Lymphopoiesis

T cells are formed in bone marrow then migrate to the cortex of the thymus to undergo maturation in an antigen-free environment for about one week where a mere 2–4% of the T cells succeed. The remaining 96–98% of T cells die by apoptosis and are phagocytosed by macrophages in the thymus. So many thymocytes (T cells) die during the maturation process because there is intensive screening to make sure each thymocyte has the ability to recognize self-peptide:self-MHC complex[16] and for self tolerance. The apoptosed thymocyte dies and is quickly recycled.

Upon maturity, there are several forms of thymocytes including [17]

• T-helper (needed for activation of other cells such as B cells and macrophages),
• T-cytotoxic (which kill virally infected cells),
• T-memory (T cells that remember antigens previously encountered), and
• T-suppressor cells (which moderate the immune response of other leukocytes). Also called T-regulatory cells (T_reg)

When T cells become activated they undergo a further series of developments. A small, resting T lymphocyte rapidly undergoes blastogenic transformation into a large lymphocyte (13–15 μm). This large lymphocyte (known in this context as a lymphoblast) then divides several times to produce an expanded population of medium (9–12 μm) and small lymphocytes (5–8 μm) with the same antigenic specificity.[18] Final activated and differentiated T lymphocytes are once again morphologically indistinguishable from a small, resting lymphocyte. Thus the following developmental states may be noticed in sequence in blood tests:

1. Prolymphocyte
2. Large lymphocyte
3. Small lymphocyte

Basic Map of T Cell lymphopoiesis

This basic map of T Cell formation in sequence, is simplified and is akin to textbook descriptions, and may not reflect latest research.(Medical Immunology, p. 119)

In the thymus

• MLP
• ETP
• DN1
• (B; Mφ)
• DN2
  • (DC; NK)
• DN3
  • (γδ)
• DN4
• DP
  • (TNK; CD4; CD8; T_reg)

In the Periphery

• (T_h1; T_h2)

Unlike other lymphoid lineages, T cell development occurs almost exclusively in the thymus. T-lymphopoiesis does not occur automatically but requires signals generated from the thymic stromal cells. Several stages at which specific regulators and growth factors are required for T cell development to proceed have been defined. Later in T cell development and its maturation these same regulatory factors again are used to influence T cell specialization.

T cells are unique among the lymphocyte populations in their ability to further specialize as mature cells and become yet more mature. And T cells come in many flavors, for example: the conventional TcRαβ T cells; the so-called unconventional TcRγδ T cells; NKT cells; and T regulatory cells (T_reg). Details regarding the developmental and life cycle of the unconventional T cells are less well-described compared to the conventional T cells.

Stages of T cell maturation

Stage One: Thymic Migration

Multipotent lymphoid progenitors (MLP) enter the T cell pathway as they immigrate to the thymus. The most primitive cells in the thymus are the early thymocyte progenitors (ETP), which retain all lymphoid and myeloid potential but exist only transiently, rapidly differentiating into T and NK lineages. (Medical Immunology, p. 118)

Stage Two: Proliferative Expansion and T Lineage Commitment

Final commitment to the T cell lineage occurs within the thymic microenvironment, the microscopic structures of the thymus where T cells are nurtured. The most primitive T cells retain pluripotential ability and can differentiate into cells of the myeloid or lymphoid lineages (B cells, DC, T cells, or NK cells).

More differentiated double negative T cells (DN2 cells) have more limited potentiality but are not yet fully restricted to the T cell lineage (they can still develop into DC, T cells, or NK cells). Later on, they are fully committed to the T cell lineage- when thymocytes expressing Notch1 receptors engage thymic stromal cells expressing Notch1 ligands, the thymocytes become finally committed to the T-cell lineage. See Gallery Image "Double Negatives"

With the commitment to the T cell lineage, begins a very complex process known as TCR gene rearrangement. This creates an enormous diversity of T cells bearing antigen receptors. Afterward some T cells leave the thymus to migrate to the skin and mucosae.

Stage Three: β-Selection

Stage Four: T Cell Receptors Selection

Only 2% to 3% of the differentiating thymocytes, those that express TcR capable of interaction with MHC molecules, but tolerant to self-peptides, survive the Stage Four selection process.

Stage Five: Continuing Differentiation in the Periphery

It was previously believed that the human thymus remained active as the site of T cell differentiation only until early adulthood and that later in adult life the thymus atrophies, perhaps even vanishing. Recent reports indicate that the human thymus is active throughout adult life. Thus several factors may contribute to the supply of T cells in adult life: generation in the thymus, extra-thymic differentiation, and the fact that memory T cells are long-lived and survive for decades.

