Normal bone marrow histology

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CHAPTER 3 Normal bone marrow histology

Bone marrow structure

The constituents of the normal BM are closely packed within a hard bony ‘container’. Hemopoiesis occurs in the intertrabecular space within marrow cavities. The bony trabeculae (cancellous bone) are lined by endosteum, osteoblasts and osteoclasts. The stromal elements form an extensive, closely woven network in which the hematopoietic precursors are embedded, attached in various ways and to different components by the adhesive proteins and by other cells, such as the central macrophages in the erythroid islands. Hematopoietic precursors receive their nutrients, vitamins, hormones, regulatory factors, cytokines and modulators through the extracellular matrix (ECM), which also contributes to the regulation of the cell cycle, cellular differentiation and apoptosis. The blood supply to the BM consists of two systems: periosteal arteries, which give off branches to the BM after they penetrate the bone, and nutrient arteries. Blood drains from the BM cavity through central veins. The BM receives approximately 2–4% of cardiac output. The microvasculature of the BM comprises a network of sinusoids. Hemopoiesis only occurs in the interstital space between these sinusoids, thereby ensuring that hemopoietic progenitors are located close to the blood supply. Normal BM contains a network of fine branching reticulin fibers between parenchymal cells, which provide the extracellular matrix for the BM. There is a higher concentration of thicker fibers around arterioles and near the endosteum. The BM also has a nerve supply.

Bone marrow trephine biopsy

The process of obtaining a bone marrow trephine biopsy (BMTB) originates in the ancient procedure of trepanning.1 Prior to the advent of BMTB needles, clot preparations of aspirated marrow were prepared for diagnostic purposes. BM biopsies were obtained only as a means of diagnosis if marrow was inaspirable, a ‘dry tap’. The modification of needles by Jamshidi in the 1970s revolutionized the process of obtaining intact cores of bone and bone marrow for examination, primarily from the pelvis. The most common site biopsed is the posterior superior iliac crest. Other sites which can be biopsied include the anterior superior iliac crest, tibia, and vertebrae. Biopsy of the sternum is contraindicated due to the significant morbidity and mortality associated with this practice. The optimal length of a BMTB is 2–3 cm; shorter biopsies may not be representative and may not detect diseases that have a focal or patchy pattern of BM involvement.2

Marrow cellularity

The BMTB is particularly useful for the assessment of marrow cellularity. This is the relative amount of BM cells to adipocytes, which is assessed subjectively and should be interpreted in the context of the age of the patient. The terms normocellular (normal for age), hypercellular (increased cellularity for age) and hypocellular (reduced cellularity for age) are used. Cellularity reduces with increasing age (Table 3.2).4,5 In practice, the formula (cellularity = 100 − patient age) can be applied for adults; however, it does not correlate with cellularity at the extremes of the age range. The intertrabecular spaces adjacent to the marrow cortex tend to be hypocellular and should not be assessed when determining overall BM cellularity.

Table 3.2 Cellularity ranges for various age groups

Age Cellularity
Newborn to 3 months 80–100%
Childhood 60–80%
20–40 years 60–70%
40–70 years 40–50%
>70 years 30–40%

Marrow architecture

The BMTB enables the assessment of bone marrow architecture, the distribution of cellular elements and the bone and stromal cells. The outermost elements of the biopsy are composed of collagenous periosteal connective tissue, followed by a zone of cartilage or cortical bone (depending on the age of the patient). After this the bone breaks up into a meshwork of trabeculae, between which are the intertrabecular spaces. Hemopoietic cells are present within these intertrabecular spaces and are supported by fat cells, stromal cells, histiocytes extracellular matrix and blood vessels (Fig. 3.2). The hemopoietic cells are located within the intertrabecular spaces. The intertrabecular areas can be divided into three zones which contain different hemopoietic cell types (Fig. 3.3):

Small arteries and arterioles are often seen in the intermediate and central zones; these may be surrounded by cuffs of immature myeloid cells around them.

Hemopoiesis

The process of formation of blood elements from hemopoietic stem cells has been described in detail in Chapter 2. The BMTB enables the visualization of the spatial localization of the individual cell lineages during their development. Hemopoietic progenitors are present in cords, islands or clusters. Fully mature erythroid and granulocytic cells and platelets migrate through the sinusoidal endothelial cells to enter the bloodstream.

Erythropoiesis

Erythroid progenitors are found in small and large ‘islands’ called erythroid colonies within the intermediate and central zones of the marrow cavity. Erythroid islands are made up of concentric circles of immature erythroblasts (proerythroblasts) and a spectrum of maturing erythroid precursors leading to the late erythroblasts. Each erythroid island has a central iron-containing macrophage. The most primitive erythroid progenitor cells are present centrally around the macrophage and the maturing forms towards the periphery6 (Fig. 3.4). The central macrophage possesses dendritic processes, which extend between the maturing erythroid precursors. Its function is to support and nurture the erythroblasts, act as a source of iron and remove debris from dying cells and extruded nuclei. The central macrophage is often difficult to identify in histologic sections. Erythroid precursors are easily identified by being in distinct islands with cells of varying maturity, their almost perfectly round nuclei and by a perinuclear halo, an artifact of fixation and processing.

Proerythroblast. The earliest recognizable erythroid precursors (proerythroblasts) are medium to large round cells with minimal cytoplasm, large round nuclei with dispersed or open chromatin, many small nucleoli and a crisp nuclear membrane. A rim of weakly basophilic cytoplasm with a halo is also present (Fig. 3.5A).

Maturing erythroblast (also called normoblast). These are smaller than proerythroblasts, and differ in their nuclear and cytoplasmic characteristics. As a rule, with maturation, nuclear size reduces and the amount of cytoplasm increases. The nuclear chromatin becomes more condensed and acquires a uniform, condensed, hyperchromatic ‘ink dot’ appearance. It is this nuclear characteristic that enables late normoblasts to be distinguished from lymphocytes. As hemoglobin forms, the cells acquire rims of pale pink cytoplasm (orthochromatic erythroblast) which with further maturation acquires the crisp orangiophilia of mature RBCs (Fig. 3.5B).

Red blood cell. This is the terminally differentiated and most mature erythroid cell. Morphologically, it is an anucleate, orange biconcave disc, with an average size of about 8 µm.

Erythroid cells can be identified by IHC using antibodies to glycophorin A (CD235) or C and intracellular hemoglobin. Glycophorin A highlights both nucleated erythroid precursors and RBCs (Fig. 3.6) while hemoglobin A tends to be restricted to hemoglobinized nucleated erythroid precursors. In situ hybridization can also be performed using probes for hemoglobin A.