Erythropoiesis

Erythropoiesis is the process by which red blood cells (erythrocytes) are produced. In human adults, this usually occurs within the bone marrow. In the early fetus, erythropoiesis takes place in the mesodermal cells of the yolk sac. By the third or fourth month, erythropoiesis moves to the spleen and liver. In humans with certain diseases and in some animals, erythropoeiesis also occurs outside the bone marrow, within the spleen or liver. This is termed extramedullary erythropoiesis. The tibia and femur cease to be important sites of hematopoiesis by about age 25; the vertebrae, sternum, and ribs continue to produce red blood cells throughout life.

Erythrocyte differentiation
In the process of red blood cell maturation, a cell undergoes a series of differentiations. The following stages of development all occur within the bone marrow: 1. 2. 3. 4. 5. 6. 7. 8. pluripotent hematopoietic stem cell multipotent stem cell unipotential stem cell pronormoblast basophilic normoblast/early normoblast polychromatophilic normoblast/intermediate normoblast orthochromatic normoblast/late normoblast reticulocyte

After these stages, the cell is released from the bone marrow, and ultimately becomes an "erythrocyte" or mature red blood cell circulating in the peripheral blood. These stages correspond to specific appearances of the cell when stained with Wright's stain and examined by light microscopy, but correspond to other biochemical changes. In the process of maturation a basophilic pronormoblast is converted from a cell with a large nucleus and a volume of 900 m3 to an enucleated disc with a volume of 95 m3. By the reticulocyte stage, the cell has extruded its nucleus, but is still capable of producing hemoglobin.

Regulation of erythropoesis
A feedback loop involving erythropoietin helps regulate the process of erythropoiesis so that, in non-disease states, the production of red blood cells is equal to the destruction of red blood cells and the red blood cell number is sufficient to sustain adequate tissue oxygen levels but not so high as to cause sludging, thrombosis, or stroke. Erythropoeitin is produced in the kidney and liver in response to low oxygen levels. In addition, erythropoeitin is bound by circulating red blood cells; low circulating numbers lead to a relatively high level of unbound erythropoeitin, which stimulates production in the bone marrow. Recent studies have also shown that the peptide hormone hepcidin may play a role in the regulation of hemoglobin production, and thus effect erthropoiesis. The liver Hand Out Hematologi 1-7 1

produces hepcidin. Hepcidin controls iron absorption in the gastrointestinal tract and iron release from reticuloendothelial tissue.1 Iron must be released from macrophages in the bone marrow to be incorporated into the heme group of hemoglobin in erythrocytes.

Red blood cell

Human red blood cells Red blood cells are the most common type of blood cell and the vertebrate body's principal means of delivering oxygen from the lungs or gills to body tissues via the blood. Red blood cells are also known as RBCs or erythrocytes (from Greek erythros for "red" and kytos for "hollow", with cyte nowadays translated as "cell"). A schistocyte is a red blood cell undergoing fragmentation, or a fragmented part of a red blood cell.

Erythrocytes

From left to right: erythrocyte, thrombocyte, leukocyte. Hand Out Hematologi 1-7 2

Erythrocytes consist mainly of hemoglobin, a complex molecule containing heme groups whose iron atoms temporarily link to oxygen molecules in the lungs or gills and release them throughout the body. Oxygen can easily diffuse through the red blood cell's cell membrane. Hemoglobin also carries some of the waste product carbon dioxide back from the tissues. (In humans, less than 2% of the total oxygen, and most of the carbon dioxide, is held in solution in the blood plasma). A related compound, myoglobin, acts to store oxygen in muscle cells. The color of erythrocytes is due to the heme group of hemoglobin. The red blood cells change color due to the state of the hemoglobin: when combined with oxygen the resulting oxyhemoglobin is scarlet and when oxygen has been released, the resulting deoxyhemoglobin is a dark maroon color. The blood plasma alone is straw-colored. The keeping of oxygen-binding proteins in cells (rather than having them dissolved in body fluid) was an important step in the evolution of vertebrates; it allows for less viscous blood and longer transport ways of oxygen.

