Desenvolvimento de Linfócitos T alfa/beta

Question 1

Recetor da célula T (TCR; T cell receptor) alfabeta

Answer 1

The T-cell receptor heterodimer is composed of two transmembrane glycoprotein chains, alfa and beta. The extracellular portion of each chain consists of two domains, resembling immunoglobulin V and C domains, respectively.

Both chains have carbohydrate side chains attached to each domain. A short stalk segment,
analogous to an immunoglobulin hinge region, connects the Ig-like domains to the membrane and contains the cysteine residue that forms the interchain disulfide bond.

The transmembrane helices of both chains are unusual in containing positively charged (basic) residues within the hydrophobic transmembrane segment. The alfa chain carries two such residues; the beta chain has one.

Cadeias gama e delta no recetor das células T (TCR) gamadelta

Question 2

O complexo TCR alfabeta -CD3

Answer 2

Ligação não-covalente

ITAM - Immunoreceptor Tyrosine-based Activation Motif

O complexo TCR gamadelta-CD3 é igual, com exceção da presença das cadeias gama e delta que substituem as cadeia alfa e beta do TCR

The functional T-cell receptor (TCR) complex is composed of the antigen-binding TCR alfa:beta heterodimer associated with six signalling chains: two epsilon, one delta, and one gama collectively called CD3, and a homodimer of zeta. Cell surface expression of the antigen-binding chains requires assembly of TCR a:β with the signalling subunits. Each CD3 chain has one immunoreceptor tyrosine-based activation motif (ITAM), shown as a yellow segment, whereas each zeta chain has three.

Question 3

Rearranjo dos genes que codificam para as cadeias alfa e beta do TCR

Answer 3

The TCR alfa- and beta-chain genes are composed of discrete segments that are joined by somatic recombination during
development of the T cell.

For the a chain (upper part of figure), a Valfa gene segment rearranges to a Jalfa gene segment to create a functional V-region exon. Transcription and splicing of the VJalfa exon to Ca generates the mRNA that is translated to yield the T-cell receptor a-chain protein.

For the β chain (lower part of figure), like the immunoglobulin heavy chain, the variable domain is encoded in three gene segments, Vbeta, Dbeta, and Jbeta. Rearrangement of these gene segments generates a functional VDJbeta V-region exon that is transcribed and spliced to join to Cbeta; the resulting mRNA is translated to yield the Tcell
receptor beta chain.

The alfa and beta chains pair soon after their synthesis to yield the alfa:beta T-cell receptor heterodimer.

Not all J gene segments are shown, and the leader sequences preceding each V gene segment are omitted for simplicity

Question 4

As células T diferenciam-se no timo, migrando depois para a periferia onde são ativadas por Ag exógenos

Answer 4

T-cell precursors migrate from the bone marrow to the thymus, where they commit to the T-cell lineage following Notch receptor signalling.

In the thymus, T-cell receptor genes are rearranged (top first panel); alfa:beta T-cell receptors that are compatible with self MHC molecules transmit a survival signal on interacting with thymic epithelium, leading to positive selection of the cells that bear them. 
Selfreactive receptors transmit a signal that leads to cell death, and cells bearing them are removed from the repertoire in a process of
negative selection (top second panel). 

T cells that survive selection mature and leave the thymus to circulate in the periphery; they repeatedly leave the blood to migrate through the peripheral lymphoid organs, where they may encounter their specific foreign antigen
and become activated (top third panel).

Activation leads to clonal expansion and differentiation into effector T cells. Some of these are attracted to sites of infection, where they can kill infected cells or activate macrophages (top fourth panel); others are attracted into Bcell areas, where they help to activate an antibody response (not shown).

Question 5

Desenvolvimento de células T - W Fragen

Answer 5

• Objetivo?
  • Produzir células dotadas de um TCR funcional, e por isso capazes de reconhecer antigénios e montar uma resposta imunológica celular.
• Onde?
  • No timo. (Notar diferença em relação a células B: medula óssea)
• Como?
  • Através da diferenciação a partir de células pluripotentes, as quais
  sofrem alterações específicas de expressão génica que determinam a linhagem T.

