Apoptosis in the Pathway of T cell Development and Differentiation - Term Paper Example

 «Apoptosis in the Pathway of T cell Development and Differentiation» Apoptosis, also known as programmed cell death, is a genetically controlled, orchestrated event, where the cell participates in its own demise. Cell death by apoptosis is determined by many factors, including cell type, developmental and physiological state of the cell. Apoptosis employs an energy-dependent cascade of events at the molecular level. There are two main pathways: the extrinsic or death receptor pathway, and the intrinsic or mitochondrial pathway. Both pathways involve a signal transduction pathway in response to specific stimuli. Transduction of the signal results in formation of cellular structures that trigger the activation of caspases (cysteine-aspartic acid proteases). The two pathways converge on the execution pathway of the cell, involving the action of execution caspases, endonucleases, and proteases, to degrade DNA and proteins. A parallel pathway, the granzyme A pathway, which is activated by cytotoxic T cells, shows caspase-independent cell death via single stranded DNA damage (Martinvalet, Zhu, & Lieberman, 2005). The main morphological features of apoptosis are chromatin condensation and nuclear fragmentation (Galluzzi, et al., 2007). Progressive condensation and budding result in the formation of apoptopic bodies, with ligands for phagocytic receptors. The bodies are engulfed through phagocytosis by macrophages, neoplastic cells, and parenchymal cells (Elmore, 2007). Apoptosis does not cause an inflammatory response because the apoptotic cells do not leak their cellular constituents into the surrounding tissue fluids. Moreover, the quick phagocytosis prevents secondary necrosis, and the engulfing phagocytes actually downregulate the immune and inflammatory responses (Ren & Savill, 1998). Compared to apoptosis where the cell plays an active role in its death, necrosis is characterized by passive cells where death is accompanied by cell inflammation. The inflammatory response over an area of surrounding cells distinguishes necrosis from apoptosis. Two mechanisms, damage to cell membranes, and the reduction in the cell’s energy supply, result in necrotic cell injury. Some of major morphological changes during necrosis are cell swelling (oncosis), formation of blebs, swelling of endoplasmic reticulum; mitochondrial irregularities; disrupted organelle membranes; swollen and ruptured lysosomes; and eventually disruption of the cell membrane. Loss of cell membrane integrity releases the cytoplasmic contents into the surrounding tissue, sending chemotatic signals that are involved in the eventual recruitment of the inflammatory response. There are no biochemical markers that can indicate necrotic events, which can only be observed with electron microscopy. Necrosis is considered harmful because inflammation can encourage the growth of local tumours (Galluzzi, et al., 2007). Although the mechanisms and morphologies of apoptosis and necrosis differ, there is overlap between these two processes. Evidence indicates that necrosis and apoptosis represent morphologic expressions of a shared biochemical network described as the “apoptosis-necrosis continuum” (Zeiss, 2003). Decreases in caspases and intracellular ATP during an ongoing apoptopic process will convert apoptosis to a necrotic event. There are two other types of cell death: autophagy and mitotic catastrophe (Galluzzi, et al., 2007). Autophagy is characterized by lack of chromatin condensation and formation of vacuoles in the cytoplasm. Compared to apoptosis, which is rapid, autophagy is a slow process where components of the cytoplasm are sequestered in vacuoles and hydrolyzed by digestive enzymes. Mitotic catastrophe, occurring during or after a failed mitosis, is characterized by excessive formation of nuclei. 1. Discuss the role of apoptosis in the development of haemopoiesis and then specifically in T cell development. Haemopoiesis is the process of the lifelong production of the different blood cells. Blood cells are derived from stem cells in the adult bone marrow and foetal embryos. The process of haemopoiesis starts with stem cell division resulting in two daughter cells where one cell serves for self –renewal, while the other undergoes differentiation to form the myeloid and the lymphoid progenitors. From the myeloid progenitor branch, arise dendritic cells and other committed blood progenitor cells: granulocyte-monocyte, eosinophils, basophils, megakaryocytes, and erythroids. These produce the corresponding cells: macrophages, neutrophils, eosinophils, basophils, platelets and erythrocytes (Abbas & Lichtman, 2003). Commitment to differentiate to a specific cell lineage is affected by chance and the external signals received by the cells. Several transcription