Hemopoiesis

Introduction and Key Concepts for Hemopoiesis

All formed elements, with the exception of some lymphocytes, have a finite life span in circulation, so there must be an ongoing replacement throughout the life of an individual. The magnitude of the task for a given cell type can be appreciated if the approximate required daily production rate is calculated from estimates of the total number in circulation and the rate of turnover for the cell type. For erythrocytes, with a life span of about 120 days, the daily production rate is roughly 250 billion, for neutrophils, with a time in circulation of less than a day, the daily production rate is normally roughly 60 billion, and for platelets, with a life span of about 10 days, the daily production rate is approximately 150 billion. The development of each type of blood cell involves numerous cell divisions and a series of differentiation steps so that a small number of completely undifferentiated stem cells produce enormous numbers of cells that have the specific equipment necessary for the particular mature cell to perform its functions. Although all blood cells originate from a common pluripotential hemopoietic stem cell, each blood cell type has its own lineage of cell generations committed to proliferate and, at the same time, differentiate only into that cell type. Cells that can be recognized morphologically as undertaking differentiation into a particular blood cell are called precursor cells. Precursor cells are produced by cells that are committed to a specific lineage (i.e., they are determined to give rise to, e.g., only erythrocytes) but show no morphological signs of differentiation. These are called progenitor cells. Some progenitor cells are not restricted in potential to just one blood cell lineage but rather to two lineages. Progenitor cells are also termed colony-forming cells (CFCs). A commonly used abbreviation system for designating specific progenitor cells uses the first letter of the blood cell name after the letters CFC, for example, CFC-E for erythrocyte colony–forming cell and CFC-B for basophil colony–forming cell. Development of blood cells occurs mostly in the specialized environment of the bone marrow. Because extensive cell proliferation is required, the process is very vulnerable to irradiation, so protection within the cores of bones is clearly advantageous. The development of each type of blood cell involves a series of precursor cells that can be recognized in smears of the red bone marrow that have been stained with the same procedures used for peripheral blood smears. Lymphocytes and monocytes are little differentiated, so the morphological appearance of their precursors (lymphoblasts and promonocytes, respectively) is not easily distinguished. By contrast, the precursors of erythrocytes, granulocytes, and platelets exhibit relatively distinct features as they differentiate in a series of steps that involve predictable changes and identifiable stages. The red bone marrow is a hemopoietic compartment where blood cells (except lymphocytes) develop and mature (Fig. 8-15C).

ERYTHROCYTE DEVELOPMENT is called erythropoiesis. The appearances of the precursors of erythrocytes reflect the processes that must take place to generate, from an undifferentiated cell, a cell that is essentially a plasmalemma bag of hemoglobin. In the initial stage, the proerythroblast, the main event is generation of free ribosomes that will be needed to synthesize the globin that will combine with heme to form hemoglobin. Therefore, the intense basophilic staining of the cytoplasm in the next stage, the basophilic erythroblast, results from a peak concentration of free ribosomes that begin translation of globin mRNAs. In the polychromatophilic

erythroblast, enough hemoglobin has accumulated to confer some eosinophilia to the cytoplasm, whereas the concentration of ribosomes has decreased from dilution that accompanies cell division. Continuation of cell division, dilution of ribosomes, and accumulation of more hemoglobin account for the strong eosinophilic staining of the cytoplasm in the orthochromatophilic erythroblast

(normoblast). Concurrent with the successive changes in staining of the cytoplasm during erythrocyte development in the cytoplasm are changes in the appearance of the nucleus. Production of ribosomes and transcription of mRNA for globin and other proteins are manifested by a large euchromatic nucleus with prominent nucleoli in the proerythroblast; subsequent stages have progressively smaller, less active nuclei, and the nucleus is ultimately extruded at the end of the orthochromatophilic erythroblast stage. The mitochondrion, another key organelle, is required to synthesize protoporphyrin and combine it with iron to form the heme of hemoglobin (Figs. 8-9A and 8-10A to 8-11C).