• Cada uma dos muitos milhões de células T (e B) que circula no nosso corpo expressa um recetor de antigénio único: células que entram no timo não estão ainda comprometidas para a linhagem T e não expressam TCR. Células T que saem do timo são funcionais e expressam um TCR único.

Question 6

Celulas T na ausência de timo?

Answer 6

Ausência de timo –> Ausência de células T!!

• Timectomia ao nascimento (ratinhos)
• Síndrome de DiGeorge (humanos)- deleção 22q11.2; Tbx1 aplasia tímica

Mutação do gene Foxn1 (Forkhead box n1), fator de transcrição fundamental para a diferenciação e manutenção das células epiteliais do timo (TEC)

Question 7

scid/scid mouse vs nu/nu mouse

Answer 7

–scid/scid mouse:
Lymphocyte defect

deficiência em Prkdc
(envolvida na recombinação VDJ); Ausência de células T e B

–nu/nu mouse:
Thymus defect

Deficiência em Foxn1
(atímico); Ausência de células T

scid/scid com bone marrow & stem cells do nu/nu: grafted cells repopulate normal thymus

nu/nu com thymus graft in kidney do scid/scid: normal cells repopulate grafted thymus

=> Analyze spleen cells: T cell numbers after graft == para os 2 tipos de ratos

Question 8

Timo

Answer 8

The thymus is an organ surrounded by a fibrous capsule that encloses multiple lobes, each of which is separated into two major regions: the outer cortex (cortical epithelial cells, macrophages) and inner medulla (DCs, Medullary epithelial cells)

Question 9

Desenvolvimento de células T no timo

Answer 9

T-cell precursors from the bone marrow travel to the thymus via the bloodstream, undergo development to mature T cells, and are exported to the periphery, where they can undergo antigen-induced activation and differentiation into effector cells and memory cells. Each stage of development occurs in a specific microenvironment of the thymus and is characterized by specific intracellular events and distinctive cell-surface markers.

Thymocytes enter the thymus via blood vessels at the corticomedullary junction and then travel into the cortex, proliferating first in the region just below the capsule (the subcapsular cortex). As they mature they migrate from the cortex into the medulla and ultimately exit via vessels in the corticomedullary junction. This schematic summarizes the developmental progression in more detail. (VER!! thymic setting precursors –> DN1 –> DN2 (both DN1 and DN2 still multipotent and can give rise to myeloid and NK cells) –> commitment to Tcell lineage: DN3 –> TCR gene rearrangement –> DN3 (ou então gamadelta T cell??)–> DN4 –> alfabetaTCR DP -(Tregs, IEL, NKT??)-> CD4’+ SP ou CD8+ SP ou cell death

The most immature, CD4-CD8- (double negative, DN) thymocytes pass through
several stages (DN1-DN4), during which they commit to the T-cell lineage and begin to rearrange their T-cell receptor (TCR) gene loci. Those that successfully rearrange their TCRβ chain proliferate, initiate rearrangement of their TCRa chains, and become CD4+CD8+ (double positive, DP) thymocytes, which dominate the thymus. 

DP thymocytes undergo negative and positive selection in the thymic cortex.

Positively selected thymocytes continue to mature and migrate to the medulla, where they are subject to another round of negative selection to self antigens that include tissuespecific proteins. Mature T cells express either CD4 or CD8 (single positive, SP) and leave the thymus with the potential to initiate an immune response.

Although most thymocytes develop into conventional αβ TCR CD4 or CD8 T cells, some thymocytes develop into other cell lineages, including lymphoid dendritic cells, γδ TCR T cells, natural killer T (NKT) cells, regulatory T cells, and intraepithelial lymphocytes (IELs), each of which has a distinct function.

