9 Blood and bone marrow

Blood is a liquid connective tissue that circulates throughout the body in the blood vessels, delivering many substances critical for life to their cellular destinations. Blood carries electrolytes such as sodium, potassium, calcium, magnesium and other ions; glucose, amino acids, fats, vitamins, antibodies, hormones and hundreds of other molecules. Only multicellular animals use blood to distribute all of these important molecules to each and every cell within the organism. Single-celled organisms do not need blood; they receive these substances directly from the environment. But that also means they can only survive in water. We humans carry our water around within our circulatory system, and that allows us to lead a more independent existence.

Composition of blood

As with any connective tissue, blood is composed of cells and extracellular matrix. Cells of the blood include:

  • red blood cells (a.k.a. erythrocytes)
  • white blood cells (a.k.a. leukocytes)
  • platelets (a.k.a. thrombocytes)

Platelets are not actually cells but fragments detached from large cells called megakaryocytes that reside in the bone marrow. Cells and cell fragments of blood are collectively called the formed elements of blood.

The extracellular matrix of blood is called plasma and is a composition of water, blood proteins and all inorganic and organic substances the blood carries.

Blood drawn from the body can be fractionated by centrifugation and usually separates into three layers based on the density.

  • The bottom layer contains red blood cells. It is red in color and usually constitutes 40-55% of the blood volume.
  • The middle layer contains white blood cells and platelets and is called the buffy coat. It is milky, opaque and is only a few millimeters deep. It is only about 1% of the blood volume.
  • The upper layer is plasma, and it is translucent and yellow. It is 45-60% of the blood volume.
    Figure 1: Layers of centirifuged blood

Cells of the blood

Cells of the blood, erythrocytes, leukocytes and cell fragments called thrombocytes, are produced in the red bone marrow by a process called hemopoiesis. All blood cells, both red and white, develop from the same pluripotent hemopoietic stem cell. Upon maturation, they exit to blood and either circulate in blood vessels where they perform their function (erythrocytes) or travel to their destination in the circulatory system and leave circulation to take up residence in other connective tissues of the body (white blood cells). Recall how connective tissue proper had many resident immune cells such as macrophages, lymphocytes, and other white “blood” cells.

Hemopoiesis

Hemopoiesis subchapter adapted from BC OpenStax Anatomy and Physiology book under the CC BY licence.

Hemopoiesis begins when the hemopoietic stem cell is exposed to appropriate chemical stimuli, collectively called hemopoietic growth factors, which prompt it to divide and differentiate. One daughter cell remains, a hemopoietic stem cell, allowing hemopoiesis to continue. The other daughter cell becomes one of two types of more specialized stem cells:

  • Lymphoid stem cells
  • Myeloid stem cells
Figure 2: Hemopoiesis in the bone marrow. Hemopoiesis is the production of the formed elements of blood.

Lymphoid stem cells give rise to a class of leukocytes known as lymphocytes, which include the various T cells, B cells and natural killer (NK) cells, all of which function in immunity. However, hemopoiesis of lymphocytes progresses somewhat differently from the process for the other formed elements. In brief, lymphoid stem cells quickly migrate from the bone marrow to lymphatic tissues, including the lymph nodes, spleen and thymus, where their production and differentiation continues. B cells are so named because they mature in the bone marrow, while T cells mature in the thymus.

Myeloid stem cells give rise to all of the other formed elements, including the erythrocytes; megakaryocytes that produce platelets; and a myeloblast lineage that gives rise to monocytes and three forms of granular leukocytes:  neutrophils, eosinophils, and basophils.

Lymphoid and myeloid stem cells develop into mature blood cells in several intermediate steps of precursor cells that have names ending with -blast. For instance, megakaryoblasts are the precursors of megakaryocytes, and proerythroblasts become reticulocytes, which eject their nucleus before maturing into erythrocytes. Notice that erythrocytes and white blood cells called granulocytes develop from myeloid stem cell line while lymphoid stem cell line leads only to one type of white blood cells, the lymphocytes.

Hemopoietic Growth Factors

The path the stem cells take depends on the hemopoietic growth factors. These include the following:

  • Erythropoietin is a hormone secreted by the the kidneys in response to low oxygen levels. It prompts the production of erythrocytes.
  • Thrombopoietin, is a hormone produced by the liver and kidneys. It triggers the development of megakaryocytes into platelets.

Erythrocytes

Erythrocytes are the most numerous blood cells. Erythrocytes are uniform in size, 7 to 8 μm in diameter. Because their size is relatively consistent, they can be used to estimate the size of other cells and are referred to as the “histological ruler.”

