Pathology 104 M. Hall
Dental Histology
Fall 2003
BONE MARROW
(G & H, Chap. 10, pp 237-250)
The cells that are seen in peripheral blood, both white and red, have to be synthesized somewhere else and released into the blood. A steady supply of these cells must be maintained in order to replace those that are destroyed, die of old age, or die while performing their daily duties.
The medullary cavities of long bones, and the spaces between the trabeculae of spongy bone, is where the process of hemopoiesis (blood formation) takes place. Bone has a rich vascular supply which gives rise to an extensive network of large sinusoids (sinusoidal capillaries) in the marrow cavities. The spaces between the sinusoids are filled with islands of hemopoietic cells where the various types of blood cells are continuously formed. Mature blood cells enter and leave the bone marrow through these sinusoids.
The cells in red bone marrow tissue are very diverse, because they comprise the different stages in the development of seven types of blood cells—erythrocytes, neutrophils, basophils, eosinophils, lymphocytes, monocytes and megakaryocytes (which form platelets). Additionally numerous macrophages are found which destroy malformed cells, as well as the extruded nuclei of erythrocyte precursor cells. A histological smear of bone marrow, which you shall study in the laboratory, shows a complex jumble of these cells. Bone marrow also contains large numbers of reticular cells which accumulate fat in their cytoplasm, and thus resemble adipose cells. At birth, bone marrow is red, due to the large number of erythrocytes being produced there, and is called red marrow. As we age, the diaphyses of long bones accumulate more and more fat in the reticular cells, while hemopoiesis slows down, and the marrow cavity is filled with yellow marrow. Hemopoiesis then continues in spongy bone throughout life.
All blood cells arise from a common pluripotential hemopoietic stem cell (PHSC). When these divide they give rise to two types of multipotential hemopoietic stem cells (MHSC’s), CFU-S and CFU-Ly, which then divide to give rise to unipotential progenitor cells. Progenitor cells divide to give rise to precursor cells, which differentiate into a single mature blood cell type.
Stem Cells have the appearance of small lymphocytes, with small round nuclei and very little cytoplasm, and in this form they do not divide. However, when conditions are right, this inactive stem cell can grow back into a large blast form after which it can divide and carry out its specific function. PHSC’s comprise less than 0.1% of the cells of bone marrow. When they are stimulated to divide they give rise to more PHSC’s as well as to two types of MHSC’s—those that give rise to T and B lymphocytes (CFU-Ly) and those that give rise to all the other blood cells (CFU-S).
Colony Forming Units (CFU’s). If bone marrow cells are spread across a culture dish under conditions that promote cells division and growth, some cells give rise to colonies of cells. These cells are called “colony forming units” or CFU’s. Some CFU’s are true stem cells (CFU-LY and CFU-S). They can divide indefinitely and produce colonies containing all the types of blood cells. Other CFU’s give rise to only one lineage of blood cells (e.g erythrocytes, neutrophils etc.), and these CFU’s are called progenitor cells. Thus a CFU-E produces only erythrocytes while a CFU-N produces only neutrophils.
Progenitor Cells also resemble small lymphocytes and divide only now and then to replenish the dwindling supply of precursor cells. However they are already committed to a single blood cell lineage—they are unipotential. Progenitor cells are programmed to enter a specific lineage long before they begin to mature into that final cell type
Stem cells and progenitor cells are inactive most of the time, instead of continuously dividing. Most cell division is done by precursor cells on their way to becoming mature blood cells
Precursor Cells. When progenitor cells divide, they give rise to precursor cells, such as proerythroblasts, myeloblasts etc, which are incapable of self renewal. Precursor cells divide actively, but they continue to steadily mature, thus removing themselves from the pool. For example, a pool of dividing erythroblasts will disappear within 10 days unless stem cells add new proerythroblasts to the pathway.
Generally cells become more distinctive as they approach their final forms, and thus the most mature precursor cells are the easiest to recognize. The early precursor cells (myeloblast and promyelocyte) of the granulocyte lineage, look very similar to the untrained eye and it is impossible to tell which lineage one of these early cells belongs to.