Question 10

Função e expressão de TCR durante o processo de diferenciação T no timo

Answer 10

DN1 (CD44+ CD25-) and DN2 (CD44+ CD25+) thymocytes do not express any T-cell receptor proteins on their surface.

The pre-TCR is assembled during the transition from the DN2 to the DN3 (CD44- CD25+) stage of development, when a successfully rearranged TCRbeta chain dimerizes with the nonvariant pre-Talfa chain. Like the mature alfabeta TCR dimer,
the pre-TCR is noncovalently associated with the CD3 complex. Successful assembly of this complex results in intracellular signals at the DN3 stage that induce a variety of processes, including the maturation to the DP stage and rearrangement of the TCRa chain.

Once a thymocyte has successfully rearranged a TCRalfa chain (in transition between the DN4 and DP stages), this chain dimerizes with the TCRbeta chain, replacing the pre-Talfa chain and generating a mature alfabeta TCR. The αβ TCR expressed by DP thymocytes also complexes with CD3 and can generate signals that lead to either positive or negative selection (differentiation or death, respectively), depending on the affinity of its interaction with the MHC/peptide complexes it encounters in
the thymic cortex and medulla. Although the αβ TCR/CD3 complex expressed by mature SP T cells is structurally the same as that expressed by DP thymocytes, the signals it generates are distinct. It responds to high-affinity engagement not by dying, but by initiating cell proliferation, activation, and
the expression of effector functions. Low-affinity signals generate survival signals. The basis for the differences in consequence of signals generated by DP and SP TCR complexes is still unknown.

Question 11

Recetor da célula pré-T (pre-TCR)

Answer 11

The pre-TCR is assembled during the DN stage of development when a successfully rearranjed TCRb chain dimerizes with the nonvariant pre-Talfa chain. Like the mature TCR dimer, the pre-TCR is non-covalently associated with the CD3 complex.

Expresso nos estadios DN3 e DN4

pre-TCR –> signals –> cell becomes permisse for TCR alfa-chain locus arrangement OR stimulates expression of CD4 and CD8 coreceptors OR stimulates proliferation OR Stops additional TCR beta-chain locus arrengements (allelic exclusion)

Question 12

Estrutura e atividade do TCR durante o desenvolvimento T

Answer 12

The mature ab TCR is expressed at the DP stage of development once a DP has successfully rearranged a TCRa chain that will dimerize with the TCRb, replacing the pre-TCR a chain. This mature TCR generates signals that lead to either positive or negative selection (differentiation or death, respectively), depending on the affinity of the interaction (intermediate or high with costimulatory signals, respectvely)

Although the ab TCR/CD3 complex expressed by mature SP T cells is structurally the same as that
expressed by DP thymocytes, the signals it generates are distinct. It responds to high affinity engagement not by
dying, but by initiating proliferation, activation, and expression of effector functions. Low-affinity signals induce survival- The basis for the differences in signals generated by DP and SP TCR complexes is still unknown

Question 13

em suma subsets cels T no timo

Answer 13

Em vertebrados são produzidos 2 subsets de células T no timo:

• TCR aβ- subset dominante em respostas imunológicas adquiridas

• TCR γδ- proteção das mucosas; primeiras células T a desenvolver-se durante o desenvolvimento fetal; diferenciação deste subset é
reduzido após nascimento

Question 14

colonização do timo durante o periodo embrionário

Answer 14

Durante o período embrionário o timo é primeiro colonizado por timócitos γδ e posteriormente por timócitos αβ que predominam no adulto

Time course of appearance of γδ thymocytes and αβ thymocytes during mouse fetal development. The graph shows the percentage of cells in the thymus that are double negative (CD4-CD8- ) and bear the γδ T-cell receptor (black line) or are double positive (CD4+ CD8+) and bear the αβ T-cell receptor (blue line). Fetal animals generate more γδ T cells than αβ T cells, but the proportion of γδ T cells generated drops off dramatically after birth. This early dominance of γδ TCR cells may have adaptive value: a large portion of these
cells express nondiverse TCR specificities for common pathogen proteins and can mount a quick defense before the more traditional adaptive immune system has fully developed.