Human erythrocytes
The diameter of a typical human erythrocyte disk is 68 m, much smaller than most other human cells. A typical erythrocyte contains about 270 million hemoglobin molecules, with each carrying four heme groups. Adult humans have roughly 23 1013 red blood cells at any given time (women have about 4 million to 5 million erythrocytes per cubic millimeter (microliter) of blood and men about 5 million to 6 million; people living at high altitudes with low oxygen tension will have more). Red blood cells are thus much more common than the other blood particles: There are about 4,00011,000 white blood cells and about 150,000 400,000 platelets in a cubic millimeter of human blood. The red blood cells store collectively about 3.5 grams of iron, more than five times the iron stored by all the other tissues combined.

Life cycle
The process by which red blood cells are produced is called erythropoiesis. Erythrocytes are continuously being produced in the red bone marrow of large bones, at a rate of about 2 million per second. (In the embryo, the liver is the main site of red blood cell production.) The production can be stimulated by the hormone erythropoietin (EPO), synthesised by the kidney; which is used for doping in sports. Just before and after leaving the bone marrow, they are known as reticulocytes which comprise about 1% of circulating red blood cells. Erythrocytes develop from stem cells through reticuloctyes to mature erythrocytes in about 7 days and live a total of about 120 days. The aging cells swell up to a sphere-like shape and are engulfed by phagocytes, destroyed and their materials are released into the blood. The main sites of destruction are the liver and the spleen. The heme constituent of hemoglobin is eventually excreted as bilirubin.

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Diseases and diagnostic tools

Affected by Sickle-cell disease, red blood cells alter shape and threaten to damage internal organs. Blood diseases involving the red blood cells include:

Anemias (or anaemias) are diseases characterized by low oxygen transport capacity of the blood, because of low red cell count or some abnormality of the red blood cells or the hemoglobin. o Iron deficiency anemia is the most common anemia; it occurs when the dietary intake or absorption of iron is insufficient, and hemoglobin, which contains iron, cannot be formed o Sickle-cell disease is a genetic disease that results in abnormal hemoglobin molecules. When these release their oxygen load in the tissues, they become insoluble, leading to mis-shaped red blood cells. These sickle shaped red cells are rigid and cause blood vessel blockage, pain, strokes, and other tissue damage. o Thalassemia is a genetic disease that results in the production of an abnormal ratio of hemoglobin subunits. o Spherocytosis is a genetic disease that causes a defect in the red blood cell's cytoskeleton, causing the red blood cells to be small, sphereshaped, and fragile instead of donut-shaped and flexible. o Pernicious anemia is an autoimmune disease wherein the body lacks intrinsic factor, required to absorb vitamin B12 from food. Vitamin B12 is needed for the production of hemoglobin. o Aplastic anemia is caused by the inability of the bone marrow to produce blood cells. o Pure red cell aplasia is caused by the inabilty of the bone marrow to produce only red blood cells. o Hemolysis is the general term for excessive breakdown of red blood cells. It can have several causes. The malaria parasite spends part of its life-cycle in red blood cells, feeds on their hemoglobin and then breaks them apart, causing fever. Both sickle-cell disease and thalassemia are more common in malaria areas, because these mutations convey some protection against the parasite. 4

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Polycythemias (or erythrocytoses) are diseases characterized by a surplus of red blood cells. The increased viscosity of the blood can cause a number of symptoms. o In polycythemia vera the increased number of red blood cells results from an abnormality in the bone marrow. Several microangiopathic diseases, including disseminated intravascular coagulation and thrombotic microangiopathies, present with pathognomonic (diagnostic) RBC fragments called schistocytes. These pathologies generate fibrin strands that sever RBCs as they try to move past a thrombus.

Several blood tests involve red blood cells, including the RBC count (the number of red blood cells per volume of blood) and the hematocrit (percentage of blood volume occupied by red blood cells). The blood type needs to be determined to prepare for a blood transfusion or an organ transplantation.