Colony Stimulating Factors (CSF)
Hemopoiesis is regulated by numerous colony stimulating factors (hormones) produced by a variety of cell types within and outside of the bone marrow. These factors are glycoproteins. Other stimulating factors are produced by stromal cells of the bone marrow, inserted into their plasma membranes and thus require contact between stem cells and the stromal cells for activation to occur. Each of the many factors that have been identified acts on specific stem cells, progenitor cells and precursor cells, inducing mitosis, differentiation or both. Thus CSF’s regulate the number of new stem cells, progenitor cells and precursor cells that enter into the developmental pathway, as well as their differentiation, or both. (A much abused CSF is erythropoietin (EPO), which stimulates the production of erythrocytes. Endurance athletes, such as marathon runners, will inject erythropoietin in order to increase the number of erythrocytes and thus the oxygen-carrying capacity of the blood, thus increasing their endurance. This can also be achieved by training at high altitude, which also results in an increase in the number of erythrocytes/ml of blood).
Megakaryocytes are the terminally differentiated forms of the unipotential platelet progenitor, CFU-Meg. These are very large cells (up to 100µM) which are formed by repeated division of the nucleus, without accompanying cell division. Thus the nucleus becomes polyploid, as much as 64N. Megakaryocytes bud off parts of their cytoplasm into the sinusoids, forming platelets, which play an important role in blood coagulation. Platelets are small (2 – 4 µM),and anucleate, but contain all the other normal cytoplasmic organelles. Platelets function in limiting hemorrhage in case of injury. If the endothelial lining of a blood vessel is disrupted, platelets become activated, release the contents of their granules, adhere to the damaged region of the vessel wall and to each other. Interactions of tissue factors, plasma-borne factors and platelet-derived factors form a blood clot.
Proerythroblast ® erythroblast ® normoblast ® reticulocyte ® erythrocyte
Proerythroblasts are the earliest recognizable cells that are committed to become erythrocytes. (Remember that they are derived from progenitor cells). They have intensely basophilic cytoplasm due to the large numbers of ribosomes that will become involved in the synthesis of hemoglobin.
Proerythroblasts actively divide and differentiate with the following changes:
Cell volume decreases
Nucleus shrinks and becomes denser
Basophilia decreases due to decrease in number of ribosomes
Hemoglobin content increases resulting in increase in acidophilia
Early erythroblasts are called ‘basophilic erythroblasts’ while later stages are called ‘polychromatophilic (many colored) erythroblasts’. The change in the color of erythroblasts results from a decrease in the number of ribosomes and an increase in the content of hemoglobin as the cell matures. All erythroblasts have a round, so-called ‘checkerboard’ nucleus.
Normoblasts (orthochromatophilic erythroblasts) have stopped dividing, but continue to differentiate. They have a small, darkly stained nucleus and reddish/yellow cytoplasm.
Reticulocytes have no nucleus, bur contain remnants of RNA as well as mitochondria, lysosomes, vesicles etc. They destroy these over the following 3 days to become erythrocytes, which enter the blood stream through the sinusoids.
Erythrocytes have no RNA or intracellular organelles. They circulate in the blood for 120 days and then are removed by macrophages in the spleen and liver.
Myeloblast ® promyelocyte ® myelocyte ® metamyelocyte ® granulocyte
Myeloblasts have a generalized ‘blast’ appearance, with a large, pale, round nucleus, several nucleoli and pale bluish cytoplasm with no granules or visible features.
Promyelocytes resemble myeloblasts but have a few non-specific granules in their cytoplasm. It is not certain whether there is a common promyelocyte that gives rise to all of the granulocytes, or whether there are specific promyelocytes for each lineage. They divide actively. (Here is one of the many ‘exceptions to the rule’ that you will encounter in histology—‘blasts’ divide and ‘cytes’ do not—“yeah sure”!!).
Myelocytes are lineage specific. They can be recognized by the presence of specific granules in their cytoplasm. They also divide actively. (yet another exception to the rule!).
Metamyelocytes are post mitotic (whew!). Their nucleus has begun to take on bean-shaped/horse-shoe-shaped appearance, and their specific granules are very visible.
Granulocytes (mature cells) generally have a multi-lobed nucleus, and contain large numbers of specific granules They are stored in the marrow and move out into the circulation when needed, from where they find their way into the connective tissues.
Monoblasts and lymphoblasts
Monoblast ® promonocyte ® monocyte ® macrophage
Lymphoblast ® prolymphocyte ® lymphocyte
Cells of these developmental series can only be recognized in bone marrow with special stains. However, they are easily recognizable in peripheral blood and in connective tissues.