PLATELET DEVELOPMENT is called thrombopoiesis. Although platelets are small (2–4 μm in greatest diameter) fragments of highly organized cytoplasm, they are produced by very large cells called megakaryocytes. These cells measure up to 100 μm or more in diameter. Megakaryocytes develop from precursor cells (called megakaryoblasts) through a series of incomplete cell cycles (endomitosis) that do not include division of the nucleus or cytoplasm. The result is that the nucleus of a mature megakaryocyte has up to 64N chromosomes, instead of the usual 2N chromosomes. The nucleus is large and lobular, but it remains one nucleus. The cytoplasm develops numerous mitochondria, a variety of granules, and microfilaments and microtubules. As the megakaryocyte reaches maturity, its cytoplasm becomes cordoned off by an elaborate system of membranes, called demarcation membranes or channels, which subdivide the cytoplasm into platelet zones—like perforations in a sheet of stamps (Figs. 8-9B and 8-12A–C).

GRANULOCYTE DEVELOPMENT (GRANULOCYTOPOIESIS) proceeds through an orderly sequence of events that result in cytoplasm that is packed with granules containing a wide variety of substances related to inflammation and destruction of pathogenic organisms. As is the case with erythrocyte development, the initial discernible event is generation of components (ribosomes and RNA) needed for protein synthesis, but in granulocyte development, the proteins will be packaged in vesicles (granules). Accordingly, packaging of the proteins requires development of an extensive endoplasmic reticulum and Golgi complex. These events are prominent in the myeloblast and promyelocyte stages, both of which have relatively large, active nuclei with nucleoli and cytoplasm that is basophilic owing to its content of ribosomes. Granule generation occurs sequentially, the nonspecific (lysosomal) granules first, in the promyelocyte stage, and the specific granules second, in the myelocyte stage. Because specific granules first appear in the myelocyte stage, this is the earliest stage at which the precursors of the three granulocytes can be distinguished from each other. Other notable changes during granulocyte maturation are progressive condensation, elongation, and segmentation of the nucleus (Figs. 8-13A to 8-15B).

Proerythroblast

D. Cui

Nucleoli

A

Figure 8-10A. Proerythroblasts, bone marrow smear.

Wright stain, 710; inset 1,569

The proerythroblast is a relatively large cell with a large, round nucleus containing two to three nucleoli. The cytoplasm appears basophilic (blue) because of the presence of a large number of ribosomes. At this stage, the cell is beginning to accumulate the necessary components for the production of hemoglobin. Proerythroblasts are precursor cells, which develop from two functionally identifiable progenitor cells: burst-forming unit–erythroid (BFU-E) cells, which take about a week to mature to become colony-forming unit–erythroid

(CFU-E) cells and another week to become proerythroblasts. Proerythroblasts undergo mitosis to produce two daughter cells that will develop the features of basophilic erythroblasts.

Basophilic erythroblast

B

Figure 8-10B. Basophilic erythroblasts, bone marrow smear. Wright stain, 710; inset 1,569

The basophilic erythroblast is smaller than the proerythroblast, and its cytoplasm is deep blue because of a high content of tightly packed ribosomes. Compared to proerythroblasts, basophilic erythroblasts have smaller nuclei with a coarser texture because most of the chromatin is in the heterochromatin form. At this stage, nuclei are less active than in proerythroblasts, and their nucleoli disappear. These cells undergo mitosis and divide into daughter cells, which mature to become polychromatophilic erythroblasts. (OE, orthochromatohilic erythroblast; PoE, polychromatophilic erythroblast; BE, basophilic erythroblast.)

D. Cui

Polychromatophilic

erythroblast Polychromatophilic erythroblast

Figure 8-10C. Polychromatophilic erythroblasts, bone marrow smear. Wright stain, 710; inset 1,569

The polychromatophilic erythroblast is smaller than its parent cell (basophilic erythroblast). The nucleus is smaller and it has no nucleoli. The condensed nucleus is densely stained, and it displays a patchy pattern because of condensation of chromatin. The cytoplasm of the cell is usually grayish in overall color because, at this stage, significant amounts of hemoglobin have been produced and accumulated in the cytoplasm, so that the staining of the cytoplasm reflects the presence of both ribosomes (basophilic) and hemoglobin (eosinophilic). Therefore, the cytoplasm is mottled with mixed patches of blue and pink (polychromatophilic means “attracting multiple colors”). Polychromatophilic erythroblasts undergo mitosis and divide into daughter cells that develop into orthochromatophilic erythroblasts.

