BONE MARROW: 1 GENERAL CONSIDERATIONS The assessment for a possible bone marrow exceeding one billion per kilogram per day for (BM) neoplasm typically begins with a review life. As expected, exquisite regulation of cell of the complete blood count (CBC) and the production is required via the coordinated inter- peripheral blood smear. Such an assessment is play of promotional and inhibitory cytokines, based on a knowledge of normal blood and BM largely created by the progeny of mesenchymal parameters, normal hematopoiesis, and normal stem cells such as fbroblasts, endothelial cells, BM architecture in terms of all hematolymphoid osteoblasts, and adipose cells (Table 1-1; fg. lineages, BM stroma, and bony trabeculae. Also 1-1) (1–14). required is at least a basic understanding of Hematopoietic stem cells are recruited to regulatory factors, including transcriptional specialized loci by mesenchymal cells as well regulators, cell surface receptors, growth factors, as the products of mesenchymal cells. These and other cytokines and receptors (1). hematopoietic stem cell niches are highly pro- The BM is the site of the massive produc- tected sites in which various regulatory factors tion of erythrocytes, platelets, and neutrophils, impact stem cell proliferation and maturation Table 1-1 OVERVIEW OF HEMATOPOIESISa BMb Microenvironmental Niches Mesenchymal stem cells – asymmetric cell division with self-renewal and multilineage mesodermal maturation (osteoblasts, fbroblasts, chondrocytes, endothelial cells, adipocytes) Bony trabeculae lined by osteoblasts create niches; produce factors to recruit, retain, and regulate HSC Sinusoidal vessels create niches; produce factors to recruit, retain, and regulate HSC Support matrix of reticulin fbers, extracellular matrix with soluble regulatory factors Macrophages (HSC derived) Regulatory Factors Produced locally by BM microenvironmental cells and macrophages Some produced systemically Bind to receptors on cell membrane Ligand-receptor binding initiates cascade of intracellular events resulting in increased proliferation of immature receptor-bearing cell, or enhanced functional activity of mature receptor-bearing cell Early acting multilineage cytokines include GM-CSF, IL-1, IL-3, IL-6 and FLT3 ligand Lineage specifc cytokines include EPO, TPO, and G-CSF Inhibitory factors include TGF-b and IFN-a Hematopoietic Stem Cells Very rare in BM; not morphologically recognizable Asymmetric cell division with self-renewal and multilineage progenitor cell production Common myeloid progenitor cell; all hematopoietic/dendritic cell lineages and osteoclasts Common lymphoid progenitor cell; all B, T, NK, and some dendritic cells Numerous transcription factors, microRNA and other factors critical in the regulation of hematopoiesis and lineage differentiation aData from references 1–7, 9–15, and 46. bBM = bone marrow; HSC = hematopoietic stem cell; GM-CSF = granulocyte macrophage colony-stimulating factor; IL = interleukin; FLT3 = fbroblast growth factor 3; EPO = erythropoietin; TPO = thrombopoietin; G-CSF = granulocyte colony- stimulating factor; TFG = transforming growth factor; IFN = interferon; CSF = colony-stimulating factor; M-CSF = macro- phage colony-stimulating factor. 1 Tumors of the Bone Marrow Figure 1-1 SCHEMATIC OF HEMATOPOIETIC STEM CELL NICHE The hematopoietic stem cell niche is often close to blood vessels. Stem cells are recruited to this site by mesenchymal stromal cells and the products of mesenchymal stromal cells (MSC = mesenchymal stem cell; HSC = hematopoietic stem cell; TPO = throm- bopoietin; G-CSF = granulocyte colony-stimulating factor; IL = interleukin; Angpt = angiopoietin; SCF = stem cell factor; SDF = stromal cell-derived factor). (Fig. 1.3 from Park C, Kharas M. The hematopoietic system and hematopoiesis. In: Orazi A, Foucar K, Knowles D, Weiss L, eds. Knowles’ Neoplastic Hematopathology, 3rd ed. Philadelphia: Lippincott Williams & Wilkins; 2014:13.) (1,9,10). Neoplastic cells may usurp normal discussed later in this chapter, the morpho- hematopoietic stem cell niches and infuence logic stages of maturation are well-established mesenchymal cells, and thus successfully efface for the myeloid and erythroid lineages. The normal BM and elude chemotherapy (15). None maturation of lymphoid cells is best assessed of this elaborate interplay of hematopoietic stem by immunophenotyping and the maturation of cells and mesenchymal stem cells and progeny is megakaryocytes by the extent of nuclear lobula- apparent by light microscopy, except for major de- tion. Immunophenotypic studies are required fects such as the loss of hematopoietic lineage(s) for megakaryoblast identifcation. or extensive fbrosis of the BM stroma. INDICATIONS FOR BONE HEMATOPOIESIS AND MARROW EXAMINATION