Question 15

Seleção tímica

Answer 15

• Seleção positiva
Sobrevivência de timócitos cujos TCRs reconhecem moléculas de MHC do próprio
Garante restrição ao MHC

• Seleção negativa
Elimina timócitos cujos TCRs interagem com elevada afinidade com complexos MHC-péptido

• Seleção de um repertório de células T capazes de reconhecer Ag exógenos apresentados por moléculas de MHC do próprio (células
T funcionais)

• Eliminação da maioria das células autoreativas (ativadas por complexos MHC-péptido do próprio)

Question 16

Visão global do desenvolvimento de Linfócitos T no timo

Answer 16

Precursors of T cells travel from the bone marrow through the blood to the thymus. The progenitors of αβ T cells are double-negative (DN) T cells. In the thymic cortex, these cells begin to express TCRs and CD4 and CD8 coreceptors.
Selection processes eliminate self-reactive T cells in the cortex at the double-positive (DP) stage and also eliminate singlepositive (SP) medullary thymocytes. They promote survival of thymocytes whose TCRs bind self MHC molecules with low
affinity.

Functional and phenotypic differentiation into CD4+CD8− or CD8+CD4− SP T cells occurs in the medulla, and mature T cells (CD4+, CD8+, Treg) are released into the circulation.

Some double-positive cells differentiate into CD4+CD8− regulatory T cells (Treg). The development of γδ T cells is not shown.

Question 17

The affinity model of T-cell positive and negative selection.

Answer 17

O modelo de seleção de células T no timo baseado na afinidade de interação TCR: MHC-péptido

Random TCRa and β chain gene rearrangements generate a large pool of immature thymocytes expressing a varied repertoire of specificities.

The T-cell receptors on many of these cells fail to have sufficient binding strength to the self peptide:self MHC complexes on thymic epithelium and so receive no signals. These cells die by neglect.

Another fraction of immature thymocytes are positively selected because their T-cell receptors bind with sufficient strength to the self peptide:self MHC complexes on thymic epithelium to generate T-cell receptor dependent survival signals. From this cohort of positively selected thymocytes, negative selection removes those thymocytes whose receptors have excessively strong reactivity to self peptides complexed with self MHC molecules (resulting in clonal deletion), thereby establishing self-tolerance of the mature T-cell population.

A small subset of positively selected cells receiving signals slightly weaker than those inducing negative selection differentiate into regulatory T cells (Tregs), a process referred to as agonist selection.

Question 18

Effect of class I or II MHC deficiency on thymocyte populations

Answer 18

Cell Type: CD4- CD8-
Control mice: +
Class I deficient: +
Class II deficient: +

Cell Type: CD4+ CD8+
Control mice: +
Class I deficient: +
Class II deficient: +

Cell Type: CD4+
Control mice: +
Class I deficient: +
Class II deficient: -

Cell Type: CD8+
Control mice: +
Class I deficient: -
Class II deficient: +

Question 19

Seleção no Timo: Expressão ectópica de autoantigénios (?)

Answer 19

esquema? brain, intestine, liver, leukocytes, epidermis, …….

Autoimmune Regulator: AIRE
expressão em mTEC

Question 20

Papel da expressão tímica de AIRE

Answer 20

Model of the function of Aire in the thymus.

A) Aire appears to help mediate the transcription of many selfantigens in mTECs in the thymus.

B) Impact of Aire on T-cell selection. These self-antigens are then presented in the thymus to developing thymocytes in the medulla, and this results in the deletion of self-antigen specific thymocytes in this compartment.
In the absence of Aire, the selfantigens fail to be generated by these mTECs, and self-antigen specific T cells mature and escape the thymus and migrate into the peripery and promote autoimmune responses.

