally even though the function is not affected. | View Page |
| The neutrophils illustrated in this photograph are representative of those seen in the smear. The total WBC was 28,500 cells/cumm. The appropriate report to be issued following a morphology consultation would be: | View Page |
| Typical cells on a peripheral blood smear as photographed here were repeatedly encountered as the smear was reviewed. The peripheral white blood cell count was 51,000/ml with an orderly maturation sequence. The comment "leukemoid reaction" may properly be appended to the report. | View Page |
| A peripheral blood smear with many myeloid cells (photograph) was presented for morphology review. Toxic vacuoles in the neutrophil and monocyte most likely represent: | View Page |
| Alder- Reilly Anomaly Large inclusions in leukocyte cytoplasm appear with Alder-Reilly syndrome. Inheritance patterns are not completely clear. The condition is characterized by larger than usual azurophilic and deeply violet staining granules clustered throughout the cytoplasm (even covering the nucleus)in all granulocytes. There are variations in which some lymphocytes and monocytes may be affected. These inclusions represent partially degraded mucopolysaccharides within lysosomes.Alder-Reilly bodies may be found independently of genetic mucopolysaccharidoses as an inherited anomaly (Jordan's anomaly). Cytoplasmic vacuoles of toxic origin are not present in Alder-Reilly cells. The background condition in Alder-Reilly syndrome is mucopolysaccharidosis with various types of bone and cartilage disorders, reported first in gargoylism, then in Hunter and Hurler syndromes. Accompanying conditions are hepatosplenomegaly, corneal opacities, and mental retardation. Reference: Brunning, Richard D. Morphologic Alterations in Nucleated Blood and Marrow Cells in Genetic Disorders. Human Pathol: 99-124, March, 1970 | View Page |
| Auer Rod Illustrated in the photograph is a immature granulocyte with a distinct rod-shaped intracytoplasmic inclusion. This inclusion is known as an Auer rod, which is seen in up to 10% of blast cells in patients with acute myelogenous leukemia. An Auer red is the fusion of primary granules into rod-like inclusions. | View Page |
| The May -Hegglin anomaly Illustrated in the upper photograph is a poorly defined cytoplasmic inclusion somewhat resembling a Doehle body. Note, however, that this inclusion is well defined and there is no evidence of toxic granulation in the cytoplasm.When Doehle-like bodies are identified, May-Hegglin anomaly should be considered in the differential diagnosis even though this entity is rare.The May-Hegglin anomaly is an inherited dominant condition in which large 2 - 5 um, basophilic and pyronophilic inclusions are present in granulocytes, including neutrophils, eosinophils, basophils, and monocytes.Similar to Doehle bodies, the May-Hegglin inclusions also are composed of RNA, probably derived from the rough endoplasmic reticulum. May-Hegglin anomaly includes giant platelets containing few fine granules (lower photograph).Sometimes the platelets have bizarre shapes and variable sizes. Variable degrees of thrombocytopenia complicated by mild bleeding problems and purpura may accompany the aberrant platelets. | View Page |