CLINICAL CORRELATION

C

Erythrocyte

Reticulocytes

Figure 8-11C. Reticulocytosis, Peripheral Blood Smear. New methylene blue stain, 1,020

Reticulocytosis is a condition characterized by an increased number of reticulocytes. Reticulocytes are premature red blood cells. The normal percentage of reticulocytes is 0.5% to 1.5%. Hemolytic anemia usually increases erythropoietin production, which in turn causes the bone marrow to produce more red blood cells, resulting in a reticulocyte percentage of above 4% to 5%. An increased number of reticulocytes in peripheral blood is an important indication of hemolysis (red blood cell rupture) or bleeding. It can also be the consequence of treating the anemia of chronic kidney disease with erythropoietin. This illustration shows the increased number of reticulocytes with new methylene blue stain after hemolytic anemia.

A

Megakaryoblast

Figure 8-12A. Promegakaryocytes (immature megakaryocytes), bone marrow smear. Wright stain, 754; inset 1,605

Promegakaryocytes develop from megakaryoblasts in the bone marrow. Megakaryoblasts lose their ability to undergo cytokinesis but undergo a series of incomplete cell cycles called “endomitosis” that results in replication of DNA up to 64N without division of nuclei or cytoplasm. Each megakaryoblast has a large nucleus, multiple nucleoli, and basophilic cytoplasm (Fig. 8-9B). Promegakaryocytes can be recognized by their large size, round (or oval) nuclei, and large amount of cytoplasm. Development of a demarcation membrane system is an important feature of thrombopoiesis and begins at a very early stage. The demarcation membrane system is produced by the invagination of plasma membranes to form branched interconnected channels through the cytoplasm. This system may play an important role in later subdivision of the cytoplasm into platelet zones.

Neutrophils

Figure 8-12B. Megakaryocytes, bone marrow smear. Wright stain, 754; inset 2,827

Because of their very large size, megakaryocytes are sometimes called giant cells. They have a large indented (partially lobulated) nucleus and an extremely voluminous cytoplasm containing a variety of granules. In this stage, the demarcation membrane system is complete, and the megakaryocytes are ready to release platelets. Megakaryocytes tend to move toward and attach to the marrow sinusoids (capillaries in the bone marrow) after they become mature. The platelets are released into the blood circulation through the wall of the marrow sinusoids. Note the neutrophils on the surface of the megakaryocyte for size comparison. The inset shows incipient platelets (fragments of cytoplasm beginning to become platelets) in the surface region of the megakaryocyte that are ready to be released. Thrombopoietin, a humoral factor produced in the liver, is believed to regulate megakaryocytes and the production of platelets.

CLINICAL CORRELATION

C

Megakaryocytes

Megakaryocyte

Figure 8-12C. Essential Thrombocytosis, Bone Marrow Smear. Wright stain, 304

Essential thrombocytosis, also called essential thrombocythemia, is one of the myeloproliferative disorders, characterized by overproduction of platelets by megakaryocytes in the bone marrow without an identifiable cause. The platelet counts exceed 600,000/μL, but the platelets do not function properly. Symptoms and signs include headache; bleeding from gums, nose, and gastrointestinal tract; throbbing and burning pain of the hands and feet caused by thrombosis of small arterioles; and splenomegaly (enlarged spleen). Treatment includes using low-dose aspirin to control headache and other vasomotor symptoms and anticancer agents such as hydroxyurea to maintain proper platelet count. Bone marrow smears show increased numbers of megakaryocytes. Large platelets, similar in size to red blood cells, may be found in the peripheral blood.

