Understanding T Cells: T Cell Types, the T Cell Receptor and T Cell Activation, Differentiation and Functions

When a pathogen enters the body, it is first met by the innate immune response. This is a non-specific response that identifies pathogens from the presence of general microbial-associated molecular patterns (MAMPs). In addition to acting as the first line of defense, the innate immune system also activates the adaptive immune response. The adaptive immune response is made up of cellular and humoral immunity and relies on the recognition of specific pathogen signatures called antigens to remove threats. This article focuses on cellular immunity and the T cells that comprise it. We discuss the different types of T cells, their development and the consequences of defective T cells.

T cells, also known as T lymphocytes, are the immune cells that drive the cellular immune response. There are several different types of T cells, each playing a distinct role. The functions of T cells include killing pathogens and damaged cells, moderating the immune response and facilitating the actions of other immune cells. 

T cells are distinguished by the presence of the T-cell receptor and the cluster of differentiation 3 (CD3), which are found on all T cells, regardless of their role. T cells can further be categorized into subtypes by the expression of other markers including CD4and CD8.¹ 

Following the identification of B cells in the 1950s, research by Jacques Miller in the early 1960s showed that the removal of an organ called the thymus in mice resulted in severe immunodeficiencies.² However, it was not until the late 1960s that lymphocytes produced by the thymus were designated as “T cells” and were seen to facilitate the production of antibodies by B cells.²

Since then, T cells have been studied in depth, and we now have a thorough understanding of their subtypes, development, functions and interactions with antigens. Even so, rare types and functions of T cells are still being discovered.³

Like other lymphocyte cells, T cells begin development in the bone marrow as hematopoietic stem cells (Figure 1). However, while B cells and natural killer (NK) cells typically remain in the bone marrow until they are fully mature, immature T cells – known as thymocytes – leave the bone marrow and travel to the thymus to mature there. Once fully mature, T cells leave the thymus and enter peripheral circulation. By entering circulation and residing in immune organs such as lymph nodes, T cells increase their chances of encountering the specific antigen that will activate them.

T cells cannot interact directly with pathogens. The pathogens must be processed into short peptide chains called antigens inside cells and presented to the T cells on major histocompatibility complexes (MHCs). The interaction of MHC–antigen combinations with T-cell receptors (TCRs) activates the T cell and initiates its effector functions. Activated T cells proliferate rapidly, producing thousands of clones that all target cells expressing the same antigen. T-cell effector functions include:

• Directing and managing other arms of the immune response
• Directly killing infected or damaged cells
• Downregulating the immune response once the infection is cleared

Figure 1: The differentiation pathway for the development of T cells, and the different types of T cells. Credit: Technology Networks.

There are several different types of T cells, all of which play an important role in the adaptive immune response. T cells are categorized according to the markers on their surface, the structure of their T-cell receptor and their effector functions (Figure 1).

The two main types of effector T cells are:

• CD4+ T cells, also known as “helper” cells
• CD8+ T cells, also known as cytotoxic T lymphocytes (CTLs) or “killer” T cells

Helper T (Th) cells direct and facilitate the immune response, while CTLs target and destroy infected and damaged cells. Regulatory T cells (Tregs), which are a subtype of CD4+ T cells, suppress and regulate the cellular immune response. Finally, memory T cells are antigen-specific cells (either CD4+ or CD8+), which survive for long periods following the clearance of infection or damage and are able to proliferate rapidly to produce large numbers of effector cells, if that specific antigen is ever encountered again. 

The presence of CD4 or CD8 on Th cells or CTLs determines what kind of antigen will activate the cell. TCRs on CD4+ Th cells interact with MHC class II molecules on professional antigen-presenting cells (APCs) loaded with antigens from extracellular sources such as bacteria or parasites. In comparison, CD8+ CTLs are activated by MHC class I molecules presenting endogenous or intracellular antigens, such as those produced from a viral infection or tumor cells. 