• Unconventional T cells

The thymus also gives rise to the so-called ‘unconventional T cells’ such at γδ T cells, natural killer T cells (NKT) and regulatory T cells (T_reg).

• γδ T cells

γδT cells represent only 1% to 5% of the circulating T cells, but are abundant in the mucosal immune system and the skin, where they represent the dominant T cell population. These ‘non-MHC restricted T cells’ are involved in specific primary immune responses, tumor surveillance, immune regulation and wound healing.[19]

Several differences between αβ and γδ T cell development have been described. They emigrate from the thymus in "waves" of clonal populations, which home to discrete tissues. For example, one kind is found in the peripheral blood while another predominates in the intestinal tract.

• Natural Killer T cells

Human NKT cells are a unique population and are thought to play an important role in tumor immunity[20] and immunoregulation.

• T Regulatory cells

T_reg cells are considered as naturally occurring regulatory T cells. Tregs comprised about 5% of the circulating CD4⁺ T cells. These cells are thought to possess an important autoimmunity property by regulating 'autoreactive' T cells in the periphery. (Medical Immunology, p. 117-122)

B cells are formed and mature in bone marrow (and spleen).

It is a good mnemonic aide that B cells are formed in the bone marrow, but it is a mere coincidence since B cells were first studied in the chicken's bursa of Fabricius and it is from this bursa that B cells get their name.

These B cells then leave the bone marrow and migrate via bloodstream and the lymph to peripheral lymphoid tissues, such as a spleen, lymph nodes, tonsils and mucosal tissues. Once in a secondary lymphoid organ the B cell can be introduced to an antigen that it is able to recognize.

Through this antigen recognition and other cell interactions the B cell becomes activated and then divides and differentiates to become a plasma cell. The plasma cell, a B cell end product, is a very active antibody-secreting cell that helps protect the body by attacking and binding to antigen.

Even after many decades of research, some controversy remains as to where B cells mature and 'complete their education', with the possibility remaining that the site may also partially be peri-intestinal lymphoid tissues.[21]

B lymphopoiesis occurs exclusively in the bone marrow and B lymphocytes are made continuously throughout life there in a 'microenvironment' composed of stromal cells, extracellular matrix, cytokines and growth factors, which are critical for proliferation, differentiation, and survival of early lymphocyte and B-lineage precursors.

The relative proportion of precursor B cells in the bone marrow remains rather constant throughout the life span of an organism. There are stages such as Pre-B-I cells (5% to 10% of the total); Pre-B-II cells (60% to 70%) while the remaining 20% to 25% are immature B cells. Most textbooks say that B Cells mature in the bone marrow but, generally, immature B cells migrate to the spleen for 'higher education' of some sort where they go through transitional stages before final maturation.(Medical Immunology, p. 136)

B lymphocytes are identified by the presence of soluble immunoglobulin G (IgG). This is the most common protective immunoglobulin in the adult body. After antigenic stimulation, B cells differentiate into plasma cells that secrete large quantities of soluble IgG. This is the final stage of B lymphopoiesis but it is the clincher because the plasma cells must either issue antibody close to a source of infection, or disseminate it in the blood to fight an infection at a distance or in an inaccessible part of the body.

Basic map of B cell lymphopoiesis

Early B cell development: from stem cell to immature B cell

Transitional B cell development: from immature B cell to MZ B cell or mature FO B cell.

A generally regarded valid map of B cell lymphopoiesis is as follows in sequence, in two parts with the first being in the bone marrow and the second in the spleen:.[22] The development process in the bone marrow occurs in germinal centers

In the bone marrow

• Pro-B
• Pre-B-I
• Pre-B-II large
• Pre-B-II small
• Imm(ature)

In the spleen

• T1
• T2/T3
• (Marginal Zone (MZ); B-1; B-2)
• B-2 further differentiate into:
  • (Germinal Center (GC); Memory; Plasma )

NK cells, which lack antigen specific receptors, develop in the bone marrow. After maturation and release from the marrow they circulate in the blood through their lifetime seeking opportunity. The opportunity they seek is to encounter and recognize and then kill abnormal cells such as cancer or virally infected cells. It is well known that lymphocytes never have granules or at least not granules that are readily visible even upon staining. However NK cells are the exception. They do have numerous granules which provide their ability to kill cells and these granules are why NK cells have an alternate name- Large Granular Lymphocytes (LGLs).

