MS Pathophysiology (Continued)

When a CD4⁺ T_N cell finally contacts a cognate antigen–expressing DC, it is either activated or tolerized. Both outcomes are germane to MS. Both require contact between the CD4⁺ T_N cell receptor and an MS-relevant small peptide fragment cleaved from an ingested protein and inserted into a cleft in a human leukocyte antigen (HLA) molecule that then is expressed on the DC surface. Cognate antigen contact lasts longer than contact with DCs expressing irrelevant peptides, and contact is of even longer duration if the CD4⁺ T_N cell is being activated rather than tolerized.

DC maturation is critical for activation of a CD4⁺ T_N cell. Maturation involves increased expression of costimulatory molecules (e.g., CD80/86 and CD40) and reduced expression of tolerizing molecules (e.g., immunoglobulin-like transcript 3 [ILT3], transforming growth factor-beta 1 [TGF-beta1], and interleukin-10 [IL-10]). Full activation of CD4⁺ T cells by DCs requires two signals. The first signal is delivered by T cell receptors following contact with their cognate antigen presented to them at an immunologic synapse by a MHC class II molecule. The second set of signals is delivered by a network of interactive costimulatory molecule pairs with CD80/86, expressed initially at low level by DCs, and CD28 expressed constitutively by T cells, providing one prototypic pair and CD40 expressed by DCs in the early stage of their activation and CD40 ligand (CD40L/CD154), induced on T cells in the early hours of their activation, providing a second.

Both cross-communicating pairs upregulate synthesis of their counter ligand, and both pairs, acting in concert, transduce additional activating signals via intracellular second messengers. Each pair synergistically reinforces the actions of the other. CD28 is critical for the initiation of T cell activation, while CD40L has a key role in sustaining it. CD40L also promotes a Th1 cell bias and an even more marked Th17 cell bias. T cells, once activated, proliferate within the LN to gener-ate an up to 10⁴-fold increase of CD4⁺ effector T cells (T_E cells).

Cytotoxic T lymphocyte antigen-4 (CTLA-4), a homologue of CD28, has a critical role in the prevention of autoimmunity. CTLA-4 is expressed by activated T cells and by regulatory T cells. CTLA-4 is focused at the immunologic synapse where it opposes CD28. Both CD28 and CTLA-4 bind CD80/86 but because of its higher affinity CTLA-4 can out-compete CD28 and reverse or blunt the T cell activating actions of CD28. DCs downregulate their expression of CD80/86 in response to the potent negative signal provided by CTLA-4 when it binds to CD80/86. In this way CTLA-4 promotes tolerance.

CTLA-4 is not expressed by resting T cells but comes to be expressed by activated T cells as an immune response evolves. CTLA-4 tempers what would otherwise be an excessive T cell expansion. CTLA-4 is constitutively expressed, and highly so, on the surface of regulatory T cells and, additionally, as a released biologically active soluble splice variant. Regulatory T cells have a major role in autoimmunity prevention. Even when prevention fails, as in MS, regulatory T cells can lessen MS relapse frequency and the severity of those relapses that do occur. Regulatory T cells also participate in the processes that end a relapse (see later). Agents that duplicate actions of CTLA-4 are of interest as MS treatments.

CD4⁺ T_E cells are divided into subtypes known as Th1 cells, Th17 cells, and Th2 cells. Broadly viewed, Th1 cells protect against intracellular organisms, Th17 cells protect against fungi, whereas Th2 cells protect against helminths, certain other extracellular pathogens, and allergens. DCs, over the course of their activation of naïve T cells, can polarize T-cell development along paths that lead to the preferential expansion of a single Th-cell subset. Th-cell subsets interact; Th2 cells inhibit Th1 cells and vice-versa. Th1 cells and Th17 cells cause damage in MS, while Th2 cells protect because they inhibit Th1 and Th17 cells. Thus any drug (e.g., glatiramer acetate, teriflunomide) or mechanism that shifts polarization from a Th1 to Th2 dominance might be expected to prove beneficial in MS.

The CD4⁺ T_E-cell population in MS contains both interferon-γ–secreting Th1 cells and interleukin-17– secreting Th17 cells. Deep cervical LN-generated MSrelevant CD4⁺ T_E cells migrate to the LN medulla, express S1P-1, move via efferent lymph into the blood and then to the CNS to participate in an MS relapse (see later).

As a relapse ends, most CNS-infiltrating Th1 and Th17 T_E cells die in situ by apoptosis, but perhaps 5% survive as T-effector memory (T_EM) cells that remain in the periphery to provide a prompt defense against a subsequent challenge. An additional 5% to 10% remain as CD4⁺ T central memory (T_CM) cells that express the same adhesion molecules as T_N cells so that they too reenter LNs via high endothelial venules, sample DCs, and recirculate via efferent lymph and then the blood to other LNs. Unlike T_N cells, MS-relevant T_CM cells also survey the tissues and other body compartments, including the CSF, seeking an APC loaded with the MS-relevant peptide they are programmed to recognize. T_CM cells outnumber T_N cells with the same specificity so that a secondary CD4⁺ T_CM-cell response in a LN is usually more rapid and robust than a CD4⁺ T_N-cell response. Activation of CD4⁺ T_N cells requires peptide presentation by mature DCs, but requirements for CD4⁺ T_CM-cell reactivation are less stringent; they can respond to some extent to antigenic peptide–presenting macrophages and to immature DCs. CD4⁺ T_CM-cell reactivation leads to generation of a new CD4⁺ T_E-cell population that again migrates to the circulation and then to the CNS.