The majority of these cells have conventional TCRs formed by α and β peptide chains. However, a small percentage of T cells develop an unconventional TCR structure, consisting of γ and δ chains. This allows them to interact with a wide range of antigens – both endogenous and exogenous, self- and foreign antigens – in an MHC-independent manner. In contrast to αβ T cells, γδ T cells exhibit both innate and adaptive characteristics, allowing for early immune responses and surveillance at mucus membranes.⁴

Th cells express the CD4 marker and facilitate the functions of other immune components in three main ways:

• Increasing the phagocytic function of innate immune cells such as macrophages and neutrophils
• Activating antibody production by B cells
• Guiding CTL cytotoxicity. 

Th cells are activated by the presentation of a specific antigen in an MHC class II molecule on a professional APC to a CD4+ T cell. 

There are several different subsets of Th cells, which produce different, characteristic sets of cytokines, suitable for tackling different types of infection. Once the Th cell is activated, cytokines released by innate immune cells and APCs early on in infection or damage direct development into the most appropriate Th subset for the situation (Figure 2). 

Once the Th cell commits to a specific profile, a positive feedback loop ensures more of the same subset are produced. For example, Th1 cells are generated during a viral or bacterial infection. These cells produce effector cytokines including tumor necrosis factor (TNF), interferon (IFN)-γ and interleukin (IL)-2, which drive cytotoxic T-cell responses, T-cell proliferation and phagocytosis by neutrophils.^{5, 6} In comparison, an extracellular parasitic infection is more likely to result in a Th2 response and the production of IL-4, IL-5 and IL-13. These cytokines are key for defense against large or extracellular pathogens, as they support antibody production, eosinophilic responses and contribute to wound healing following infection clearance.⁷ The Th17 response is induced by fungal and extracellular bacterial infections, and it results in the production of highly inflammatory cytokines that support neutrophil functions, induce production of antimicrobial proteins and maintain the integrity of epithelial barriers.⁸

Figure 2: The differentiation profiles of CD4+ T cells, including some of the main Th subsets and their functions. Credit: Technology Networks.

Killer T cells, also known as cytotoxic T cells, cytotoxic T lymphocytes (CTLs) or CD8+ T cells, are T cells that express CD8 markers and use cytolytic mechanisms to kill damaged, cancerous or infected cells (Figure 3). Like CD4+ Th cells, CD8+ T cells will also differentiate into various subtypes with different effector functions. While the most well-known and well-characterized subset is Tc1, recent research has shown additional CD8+ T-cell subsets that more closely mirror their CD4+ Th cell counterparts. For example, when activated, Tc2 subset CD8+ T cells produce IL-4 and IL-5 and stimulate antibody responses.⁹

CTLs are activated by the presentation of a specific endogenous antigen in an MHC class I complex. Once activated, CTLs release granules that contain perforin and granzyme enzymes. Perforin is a membrane disrupter that creates holes in the target cell wall, while granzymes cleave proteins inside the target cell and initiate cell death by apoptosis. In addition, CTLs also produce large amounts of the cytokine IFN-γ to support their own functions further, including increasing CTL motility, promoting the recruitment of neutrophils and macrophages and increasing MHC class I expression on target cells.¹⁰

Figure 3: CTL killing mechanism. Credit: Technology Networks.

Although they may have similar subsets, there are clear differences between the functions and mechanisms of CD8+ T cells and CD4+ T cells. CD4+ T cells are activated by exogenous antigens on MHC class II molecules, which are only expressed on professional APCs (i.e., dendritic cells, macrophages and B cells). In comparison, CD8+ cells can be activated by MHC class I molecules (expressed on almost all cells), which present endogenous antigens. 

The other main difference between the types of T cells is the effector functions. The main function of CD8+ T cells is to kill infected or cancerous cells directly by releasing lytic enzymes. However, CD4+ T cells do not directly destroy infected cells, but support the lytic action of CD8+ T cells, phagocytosis by macrophages and antibody production by B cells. 

