Myeloid Cells in Health and Disease: A Synthesis

Between 1878 and 1880, Paul Ehrlich (1854-1915), a medical student and then assistant physician at the Charité Hospital, Berlin, demonstrated, using acid and basic aniline coal tar dyes, that the different types of blood leukocytes could be distinguished on the basis of the staining properties of their granules ( Fig. 1 ). Ehrlich’s technique for staining blood films and his method of differential blood cell counting ended years of speculation regarding the classification of white cells. His discoveries were among the greatest advances in modern hematology, and the principles surrounding his methodology are applied to this day. Nevertheless, prior to Ehrlich there were several notable landmarks that led to a fuller understanding of white cells, their origin, and possible function. This chapter highlights many of the important achievements of Paul Ehrlich and others. It is not intended to be comprehensive. The priority of many of the observations described, particularly in pre-Ehrlich time, remains controversial to this day.

The life and work of Elie Metchnikoff are a rich source of inspiration to anybody interested in the biology and pathophysiology of myeloid cells. He made the fundamental discoveries that subsequently shaped the development of the field and that represent, still today, the basis of our knowledge. First and foremost, he defined these cells by their function (i.e., “phagocytosis”), a definition that suits better than any other designation the nature of these cells, including perhaps the term “myeloid” itself. Metchnikoff described for the first time a number of crucial features of phagocytic cells, including (i) phagocyte-mediated host protection; (ii) active internalization of live, in addition to dead, organisms; (iii) uptake of senescent or damaged host cells; (iv) destruction of internalized particles; (v) bacterial killing by virtue of enzymes (“cytases”); (vi) vacuolar acidification; (vii) distinction between microphages (polymorphonuclear leukocytes) and macrophages; (viii) inflammatory recruitment of phagocytes; (ix) chemotaxis; and (x) diapedesis. For these reasons, Metchnikoff is unanimously considered the founding father of the field of phagocyte biology.

Most hematologists accept that hematopoiesis is initiated by multipotential hematopoietic stem cells. These cells are regarded as being few in number but diverse in properties and as being usually noncycling in normal health. It is well documented that, when myeloid populations are damaged by irradiation or chemotherapy, hematopoietic repopulation is initiated by such stem cells.

The drive to clear dying cells is evident even in the deepest branches of the Archaea and Bacteria phylogeny, where colonial biofilms appeared as a mode to promote survival in diverse environments ( 1 ). These ancient colonies (earliest fossil record ∼3.25 billion years ago) already displayed attributes of multicellular organism specialization, removing nonfunctional spent cells to recycle nutrients and maintain the integrity of the colony ( 1 , 2 ). Multicellularity followed shortly after and introduced a requirement for removal of nonself ( 3 , 4 ). Phagocytosis provided an elegant answer to both challenges, and has since served as a primary tool for cell turnover and removal of foreign invaders across all animal groups. It therefore provides a good stage to examine the evolution of myeloid cells through their contributions to homeostasis and host defenses ( Fig. 1 ). This chapter first focuses on ancestral phagocytes and examines their progression from primarily homeostatic cells to multifaceted effectors and regulators of immunity. The literature provides some insight into macrophage and lower metazoan hemocyte function as far back as echinoderms and urochordates. Further examination of gene marker conservation (e.g., apoptotic genes) in sponges and other colonial organisms allows us to dig deeper to examine the factors that led to the phylogenetic origins of cell clearance mechanisms and their continued evolution across newly developing animal branches. Subsequently, we focus on key challenges encountered by higher vertebrate myeloid cells as they manage increasingly complex mechanisms of immunity while maintaining a strict balance between proinflammatory and homeostatic cellular responses. In one example, we examine the impact of specialization through the diverging contributions of macrophages and neutrophils. We then consider the continued specialization of the myeloid lineage through the eyes of the dendritic cell (DC), which, through antigen presentation, effectively integrated new adaptive features into well-established and robust innate mechanisms of immunity. Indeed, documenting the multiple facets that comprise the life history of myeloid cells across evolution would not be possible in a single chapter. However, by focusing on their origins as phagocytes, we can appreciate the continued struggle of a host to develop novel and effective strategies to combat invading pathogens while ensuring the continued maintenance of tissue integrity and homeostasis.

Innate immunity shields all metazoans against infections. Its main features, including sensing, signaling, and effector mechanisms, are conserved from invertebrates to vertebrates. The hallmark of innate immunity is its reliance on a limited set of non-clonally-distributed receptors, which detect signature molecules of microbial origin and activate subsequent effector mechanisms. This concept, coined by Charles Janeway in 1989 as the self-versus-microbial-nonself discrimination system, has opened a large field of research for the so-called pattern recognition receptors (PRRs) and their cognate microbial elicitors, the pathogen-associated molecular patterns ( 1 ). Drosophila has rapidly emerged as a particularly suitable model organism for this research. Indeed, like all invertebrates, Drosophila exclusively relies on an innate immune system, which fends off infections in highly contaminated environments. Most importantly, Drosophila has benefited from more than a century of laboratory-use experience, yielding a wide array of molecular and genetic tools. Investigations on the defense reactions in flies rapidly provided valuable insights into the evolutionary conservation between insects and mammals, including humans, of the signal transduction pathways that control the innate immune system ( 2 ). Most prominent is the seminal finding in 1996 of the chief role of the Toll signaling pathway in the control of fungal infections in Drosophila ( 3 ). This study paved the way for the identification of the first mammalian PRR, Toll-like receptor 4 (the launching member of the TLR family), and the understanding of the innate immune system’s molecular mechanisms for sensing, signaling, and activation of adaptive immunity ( 4 – 6 ). Following more than 2 decades of in-depth analysis exploiting several infection models combined with genetic and genomic approaches, research on the Drosophila immune system revealed complex interconnected humoral and cellular processes, both of which show striking similarities with those of mammals. In this review, we provide a global view of the Drosophila host defense while drawing particular attention to the role of its monocyte-macrophage-like cells, the plasmatocytes. We provide general insights on the recent advances in Drosophila hematopoiesis and give a comprehensive summary on the so-far identified receptors involved in microbial detection, binding, and the ensuing internalization processes.

Monocytes, macrophages, and dendritic cells (DCs) are populations of myeloid mononuclear cells (MMCs) that provide critical sensing functions in innate immunity and a bridge to adaptive immunity through antigen presentation. They also perform important effector functions and contribute to chronic inflammation and healing. Collectively they have been described as the “mononuclear phagocyte system” (MPS) ( 1 ). As originally conceived, the MPS had a single blood-borne precursor, the monocyte. It is now appreciated that the development and homeostasis of MMCs is considerably more complex. In this chapter, we discuss the ontogeny of these diverse cells and reflect on ways in which ontogeny is linked to functional specialization, plasticity, and immune regulation.