factors that are expressed at low levels have been found to regulate differentiation of the lineages. Growth factors also direct the differentiation to a cell lineage (Gordon, 2007). Continuous self-renewal of stem cells produces homogeneous populations of daughter stem cells, showing that the behaviour of haemopoietic stem cells is quite predetermined (Müller-Sieburg, Cho, Thoman, Adkins, & Sieburg, 2002). Role of apoptosis in haemopoiesis During an organism’s lifetime, almost every cell is replaced many times. Red blood cells are replaced at the rate of 3000 cells per second. Because some blood lineages have half-life of many years (others have half-life of a few hours only), it is possible that overproduction of blood cells could occur. Uncontrolled and continuous increase in the number of blood cells could lead to disease. With normal and functional apoptopic machinery, the number of blood cells remains balanced, and cells damaged by internal and external stresses are removed or deleted (Abbas & Lichtman, 2003). Apoptosis has direct roles in controlling the number of in vivo haemopoietic stem cells, which maintains homeostasis and the number of cells constant at any given time. If apoptosis is prevented, there is an uncontrolled increase and proliferation of haemopoietic stem cells, increased progenitor lines and the development of malignancies and autoimmune disease (Domen, Cheshier, & Weissman, 2000). The withdrawal and inhibition of growth factors (glycoprotein hormones) and cytokines during the development of haemopoietic stem cells act as signals that the cells are to undergo apoptosis (Cowling & Dexter, 1994). Growth factors are survival signals that affect cell processes through polypeptide receptors that are bound to the membrane. These signals share features of signal transduction with proliferative responses. When growth factors are inhibited in the cytoplasm, the cells will express Bcl-2, an apoptosis inhibitor. Bcl-2 was the first mitochondrial protein found to inhibit apoptosis through its inhibitory action on the release of cytochrome c from the mitochondria to the cytoplasm, where it can bind to a protein that can activate the caspases that initiate cellular demise. The studies suggest a model where the major role of haemopoietic growth factors was to suppress apoptosis and mitogen action (Fairbain, Cowling, Reipert, & Dexter, 1993). Thus, cell death could be the underlying cause of some haematological diseases (Cowling & Dexter, 1994). As proof, studies in transgenic mice that overexpress Bcl-2 (and therefore, apoptosis is inhibited) show perturbations in cell populations and result in malignancies and tumorigenesis (Domen, Cheshier, & Weissman, 2000). The absence of growth factors also represses the expression of Mcl-1, also a member of the Bcl-2 family. Mcl-1 is required for the survival of haemopoietic stem cells (Opferman, et al., 2005). In mice, deletion of Mcl-1 resulted in the loss of bone marrow progenitor cells. But growth factors increased transcription of the Mcl-1 gene, showing that Mcl-1 is a critical and specific regulator important for the homeostasis of hematopoietic stem cells. Roles of apoptosis in the development of T lymphocytes Apoptosis has a central role in lymphocyte development and homeostasis. The phenomenon of cell death has to be finely tuned during T cell development to prevent uncontrolled apoptosis, which can cause lack of cells and lead to immunodeficiency. Lack in apoptosis, on the other hand, leads to lymphomas or autoimmune disorders (Rathmell & Thompson, 2002). There are several stages of T cell (also called T lymphocytes) development from the progenitor lymphoid cells to the mature phenotype. At each stage, apoptosis is present (Figure 1). During the early development stage, growth factor signalling regulates the survival of progenitors (haemopoietic stem cells discussed previously in this paper). As the lymphoid cells differentiate towards maturity, the role of apoptosis is to remove the lymphocytes that are autoreactive or those that have affinity for self-antigens. Another role for apoptosis is to ascertain that the functional antigen receptors are expressed (Opferman, 2008). Apoptosis during T cell development is regulated by the Bcl-2 protein family, which has both pro and anti-apoptopic proteins (Opferman & Korsmeyer, 2003); and the tumour necrosis factor (TNF) death receptors (Holtzman, Green, Jayaraman, & Arch, 2000). Understanding the role of apoptosis in the development of T lymphocytes requires the knowledge of the T cell formation pathway. The T cells originate from lymphoid progenitors of haemopoietic stem cells. In the thymus, they undergo a series of differentiation stages before maturing into helper T cells and cytotoxic T cells. After maturation, they are released into peripheral and circulation systems as active