Question 21

O processo de desenvolvimento T é muito “custoso”:

Answer 21

• 98% de todos os timócitos não terminam o seu processo de maturação, morrendo por apoptose no timo
• No entanto, este processo garante a diferenciação de células T funcionais, com reduzido potencial auto-reativo

Question 22

Células T reguladoras:

Answer 22

• Diferenciam-se no timo (tTreg) mas também na periferia (pTreg)
• Expressam FOXP3 e CD25 (cadeia alfa do recetor da IL-2)
• Inibem a proliferação de outras células T in vitro e in vivo
• Crucias para a manutenção da tolerância periférica: Previnem o desenvolvimento de doenças autoimunes

Possíveis mecanismos de ação. These may all contribute to quelling immune responses in vivo.

(1) Cytokine deprivation: Tregs
express relatively high levels of high-affinity IL-2 receptors and can compete for the cytokines that activated T
cells need to survive and proliferate.

(2) Cytokine inhibition: Tregs secrete several cytokines, including IL-10 and TGF-β, which bind receptors on activated T cells and reduce signaling activity.
(3) Inhibition of antigen-presenting cells: Tregs can interact directly with MHC class II–expressing antigen presenting cells and inhibit their maturation, leaving them less able to activate T cells.
(4) Cytotoxicity: Tregs can also display cytotoxic function and kill cells by secreting perforin and granzyme.

Question 23

Indução da migração de timocitos do timo para a periferia

Answer 23

A migração de timócitos do timo para a periferia é
induzida por sinalização via S1PR1

• 3-4 days de maturação na medula após seleção

• Migração por gradiente de S1P (Sphingosine 1-
phosphate)

Recent thymic emigrants
(células T recentemente emigradas do timo)

CD4 and CD8 single-positive thymocytes that have successfully survived positive and negative selection are found in the medulla but
are not yet fully mature.

At the termination of the maturation process, which takes 3–4 days, the CD4 and CD8 single-positive thymocytes upregulate the sphingosine 1-phosphate (S1P) receptor, known as S1PR1.

S1PR1 is a G-protein coupled receptor that promotes chemotaxis of the cells toward the ligand S1P.

Due to the high levels of
S1P in the blood, single positive thymocytes are induced to leave the thymus by entering the blood, where they become part of the recirculating naïve T-cell population

Question 24

Fatores de transcrição envolvidos no comprometimento para a linhagem T

Answer 24

Notch1 –> Tcell factor (TCF)1
GATA-3

–> expressão de genes específicos da linhagem T (complexo CD3, Rag)

Bcl11b –> bloqueio de todas as linhagens não T
Ativação completa do programa T

Notch signalling induces the expression of 2 transcription factors, T-cell factor-1 (TCF1) and GATA3, each of which is required for T-cell development. Together, TCF1 and GATA3 initiate expression of several T-lineage-specific genes, such as those encoding
components of the CD3 complex, as well as Rag1, a gene required for T-cell receptor and B-cell receptor gene rearrangements.

However, TCF1 and GATA3 are not sufficient to induce the entire program of T-cell-specific gene expression. A third transcription factor, Bcl11b, is required to induce T-lineage commitment by restricting progenitor cells from adopting alternative fates; this final phase of T-cell commitment is a necessary prerequisite for activating the complete T-cell gene expression program

Question 25

Modelos propostos para determinação da linhagem CD4 ou CD8 no timo

Answer 25

• instrutivo
• estocástico
• sinalização cinética

Runx3: necessário para a diferenciação de CD8+ –> ativa expressão de CD8;
inibe expressão de CD4 e ThPOK

ThPOK: necessário para a diferenciação de CD4+ –> ativa expressão de CD4;
inibe expressão de CD8 e Runx3

(a) According to the instructive model, interaction of a coreceptor with the MHC molecule for which it is specific results in down-regulation of the other coreceptor.
(b) According to the stochastic model, down-regulation of CD4 or CD8 is a random process.