Human erythrocytes have the shape of biconcave disks that are thinner in the middle than at the periphery. The biconcave shape provides a greater surface for gas exchange and makes it easier for erythrocytes to bend as they pass through narrow capillaries that are frequently half the size of the erythrocyte. The shape of the erythrocyte is maintained by spectrin, a cytoskeletal fiber that provides flexibility and mechanical strength.

Erythrocytes lack nuclei and most organelles, and they are filled by hemoglobin, an iron-bound protein that binds oxygen and gives them their red color. Erythrocytes eject the nucleus and lose most of their other organelles before they enter circulation. However, especially when the production of erythrocytes is sped up due to a change in oxygen concentration or bleeding, immature erythrocytes, known as reticulocytes, can contain remnants of organelles, which appear as delicate smudges resembling a net (reticulum). Reticulocytes lose the remaining organelles within 1-2 days in circulation. Reticulocytes should not exceed 1-2% of the erythrocyte count.

Figure 3: Erythrocytes in the blood smear. Reticulocytes with visible net like remnants of the organelles are marked with arrows.

Erythrocytes are naturally red but also stain pink with eosin, which augments their color in histological slides. Because of their shape, the middle of the disc seems to have a lighter color than the ring found on the periphery.

Leukocytes

The leukocytes, also known as white blood cells, are a major component of the immune system. Although leukocytes and erythrocytes both originate from hematopoietic stem cells in bone marrow, they are very different from each other.

  1. Leukocytes are less numerous in circulation as compared to erythrocytes, but that does not mean there are less of them in the body. The majority of white blood cells reside in connective tissues, where they defend the organism against infection or destroy the organism’s own defective cells, and in secondary lymphatic organs such as the spleen and lymph nodes.
  2. In contrast to erythrocytes that are uniform, there are several different categories of leukocytes, and they even develop from two separate stem cell lines that split early in the process of hemopoiesis. The neutrophils, basophils and eosinophils, as well as monocytes, originate from the myeloid cell line, while lymphocytes, including a special lymphocyte called a Natural Killer cell (NK), arise from the lymphoid stem cell line, as seen in Figure 3.
  3. All leukocytes have nuclei, and most of them have granules in the cytoplasm. Leukocytes are practically invisible in microscopy and need to be stained. Historically, leukocytes were named based on the presence or absence of granules and their preference for absorbing dyes. That’s why they have names such as basophils (cells that have granules that like and absorb basic dyes) or acidophils (if they like and absorb acidic dyes).

Based on the presence of granules, and how they absorb stains, leukocytes are subclassified into:

  • Granulocytes – cells with prominent granules
    • neutrophils
    • eosinophils
    • basophils
  • Agranulocytes – cells that lack specific granules
    • monocytes
    • lymphocytes (remember that lymphocytes derive from another stem cell line as well)

Agranulocytes might have some visible spots (granules) that are smaller and stain azure, which are lysosomes. They are easy to distinguish from large granules containing active substances that granulocytes secrete to fight invaders.

How to recognize blood cells under the microscope

Blood smear

Blood cells can be observed under the microscope as a blood smear, that is, a droplet of blood spread on the glass slide to the thickness of a single layer of cells. The most abundant elements are erythrocytes. Figure 4 shows the normal distribution of cells found in blood. While there are numerous erythrocytes, only two white blood cells are visible in the image, along with a few tiny purple dots visible between erythrocytes, which are platelets.

Figure 4: Blood smear at low magnification.

A complete blood count (CBC) is the most commonly ordered laboratory test. A typical CBC laboratory panel includes the following:

  • White blood cell (WBC) count
  • Leukocyte types count (WBC differential). The white blood cells counted are neutrophils, eosinophils, basophils, lymphocytes and monocytes.
  • Erythrocytes (red blood cell, RBC) count
  • Hematocrit (HCT; also called packed cell volume PCV). Hematocrit indicates the percentage of erythrocytes in the blood sample
  • Hemoglobin (Hgb) concentration
  • Erythrocyte indices such as, mean corpuscular volume (MCV), which relates to the size of the red blood cells; mean corpuscular hemoglobin (MCH), which shows the amount of hemoglobin in an average erythrocyte
  • Thrombocyte (platelet) count

White Blood Cell Differential

Figure 5: Illustration of the shapes and relative sizes of white blood cells

Neutrophils

Neutrophils are the most numerous white blood cells as well as the most common granulocytes. Neutrophils measure 10 to 12 μm in diameter in blood smears and are obviously larger than erythrocytes. They are easily identified by their multi-lobed nucleus. Mature neutrophils have two to four lobes joined by strands, but it’s important to know that the nucleus arrangement changes in living neutrophils, including the position or even the number of lobes. They have a wide ring of almost colorless cytoplasm due to their lack of dye absorption because they are neutral toward both stains.