NK cells are the only lymphocytes considered part of the innate immune system (in contrast to the adaptive immune system. Yet they are much more closely related to T cells (part of the adaptive immune system) than to other cells of the innate immune system. NK cells not only share many surface markers, functions and activities in common with T Cells, they also arise from a common T/NK progenitor. The T/NK precursor is also believed to be the source of a subpopulation of lymphoid DC. (Medical Immunology, p. 121)

NK cells have a definition 'barcode' as CD3, CD16+, CD56t lymphocytes. (See Barcode Section of this article). NK progenitors can be found mainly in the thymus (mouse), but the thymus is not absolutely required for NK development. Probably NK cells can develop in a variety of organs but the major site of NK cell development is not known.

In humans, the majority (85–90%) of the NK cells have a high cytolytic capacity (the ability to lyse cells). A smaller subset (10–15%) called NK 'CD56 bright' is chiefly responsible for cytokine production and has enhanced survival. Traveling to lymph nodes the 'CD56 bright' NK cells differentiate again into mature NK cells which express killer cell immunoglobulin-like receptors (KIR), natural cytotoxicity receptors (NCR), and critical adhesion molecules. (Medical Immunology, p. 122)

The process by which CLP cells may differentiate to generate dendritic cells of lymphoid lineage is not yet well defined.[23]

DCs are highly specialized and efficient antigen-presenting cells. Cells identical in appearance come both from a myeloid lineage (referred to as myeloid dendritic cells) and also from a lymphoid lineage (referred to as plasmacytoid dendritic cells).

The development and regulation of DC is not well-characterized. While the DC precursors have been identified in the human fetal liver, thymus, and bone marrow, during adult life DC are thought to be produced only from the bone marrow and released into the blood to wander and settle down. Overall a large number of DC of varying types are dispatched throughout the body, especially at epithelia such as skin, to monitor invaders and nibble their antigens. (Medical Immunology, p. 122)

Lymphocytes have a number of alarming properties such as the ability to wander around the body and take up lodging almost anywhere, and while on the way issue commands in the form of cytokines and chemokines and lymphokines, commands that affect many cell types in the body and which may also recursively induce further lymphopoiesis. One strong behavior pattern that captivates researchers and the public alike is the ability of lymphocytes to act as police, judge and executioner to kill other cells or demand that they suicide, a command that is usually obeyed. There seems to be no other sentencing option available.

Killers are distinguished from cells such as macrophages that eat other cells or munch debris by a method called phagocytosis. Killers do not use phagocytosis, they just kill and leave the clean-up to other cells.

Killers are known to attack virus-infected cells and cells that have become cancerous. Because of these abilities much research has been done into transforming these qualities into medical therapy but progress has been slow.

Here is the parade of killers and how they work:

• Cytotoxic T cells

(also called Tc or antigen-specific cyto -lytic or -toxic T lymphocytes (CTL)).[24] Tc kill by apoptosis and either splash their target with perforin or granzymes or else use Fas-Fasl Interaction to command target elimination. This kills cells that are infected and display antigen.

• NK cells (also called LGL (large granular lymphocytes))

These kill with exactly the same methods as Tc, but have no interaction with any antigen. They select their targets based on typical molecules displayed by cells that are under stress by viral infection. NK Cells mainly are in the circulation (5-15% of the circulating lymphocytes) yet are also distributed in tissues everywhere.[25]

• LAK cells (Lymphokine-activated killer) are a laboratory/clinical subset of NK Cells promoted by IL-2 to attack tumor cells.[26]
• NKT cells see Natural Killer T cell main article

Natural Killer T Cells. Human NK T cells are a unique population (which express NK cell markers such as CD56 and KIR). NKT cells are thought to play an important role in tumor immunity and immunoregulation. (Medical Immunology, p. 135), yet little is known. Recent evidence suggests a role working together with hepatic stellate cells being a liver-resident antigen-presenting cell that presents lipid antigens to and stimulates proliferation of NKT cells.

• Natural killer-like T cells

A heterogeneous group with ill-defined properties.

However, in summary there is no known cell or set of cells that is capable of killing cancerous cells in general.

Because all WBCs are microscopic, colorless and often seemingly identical in appearance they are individually identified by their natural chemical markers, many of which have been analyzed and named. When two cells have the same markers, the reasonable assumption is made that the cells are identical at that time. A set of markers is colloquially described as the barcode for that cell or that cell line.

• Here is an example of how a barcode can come to be, for the all-important hematopoietic stem cell (HSC) as an example.

HSCs are technically described as: lacking FMS-like tyrosine kinase 3 (Flt3) and lacking the markers specific to discrete lymphoid lineages (Lin), but expressing high levels of Sca1 and c-kit; HSC also express CD44, low levels of Thy1.1 (CD90), but no IL-7Ra or CD27.