(c) According to the kinetic signalling model, the decision to commit to the CD4 or CD8 lineage is based on the continuity of the TCR signal that a thymocyte receives. Positive selection results in down regulation of CD8 on all
thymocytes. This will not alter the intensity of a TCR/CD4/MHC class II signal, and cells receiving this signal
will continue development to the CD4 SP lineage. However, down-regulation of CD8 diminishes (interrupts) a
TCR/CD8/MHC class I signal, an experience that sends a cell toward the CD8 lineage. IL-7 signals are required to “seal”

Question 26

Função tímica com a idade!

Answer 26

Declínio!!!

Tamanho do timo diminui com a idade: 20 anos ~80% do timo é tecido linfoide; 40 anos ~5% do timo é tecido linfoide

The size of the thymus decreases with increasing age. Until about 20 years of age, more than 80% of the thymus is composed of lymphoid tissue. With age, that proportion declines continuously until, from the age of about 40 onwards, only
about 5% of the thymus is morphologically lymphoid. Loss of lymphoid tissue — called thymic involution — seems to occur at the level of the thymic epithelium because, in mice, a young thymus can thrive when grafted into an old host, indicating that T-cell progenitors are active in aged bone marrow.

Histological examples of thymus sections. The aged thymus is massively infiltrated by non-thymic tissue, mostly fat, but still harbours islands of lymphoid tissue, including both cortical and medullary ‘units’.

Question 27

Função tímica com a idade! (cont)

Answer 27

Redução da atividade tímica com a idade Mas… Há manutenção de alguma atividade tímica, mesmo aos 80 anos!

In the aged thymus, the perivascular space (P) is increased in size and the thymopoietic cortex (C) and medulla (M) are significantly
constricted.

A clear reduction in double positive (DP) CD4+CD8+ T cells be seen in aged
individuals.

(b) sjTREC (single joint T-cell receptor excision circle)
measurement per microgram of circulating peripheral blood mononuclear cell (PBMC) DNA from healthy human donors ranging from <1 to ∼80 years
of age demonstrates a steady decline of naïve T cell output with age.

(d) Islands of cortical and medullary tissue in aged thymus (78-year-old
female) show signs of active thymopoiesis by CD1a and Ki67 immunostain.

Question 28

Rearranjo de um segmento Va com Ja no locus da

cadeia a do TCR induz deleção do locus da cadeia d

Answer 28

TCR Excision Circle harbouring signal joint
(sjTREC)

Este DNA epissomal resultante da deleção do locus d do TCR é mantido nas células T após migração do timo para a periferia. Os TRECs são estáveis e não
se dividem, mas são diluídos por divisão celular.

Deletion of the TCRδ locus is induced by rearrangement of a Va to Ja gene segment. The TCRδ locus is entirely contained within the chromosomal region containing the TCRa locus. When any V region in the Va /Vδ region rearranges to any one of the Ja segments, the intervening region, and the entire Vδ locus, is deleted. Thus, Va rearrangement prevents any continued expression of a Vδ gene and precludes lineage development down the γ:δ
pathway.

O locus que codifica a cadeia delta está localizado inteiramente dentro do locus da cadeia alfa

Question 29

Conceitos chave

Answer 29

• Progenitores não comprometidos para a linhagem T migram da medula óssea para o timo. Nestes, a sinalização Notch é necessária para o comprometimento para a linhagem T.

• As células T imaturas são designadas por timócitos, e a sua maturação progride ao longo de diferentes estadios de desenvolvimento genericamente definidos pela expressão dos co-recetores CD4 e CD8. A subpopulação mais imatura de timócitos não expressa CD4 nem CD8 (DN, double negative). O desenvolvimento de timócitos DN progride a través de
quatro estadios (DN1 a DN4) definidos pela expressão de CD44 e CD25. Durante esta progressão, estes proliferam e rearranjam o TCR. Os timócitos maturam em células T que expressam um TCRaβ ou γδ.
Timócitos que rearranjam as cadeias δ e γ diferenciam-se em linfócitos T γδ, ao passo que timócitos que rearranjam eficientemente a cadeia β do TCR ficam comprometidos com a linhagem aβ.