B

Promyelocyte

Promyelocyte

D. Cui

Promyelocyte

Figure 8-14A. Promyelocytes, bone marrow smear. Wright stain, 710; inset 1,569

Promyelocytes (also called progranulocytes) have a round or oval-shaped nucleus with one to three nucleoli. The cytoplasm is light blue, containing some dark purple granules (azurophilic granules). At this stage, specific granules have not been produced. Promyelocytes vary in size and are produced by the division of myeloblasts. Their nuclei have a smooth, fine texture because of the fact that most of the chromatin is euchromatin, which is delicately dispersed.

Promyelocytes divide to form myelocytes.

Figure 8-14B. Myelocytes, bone marrow smear. Wright stain, 710; inset 1,569

The myelocyte has an oval or kidney-shaped nucleus that has no nuclei and a coarse texture because of increasing heterochromatin content. The cytoplasm contains azurophilic granules and specific granules. In this stage, the cell has stopped producing azurophilic granules. In addition, these granules have also been diluted during cell division, so they are not as prominent as in the promyelocyte stage. At the same time, the cell has begun to make and accumulate specific granules so that the cytoplasm starts to take the characteristic appearance of a mature granulocyte. The specific granules in neutrophilic myelocytes are small and appear pinkish, granules in eosinophilic myelocytes are large and stain red, and granules in basophilic myelocytes are blue-purple. Basophilic myelocytes are least numerous and, therefore, difficult to find. In general, myelocytes are smaller than promyelocytes, and their nuclear shape and size may vary. This stage is the longest in granulocytopoiesis.

D. Cui

Eosinophilic metamyelocyte

Figure 8-14C. Metamyelocytes, bone marrow smear.

Wright stain, 710; inset 1,569

Metamyelocytes are small cells. They have condensed nuclei, which are elongated with various degrees of indentation and contain clumped chromatin. Metamyelocytes are unable to divide (myelocytes are the last stage in cell division). The cytoplasm of metamyelocytes contains both types of granules. Their cytoplasm and granules are similar to those of mature granulocytes. At this stage, cells have their distinguishing features, but their nuclei have not yet become segmented.

Eosinophil

Figure 8-15B. Bone marrow cells, bone marrow smear. Wright stain, 710

This is an example of blood cells at various stages of development in the bone marrow, which includes both the erythrocyte and the granulocyte series. These cells, at various stages of development, are densely packed together and can be found randomly distributed in the bone marrow. During the maturation process, the cell size becomes smaller and nuclei become denser. In the erythropoiesis series, the cytoplasm of cells becomes light blue and then more pink, and nuclei become much denser and smaller and finally disappear. In the granulocytopoiesis series, the cytoplasm becomes less blue, primary (nonspecific) granules are produced, and then specific granules are produced and are present in myelocytes, which gives these cells the appearance characteristic of their identity as a neutrophil, eosinophil, or basophil. In other changes, nuclei become progressively denser, and the shape changes from round to oval, elongated, indented, and then lobed (segmented).

Blood cells

Reticular tissue

Adipocyte

spaces

C

Figure 8-15C. Bone marrow, bone marrow smear. Wright stain, 35; inset 184

Bone marrow is a specialized example of a reticular connective tissue, a loose connective tissue in which numerous cells are supported by a delicate network of reticular fibers. It resides in cavities within bones (see Figs. 5-8, 5-10, and 5-11). Bone marrow can be categorized into red bone marrow and yellow bone marrow. The term red bone marrow denotes active hematopoiesis; yellow bone marrow refers to a marrow composed chiefly of adipocytes (fat cells). Pictured is a smear of red bone marrow, which contains many developing blood cells, a few adipocytes, and some thin-walled blood vessels (sinusoidal capillaries). The red bone marrow is organized into a hemopoietic compartment and a vascular compartment. The hemopoietic compartment is a network of reticular fibers in which immature and mature blood cells are suspended. The vascular compartment is composed of mainly sinusoidal capillaries, which allow mature blood cells to enter the blood circulation.