Few, if any, individual cells survive throughout the life of the animal, an observation that sets up the critical concepts of cell life span, turnover, and removal and maintenance of homeostatic cell numbers. These issues are of special interest for understanding the properties of the myeloid cell lineage, which includes cells such as neutrophils, which may exhibit in the normal naive adult mammal the shortest life span of all but yet are maintained in relatively constant numbers within the circulation. However, our understanding of the underlying mechanisms for myeloid cell maintenance and removal is still substantially limited and also requires reexamination in light of new ideas about the ontogeny, characterization, and distribution of the myeloid cells in general. Accordingly, this essay will focus on the concepts and questions that, we argue, are in need of exploration, rather than providing a detailed review of what is a huge literature. By focusing on four of the myeloid-lineage cell types (neutrophils, monocytes, macrophages, and dendritic cells [DCs]), we will also be able to bring to the fore many of the key issues that characterize this set of questions.

The mononuclear phagocyte system (MPS) was originally defined by van Furth and Cohn ( 1 ) as a family of cells of the innate immune system derived from hematopoietic progenitor cells under the influence of specific growth factors ( 2 , 3 ). Differentiated cells of the MPS, monocytes and macrophages, are effectors of innate immunity, engulfing and killing pathogens. They are also needed for tissue repair and resolution of inflammation and for the generation of an appropriate acquired immune response. Their biology and differentiation have been reviewed by a number of authors ( 2 , 4 – 8 ). The original definition of the MPS considered an essentially linear sequence from pluripotent progenitors, through committed myeloid progenitors shared with granulocytes, to promonocytes and blood monocytes, and thence to tissue macrophages ( 2 , 4 – 8 ). Resident macrophages differ in function between tissues, and within tissues they occupy a specific niche ( 9 ). In some locations, for example, associated with epithelia, they clearly have individual identifiable territories that form a regular pattern ( 2 , 3 ).

Monocytes are a conserved population of leukocytes that are present in all vertebrates, with some evidence of a parallel cell population in fly hemolymph ( 1 ). Monocytes are defined by their location in the bloodstream, their phenotype and nuclear morphology, as well as by their characteristic gene and microRNA expression signatures ( 2 – 5 ). In mice, monocytes represent 4% of the nucleated cells in the blood, with considerable marginal pools in the spleen and lungs that can be mobilized on demand ( 6 , 7 ). Within the blood, monocytes, and in particular the classical Ly6C+ mouse subset, exhibit a characteristically short half-life of 20 h ( 8 ), akin to that of similar ephemer neutrophils ( 9 ).

Ralph Steinman was posthumously awarded a Nobel Prize in 2011 “for his discovery of the dendritic cell and its role in adaptive immunity.” He first coined the term “dendritic cell” in 1973 while working with Zanvil Cohn. His findings laid the foundations for a new field of immunology. This has enormous therapeutic potential for new vaccination strategies against cancer and infectious diseases. It also holds promise for new therapies for autoimmune disease, allergy, and transplantation reactions.

Major histocompatibility complex class I molecules (MHC-I) and class II molecules (MHC-II) are transmembrane glycoproteins that share the property of binding short peptides that are produced by the cells that express them. The generation of peptides and their subsequent association with MHC molecules is referred to as antigen processing. Antigen processing by myeloid cells, particularly dendritic cells (DCs), and the presentation of antigen-derived peptides to CD4+ and CD8+ T cells by MHC-I and MHC-II expressed on these cells are critical steps for effective adaptive immune responses. However, the mechanisms involved in antigen processing for MHC-I and MHC-II are different ( Fig. 1 ). For recognition by mature effector CD4+ T cells MHC-II-associated peptides are generated and bind within the endolysosomal system, while for recognition by mature CD8+ T cells MHC-I-associated peptides are generated in the cytosol from newly synthesized proteins and bind to MHC-I molecules in the endoplasmic reticulum (ER). For priming naive CD4+ T cells, the MHC-II processing pathway used by DCs also relies on peptide generation and binding in the endolysosomal system. However, priming CD8+ T cells requires endocytosis of antigens by the DCs followed by their transfer into the cytosol for proteolysis into peptides that ultimately bind to MHC-I molecules, a process known as cross-presentation or cross-priming. In this chapter we will discuss both general and myeloid-specific mechanisms of both MHC-I- and MHC-II-restricted antigen processing and presentation, phenomena that are intimately involved with the biosynthesis of the MHC glycoproteins.

Microglia are the resident macrophages of the brain parenchyma ( 1 ). Although it has long been known that microglia are of myeloid lineage, based on immunocytochemical detection of macrophage-restricted antigens ( 2 ), it has only relatively recently been shown, by fate mapping studies, that these cells are of yolk sac origin and enter the developing neuroepithelium of the central nervous system (CNS) in the embryo ( 3 ). They are present throughout the length of the neuraxis, characterized by their fine processes emanating from a small cell body, and each cell appears to occupy its own territory. The morphology of microglia and their territorial behavior is well illustrated in retina whole mounts ( Fig. 1 ). The density and morphology of the microglia vary between distinct functional divisions of the CNS, with the lowest density found in the cerebellum and perhaps the highest density in the substantia nigra ( 4 ). These regional differences have been well studied in rodents, the most common experimental animal models, and although similar regional differences are seen in the human brain, there are some notable differences. In the rodent brain, the microglia are denser in gray matter than in white matter, while in the human brain, the microglia are denser in the large-fiber tracts that dominate the larger brain ( 5 ).

Although bone is one of the hardest tissues in the body, necessary for its structural and protective roles, this organ is not static. Bone matrix must be renewed over time in order to maintain its mechanical properties, and myeloid lineage cells called osteoclasts (OCs) are the specialized cells that perform this critical function. Since bone is the major storage site for calcium, OCs play an important role in the regulation of this signaling ion by releasing it from bone. In this process, OCs respond indirectly to calcium-regulating hormones such as parathyroid hormone and 1,25(OH)2 vitamin D3. Growth factors such as insulin-like growth factor-1 (IGF-1) and transforming growth factor β (TGF-β) are also incorporated into bone matrix and released by OCs, affecting the coupling of bone formation to bone resorption and potentially targeting other cells in the microenvironment, such as metastatic tumors. Lastly, OCs retain features of other myeloid cells, such as antigen presentation and cytokine production, which afford them the potential to affect immune responses. Thus, the OC plays many roles in health and disease.