Regulatory T cells (Tregs) are T cells that control and modulate the immune response, by inhibiting T-cell responses to self-antigens (i.e., antigens on the body’s own cells) and suppressing immune responses once an infection has been cleared. Tregs start life as CD4+ T cells, but self-antigens activate their TCRs during development. Instead of being destroyed like other self-reactive lymphocytes, they express Foxp3 and become Tregs.¹¹

Foxp3 is an essential marker in Treg identity, but several other key markers also enable Tregs to carry out their immunosuppressive functions via a variety of methods (Figure 4). For example, CD25 is a competitive receptor for IL-2 – an important survival and proliferation signal for T cells. The CD25 on Tregs sequesters IL-2 away from other T cells, preventing replication. Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death receptor protein 1 (PD-1) inhibit T-cell activation by blocking the costimulatory receptors that help activate the TCR.¹¹ 

Figure 4: Treg markers for immunosuppressive functions. Credit: Technology Networks.

In addition, Tregs produce high levels of immunosuppressive cytokines such as IL-10 and transforming growth factor-β (TGF-β), which downregulate inflammation and immune cell proliferation. 

In recent years, a new type of Treg, CD8+ Tregs, has been discovered. These are distinct from conventional Tregs, which are derived from Th cells, but are also thought to play a role in immune regulation and homeostasis. They have been shown to suppress T-cell proliferation, inhibit autoimmunity and are thought to suppress or remove activated Th cells, though multiple questions about their function and origins still remain.¹²

When naïve T cells are activated by their specific antigen and begin to proliferate, some of the resulting population of clones become memory cells. These cells survive for long periods – years or even decades – following infection and can be reactivated by a repeat encounter with the same antigen. Memory T cells respond much faster than naïve T cells, and so can mount a faster immune response, meaning that any subsequent infections with the same pathogen are milder, or prevented entirely. 

There are several subsets of memory T cells, which are classified according to the locations they patrol while waiting for antigen encounters. Resident memory T cells (T_RM) remain at the primary site of infection or damage, where the original immune response occurred. Central memory T cells (T_CM) circulate through secondary lymphoid tissues such as the lymph nodes, while effector memory T cells (T_EM) circulate through non-lymphoid tissues.¹

The T-cell receptor is essential for T-cell antigen recognition and activation. It interacts with antigens presented on MHC class I or MHC class II molecules and co-receptors in order to trigger T-cell activation, proliferation and effector functions. 

Figure 5: The T-cell receptor interacts with MHC class II on a professional APC. Credit: Technology Networks.

T-cell receptors consist of two polypeptide chains – TCRα and TCRβ – which form a heterodimer (Figure 5). Each polypeptide chain has a conserved, constant region and a hypervariable region. The TCR is also accompanied by a CD3 co-receptor, which is found on all T cells, regardless of their other markers. The CD3 co-receptor consists of four different polypeptide chains: one δ, one γ, two ε and two ζ. CD3 also helps to stabilize the TCR and transduce the activation signal to the nucleus to begin production of effector proteins. 

Interaction with MHC class II/antigen complexes on an APC alone isn’t sufficient for T-cell activation. Complete signal transduction requires co-stimulatory signals from co-receptors. On the T cell, this receptor is CD28, which binds to CD80 or CD86, the corresponding co-receptor on the APC. Co-stimulatory receptor expression is upregulated by innate immune responses, but if an antigen is presented to a T cell without the presence of co-receptors, the T cell becomes anergic (non-responsive). These measures prevent the unnecessary activation of T cells, which could result in destructive, unneeded immune responses. 

There are an enormous number of potential antigens that T cells must be able to recognize as markers for infection. The number of individual genes needed to encode for all these potential receptors individually would be far more than the cellular DNA could cope with. Therefore, T cells use a process called gene rearrangement to meet this demand instead. 

TCR genes are organized in three separate gene loci: variable (V) and joining (J) regions, which code for the hypervariable regions of both the TCRα and TCRβ polypeptides, and diversity (D), which codes for the constant region of the TCRβ. During T-cell development in the thymus, one of each of these gene segments are brought together, and the unneeded extra segments are removed by RNA splicing. The random selection of V, J and (if necessary) D segments accounts for some of the diversity of TCRs. The rest is provided by junctional diversity during splicing, which adds or deletes short sections of nucleotides at the junctions between segments. 