Stem cell

Stem cells are primal cells common to all multi-cellular organisms that retain the ability to renew themselves through cell division and can differentiate into a wide range of specialized cell types. Research in the human stem cell field grew out of findings by Canadian scientists Ernest A. McCulloch and James E. Till in the 1960s.[1][2] The three broad categories of mammalian stem cells are: embryonic stem cells, derived from blastocysts, adult stem cells, which are found in adult tissues, and cord blood stem cells, which are found in the umbilical cord. In a developing embryo, stem cells are able to differentiate into all of the specialized embryonic tissues. In adult organisms, stem cells and progenitor cells act as a repair system for the body, replenishing specialized cells. As stem cells can be readily grown and transformed into specialised cells with characteristics consistent with cells of various tissues such as muscles or nerves through cell culture, their use in medical therapies has been proposed. In particular, embryonic cell lines, autologous embryonic stem cells generated through therapeutic cloning, and highly plastic adult stem cells from the umbilical cord blood or bone marrow are touted as promising candidates.[3]

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Stem cell properties

Defining properties
The rigorous definition of a stem cell requires that it possesses two properties:

Self-renewal - the ability to go through numerous cycles of cell division while maintaining the undifferentiated state. Unlimited potency - the capacity to differentiate into any mature cell type. In a strict sense, this makes stem cells either totipotent or pluripotent, although some multipotent and/or unipotent progenitor cells are sometimes referred to as stem cells.

These properties can be illustrated in vitro, using methods such as clonogenic assays, where the progeny of single cell is characterized.[4][5] However, in vitro culture conditions can alter the behavior of cells, making it unclear whether the cells will behave in a similar manner in vivo. Considerable debate exists whether some proposed adult cell populations are truly stem cells.

Potency definitions

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Pluripotent, embryonic stem cells originate as inner mass cells with in a blastocyst. The stem cells can become any tissue in the body, excluding a placenta. Only the morula's cells are totipotent, able to become all tissues and a placenta. Potency specifies the differentiation potential (the potential to differentiate into different cell types) of the stem cell.

Totipotent stem cells are produced from the fusion of an egg and sperm cell. Cells produced by the first few divisions of the fertilized egg are also totipotent. These cells can differentiate into embryonic and extraembryonic cell types. Pluripotent stem cells are the descendants of totipotent cells and can differentiate into cells derived from the three germ layers. Multipotent stem cells can produce only cells of a closely related family of cells (e.g. hematopoietic stem cells differentiate into red blood cells, white blood cells, platelets, etc.). Unipotent cells can produce only one cell type, but have the property of selfrenewal which distinguishes them from non-stem cells.

Embryonic stem cells

Main article: Embryonic stem cell Embryonic celtic cell lines (ES cell lines) are cultures of cells derived from the epiblast tissue of the inner cell mass (ICM) of a blastocyst. A blastocyst is an early stage embryo - approximately 4 to 5 days old in humans and consisting of 50-150 cells. ES cells are pluripotent, and give rise during development to all derivatives of the three primary germ layers: ectoderm, endoderm and mesoderm. In other words, they can develop into each of the more than 200 cell types of the adult body when given sufficient and necessary stimulation for a specific cell type. They do not contribute to the extraembryonic membranes or the placenta. When given no stimuli for differentiation, ES cells will continue to divide in vitro and each daughter cell will remain pluripotent. The pluripotency of ES cells has been rigorously demonstrated in vitro and in vivo, thus they can be indeed classified as stem cells. Because of their unique combined abilities of unlimited expansion and pluripotency, embryonic stem cells are a potential source for regenerative medicine and tissue replacement after injury or disease. To date, no approved medical treatments have been derived from embryonic stem cell research. This is not surprising considering that many nations currently have moratoria on either ES cell research or the production of new ES cell lines.