Eosinophils represent a minor component of circulating leukocytes and are generally considered to be terminally differentiated as postmitotic cells, yet it is now appreciated that they can be long-lived, multifaceted granulocytes involved in a variety of regulatory functions. Like other granulocytes, eosinophils develop and differentiate in the bone marrow. Under homeostasis, eosinophils are distributed in the blood, lung, thymus, uterus, adipose tissues, mammary gland, spleen and the lamina propria of the gastrointestinal (GI) tract ( 1 ), indicating a physiological function in each organ. Although eosinophils outside of the bone marrow are deemed as mature, recent evidence suggests the existence of multiple tissue-specific subtypes on the basis of distinct cell surface marker expression and functions ( 2 – 4 ). Driven by eosinophil-specific chemokines (primarily eotaxins) produced at baseline and markedly upregulated after a variety of stimuli ( 5 ), mature eosinophils are recruited from the circulation into their physiological locations and inflammatory sites, respectively. The cytokine interleukin-5 (IL-5), produced primarily by type 2 T helper (Th2) cells ( 6 ) and type 2 innate helper lymphoid cells (ILC2) ( 7 ), is crucial for eosinophil differentiation, priming, and survival ( 8 ). Conversely, eosinophils also serve as a source of a variety of cytokines and growth factors closely associated with multiple immunomodulatory functions to be discussed later. Through their vast cytokine arsenal and engagement of cell contact, eosinophils modulate immune responses through an array of interactive and orchestrated mechanisms, in trans and cis fashions, by cellular and humoral mediators, in both innate and adaptive immune responses. Recently, a burgeoning body of evidence has uncovered several underappreciated roles for eosinophils that could modulate both the adaptive and innate arms of immunity. An essential goal of this chapter is to summarize the role of eosinophils in physiological and inflammatory processes in human and small mammal models in order to identify novel pharmacological targets for specific disease management.

The first documented experiments using intravital microscopy were performed in the 19th century, in which very thin translucent tissues were used so that light could penetrate through the tissue and leukocyte trafficking could be observed ( 1 ). Neither human tissues nor solid organs in animal models could be used at the time. As such, tissues like the rodent mesentery, cremaster muscle, and ear and the bat wing were the preparations of choice for the next century. This type of imaging unveiled the very dynamic interaction of immune cells with vessel walls. The experimentalists tried to keep the conditions as close to the natural environment as was feasible. The bat wing and ear vasculatures required no surgery, making them likely the least perturbed approach. The mesentery and cremaster, which required only minor surgery, likely did induce a nonphysiologic baseline of leukocyte-vessel wall interactions. However, this came with the benefit of being able to examine cellular functions and behaviors under shear forces associated with blood flow as well as the surrounding architecture of capillaries and venules that was impossible to replicate in vitro. Indeed, as diligent as experimentalists were, in vitro settings could not completely replicate the behavior of immune cells as they interacted with each other, red blood cells and platelets in capillaries and postcapillary venules surrounded by pericytes and with macrophages, mast cells, and the myriad of other resident immune and parenchymal cells that constitute a living organ. Moreover, interorgan and neural communications were also not possible in vitro. However, it is always critical to remember that rodents, bats, and fish are not humans, and so all interpretations must be made with this in mind. It is also worth mentioning that many of the in vivo discoveries were made hand in hand with key in vitro experiments that allowed simplification of the complex model to elucidate cellular and molecular events.

The existence of extracellular pattern recognition was appreciated early on after formation of the pattern recognition receptor (PRR) theory ( 1 ) by the identification of Toll in Drosophila and cloning of the lipopolysaccharide sensor Toll-like receptor 4 (TLR4) in mammals (see reference 2 for a historical overview). However, it was evident that intracellular bacterial pathogens also induce inflammatory responses in infected cells, although the dedicated receptors for sensing of such threats remained elusive for some time. Analysis of invasive pathogenic bacteria and viruses showed that indeed such receptors exist. In recent decades, these cytosolic PRRs and their cognate microbe-associated molecular patterns (MAMPs) were identified.

Inflammation is the body’s response to injury, pathogen exposure, and irritants. Pattern recognition receptors allow our body to recognize a diverse array of patterns generated during exposure to these insults. In 2002, the nucleotide-binding domain leucine-rich repeat-containing (NLR, also known as NOD-like receptor) gene family of pattern recognition receptors was discovered ( 1 – 3 ). While several members were already recognized at that point, reports of the entire NLR family provided a global view. In the past 15 years of research, the physiological relevance of these genes has been revealed to include a diverse variety of functions. Gene mutations in some of the family members have been linked to autoinflammatory diseases in humans ( Fig. 1 ). This association of mutations in NLR genes to autoinflammatory diseases indicates critical functions in the regulation of immunity and inflammation.

Multicellular organisms have had to develop a rapid response to infection and tissue injury. In all animals on our planet, this involves mobilization of specialized cells to the focus of the infection or injury. This important insight was beautifully illustrated by Elie Metchnikoff, whose detailed drawings of cells being recruited to the site of injury caused by a rose thorn in a starfish embryo gave us the first glimpse of cells he termed “macrophages,” and neutrophils, which he termed “microphages.” If not the first person to observe phagocytosis and leukocyte diapedesis, Metchnikoff was probably the first person to fully appreciate the role of these two important cellular processes in “natural” or innate immunity ( 1 ).

Before we discuss lipids and their role in homeostasis and host defense, we will recount the essence of the inflammatory response. Inflammation is a reaction of the microcirculation; it’s a protective response initiated after infection or injury. While both local and systemic responses can be activated, inflammation is an essential biological process with the objective of eliminating the inciting stimulus, promoting tissue repair/wound healing, and, in the case of infection, establishing memory such that the host mounts a faster and more specific response upon a future encounter. The acute inflammatory response is a complex yet highly coordinated sequence of events involving a large number of molecular, cellular, and physiological changes. It begins with the production of soluble mediators (complement, chemokines, cytokines, eicosanoids—including prostaglandins [PGs], free radicals, vasoactive amines, etc.) by resident cells in the injured/infected tissue (i.e., tissue macrophages, dendritic cells, lymphocytes, endothelial cells, fibroblasts, and mast cells), concomitant with the upregulation of cell adhesion molecules on both leukocytes and endothelial cells that promote the exudation of proteins and influx of granulocytes from blood ( 1 ). Upon arrival, these leukocytes, typically polymorphonuclear leukocytes (PMNs) in the case of nonspecific inflammation or eosinophils in response to allergens, function primarily to phagocytose and eliminate foreign microorganisms via distinct intracellular (superoxide, myeloperoxidase, proteases, and lactoferrins) and/or extracellular (neutrophil extracellular traps) killing mechanisms ( 2 ). It is likely that the magnitude of the infectious load and its eventual neutralization signal the next phase of active anti-inflammation and proresolution ( 3 ).