Both B and T cells begin development from hematopoietic stem cells in the bone marrow. However, where B cells complete the entirety of their development in the bone marrow, immature T cells known as thymocytes leave the bone marrow and travel to the thymus to complete maturation. 

Figure 6: T-cell development. Credit: Technology Networks.

Thymocytes arrive in the thymus as pro-T cells, which do not express TCRs or CD4/CD8 (Figure 6). The lack of CD4 or CD8 at this stage denotes the thymocytes as “double negative”. Gene rearrangement begins at the pro-T-cell stage, with the TCRβ chain rearranged first. If this recombination is successful, the β chain is transported to the cell surface, and the cell becomes a pre-T cell, suppressing any further β chain rearrangement. The formation of the pre-T cell also mediates signals to begin α chain rearrangement, completing the TCR complex. All cells now express both CD4 and CD8 markers, so they are termed “double positive”. 

At this stage of development, the thymocytes are also tested for tolerance. The TCRs must only recognize foreign antigens – recognition of normal, healthy self-antigens can result in autoimmune disease. To test tolerance, the thymocytes are exposed to epithelial cells in the thymus that express a wide variety of self-antigens. Strong recognition of self-antigens results in negative selection, and the cells are triggered to die by apoptosis. No recognition of antigens at all means the thymocyte dies by neglect. 

Thymocytes that weakly recognize self-antigens (therefore indicating that the TCR is functional, but the signal would not be sufficient to activate a mature T cell) are positively selected and continue development. Positively selected cells mature into single positive immature T cells, which express either CD4 or CD8. Finally, the thymocytes move through the final layer of the thymus – the medulla – and leave the thymus as fully mature T cells. 

Mature T cells circulate in peripheral lymphoid tissues until they encounter their specific antigen and become activated. However, T cells cannot interact directly with antigens or the pathogens from which they are derived. Pathogens are processed inside cells into short peptide sequences and presented on the MHC to the T cells. This is achieved through one of two pathways, depending on the source of the antigen. MHCs are coded for by the human leukocyte antigen (HLA) genes. These genes are both polygenic and polymorphic – meaning the complex is transcribed from three genes, each of which have hundreds of potential alleles. This gives rise to high levels of diversity to allow for the presentation of an extremely wide range of antigens; it is estimated that the human population as a whole expresses 8–9 million HLA genes for MHC class I.¹³  

MHC class I molecules are found on most cell types in the body and present endogenous antigens to CTLs. Intracellular proteins (e.g., from viral or other intracellular pathogen infections) are digested into peptides in the proteasome and then transported to the endoplasmic reticulum (Figure 7A). Here, short peptides of 9–11 amino acids are loaded into MHC class I molecules and transported to the surface for presentation to the TCRs of CTLs.¹⁴ 

Figure 7: A) The MHC class I pathway. B) The MHC class II pathway. Credit: Technology Networks.

MHC class II molecules are found only on APCs and present exogenous antigens to Th cells. In this pathway, extracellular pathogens such as bacteria are phagocytosed or endocytosed by APCs into endosomes (Figure 7B). The endosome releases lytic enzymes that digest the pathogens into short peptides of 13–25 amino acid. These peptides are loaded into MHC class II molecules and transported to the cell surface for presentation to Th cells. To prevent them from binding to cellular proteins, an invariant chain protein (CLIP) sits in the MHC binding groove, blocking it. The chain is cleaved, and the CLIP fragment is removed in the endosome with the exogenous antigens.