• A maioria dos timócitos diferenciam-se em linfócitos T aβ. No entanto, os linfócitos T γδ são as primeiras células a diferenciar-se durante o desenvolvimento fetal, colonizando a periferia por ondas. As células T γδ e T aβ cells são distintas funcionalmente. A maioria das células T γδ possuem um repertório limitado. Estas são muito importantes na defesa
contra microrganismos patogénicos nas mucosas e pele

Os linfócitos T aβ são muito mais diversos, possuem uma distribuição na periferia muito mais ampla e desempenham também funções muito diversas.

• Timócitos que rearranjam a cadeia β do TCR são submetidos a um processo designado por seleção β. A seleção β é iniciada pelo assembly da cadeia β do TCR com uma cadeia a invariante designada por pré-Ta, e com as proteínas do complexo CD3, formando o pré-TCR. Sinalização via pré-TCR resulta no comprometimento para a linhagem T aβ, promoção da proliferação destes timócitos, maturação para o estadio DP (double positive) e iniciação do rearranjo da cadeia a do TCR.

• Os timócitos DP são a subpopulação mais abundante no timo e são as primeiras células a expressar um TCR aβ completamente rearranjado.
Neste estadio no cortex, esta subpopulação é submetida ao processo de seleção positiva e negativa, responsáveis pela restrição ao MHC e tolerância ao próprio, respetivamente. Dependendo da afinidade do seu TCR para complexos MHC-péptido expressos por células tímicas estromais, 3 resultados distintos podem ocorrer. A maioria dos timócitos DP morrem por falhar reconhecer complexos MHC-péptido com afinidade suficiente (falham o processo de seleção positiva, sendo designado este processo “death by neglect”). Aproximadamente 2 a 5% dos timócitos DP morrem por reconhecerem complexos MHC-péptido com elevada afinidade (seleção negativa).

Finalmente, uma percentagem semelhante destes timócitos (2 a 5%) são selecionados positivamente, progredindo no processo de desenvolvimento, graças ao reconhecimento de complexos MHC-péptido com afinidade intermédia.

• Interações TCR /MHC-péptido com elevada afinidade resultam em seleção negativa, tipicamente por indução de apoptose destes timócitos autoreativos (deleção clonal). Células epiteliais tímicas medulares (mTECs) expressam o fator de transcrição AIRE, que é largamente responsável pela expressão de Ag restritos a tecidos periféricos. Outras
APCs tímicas, incluindo DCs e linfócitos B, são também capazes de mediar seleção negativa no cortex e na medula tímica. Assim, o processo de seleção negativa pode ocorrer tanto no cortex como na medula. O processo de seleção positiva ocorre no cortex. Timócitos selecionados positivamente aumentam a expressão do recetor de quimiocina CCR7, que lhes permite migrar do cortex para a medula tímica. Timócitos na transição do estadio de desenvolvimento DP para SP (Single positive), podem ser selecionados negativamente por Ag de tecidos periféricos,
apresentados por mTECs.

• Os fatores de transcrição ThPOK e Runx3 desempenham um papel fundamental no comprometimento para CD4 e CD8, respetivamente.
• Numa pequena percentagem de timócitos, interações TCR/MHC-péptido de elevada afinidade não promovem deleção, mas antes promovem o seu desenvolvimento em linhagens especializadas, incluindo em células NKT, IELs e células T reguladoras.
• Timócitos SP que sobreviveram o processo de seleção negativa iniciam um programa celular que leva ao aumento da sua sobrevivência e promove aumento da expressão de recetores que facilitam a sua migração do timo. A saída de timócitos SP do timo depende da expressão do recetor de S1P. Timócitos que acabam de sair do timo são designados por “recent thymic emigrants”.
• Os mecanismos de tolerância central não removem todas as células T autoreactivas do repertório de linfócitos T. Mecanismos de manutenção de tolerância ao próprio na periferia existem para colmatar esta limitação.