3.1.2 Positive and negative selection and B-cell receptor reformatting

During their development, B-cell and T-cell undergo a dual process of positive and negative selection in which cells that react with high affinity against selfantigens are eliminated because they constitute a significant danger of triggering autoimmune responses.

In the case of B-cells, outside the negative selection mechanisms, cell survival depends essentially on their ability to compete for survival factors such as the B-cell activating factor (BAFF, also known as B lymphocyte stimulator (BLyS), a subset of the tumor necrosis factor (TNF) tumor necrosis factor ligand superfamily member 13B), which is present in the circulation and produced by resident cells within secondary lymphoid organs [24]. Positive selection mainly corresponds to BCR functionality test and depends on a moderate response to the self-antigen, which stimulates cell maturation and survival. When self-reactivity exceeds a certain level, a process so-called receptor reformatting is triggered by inducing a new cycle of gene rearrangements, principally in the IG L chain loci (see Chapter 2). If the receptor reformatting fails, or if the developing B-cell leaves the bone marrow and encounters the self-antigen soon after it arrives in the spleen, this cell will be eliminated [21].

### 3.1.3 Immature B-cell generation

The differentiation of hematopoietic stem cell (HSC) into immature B-cell passes through four successive steps, which could be identified by the presence of certain markers, corresponding to the different stages of rearrangement of Ig genes: a. Early pro-B-cell. These cells appear before the start of rearrangements. They are identified by CD45R, major histocompatibility complex class II (MHC II), and CDI9 and CD38 markers.

system, but, paradoxically, they can also promote the persistence of many cancers and various immune-mediated diseases. The main non-hematopoietic stromal cells involved in immunity include fibroblasts, myofibroblasts, endothelial cells, pericytes, smooth muscle cells and mesenchymal

• Bone marrow stromal stem cells (BMSC) are almost all of mesenchymal origin and, therefore, are also known as skeletal or mesenchymal stem cells. They influence the microenvironment surrounding Bcell precursors, and thus exert local effects on their development through cytokines and chemokines.

HSCs are multipotent cells; i.e., they are able to differentiate into each of the types of blood cells. After each division into two cells, one of them maintains the HSCs stock and the other one differentiates into one

The immature B-cell leaves the bone marrow parenchyma and passes through an endothelial barrier and enters the blood sinusoids, where they are retained, before finally being released into the peripheral blood [29], which allows them to migrate to the spleen where they complete their development [30]. The transition from the immature B-cell to the mature B-cell takes place in a few days and leads, via alternative splicing of long primary mRNA transcripts from the IG heavy (IGH) locus [31], to the coexpression on naïve mature B-cells of membrane IgM and IgD that share the same antigenic specificity. Only about 5% of immature B-cells will sustainably give rise to peripheral B-cells; however, most newly formed ones dis-

A so-called BCR-mediated positive signaling tonic signal and activation of Notch pathway, more precisely of Notch2, which is induced following the interaction with delta-like canonical Notch ligand 1 (DLL-1) expressed on splenic endothelial niches and required for marginal zone (MZ) B-cell development [32], allow the triggering of a first differentiation by a lineage cell choice that does not involve the antigenic

3.1.4.2 Intensity of tonic signal and differentiation of lymphoid follicular B-cells or MZ

are produced by a lineage choice during early B-cell differentiation.

differentiation of lymphoid follicular B-cells. Recall that these are structured by specialized stromal cells and follicular dendritic cells (FDCs), which play an important role in immune response by their ability to retain

b. Strong tonic signal, without Notch2 commitment. These lead to

a. Low tonic signal and Notch2 commitment. These will lead to differentiation into MZ B-cells. The latter forms a ring around the follicle and has a more complex structure in humans, also surrounded by a perifollicular zone [PFZ] that surrounds the red pulp. It is a macrophage-rich area (MZ macrophages [MZMs] and metallophilic macrophages [MMMs]) that will efficiently retain antigens from the bloodstream. In mice, the MZ B-cells are CD21++CD23-CD11 ++CD11c++, IgM++IgD+/. In humans, it is not yet known whether MZ B-cells are derived from the germinal center (CG), by an early exit at the IgM stage, or

of three types of blood cell, i.e., leukocyte, red blood cell, or platelet.

3.1.4 Transition from immature B-cell to mature B-cell

stromal cells.

Introductory Chapter: B-Cells

DOI: http://dx.doi.org/10.5772/intechopen.90636

appear within a few days.

specificity.