CELL IDENTIFICATION Bone marrow examination (BME) of the pos- The highlights of hematopoiesis that are terior iliac crest is typically performed on almost listed in Table 1-1 simplify the complexity of all patients with a suspected hematolymphoid this cellular system. Most of the complex inter- neoplasm in which the BM is the likely primary actions and submicroscopic localizations critical site of disease, including all myeloid neoplasms, for hematopoiesis do not play a major role in precursor lymphoid neoplasms, and leukemic the clinician’s approach to blood and BM assess- chronic lymphoid neoplasms. Exceptions occur ment. From the diagnostician’s perspective, the when all necessary diagnostic and prognostic most immature cell populations of relevance tests can be performed on the involved blood, are recognizable morphologically and can be foregoing the need for upfront BME. enumerated on differential cell counts, even if BME is also commonly performed for stag- the actual lineage of these blast cells requires ing purposes in patients with extramedullary specialized techniques for confirmation. As neoplasms (17–19). The frequency of routine 2 Bone Marrow: General Considerations staging BME has declined in recent years due phologic/cytologic diagnostic information (see to the improvement in other noninvasive, ra- below) (26). diographic staging modalities (18–20). BME is useful in establishing a defnitive di- SPECIALIZED TESTING agnosis when the assessment of extramedullary Complementary specialized testing is often pathologic sites has not resulted in a clear neo- essential for delineation of lineage and stage of plastic diagnosis. It is also commonly performed maturation of abnormal cells, defnitive diag- in patients with unexplained cytopenias/cytoses nosis, risk stratifcation/prognostication, and or other unexplained laboratory or clinical future minimal residual disease assessment (Table fndings. Although BME results may reveal a 1-2) (26,28–30). The acquisition of adequate non-neoplastic disorder in such patients, BM specimens for specialized testing is essential: neoplasms are also commonly detected (21). anticoagulated syringes are required for fow cy- Once a diagnosis of a BM neoplasm is es- tometry, karyotyping, and many molecular tests; tablished, ongoing BME monitoring is used to air dried, unfxed aspirate slides are required for assess response to therapy, repopulation of the cytochemical staining and iron stains. Decisions BM with normal hematopoietic cells, persistent/ to perform various specialized tests should be residual disease, or secondary complications. undertaken on a case by case basis, factoring in morphologic features, specimen quality, and COMPLICATIONS OF BONE evidence-based diagnostic utility (29,30). MARROW EXAMINATION The posterior iliac crest is the universally BONE MARROW LINEAGES preferred site for BME, and complications from Granulocytic Lineage BM aspiration and core biopsy procurement from this site are rare, and typically consist of The key features of normal granulocytic, minor local hemorrhage or localized infections erythroid, and megakaryocyte lineage matura- of skin and adjacent soft tissue (22). Less com- tion and architectural localization are listed mon complications include neuropathy, more in Table 1-3 (28,31). Granulocytic cells pre- significant hemorrhage with compartment dominate in the BM of patients of all ages; the syndrome, osteomyelitis, and instrument is- myeloblast percentage varies somewhat by age sues such as breaking of an imbedded trephine but is typically less than 2 percent. The mor- biopsy needle (22,23). phologic stages of maturation from myeloblast Sternal BM aspirations have life-threatening to neutrophil, the predominant granulocytic complications, due to the potential for pen- cell, are well-defned, although this maturation etrating the sternal plate inducing pericardial sequence is complex at the molecular/genetic tamponade (24). Consequently, there are few, level, requiring coordinated transcription pro- if any, situations in which a sternal procedure is grams (fg. 1-2) (32,33). performed in lieu of the posterior iliac crest. The primary growth factor driving granulo- cyte production is granulocyte colony-stimu- SPECIMEN PROCUREMENT/ lating factor (G-CSF). Mature granulocytic BASIC PREPARATIONS cells predominate on aspirate smears; they are Details regarding the BME procedure are centrally located within the BM hematopoietic beyond the scope of this book and are well-re- cavity to facilitate rapid release into the circu- viewed in other sources (25–28). Careful con- lation (fg. 1-3). Less mature granulocytic cells sideration of the types of specimens required (promyelocytes and myelocytes) are normally for the diagnosis of a BM neoplasm is