In both the MHC class I and MHC class II pathways, binding and recognition of a presented antigen as non-self by the TCR, combined with a co-stimulatory signal through co-receptors, is sufficient to activate the T cell. The bound TCR transduces a signal across the T-cell membrane, which initiates a signaling cascade inside the cell, triggering expression of genes for cytokine production, proliferation and effector functions. Activated T cells express high levels of IL-2, along with its receptor IL-2RΑ. Together, these markers ensure growth, survival and proliferation of activated cells only, resulting in the expansion of antigen-specific clones. The vast majority of clones produced will be Th and CTL effector cells, which work to control the infection. However, some will become memory cells, persisting after infection is cleared, in preparation for further encounters with the same antigen. This results in a faster, stronger immune response that either reduces disease severity or completely prevents symptomatic infection.

Once the issue – whether infection, damage or malignant cells – is cleared, Treg cells release anti-inflammatory cytokines and inhibit further effector cell replication to prevent unnecessary damage and return the immune system to homeostasis. 

T cells and B cells both begin life as hematopoietic stem cells in the bone marrow, but diverge in function, appearance and location as they develop (Figure 8). B cells direct the humoral response, while the effector functions of T cells are known as the cellular response. Both cells are activated by interaction with specific antigens, but T cells require antigen presentation, while B cells can interact with antigens directly. 

Figure 8: The differences between B cells and T cells. Credit: Technology Networks.

Although T cells are an essential part of a healthy immune response, incorrect function or lack of function can result in severe, even life-threatening issues. Malfunctioning T cells can result in several outcomes, including immunodeficiency, autoimmunity and hypersensitivity (Table 1). T cells can also become cancerous, resulting in T-cell malignancies. Let’s consider these issues in more detail.

Table 1: Outcomes of a malfunctioning immune system.

T cell immunodeficiencies

Immunodeficiencies arise when the immune system’s ability to fight pathogens is reduced or absent. Although T-cell-based primary immunodeficiencies (those caused by genetic abnormalities and present from birth) are extremely rare, absent or defective T-cell responses can have wide-ranging, severe effects on wider immunity. 

One such example is X-linked severe combined immunodeficiency (SCID), caused by mutations in the IL-2R γ chain (IL2RG) gene.¹⁵ This gene encodes part of the receptor for several cytokines essential for T-cell development and maturation, and its mutation results in an absence of functioning T cells and NK cells. Though B cell numbers may be normal or even increased, they are often only partly functional.¹⁶ X-linked SCID patients are highly at risk for severe infection, and life expectancy is short without effective treatment. Current treatment options include hematopoietic stem cell transplants, or gene therapy, in which the patient’s own hematopoietic stem cells are collected, transfected with a functional IL2RG gene and reintroduced into the patient.¹⁶

While primary immunodeficiencies are congenital, secondary immunodeficiencies can develop over time as a result of external influence. One such example that results in T-cell deficiency is the human immunodeficiency virus (HIV), which affects around 40 million people worldwide.¹⁷ HIV accelerates both the production and destruction of CD4+ Th cells, resulting in a significant reduction of Th cells over time, which also affects the normal function of both cellular and humoral adaptive immune responses. When blood CD4+ T cell levels drop below 200 cells/µl, acquired immunodeficiency syndrome (AIDS) is diagnosed, and patients become highly susceptible to opportunistic infections.¹⁸ However, antiretroviral therapies are now so effective that undetectable viral levels can be achieved. 

T cell autoimmune diseases

Autoimmune diseases occur when tolerance breaks down, and immune cells attack our body’s own cells, after failing to recognize them as “self”. 

One example of a T-cell-driven autoimmune disease is type I diabetes (T1D). Although the exact causes of T1D are unknown, there do appear to be environmental factors (e.g., congenital rubella infection) and genetic predispositions (e.g., particular HLA alleles) that can increase the chance of developing the disease.¹⁹ T1D manifests following the natural or infection-induced death of insulin-producing beta cells in the pancreas. Dendritic cells take up the autoantigens released by beta cell death and present them to Th cells. The Th cells are mistakenly activated and trigger self-reactive CD8+ CTLs, auto-antibody production from B cells and inflammatory cells. Primed CTLs accumulate in the pancreas, destroying beta cells and preventing blood glucose levels from being managed appropriately. 