Inflammation is the organism’s response to local injury in vascularized tissues programmed to traffic leukocytes and plasma delivery to an injured site or point of bacterial invasion ( 1 ); this protective response, when uncontrolled in humans, is associated with many widely occurring diseases. These include cardiovascular, metabolic, and the classic inflammatory diseases, e.g., arthritis and periodontal disease, along with cancers (reviewed in reference 2 ). Nonresolving inflammation is now widely acknowledged as a major driver in most of these diseases (for a review, see reference 3 ). The classical view of the resolution phase of the acute inflammatory response as understood and presented in pathology textbooks ( 1 , 4 ) as well as in medical dictionaries ( 5 ) was that local inflammatory chemical messengers and cells were passively diluted at the site (dilution of chemotactic gradient), hence halting further leukocyte recruitment and resolving the exudate or battlefield of inflammation ( 6 , 7 ). The historical perspective on the origins and concepts in the medical community regarding the resolution of inflammation apparently trace back as early as 11th-century Europe, and interested readers can refer to a recent review ( 8 ).

Wound repair is a complex and dynamic process that aims to restore cellular structures and tissue layers to damaged organs. Damage to the anatomical barriers against the environment, including the skin and gastrointestinal and respiratory systems, opens the organism to microbial invasion, and so the first function of the repair process is to temporarily seal this breach with a platelet plug and to counter infection as rapidly as possible. In skin wounds in healthy adults, barrier function is efficiently restored; however, repair of deeper dermal structures culminates in scar formation with loss of the original tissue structure and function. Tissue damage can be inflicted to a variety of organs by diverse stimuli, including ischemia (heart attack), burns (chemical, heat, or electrical), trauma, surgery, or infection. Although the anatomical sites injured may be distinct, for example, the immune-privileged cornea versus the gut with its complex microbiome, or the heart with its propensity for fibrosis, it is generally acknowledged that repair of all tissues shares basic commonalities. The wound-healing response can thus be typically divided into four overlapping phases: hemostasis, inflammation, cell migration/proliferation, and remodeling ( Fig. 1 ) ( 1 ).

A key determinant for the survival of an organism is the ability to recognize and respond to invading pathogens without damaging host tissues. This is accomplished largely by the concerted activity of the innate and adaptive branches of the immune system, which efficiently eliminate invading pathogens and restore tissue homeostasis. An initial step in the generation of robust immune responses is the recognition of pathogens by host cells, triggering subsequent immune cell activation and induction of proinflammatory responses. This initial recognition is facilitated via pathogen-associated molecular patterns (PAMPs) that represent highly conserved molecular structures uniquely found in bacterial, viral, and fungal pathogens but not in host tissues. Such structures include peptidoglycans, zymosan, lipopolysaccharides, flagellin, double-stranded and single-stranded RNA, and CpG-containing DNA ( 1 ). So far, an ever increasing number of receptors with the capacity to sense and respond to these PAMPs have been identified and are broadly categorized into distinct receptor families: Toll-like receptors (TLRs), RIG-I (retinoic acid-inducible gene I)-like receptors, NOD-like receptors, and C-type lectin receptors ( 2 – 4 ). Following engagement, these pattern recognition receptors trigger the activation of several inflammatory pathways essential to mediate robust antimicrobial activity and induce sustained immune responses. This central role in immunity for pathogen sensing by innate immune receptors is also reflected by the emergence of pattern recognition receptors early in evolutionary history, as evidenced by the presence of highly conserved gene orthologs in invertebrate species. Additionally, genetic analysis of human TLR and NOD genes provided evidence for strong positive selection pressure in human populations, and several nonsynonymous polymorphisms influencing receptor activity have been associated with disease susceptibility ( 1 ).

Complement is a system of blood plasma proteins that play critical roles in host defense through attracting leukocytes to sites of inflammation, mediating myeloid cell uptake and destruction of microbes, and guiding B- and T-cell activation ( 1 , 2 ). Regardless of the activation mechanism, the complement cascade converges on generation of third component of complement (C3) convertases that cleave C3 to C3a and C3b. The N-terminus of the C3α subunit is the anaphylatoxin (ANA) domain that becomes C3a after cleavage. C3b consists of two subunits containing eight macroglobulin-like domains (MG1 to -8). The β subunit consists of MG1 to MG5 plus the N-terminal half of MG6. The α subunit starts with the C-terminal half of MG6; a C1r/C1s, Uegf, and bone morphogenetic protein-1 (CUB) domain and a thioester domain (TED) inserted between MG7 and MG8; followed by the “anchor” and C345C domain (the trapezoid in Fig. 1 ).

Various cell types, including macrophages, dendritic cells (DCs), neutrophils, natural killer cells, and fibroblasts, express Toll-like receptors (TLRs) that activate the immune system ( 1 , 2 ). TLRs are type I transmembrane proteins with ectodomains containing leucine-rich repeats that mediate the recognition of pathogen-associated molecular patterns (PAMPs) derived from pathogens, and damage-associated molecular patterns (DAMPs) from dying or injured cells. TLRs harbor transmembrane domains and intracellular Toll–interleukin-1 (IL-1) receptor (TIR) domains in addition to leucine-rich repeat domains, and some adaptor molecules bind to them to activate the downstream signaling pathways. Various organisms express the TLR family, especially mammals, and 13 types of TLRs have been reported. TLR1 to -9 are conserved in both the mouse and human. However, in mice, a retroviral insertion has rendered the TLR10 molecule nonfunctional. TLR11, -12, and -13 do not occur in humans. The active TLRs localize differently. TLR1, -2, -4, -5, -6, and -10 are expressed on the cell surface, whereas TLR3, -7, -8, -9, -11, -12, and -13 are expressed in the endosome ( 3 , 4 ). Studies of mice deficient in each TLR have shown that each TLR has a distinct function in terms of PAMP recognition and the immune responses.

Lectins, defined as proteins that recognize carbohydrates, perform numerous essential biological functions. Recognizing a diverse array of carbohydrate structures, vertebrate lectins have been subdivided into several structurally distinct families which can be located intracellularly (such as the intracellular M-type family of lectins, which function primarily in the glycoprotein secretory pathway), in the plasma membrane (such as some members of the C-type lectin and Siglec [sialic acid-binding immunoglobulin-type lectin] families, which are involved in pathogen recognition and immune regulation), or are secreted into the extracellular milieu (such as some members of the galectin family, which serve several homeostatic and immune functions) ( Table 1 ). We will restrict our discussion here to selected myeloid- and plasma membrane-expressed members of only two families, the C-type lectins and Siglecs. We will provide a brief overview of each family and then focus on selected illustrative and detailed examples that highlight how these lectins influence myeloid cell functioning in health and disease. For an overview on the other lectin families, the reader is referred to an excellent website (http://www.imperial.ac.uk/research/animallectins/ctld/lectins.html).