This is called the (surface) phenotype of an HSC. It can be expressed as a set (Lin2, Sca1high, c-kit high, CD44+, Thy1.1low, CD27 2, and IL-7Ra2). This set is a ‘barcode’ for the HSC, akin to the barcode label attached to your chicken-wing plastic bag for checkout at a supermarket! Scientists use these barcodes to check, categorize and accumulate cells for many purposes often using laboratory methods such as cell flow cytometry. These barcodes partially define the modern meaning of phenotype for leukocytes.

Progression of HSC differentiation and lineage commitment is indicated by changes in this phenotype. That is, as the cell changes, the markers will also change and the barcode will change.

• Typical barcodes for some cell types appearing in this article.

• Note explaining the barcode parameter details: Flt3 is a cytokine tyrosine kinase receptor thought to be important in early lymphoid development. In addition, Flt3 plays a major role in maintaining B lymphoid progenitors. CD27 plays a role in lymphoid proliferation, differentiation, and apoptosis. The acquisition of CD27 and Flt3 by the HSC coincides with the loss of long-term repopulating potential. At this stage the cells retain both lymphoid and myeloid potential and are referred to as multipotent progenitors. (Medical Immunology, p. 114)

New questions emerge in immunology continuously as though there were a stem cell for questions. For example, it was thought that the process of lymphopoiesis was a direct, orderly unidirectional sequence. But it is not clear if end-stage lymphocytes come from progenitors that are homogeneous populations or overlapping populations. Nor is it clear whether lineages of lymphocytes develop via a continuum of differentiation with a progressive loss of lineage options or whether abrupt events result in the acquisition of certain properties.[27]

Changes in cytoplasm, morphology of the cell nucleus, granules, cell internal biochemistry, signaling molecules and cell surface markers are difficult to correlate with definite stages in lymphopoiesis. The morphological differences do not just correspond to steps in mitosis (somatic cell division), but result from continuous "maturation processes"of the cell nucleus as well as of the cytoplasm and so one must not be too rigid about morphological distinctions between certain cell stages.[28]

• Models and updates on the lymphopoiesis family tree

Until recently the model of the CMP generating all myeloid cell and the CLP generating all lymphoid cells was considered necessary and sufficient to explain the known facts observed in the generation of WBCs, and it is still found in most basic textbooks. However beginning around 2000 and gaining momentum after 2005[29] in both studies in humans and mice, new complexities were noted and published in papers. These studies are important now mainly to immunology researchers but are likely to eventually lead to changes in medical treatments.

The changes were sparked by observations that lymphopoiesis did not always break into two lineages at the level of the CLP. Worse, some macrophages (long considered a myeloid lineage) could be generated by lymphoid lineage progenitors. In essence focus has been shifted away from the CLP to the MLP (lymphoid-specified progenitors), which are clearly lymphoid progenitors yet retain some myeloid potential, particularly the ability in both humans and mice to make macrophages – one of the most versatile of immune cell defenders – and also many dendritic cells, the best 'watchdogs' of antigen invaders.

However, whatever the details may turn out to be, the process of lymphopoiesis always seems to relentlessly give rise to progeny with special attributes and abilities – "superpowers" so to speak – but with progressively more restricted lymphoid developmental potential.

This model of lymphopoiesis had the virtue of relative simplicity, agreement with nomenclature and terminology, and is still essentially valid for the laboratory mouse.

1. pHSC pluripotent, self-renewing, hematopoietic stem cells [30] which give rise to
2. MPP multipotent progenitors, which give rise to
3. ELP (or PRO) Prolymphocytes, early lymphoid progenitors, and finally to the
4. CLP Common lymphoid progenitor, a cell type fully committed to the lymphoid lineage.

pHSC, MPP and ELP cells are not fully committed to the lymphoid lineage because if one is removed to a different location it may differentiate into non-lymphoid progeny. However CLP are committed to the lymphoid lineage. The CLP is the transit cell responsible for these (generally parallel) stages of development, below:

• NK cells
• Dendritic cells (lymphoid lineage; DC2 [31])
• Progenitor B cells
  • Pro-B cells => Early Pro (or pre-pre)-B cells => Late Pro (or pre-pre)-B cells
  • Large Pre-B cells => Small Pre-B cells
  • Immature B cells
  • B Cells => (B1 cells; B2 cells)
    • Plasma cells
• Pro-T cells
  • T-cells

By 2008 it was found that "the majority of early thymic progenitor [ETP] cells do not commit to becoming T cells by the time they get to the thymus gland. ETP cells retained the ability to become either T cells or myeloid cells."[32] [33]

See also :[34] [35] [36] [37] [38] [39]