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Adult stem cells

Adult stem cell

Stem cell division and differentiation. A - stem cell; B - progenitor cell; C differentiated cell; 1 - symmetric stem cell division; 2 - asymmetric stem cell division; 3 - progenitor division; 4 - terminal differentiation Adult stem cells are undifferentiated cells found throughout the body that divide to replenish dying cells and regenerate damaged tissues. Also known as somatic (from Greek , of the body) stem cells, they can be found in children, as well as adults. A great deal of adult stem cell research has focused on clarifying their capacity to divide or self-renew indefinitely and their differentiation potential.[6] Many adult stem cells may be better classified as progenitor cells, due to their limited capacity for cellular differentiation. Nevertheless, specific multipotent or even unipotent adult progenitors may have potential utility in regenerative medicine. The use of adult stem cells in research and therapy is not as controversial as embryonic stem cells, because the production of adult stem cells does not require the destruction of an embryo. In contrast with the embryonic stem cell research, more US government funding has been provided for adult stem cell research. Adult stem cells can be isolated from a tissue sample obtained from an adult. They have mainly been studied in humans and model organisms such as mice and rats.

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Lineage
To ensure self-renewal, stem cells undergo two types of cell division (see Stem cell division and differentiation diagram). Symmetric division gives rise to two identical daughter cells both endowed with stem cell properties. Asymmetric division, on the other hand, produces only one stem cell and a progenitor cell with limited self-renewal potential. Progenitors can go through several rounds of cell division before terminally differentiating into a mature cell. It is possible that the molecular distinction between symmetric and asymmetric divisions lies in differential segregation of cell membrane proteins (such as receptors) between the daughter cells, however, there is no evidence for this mechanism. An alternative theory is that stem cells remain undifferentiated from environmental cues in their particular niche. Stem cells differentiate when they leave that niche or no longer receive those signals. Studies in Drosophila germarium have identified the signals dpp and adherins junctions that prevent germarium stem cells from differentiating[7][8]. The signals that lead to reprogramming of cells to an embryonic-like state are also being investigated. These signal pathways include several transcription factors including the oncogene c-Myc. Initial studies indicate that transformation of mice cells with a combination of these anti-differentiation signals can reverse differentiation and may allow adult cells to become pluripotent.[9] However, need to transform these cells with an oncogene may prevent the use of this approach in therapy.

Treatments
Medical researchers believe that stem cell therapy has the potential to radically change the treatment of human disease. A number of adult stem cell therapies already exist, particularly bone marrow transplants that are used to treat leukaemia.[10] In the future, medical researchers anticipate being able to use technologies derived from stem cell research to treat a wider variety of diseases including cancer, parkinson's disease, spinal cord injuries, and muscle damage, amongst a number of other impairments and conditions.[11][12] However, there still exists a great deal of social and scientific uncertainty surrounding stem cell research, which could possibly be overcome through public debate and future research. Stem cells, however, are already used extensively in research, and some scientists do not see cell therapy as the first goal of the research, but see the investigation of stem cells as a goal worthy in itself. [13].

Controversy surrounding stem cell research

There exists a widespread controversy over stem cell research that emanates from the techniques used in the creation and usage of stem cells. Embryonic stem cell research is particularly controversial because, with the present state of technology, starting a stem cell line requires the destruction of a human embryo and/or therapeutic cloning. Opponents of the research argue that this practice is a slippery slope to reproductive cloning and tantamount to the instrumentalization of a human being. Contrarily, some medical researchers in the field argue that it is necessary to pursue embryonic stem cell research because the resultant technologies are expected to have significant medical potential, and that the embryos used for research are only those meant for destruction Hand Out Hematologi 1-7 9

anyway (as a product of invitro fertilisation). This in turn, conflicts with opponents in the pro-life movement, who argue that an embryo is a human being and therefore entitled to dignity even if legally slated for destruction. The ensuing debate has prompted authorities around the world to seek regulatory frameworks and highlighted the fact that stem cell research represents a social and ethical challenge.

Totipotency is the ability of a single cell to divide and produce all the differentiated cells in an organism, including extraembryonic tissues. Totipotent cells formed during sexual and asexual reproduction include spores and zygotes. Zygotes are the products of the fusion of two gametes (fertilization). In some organisms, cells can dedifferentiate and regain totipotency. For example, a plant cutting or callus can be used to grow an entire plant. Pluripotency in the broad sense refers to "having more than one potential outcome". In biological systems, this can refer either to cells or to biological compounds.