Throughout MS remissions, myelin debris–laden immature DCs and macrophages in the cervical LNs of MS patients are thought to promote tolerance. Deep cervical LNs are favored sites for tolerance induction because they are continuously bathed with products released by the commensal biota of the nasal mucosa. The immune system is programmed to tolerate commensal organisms and to eliminate pathogens. Tolerance is mediated primarily by antigen-specific regulatory (suppressor) T cells that powerfully inhibit T-cell activation. For this reason, when antigen-specific regulatory T cells are deleted, overly robust T_E-cell responses ensue. The finding points to a ceaseless positive versus negative competition for control of DC function.

Multiple types of regulatory T cells are described. CD4⁺CD25⁺ T regulatory cells (T_REGS) and CD8⁺CD28⁻ T suppressor cells (T_S) are the most studied in MS. The role of CD4⁺CD25⁺ T_REG cells in MS remains unresolved. CD8⁺CD28⁻ T_S-cell function is grossly defective during MS relapses. CD8⁺CD28⁻ T_S-cell function reverts toward normal as relapses end and is restored, although not always fully, during remissions. Agents that augment CD8⁺ CD28⁻ T_S-cell function would be expected to be beneficial in MS.

CD8⁺, CD28⁻ T_S cells are antigen-specific. The CD8⁺, CD28⁻ T_S-cell receptor makes direct contact with its cognate antigen presented by an HLA class I allele expressed on a DC. Contact between them shunts the DC into a tolerizing mode with co-stimulatory molecule (e.g., CD86) expression reduced and tolerance-inducing molecule (e.g., ILT3) expression increased. In the obverse, when DC expression of ILT3 is silenced, proinflammatory cytokine synthesis up-regulates, and generation of disease-provoking T_E cells soars. ILT3 expression and CD8⁺ CD28⁻ T_S-cell function are reduced during MS relapses. Blood levels of type 1 interferons (IFN-α and IFN-β) are low in MS. IFN-β, widely used to treat MS, increases ILT3 expression and augments CD8⁺ CD28⁻ T_S-cell function.

A single DC can present multiple peptides to CD4⁺ and CD8⁺ T cells and can interact with up to 10 T cells, each with a different specificity, at any point in time. Likewise, a CD8⁺ CD28⁻ T_S cell can interact sequentially with several DCs, provided each expresses the antigenic peptide specific for that CD8⁺CD28⁻ T_S cell. It follows that several CD4⁺ T cells with different specificities can be tolerized in a cascade by a single CD8⁺CD28⁻ T_S cell. Further, as relapses recur and additional DCs are driven to maturity, new immune responses to recently captured self-antigens may be generated. This process is known as epitope spreading.

Multiple factors released during viral infections can drive immature tolerance-inducing DCs to maturity and convert them into immune system activators. Reinforcing actions of several factors may be required to ramp up co-stimulatory molecule expression and ramp down tolerogenic molecule expression to extents that permit an override of CD8⁺CD28⁻ T_S-cell–mediated tolerance. Included are proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α) released at sites of inflammation and swept into LNs plus binding of virus components to Toll-like receptors (TLR) expressed by DCs and macrophages. Viral infections commonly shunt MS-relevant peptide-loaded immature DCs and macrophages into a CD4⁺ T-cell–activating mode, and in this way, they can provoke MS relapses. This disease-enhancing action of viruses is probably facilitated by an unmasking in MS of a defect in CD8⁺CD28⁻ T_S-cell function that is at least in part genetically determined.

Mature DCs present antigen to T cells at the onset of an immune response, but their life span is short (only ≈3 days in mice). During the later stages of an immune response, B cells often assume the dominant APC role. B-cell receptors bind unprocessed antigen rather than the short peptide sequences recognized by T-cell receptors. Nonetheless, once an unprocessed antigen bound to a B-cell receptor has been internalized, B cells can process internalized antigenic material into small peptides, insert them into the clefts of major histocompatibility complex (MHC) class II molecules, and transfer MHC class II–processed peptide complexes to the cell surface for presentation to CD4⁺ T cells.

B cells enter LNs via high endothelial venules in T-cell–dependent areas. Most B cells move quickly from the T-cell–rich paracortex into B-cell zones called follicles. Follicle entry occurs because follicle-destined B cells express the chemokine CXCR5 that binds to its CXCL13 counterligand expressed by follicular dendritic cells (FDC), a distinct population of follicle-confined stromal cells. Follicle-infringing subcapsular macrophages transport particulate material into the follicle and pass it along to FDCs that can then present it to B cells. In addition, soluble antigens percolate into the follicle, are captured by FDCs, and presented to B cells. Antigen-pulsed B cells next leave the follicle to form monogamous immunologic synapses with DC-primed CD4⁺ T cells at the T-cell–B-cell boundary. Each B cell drags its T-cell partner along the border for a time, but the B cell then separates from its T-cell partner with the B cell moving back into the follicle, only to be followed much of the time by its prior partner T cell that has now come to express the CXCR5 chemokine, a marker for so-called follicular helper T cells (T_FH).

Bidirectional interactions between CD4⁺ T_FH cells and B cells are important for (1) germinal center formation, (2) expansion of both populations, (3) hypermutation of B cells that diversifies their antigen receptors and permits affinity-driven clonal selection, (4) B-cell differentiation into long-lived plasma cells that secrete high-affinity antibody and confer long-lasting protection from secondary challenge and, (5) development of memory B cells.

Of importance, T cells can shift lineages. Antigen-presenting B cells may shift CD4⁺ T_FH cells into CD4⁺ T_CM cells, CD4⁺ IFN-γ–secreting Th1 T_E cells and CD4⁺ Th17 T_E cells in response to low doses of autoantigens. Thus B cells can contribute in a major way to MS relapse severity.