participants in the immune response (Figure 1). Originating in the thymus, the T cells, before being transferred to the peripheral and circulation systems, undergo a series of screenings to ensure that they act only on non-self antigens. Cells that fail to pass the screening are killed through types of apoptosis where death results from a loss in signal transduction components that promote survival (neglect), and death that is receptor mediated (Rathmell & Thompson, 2002). These deaths could be attributed to one or more of the following reasons: failure to express functional antigen receptors, failure to be positively selected by major histocompatibility complex (MHC) molecules in the thymus, and self- antigen induced negative selection. Figure 1. An integrated overview of the participation of different apoptosis mechanisms during T cell development from haemopoietic stem cells to memory cells. Figure was adapted and re-drawn based on Zhang et al., 2005. Apoptosis in the pathway of T cell development and differentiation Haemapoietic stem cells from the bone marrow travel to the lymphoid system to produce T cell and B cells. At the thymus, different mechanisms of apoptosis occur along every important regulatory step of the T cell development pathway (Figure 1). 1. Positive selection. Cells entering the thymus are called double negative (DN) lines because they do not express CD4 and CD8 cell surface receptors, nor the TCR (T cell receptor) genes. Thus, these cells have to undergo four stages of recombination (DN1-DN4) to the immature single positive (SP) cells. At the DN stage, the cytokine IL-7 acts as a survival signal molecule; in its absence the intrinsic pathway’s anti-apoptopic factors Bcl-2 and Mcl-1, are repressed which leads to apoptosis (Zhang, Hartig, Dzhagalov, Draper, & He, 2005). Thus, both Bcl-2 and Mcl-1 act as downstream of IL-7 in ensuring double negative thymocyte survival. At the DN3 stage, a pre-TCR signal has to be expressed, without this, the cells will undergo apoptosis through both intrinsic and extrinsic apoptopic pathways. Bcl-2 anti-apoptopic family member, Bcl2A1 or A1, downstream of the pre-TCR and protect DN cells from apoptosis (Mandal, et al., 2005). The death receptor pathway could be the mediator of apoptosis at the DN3 stage because of the expression of several TNF receptors (Newton, Harris, & Strasser, 2000). After the DN4 stage, the thymocytes should have a functional TCR and possess both CD4 and CD8 receptors. These thymocytes are now called double positive (DP) cells. DP thymocytes that still fail to produce a functional TCR undergo a process called positive selection (Jameson, Hogquist, & Bevan, 1995). A DP thymocyte with a successfully rearranged TCR molecule scans an array of self MHC-peptide complexes in the thymus. DP thymocytes with TCR having intermediate affinity for self MHC-peptide complexes are positively selected, while those with low affinity undergo death by neglect (Goldrath & Bevan, 1999; Starr, Jameson, & Hogquist, 2003). T cells that have a high affinity for self-MHC are also not selected (negative selection) to remove autoreactive T cells that can result in tumour production. Majority of cell death (90% of developing T cells) is due to death by neglect. It is believed that Bcl-2 family members (Bcl-xL, Bim, Bak, and Bax) are the main actors in death by neglect with minimal death receptor involvement (Zhang, Hartig, Dzhagalov, Draper, & He, 2005). 2. Negative selection process. Selected double positive thymocytes then undergo a second step of selection, the negative selection process (central tolerance). Central tolerance screens for auto-reactivity with self-antigens, which are the only type of antigens that are present in the thymus. In the thymus, only self-antigens are present so the negative selection process is very efficient against autoreactivity. These selection processes results in mature T cells that are self-MHC restricted and tolerant to many self-antigens. Antigen receptors that are highly autoreactive die by apoptosis. Thus, the T cells that are allowed to leave the thymus are able to recognize and bind their respective type of self-MHC that is associated with a non-self or foreign antigen. The positive and negative selection processes differ with respect to binding affinity (Risueno, Van Santen, & Alarcon, 2006) since positive selection is associated with weak binding to MHC, while negative selection is based on strong avidity to self-antigens. Negative selection is thought to be mediated by the death receptor pathway. However, until recently, the mechanism of apoptosis was not clear due to the observation that mutants of death receptors Fas, Foxp3 and IL-2 develop severe autoimmunity despite normal negative selection. A mechanism for negative selection involves the autoimmune regulator (AIRE), a requirement in thymic stromal