G protein-coupled receptors (GPCRs) have been reviewed in depth elsewhere ( 1 ); however, the following section summarizes our present knowledge of GPCR classification and mechanisms involved in GPCR-mediated signaling.

Phagocytosis culminates with the entrapment of the target particles within large vacuoles called phagosomes. Because of the multiplicity of phagocytic receptors, it is becoming apparent that a variety of different signaling cascades can be activated during the process. However, several aspects of phagocytosis appear to be conserved, distinguishing it from other mechanisms of cellular uptake such as endocytosis and macropinocytosis. First, phagocytosis can accommodate a wide variety of particle sizes, from hundreds of nanometers to tens of micrometers ( 1 , 2 ), as well as complex particle morphologies ( 3 , 4 ). Second, phagocytosis requires the progressive engagement of phagocyte surface receptors around the entire particle ( 5 ). This ratchet mechanism has been described as the “zipper” model, which contrasts with the limited number of independent receptors that need to be activated by soluble ligands to trigger macropinocytosis ( 6 ). Third, phagocytosis is an active mechanism that involves local remodeling of the actin cytoskeleton, which drives the deformation of the plasma membrane and the progression of the receptor/ligand “ratchet” around the particle ( 7 – 10 ). In addition, as the actin cytoskeleton is tightly associated with the plasma membrane, signaling mediated by phospholipids appears to be a common feature of phagocytosis. Phosphoinositides in particular play a critical role, as phosphatidylinositol 3-kinase (PI3K) is seemingly involved in virtually all known phagocytic systems ( 11 – 14 ). These different features impose a temporal progression of the phagosome formation, which can be described by the following sequence of events: (i) binding of the ligand to surface receptors; (ii) activation of receptor-mediated signaling cascades; (iii) remodeling of the actin cytoskeleton; (iv) progressive engagement of additional receptors around the particle; and (v) membrane fusion, leading to the closure of the phagosome ( Fig. 1 ). Yet despite these conserved traits, one cannot fully appreciate the molecular mechanisms involved in phagosome formation without taking into account the diversity of phagocytic receptors and the variety of signaling cascades they induce individually and cooperatively. Thus, here, we chose to focus on some of the best-characterized receptors and signaling pathways in order to give an overview of the many roads that lead to phagosome formation, whereas phagosome maturation and subsequent responses will be described elsewhere.

The intracellular cytoskeleton, consisting of filamentous actin (F-actin), microtubules (MTs), and intermediate filaments, makes up a network of dynamic polymeric structures that regulate cell shape and function ( 1 ). Chemotaxis and phagocytosis, two essential myeloid cell functions that enable them to defend the host against harmful opportunistic microorganisms, rely heavily on a highly dynamic and multifunctional cytoskeletal network ( 2 – 4 ). The importance of the cytoskeleton in myeloid cells is emphasized by human innate immune dysfunction syndromes that arise from defects in cytoskeletal proteins or the proteins that regulate cytoskeletal function ( 5 – 7 ). Although most of our knowledge of cytoskeletal structure and function comes from fibroblasts and other cell types, myeloid cells are valuable model systems that have been exploited to study the diverse roles and functions of the cytoskeleton. In this chapter, we will review the current knowledge with respect to cytoskeletal regulation of myeloid cell function, with an emphasis on neutrophils and macrophages.

NF-κB is the prototypical proinflammatory transcription factor and plays an important role in production of cytokines, chemokines, and other proinflammatory mediators during the inflammatory response ( 1 ). NF-κB activation in macrophages is best characterized in response to proinflammatory cytokines, such as tumor necrosis factor α (TNF-α) and interleukin-1β (IL-1β), or the recognition of microbial products by Toll-like receptors (TLRs) or NOD-like receptors (NLRs). Using various upstream signaling pathways, these receptors converge on activation of the IKK (IκB kinase) complex, leading to activation of NF-κB. The IKK complex consists of two kinase subunits, IKKα (CHUK) and IKKβ (IKBKB), and the ubiquitin-binding protein IKKγ (IKBKG) ( 2 ). Despite its well-established role as a proinflammatory transcription factor in many cells, the NF-κB pathway in macrophages also has important anti-inflammatory roles ( 3 ). This was revealed by studies using the targeted deletion of IKKβ in macrophages, which was shown to have proinflammatory effects in several models of inflammation ( 4 – 7 ).

Macrophages are among the most phenotypically diverse cell types of mammalian organisms ( 1 , 2 ). They inhabit all or nearly all tissues under healthy conditions, where they play important roles as sentinels of infection and injury. These functions are enabled by the expression of a multitude of cell surface and internal receptors that recognize microbial-associated molecular patterns and/or damage-associated molecular patterns, exemplified by Toll-like receptors (TLRs) ( 3 ). Engagement of these receptors by microbial components, such as bacterial lipopolysaccharide (LPS), initiates signaling cascades that lead to the activation of latent transcription factors, including NF-κB, interferon regulatory factors (IRFs), and members of the activator protein 1 (AP1) family ( 4 , 5 ). These factors, in turn, function to activate hundreds of genes that play key roles in the orchestration of the innate immune response and that influence the development of adaptive immunity ( 6 , 7 ). In addition to this sentinel function, macrophages are professional phagocytes, serving to clear bacteria, apoptotic cells, and a diverse range of host-derived and environmental debris, thereby contributing to an additional layer of immunity and tissue homeostasis ( 2 ).

Epigenetics in the context of cell differentiation and activation refers to mechanisms that regulate and potentially stabilize gene expression in response to developmental and environmental cues. Epigenetic regulation is mediated by posttranslational modification of DNA or chromatin and by noncoding RNAs. In myeloid cells, the predominant focus of research on epigenetic mechanisms has been chromatin-mediated regulation of macrophages, which will be the focus of this review ( 1 – 3 ). Differentiation of macrophages from myeloid precursors is regulated by developmental signals and pioneer transcription factors that impart an “epigenetic landscape” that helps determine macrophage phenotype and how cells respond to environmental challenges. Macrophages protect the host from pathogenic microorganisms and other environmental insults, providing a rapid response as an initial line of defense. Accordingly, recognition of pathogen-associated molecular patterns through germ line-encoded receptors initiates and subsequently amplifies the adaptive immune response through cytokine production and antigen presentation. Importantly, macrophage phenotype is plastic, and macrophages carry out their distinct roles while maintaining the ability to adapt to local or systemic environmental changes ( 4 – 7 ).