13

B-cells

3.1.4.1 Tonic BCR signal and notch activation


These stages are strictly dependent on the presence, in the fetal liver and bone marrow, of nonlymphoid stromal cells that come into contact with the B-cell precursors and provide the input of soluble factors in the cell microenvironment that are essential for differentiation, such as stem tell factor (SCF) and interleukin 7 (IL-7). It should be noted, moreover, that the probability of reaching productive/ efficient rearrangements of Ig genes encoding the H and L chains and obtaining an intact Ig expressed on the cell surface (mIg/BCR) is low. So most of the time there are nonproductive rearrangements, which leads to the deletion of the resulting Bcells. In addition, the differentiation of B-cells also depends, according on the cell development stage, on the presence of intracellular enzymes. Thus, the enzymes encoded by the recombination-activating genes (RAGs), RAG-1 and RAG-2, are active in early and late pro-B-cells and in pre-B-cells. Finally, the activity of the enzyme terminal deoxynucleotidyl transferase (TdT), which is involved in the addition of N-nucleotides, stops at the pre-B-cell stage.

## Key Point 2jStromal cells


a. Early pro-B-cell. These cells appear before the start of rearrangements. They are identified by CD45R, major histocompatibility complex class II (MHC II),

b. Late pro-B-cell. Variable (V), diversity (D), and joining (J) recombinations for Ig H chain gene segments and receptor expression for interleukin-7 (IL-

c. Pre-B-cell. There is contact between adhesion molecules and, in particular, vascular cell adhesion molecule 1 (VCAM-1/CD106), on the surface of the stromal cells and its ligand, very late antigen-4 (VLA-4 or α4β1 integrin, a dimer composed of CD49d (α4) and CD29 (β1)), on the surface of the pre-Bcells. At this point, intact μH chains are produced and exported to the cell surface in association with proteins that simulate light chains: λ5, which is similar to a Cλ domain, and VpreB (CD179a), which has an Ig V domain-like structure. The μ chains associated to λ5 and VpreB form a complex which, together with Igα and Igβ proteins, can be expressed at the cell surface and transduce signals that inhibit other rearrangements of genes encoding the H chains but stimulate the genes encoding L chains and cell proliferation. Moreover, expression of the CD20 marker can be detected at this stage.

d. Immature B-cell. Thanks to a productive Ig L chain genes' rearrangement, these cells express IgM on their surface, which triggers a phenomenon termed

These stages are strictly dependent on the presence, in the fetal liver and bone marrow, of nonlymphoid stromal cells that come into contact with the B-cell precursors and provide the input of soluble factors in the cell microenvironment that are essential for differentiation, such as stem tell factor (SCF) and interleukin 7 (IL-7). It should be noted, moreover, that the probability of reaching productive/ efficient rearrangements of Ig genes encoding the H and L chains and obtaining an intact Ig expressed on the cell surface (mIg/BCR) is low. So most of the time there are nonproductive rearrangements, which leads to the deletion of the resulting Bcells. In addition, the differentiation of B-cells also depends, according on the cell development stage, on the presence of intracellular enzymes. Thus, the enzymes encoded by the recombination-activating genes (RAGs), RAG-1 and RAG-2, are active in early and late pro-B-cells and in pre-B-cells. Finally, the activity of the enzyme terminal deoxynucleotidyl transferase (TdT), which is involved in the

• Stromal cells constitute a support tissue of an organ, i.e., a connective tissue cell that allows the multiplication of hematopoietic stem cells (HSCs) and their differentiation by creating a microenvironment adapted to the growth and differentiation of HSCs. They are also able to

• It has recently been suggested that stromal cells play a major role, not only in the functional

regulation of many tissues and organs, but also more particularly in the immune responses. Of note, non-hematopoietic stromal cells play a key role in the development and function of the immune

addition of N-nucleotides, stops at the pre-B-cell stage.

Key Point 2jStromal cells

12

differentiate into many cell types.

feedback inhibition, which blocks any new rearrangement of the genes encoding the L chains, thus allowing the expression of a single and unique Ig specificity by each B-cell clone. At this point, the cells express the CD21

and CDI9 and CD38 markers.

Normal and Malignant B-cell

7R) are induced at this stage.

marker.

system, but, paradoxically, they can also promote the persistence of many cancers and various immune-mediated diseases. The main non-hematopoietic stromal cells involved in immunity include fibroblasts, myofibroblasts, endothelial cells, pericytes, smooth muscle cells and mesenchymal stromal cells.

• Bone marrow stromal stem cells (BMSC) are almost all of mesenchymal origin and, therefore, are also known as skeletal or mesenchymal stem cells. They influence the microenvironment surrounding Bcell precursors, and thus exert local effects on their development through cytokines and chemokines.

HSCs are multipotent cells; i.e., they are able to differentiate into each of the types of blood cells. After each division into two cells, one of them maintains the HSCs stock and the other one differentiates into one of three types of blood cell, i.e., leukocyte, red blood cell, or platelet.