Mismanaged blood glucose results in short-term effects including confusion, seizures and diabetic coma, and long-term complications including neuropathy, blindness and kidney failure. Though there is no cure, T1D can be managed by continuous blood glucose monitoring and automated insulin injections.

T cells in hypersensitivities

Hypersensitivities refer to immune responses that are misdirected at innocuous antigens. There are several different types of hypersensitivities, though most of these are antibody-mediated. Only one – type IV hypersensitivity – is mediated by T cells, and functions in a similar method to a normal pathogen response. Th cells recognize the innocuous antigen through presentation on MHC class II, and an inflammatory Th1 response is triggered, characterized by high levels of cytokines such as IFN-γ and TNF. 

One such example is contact hypersensitivity, caused by the penetration of haptens into the skin, e.g., from poison ivy or nickel. Haptens are small molecules that only elicit immune responses when attached to larger carrier self-proteins. Specialized dermal dendritic cells known as Langerhans cells present the haptens to T cells, initiating a cellular immune response.²⁰ A tuberculin-type hypersensitivity can be triggered by a similar mechanism following administration of the tuberculin purified protein – a derivative of Mycoplasma tuberculosis. This test is used as a diagnostic test for tuberculosis infection and to assess prior Mycoplasma tuberculosis exposure before vaccination.²⁰

T cell lymphomas

Both B and T cells can become defective and malignant, causing lymphomas (originating in lymph nodes) and leukemias (originating in bone marrow). T-cell malignancies tend to be rarer than B cell malignancies but can range from chronic illnesses to highly aggressive diseases.

There are two main types of T-cell lymphomas: cutaneous (CTCL) and peripheral (PTCL). CTCLs are characterized by the infiltration of malignant T cells into the skin, and so result in dermal lesions and rashes.²¹ Diagnosis of CTCL can be difficult due to the slow onset of broad clinical symptoms, but can include the biopsy of dermal lesions such as scaly plaques, coupled with complete blood panels.²² There is no current cure for CTCL, and though it can be managed with treatments such as chemotherapeutic agents and photodynamic therapy, it tends to be a chronic disease that will recur if treatment is discontinued. 

PCTLs are rarer than CTCLs, but tend to have much poorer prognoses, with a 5-year survival rate of 30–40%.²³ However, they can be cured through use of the antibiotic anthracycline coupled with autologous stem cell transplants, though relapse is common.²⁴

The natural effector functions of T cells can be leveraged and improved to treat disease. Cancer is the most common (and the most successful) target for T-cell therapies, but ongoing research shows promise for their use against pathogenic, metabolic and autoimmune diseases.²⁵ 

Figure 9: The steps of CAR T-cell therapy. Credit: Technology Networks.

Chimeric antigen receptor (CAR) T-cell therapy is the most successful form of T-cell therapy to date, with six CAR T-cell products already approved for hematological malignancies.²⁶ In CAR T-cell therapy, T cells are removed from the patient’s blood and transfected with the gene for a chimeric TCR, specifically designed to target a particular cancer antigen (Figure 9). The genetically modified T cells are then proliferated and infused back into the patient. The activated CAR T cells are better able to attack and kill the cancerous cells than the patient’s normal immune cells, and ideally provide long-term protection against relapse. 

Although they have shown great success against blood cancers, solid tumors remain a challenge for CAR T-cell treatment. Tumors are difficult to infiltrate and they perpetuate a strong immune-evasive environment, down-regulating immune responses. In these cases, tumor-infiltrating lymphocyte (TIL) therapy has shown more success. In TIL therapy, T cells from the vicinity of the tumor are collected and tested. Those that show the best anti-tumor responses are collected and the populations are rapidly expanded. This results in large populations of activated T cells that are able to attack the tumor. TIL has been successfully used in patients with solid tumors, including metastatic melanomas.²⁷ Eventually, TIL T cells are also exhausted by the immunosuppressive tumor environment, and research to develop improved, longer-lasting CAR T-cell and TIL therapies is continuously ongoing.