Over the last 20 years, a great deal of progress has been made in understanding the mechanisms that control the biogenesis and secretion of the modified lysosomes found in immune cells. The picture that emerges is that of a series of very successful “variations on a theme,” with different combinations of related proteins interacting to provide different mechanisms for secretion in each specialized cell type. This allows cytotoxic T lymphocytes (CTLs) to provide a very focused secretion toward a single point, while mast cells and platelets give a generalized release all around the plasma membrane.

In 1933, Baldridge and Gerard observed the “extra respiration of phagocytosis” when dog leukocytes were mixed with Gram-positive bacteria and assumed that it was associated with the production of energy required for engulfment of the organisms. It was later shown that this “respiratory burst” was not inhibited by the mitochondrial poisons cyanide ( 1 ) or azide ( 2 ), which indicated that this is a nonmitochondrial process. The hunt for the neutrophil oxidase was then on because oxidative phosphorylation is far and away the major mechanism by which oxygen is consumed in mammalian cells, and another system that could consume oxygen at a similar rate was of considerable interest. Although many oxidases use NADH or NADPH as substrate, this oxidase was specifically called the NADPH oxidase because in the early days there was quite a controversy as to whether the substrate was NADH ( 3 ) or NADPH ( 4 ), and the matter was finally settled in favor of NADPH.

Microorganisms maintain an evolutionary advantage over their slowly replicating eukaryotic hosts. High mutation rates, rapid doubling times, and free genetic exchange between microbial species often place a considerable burden on the infected host to counter virulence escape mechanisms. This selective pressure has driven the acquisition of numerous eukaryotic defense strategies to protect host genome integrity and promote survival at the level of the individual cell ( 1 ). These cell-autonomous effector mechanisms, often considered unique to the immune cells of advanced metazoans, have in fact been largely inherited and repurposed from our eukaryotic ancestors ( Fig. 1 ). For example, phagocytosis developed as a trophic mechanism in unicellular amoebae long before its adaptation as a tool for immunity in the specialized “immune-like” cells of early invertebrates ( 2 , 3 ). Amebocytes, hemocytes, and coelomocytes present in lower organisms likewise predate professional phagocytes in animals with their ability to bind, engulf, and kill foreign microorganisms ( 4 ).

Tumor necrosis factor (TNF) is a member of the large family of cytokines; not hormones, but important local signaling molecules that transmit information from one cell to another ( 1 ). Different cytokines convey different messages, but cytokines are key players in every important biological process, including immunity, inflammation, cell growth, migration, fibrosis, vascularization, etc. So it is not surprising that abnormalities of these key mediators, molecules that are important enough to be conserved through evolution, may be involved in disease processes. What might not have been predicted was that removal of a single upregulated cytokine can make a clinical difference. This is best documented for anti-TNF ( 2 ) but is also true for blockade of interleukin-6 (IL-6), granulocyte-macrophage colony-stimulating factor (GM-CSF), and IL-1. In this review, we will summarize the current state of knowledge about cytokine expression and dysregulation in rheumatoid arthritis (RA) and other diseases and the role of TNF, the great majority of which is produced by macrophages. The knowledge gained has impacted our understanding of and therapy for other diseases also, and by focusing on cytokines, major rate-limiting steps, and hence therapeutic targets, there are opportunities for planning of therapy for many unmet needs.

Myeloid cells play significant roles in tissue remodeling, including the turnover of extracellular matrix (ECM), regulation of inflammation, and progression of cancer. Proteinases are important mediators of these processes and myeloid cells are major sources of proteinases, including the matrix metalloproteinases (MMPs). Indeed, the first cellular sources of MMPs, collagenase (MMP8) and gelatinase B (MMP9), were discovered in neutrophils ( 1 , 2 ). Papers published in the 1970s and 1980s demonstrated that macrophages also secreted collagenolytic (MMP8), gelatinolytic (MMP9), and elastinolytic (MMP12) metalloproteinases that degrade components of the ECM in the extracellular, pericellular, and lysosomal compartments ( 3 – 7 ). Taken together, these early discoveries suggested that metalloproteinases produced by myeloid cells have a major role in the remodeling of the microenvironment.

The immune system of mammalians is organized around two components: the innate immunity and the adaptive immunity. Older in terms of evolution, the innate immune system constitutes the first line of defense against microorganisms. This system is supplemented by the adaptive immunity, which is more recent in terms of evolution and provides the basis of immunological memory. Both the adaptive and innate immune systems are composed of a cellular and a humoral arm acting in a complementary and coordinated manner to regulate the innate response.

Macrophages are a central component of antimicrobial host defense, described as being crucial for both innate immune mechanisms and adaptive immunity ( 1 ). The dichotomy between the immediate antimicrobial responses seen as nonspecific and the relatively late-onset specific T- and B-cell responses has driven our understanding of host defense for more than half a century. Innate immunity reacts instantly upon an encounter with a pathogen but has been viewed as nonspecific and incapable of building immunological memory. In contrast, adaptive immune responses can specifically recognize pathogenic microorganisms and build memory capable of protection against reinfection. Macrophages are involved in both of these responses: on the one hand, macrophages have the capacity to phagocytose and kill microorganisms in a nonspecific fashion, as well as to release proinflammatory mediators that drive inflammation; but on the other hand, they can also present antigens and initiate and modulate the specific T-cell responses through expression of costimulatory molecules and specific cytokines ( 2 ).

The intestine constitutes a crucial interface for the acquisition of essential nutrients. In addition, the gut is colonized by a huge and diverse microbiota that confers several benefits to the host, including improved nutrient absorption and protection from pathogenic invasion. Thus, the intestine is the tissue of the body with the highest constitutive exposure to foreign antigen and is also a common entry portal for many local and systemic pathogens. Therefore, the local immune system, known as the gastrointestinal lymphoid tissue, has the unenviable task of balancing efficient responses to dangerous pathogens with tolerance toward beneficial microbiota and food antigens. As in most tissues, the decision between tolerance and immunity is critically governed by the activity of local myeloid cells. However, the unique challenges posed by the intestinal environment have necessitated the development of several specialized myeloid populations with distinct phenotypic and functional characteristics that have vital roles in maintaining barrier function and immune homeostasis in the intestine.