Cellular differentiation

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Cellular differentiation is a concept from developmental biology describing the process by which cells acquire a "type". The morphology of a cell may change dramatically during differentiation, but the genetic material remains the same, with few exceptions. A cell that is able to differentiate into many cell types is known as pluripotent. These cells are called stem cells in animals and meristematic cells in higher plants. A cell that is able to differentiate into all cell types is known as totipotent. In mammals, only the zygote and early embryonic cells are totipotent, while in plants, many differentiated cells can become totipotent with simple laboratory techniques. Multipotent progenitor cells can give rise to several other cell types, but those types are limited in number. An example of a multipotent stem cell is a hematopoietic cell a blood stem cell that can develop into several types of blood cells, but cannot develop into brain cells or other types of cells. At the end of the long series of cell divisions that form the embryo are cells that are terminally differentiated, or that are considered to be permanently committed to a specific function. Scientists have long held the opinion that differentiated cells cannot be altered or caused to behave in any way other than the way in which they have been naturally committed. New research, however, has even called that assumption into question. In recent stem cell experiments, scientists have been able to persuade blood stem cells to behave like neurons, or brain cells. Scientists now believe that stem cell research could reveal far more vital information about our bodies than was previously known. There is also continuing research to see if it is possible to make multipotent cells into pluripotent cells.

Development
Image Description

Pronormoblast

Basophilic normoblast

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Polychromatic normoblast (also polychromatophilic)

Orthochromatic normoblast (also orthochromatophilic)

Examples of the developing cells are shown below. Mature erythrocytes are visible in most pictures. A pure field of mature erythrocytes can also be seen by clicking here .

stem cell

late stem cell

proerthroblast (pronormoblast)

basophilic erythroblast

orthochromatic polychromatophilic orthocromatophilic erythroblast erythroblast extruding its erythroblast nucleus (Normoblast)

reticulocyte (polychromatophilic erthrocyte)

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Erythropoiesis
Erythropoiesis is the development of mature red blood cells (erythrocytes). Like all blood cells, erythroid cells begin as pluripotential stem cells. The first cell that is recognizable as specifically leading down the red cell pathway is the proerythroblast . As development progresses, the nucleus becomes somewhat smaller and the cytoplasm becomes more basophilic, due to the presence of ribosomes. In this stage the cell is called a basophilic erythroblast . The cell will continue to become smaller throughout development. As the cell begins to produce hemoglobin, the cytoplasm attracts both basic and eosin stains, and is called a polychromatophilic erythroblast . The cytoplasm eventually becomes more eosinophilic, and the cell is called an orthochromatic erythroblast . This orthochromatic erythroblast will then extrude its nucleus and enter the circulation as a reticulocyte . Reticulocytes are so named because these cells contain reticular networks of polyribosomes. As reticulocytes loose their polyribosomes they become mature red blood cells.

ERYTHROPOIESIS
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Erythropoiesis constitute 10-30% of hemopoietic cells in the bone marrow. The mature erythrocyte is derived from the stem cell, which differentiate to erythroid colony forming cells (BFU-E, CFU-E), and next to the proerythroblast, the first morphologically recognizable cell of the series. The proerythroblast matures to the basophilic normoblast, then the polichromatic normoblast, where synthesis of hemoglobin is started. At the end, the polychromatophilic normoblast matures to the orthochromatic normoblast. The orthochromatic normoblast loses its nucleus and developes into reticulocyte, which after 2-4 days develops into mature erythrocyte. The mature erythrocyte stays for ca. 4 months in the blood. Proerythroblasts Basophilic (early) normoblast

Polychromatic (intermediate) normoblas

Pycnotic (late) normoblast

Reticulocytes

Normocytes

Microcytes

Macrocytes

Megalocytes

Anisocytosis

Hypochromia

Polychromasia

Eliptocytes

Lacrymocytes

Target cells

Acanthocytes

Echinocytes

Crenated red blood cells

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Schistocytes

Stomatocytes

Spherocytes

Leptocytes

Anulocytes

Sickle cells

Poikilocytosis

Howell-Jolly bodies

Cabot rings

Basophilic stippling

Pappenheimer bodies

Normoblasts in blood

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