cell differentiation (John, Thompson, & Winoto, 2007). Based on the model proposed by John et al. (2007), central and peripheral tolerance work together with the main downstream activation of pro-apoptopic factors Bim and Nur77. AIRE expression in thymic stromal cells results in the expression of antigens that are perceived by TCR receptors resulting in activation of gene expression of pro-apoptopic factors. However, much remains to be studied in the apoptopic mechanisms during negative selection. To summarize, several critical checkpoints that exist during T cell development ensure the survival only of the thymocytes with TCRs that have intermediate affinity for self-MHC. Death by neglect is the main death pathway for those thymocytes with useless TCRs. Negative selection removes thymocytes that are highly reactive with self-MHC. Those with useless TCRs do not receive a positive selection signal and undergo death by neglect. Those with high affinity TCRs are also eliminated by negative selection. The stringent control measures carried out by the cell via apoptosis results in an estimated 95% T cell precursors being eliminated during T cell development (Chao & Korsmeyer, 1998). This underscores the very essential role of apoptosis in developing a functional T cell repertoire. 3. Apoptosis in the maintenance of cell homeostasis in the periphery. Cells that pass the negative selection process can now differentiate into mature phenotypes and enter the periphery and the circulation. In the peripheral tissues, the T cell numbers are still tightly regulated and maintained at a constant level despite the production of new lymphocytes. This is made possible by balancing the bone marrow and thymus production, and peripheral expansion, with apoptopic cell death. In the event that the immune response is summoned, there are a large number of antigen-specific lymphocytes in the peripheral tissues. Checkpoints are put in place to make sure that only antigen-specific lymphocytes are proliferating. After the immune response, apoptosis plays the role of clearing the lymphocytes that were involved. T cells become sensitive to death induced by the death receptor Fas, suggesting that encounters between antigen and receptor could prime the lymphocytes for cell death (Rathmell & Thompson, 2002). It has also been reported that the two pathways of apoptosis could mediate the clearing of antigen-specific cells after an immune response: neglect and receptor-induced. Death receptor-induced apoptosis could allow for the proliferation of cells that are facing new antigens. Other cells dying by neglect appear to do so because they have lost some of the signals for survival after the antigens have been cleared and loss in cytokines. T cells with functional TCRs enter the periphery as naive T cells. These cells are long-lived, and constantly circulate blood, lymph, and secondary lymphoid organs. Naive T cells numbers are constant except when transferred to a host that does not have T cells. Naive T cells require growth regulator-mediated, and TCR/MHC signals to survive and proliferate (Fry & Mackall, 2000). The MHC molecules signalling survival to the naive T cells are believed to similar to those that mediate positive selection in the thymus; while the Bcl-2 gene family is conceded to be major effector molecules downstream of the signalling pathways controlling naive T cell survival (Fry & Mackall, 2000). The naive T cell is activated when it is presented by antigen my mature dendritic cells expressing high levels of MHC molecules and T cell co stimulatory molecules (Lipscomb & Masten, 2002). Activated T cells proliferate, and then differentiate into effector T cells; T helper (TH) cells cytotoxic T lymphocytes (CTLs) (Abbas & Lichtman, 2003). The effector T cells produce cytokines while CTLs kill infected cells. After the T cells have resolved the infection, more than 90% are rapidly eliminated by both death receptor and intrinsic pathway mediated apoptosis (Zhang, Hartig, Dzhagalov, Draper, & He, 2005). Activation induced cell death (AICD) is generally believed to be relevant to chronic infection, and not in acute infection. The intrinsic death pathway occurs after the depletion of growth factors for the effector cells. When this happens, the deprived cells undergo apoptosis mediated by Bcl-2 gene family members Bax and Bak (Rathmell, Lindsten, Zong, R.M, & Thompson, 2002). The remaining activated T cells differentiate into memory cells for immune protection against antigen re-entry. The survival of memory cells are also dependent on IL-7 signalling, thus apoptosis is via the intrinsic death pathway when the growth signals are limited (Zhang, Hartig, Dzhagalov, Draper, & He, 2005). In conclusion, apoptosis has very essential roles in T cell development and function. 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