Intracellular bacterial pathogens cause a wide range of diseases and significantly contribute to the morbidity and mortality associated with infectious diseases worldwide ( 1 – 16 ) ( Table 1 ). These bacteria use several different strategies to replicate in host cells and influence host processes such as membrane trafficking, signaling pathways, metabolism, cell death, and survival ( 17 – 19 ). Broadly, intracellular bacteria colonize two topologically distinct regions of the host cell and are divided into cytosolic and intravacuolar bacteria according to their intracellular lifestyle. However, most intracellular bacterial pathogens have unique intracellular life cycles with features strikingly different from one another ( Fig. 1 ). It should also be noted that intravacuolar pathogens gain access to the host cytosol to some extent, and that cytosolic bacteria might spend an underestimated part of their intracellular life cycle within membrane-bound compartments ( 20 – 22 ).

While animal models of human disease have provided great insights that would have been difficult to achieve in studies in humans, our initial experiments on protection against Mycobacterium tuberculosis were designed to take advantage of transgenic knockout mice. While animal models have contributed enormously to our understanding, it must be noted that none faithfully reproduces the pathology or course of disease of human tuberculosis (TB) or leprosy. With that caveat, we explored the question of immunologically necessary conditions for protection of mice against M. tuberculosis infection. We found that mice lacking the gene for gamma interferon (IFN-γ) died from M. tuberculosis challenge in a matter of 2 to 3 weeks after intravenous challenge and within a month after aerosol challenge ( 1 ). We hypothesized that the pathology observed in the lungs would likely be mediated by local production of tumor necrosis factor alpha (TNF-α) and were quite surprised to learn that TNF-α-depleted mice succumbed with precisely the same time to death as the IFN-γ knockouts ( 2 ). Additionally, we ( 3 ) and others ( 4 , 5 ) found that mice whose major histocompatibility complex (MHC) class I presentation to cytotoxic T lymphocytes (CTLs) was deficient, e.g., β2-microglobulin deficient or TAP (transporter associated with antigen processing) deficient, succumbed to TB infection far earlier than control mice, but many weeks later than the IFN-γ- and TNF-α-deficient mice ( 1 ) ( Fig. 1 ). These results established that IFN-γ and TNF-α are necessary for initial protection in mice, likely mediated by innate immunity and cytokine-activated macrophages, and suggested that CTLs may play a role later in infection.

Asthma is clinically defined by variable airway obstruction that causes recurrent periods of shortness of breath, chest tightness, wheezing, and coughing. Patients also often have altered mucus production and have problems in expectorating sputa because of reduced viscosity of the mucus. One of the characteristic changes to lung physiology is the occurrence of bronchial hyperreactivity, which is defined as a tendency of the smooth muscle layer to contract to nonspecific stimuli like cold air or exercise, and measured in the lung function lab as increased bronchoconstriction to very low amounts of histamine or methacholine. We now realize that asthma is not one single disorder, but rather a syndrome or a spectrum of disease, characterized by endotypes that rely on distinct pathomechanisms and controlled by various adaptive or innate immune cells ( 1 – 5 ). In early life, asthma is often allergic, driven by CD4 Th2 lymphocytes and associated with allergic comorbidity like atopic dermatitis and rhinitis. On histology, target organs often contain many eosinophils. In adult-onset asthma, almost half of the cases are not associated with allergy. Some of these patients have eosinophilic airway inflammation, whereas others have a neutrophil-predominant inflammation, a mixed neutrophil-eosinophil infiltration, or even pauci-immune disease. Important comorbidities are obesity, acid-reflux disease, and chronic rhinosinusitis ( 2 , 5 ). Across all age groups, the presence of a more neutrophilic infiltrate is associated with more (therapy-resistant) severe disease, and it is possible that this disease variant relies more on interleukin-17 (IL-17)-producing Th17 cells ( 1 , 6 ).

The immune system is fundamentally divided into the innate and adaptive arms, predominantly represented by the myeloid and lymphoid lineages, respectively, and largely derived from bone marrow progenitors. This simplistic classification belies an intricate circuitry in which the innate and adaptive cells communicate, stimulate, and regulate each other throughout the course of every immune response. Hence, in every respect myeloid cell populations are instrumental to successful defense against parasitic infections.

A major goal of immunosuppressive therapy in management of chronic inflammatory diseases and allogeneic transplants has been to harness long-term tolerance processes from short-term treatments. This should limit morbidity from long-term undermining of immune mechanisms, which is the hallmark of current immunosuppression.

The cytoplasm of an activated macrophage is a dangerous place due to the accumulation of reactive oxygen species (ROS), which can perturb cytoplasmic oxidative balance. Macrophage activation through monocyte recruitment in tissues occurs in a highly regulated manner upon detection of microbial pathogen-associated molecular patterns (PAMPs), such as the Gram-negative bacterial cell wall component lipopolysaccharide (LPS), stimulating ROS production. ROS can be generated from many cellular processes, either directly or as a result of incomplete reduction of free radicals ( 1 ). One of the main sources of ROS is the NADPH oxidase (NOX), a specialized transmembrane protein complex that generates superoxide (O2 –), which has been implicated in several inflammatory diseases ( 2 ). The generation of ROS can also stem from the mitochondrial electron transport chain, due to insufficient reduction of superoxide anions. Another source is the inducible form of nitric oxide synthase 2 (iNOS). iNOS produces nitric oxide from l-arginine and is known to play key roles in macrophage function ( 3 , 4 ).

Macrophages perform a vast range of biological functions, including regulation of embryonic development; scavenging and recycling of redundant, aged, or injured cells; modulation of tissue repair; and coordination and effector activity in host defense. By recycling and storing iron from senescent erythrocytes and other damaged cells, the macrophage controls iron homeostasis, supplying most of the iron needed for hemoglobin synthesis in erythrocyte precursors, and for the much smaller but important iron requirements of other cell populations. The macrophages serve a crucial regulatory role by functioning as a regulated storage compartment for iron. In response to systemic iron requirements, the release of iron from macrophages into plasma is negatively regulated by the interaction of the hepatic hormone hepcidin with its receptor/iron exporter ferroportin. In humans, macrophages contribute most of the iron entering the plasma compartment, with the rest of the iron influx into plasma made up from duodenal iron absorption and release of stored iron from hepatocytes. During infection and inflammation, interleukin-6, and to a lesser extent other cytokines, increases hepcidin synthesis. Hepcidin binds to macrophage ferroportin, induces its endocytosis and proteolysis, and thereby causes iron sequestration in macrophages. The resulting decrease in iron availability in other tissues can limit the growth and pathogenicity of invading extracellular microbes and is as an important means of host defense. Finally, bone marrow macrophages also have an important role in supporting efficient and rapid production of erythrocytes. The involvement of macrophages in iron metabolism thus serves both trophic and host defense functions. This review addresses the role of macrophages in iron metabolism in all these contexts, and represents an update and expansion of my previous discussion of the same subject ( 1 ).