Another interesting fact is that in women, the Barr body (the condensed, single, inactive X chromosome) forms a drumstick-shaped appendage on one of the nuclear lobes. So you can even recognize the sex of the blood smear’s owner.

  • shape of the nucleus
  • neutral granules
  • twice as big as red blood cells
Figure 6: Illustration of a neutrophil
Figure 7: Neutrophil as seen under the microscope

Eosinophils

Eosinophils are named for the eosinophilic granules in their cytoplasm that appear bright pink in blood smear. Human eosinophils are about the same size as neutrophils, and they have bi-lobed nuclei. The cytoplasm of eosinophils contains numerous, slightly elongated granules that are much smaller compared with those of basophils.

Eosinophils are rare in the blood smear as they constitute only 2-4% of the leukocytes.

  • pink stained granules
  • bi-lobar nucleus
  • twice the size of red blood cells
Figure 8: Illustration of a eosinophil
Figure 9: Eosinophil as seen under the microscope

Basophils

Basophils are dark purple to blue in color because the numerous large granules in their cytoplasm stain with basic dyes. They are similar in size to neutrophils and have a single, centrally located nucleus that might be hard to see if covered by granules.

Basophils are the least numerous of the leukocytes and account for 0.5-1% of leukocytes.

  • Dark blue granules
  • twice as big as erythrocytes
  • single nucleus
Figure 10: Illustration of a basophil
Figure 11: Basophil as seen under the microscope

Monocytes

Monocytes have a diameter of 18 μm and are the largest of the white blood cells present in a blood smear. The nucleus of the monocyte is typically indented. Both the size and shape of the nucleus are useful indicators when differentiating them from lymphocytes which are about half their size and have a less indented nucleus.

Although monocytes are classified as agranular, they contain small, dense, azurophilic granules that are actually lysosomes and contain digestive enzymes.

Monocytes are not common in the blood, as they only travel from the bone marrow to the body tissues where they differentiate into macrophages and osteoclasts. Monocytes remain in the blood for only 3 days and account for approximately 3-8% of white blood cells.

  • three times as large as red blood cells
  • single nucleus
  • no granules in cytoplasm

 

Figure 12: Illustration of a monocyte
Figure 13: Monocyte as seen under the microscope

Lymphocytes

Lymphocytes are the most common agranular white blood cells. A mature lymphocyte is 6 to 15 μm in diameter, a size similar to red blood cells. Most of the circulating lymphocytes are small, and their size is a great characteristic to tell them apart from monocytes that are about three times as large.

Lymphocytes have an intensely stained, spherical nucleus. The cytoplasm appears as a very thin, pale blue ring surrounding the nucleus. In general, there are no visible granules.

Lymphocytes originate in the bone marrow from lymphoid stem cells, and travel to the peripheral lymphatic tissues. The ones seen in blood are simply in transit. Lymphocytes can divide and multiply outside of the bone marrow, and differentiate into other types of immune cells such as plasma cells that actively produce antibodies, or all kinds of helper and memory cells.

They are also different from other leukocytes because they can exit and re-enter blood vessels.

Lymphocytes account for about 30% of the total white blood cells, and their number increases when the immune response is activated.

  • the size of red blood cells
  • single dark nucleus
  • ring of cytoplasm at the periphery
Figure 14: Illustration of lymphocytes
Figure 15: Lymphocytes as seen under the microscope

Thrombocytes

Thrombocytes (platelets) are small cytoplasmic fragments circulating in the blood and involved in blood clotting. They appear as disc-like structures about 2 to 3 μm in diameter. They have a plasma membrane but do not have nuclei.

Thrombocytes are cell fragments that separated from the megakaryocyte, a very large cell located in the bone marrow. Megakayocytes are polyploid cells (their nuclei contain multiple sets of chromosomes). During platelet formation, small bits of cytoplasm separate from the periphery of the megakaryocyte to form individual platelets. After entry into the circulatory system the platelets live for approximately 10 days.

 

Figure 16: Megakaryocyte in the bone marrow (in the middle). The arrows indicate trabeculae of the spongy bone.

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Histology Copyright © by Malgosia Wilk-Blaszczak. All Rights Reserved.

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