Concepts of the pathogenesis of atherosclerosis have evolved substantially through the decades. Viewing it as an inevitable degenerative process, Sir William Osler attributed atherosclerosis to the stress and strain of modern life at the dawn of the 20th century ( 1 ). Indeed, the pathogenesis of atherosclerosis had given rise to great controversy in the middle portion of the 19th century, particularly among German pathologists. Von Rokitansky postulated a role of incorporated thrombus into the artery wall as the primary event in atherosclerosis ( 2 ). Rudolf Virchow posited a role for proliferation of medial cells, now recognized as arterial smooth muscle cells (SMCs), in the pathogenesis of atherosclerosis ( 3 ). Virchow also recognized cell death as a component of atherogenesis and observed bone formation in atherosclerotic plaques. While von Rokitansky’s notion of the incorporation of thrombus lost popularity, the concept of atherosclerosis as a proliferative disorder of SMCs received considerable attention by pathologists and cell biologists in the latter part of the 20th century. Earl Benditt provided evidence for monotypic accumulation of SMCs in atherosclerotic plaques ( 4 ). Russell Ross focused on the role of platelet products, notably platelet-derived growth factor, as a causal stimulus for SMC growth in atherosclerotic plaques ( 5 , 6 ).

That macrophages normally inhabit endocrine organs became evident from the early studies in mice made in the laboratories of Siamon Gordon and David Hume ( 1 ). They used the macrophage surface marker F4/80 to examine tissue sections by immunohistochemistry and found macrophages in all endocrine organs. In most organs, the macrophages were found associated with the vessels. To note are several important examples. In the adrenal gland, the zona glomerulosa contained abundant macrophages that “wrapped around capillaries or line vascular sinuses, but membrane processes extend into the surrounding tissue.” In the pituitary, they found them distributed differentially in the various areas. In the thyroid, many surrounded the follicles. In the testes, the macrophages were next to the Leydig cells and not inside the seminiferous tubules. In the ovaries, macrophages were abundant around follicles and usually surrounding the vessels.

Tumors are composed of heterogeneous, transformed cell populations with different morphologies and phenotypes, which are organized in a pyramidal architecture determined by self-renewal ability, differentiation grade, and tumorigenic and clonogenic potential ( 1 ). During tumor progression, cancer cells secrete tumor-derived factors (TDFs), like cytokines, chemokines, and metabolites, which promote the development of a flexible microenvironment inducing both the generation of new vessels and the modification of the immune responses ( 2 ). Tumors can escape the immune system by three main mechanisms: (i) cancer cells can veil their identity to escape recognition by immune effectors, (ii) they can directly modify antitumor immunity, or (iii) they can recruit other immune regulatory cells whose normal function is to inhibit immune reactions and prevent the unfavorable effects of uncontrolled immune stimulation ( 3 ). Probably the most pervasive and efficient strategy of “tumor escape” relies on the tumor’s ability to create a tolerant microenvironment by modification of normal hematopoiesis. In fact, cancers can induce the proliferation and differentiation of myeloid precursors into myeloid cells with immunosuppressive functions, in both the bone marrow and other hematopoietic organs such as the spleen, at the expense of additional myeloid cell subsets, such as dendritic cells (DCs) ( 4 ). Additionally, the persisting imbalance in the number and type of myeloid cells can deeply influence myeloid cell recruitment and function at the tumor site and secondary lymphoid organs. In the bone marrow, hematopoietic progenitor cells give rise to immature DCs (iDCs). To reach complete maturation, iDCs require inflammation-related stimuli because, although able to take up, process, and present antigens, they express few or none of the costimulatory molecules, such as CD80, CD86, and CD40, necessary to exert their functions ( 5 ). The higher number of iDCs found at the tumor site stems from defects in myelopoiesis rather than simply from the lack of appropriate activation signals at the tumor site. In vitro treatment of tumor-infiltrating DCs with appropriate stimuli—such as granulocyte-macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor α (TNF-α), or CD40L—was not sufficient to induce DC maturation; this evidence supports the concept that the reduced functionality of DCs is most likely due to defects in differentiation from their iDC progenitors ( 6 ). However, iDCs are not the only myeloid cell populations modified in cancer. In postnatal life, hematopoietic stem cells present in hemopoietic compartments give rise to lymphoid and myeloid multipotent precursor cells. Other pluripotent cell types originate from the myeloid precursors: the common DCs and the immature myeloid cell precursors (IMCs). The first originates iDCs and plasmacytoid DCs, and the second is the common progenitor for macrophages, granulocytes, and monocyte-derived DCs ( 7 ). In healthy mice, IMCs rapidly differentiate into their descendant lineages; consequently, they represent a relatively low percentage of circulating myeloid cells. However, under pathological conditions, including cancer, there is a partial block in IMC differentiation, leading to the accumulation of CD11b+/Gr-1+ myeloid cells with immunosuppressive function, named myeloid-derived suppressor cells (MDSCs) ( 8 ).

The hematopoietic stem cell (HSC) is a multipotent stem cell that resides in the bone marrow and has the ability to form all of the cells of the blood and immune system. As the quintessential stem cell, it has the ability to self-replicate and differentiate into progeny of multiple lineages. Hematopoiesis describes the process of differentiating from HSCs to mature, functional cell types of the blood lineages. The existence of HSCs was first hypothesized following early experiments that demonstrated that animals receiving lethal doses of irradiation could be rescued by transplanting unfractionated bone marrow cells ( 1 ). The transplanted cells repopulated the bone marrow of the recipients and gave rise to all the cells of the blood. In accordance with this observation, in 1961 Till and McCulloch showed that unfractionated bone marrow cells were able to generate mixed hematopoietic (myeloid and erythroid) colonies in the spleens of lethally irradiated mice ( 2 ). They subsequently demonstrated that these colonies were formed by single cells that were capable of multilineage differentiation ( 3 ). Given the limitations in technology at the time, they were unable to purify these cells further, and the experiment that showed clonal origin of spleen colonies did not include lymphoid cells ( 2 , 3 ), although a later experiment did ( 4 ). Years later, with the advent of monoclonal antibodies and fluorescence-activated cell sorting, these cells could be further characterized, purified, and evaluated in functional assays. Studies have now conclusively demonstrated that HSCs are a rare population of cells that give rise to all of the cells comprising the two main branches of the hematopoietic lineage: the myeloid arm and the lymphoid arm. In mice, all long-term HSCs (LT-HSCs) are Hoxb5+ ( 5 ) and located in the central marrow attached to the abluminal side of venous sinusoids.