Lecture 9 - mechanisms of T lymphocyte tolerance

What is immune tolerance?

The active inability of the immune system to mount a destructive response to specific antigens, including self-antigens, food, commensals, and environmental allergens.

Which antigen-presenting cells are key to maintaining immune tolerance?Which antigen-presenting cells play a crucial role in maintaining immune tolerance?

Dendritic cells (DCs) — immature DCs promote tolerance by inducing anergy or Treg differentiation via TGF-β.

What determines whether DCs promote tolerance or immunity?What factors determine if DCs promote tolerance or immunity?

Their maturation state:

• Immature DCs → tolerance (Treg induction, anergy)

• Mature DCs → immunity (co-stimulation, effector activation)

What are the three signals dendritic cells deliver to T cells?

1. Signal 1: Antigen via MHC II

2. Signal 2: Co-stimulation (CD80/86)

3. Signal 3: Cytokines (e.g., IL-12 or TGF-β)

Which cytokines promote peripheral Treg (pTreg) induction?

TGF-β, IL-2, retinoic acid (RA), and short-chain fatty acids (SCFAs) in mucosal tissues.

What is FoxP3 and why is it important?

A transcription factor essential for Treg development and function.
Mutations → autoimmunity (e.g., IPEX syndrome in humans, Scurfy in mice).

What distinguishes tTregs from pTregs/iTregs?

• tTregs: Thymus-derived, recognize self-antigens

• pTregs/iTregs: Peripherally induced from naïve CD4⁺ T cells in response to non-self antigens in tolerogenic environments

Do Tregs produce IL-2?

No, but they are not anergic — they proliferate in response to exogenous IL-2.

Which chemokine receptors guide Tregs to specific tissues?

• CXCR5: Germinal centers

• CCR9/CX3CR1: Gut

• CCR10: Tumor hypoxic regions

What direct mechanisms do Tregs use to suppress immunity?

• Secretion of IL-10, TGF-β, IL-35

• IL-2 consumption via CD25

• Granzyme-mediated cytolysis

• Galectin-1 → cell cycle arrest

What indirect mechanisms do Tregs use to suppress APCs?

• CTLA-4: Binds CD80/86, blocks co-stimulation

• LAG-3: Inhibits MHC II⁺ DCs

• CD39: Breaks down ATP (pro-inflammatory signal)

• Neuropilin-1: Stabilizes DC–Treg interactions

How do Tregs suppress antigen-non-specific cells?

Through bystander suppression — TCR-activated Tregs suppress nearby effector cells regardless of TCR specificity.

How do specialized Treg subsets suppress different Th responses?

They co-express Th-lineage-specific transcription factors (e.g., T-bet, GATA3, STAT3) and migrate to sites using matching chemokine receptors.

What causes Treg-to-Th17 conversion in autoimmune arthritis?

• IL-6 from synovial fibroblasts

• Converts CD25^lowFoxP3⁺ Tregs into IL-17–producing Th17 cells

• Enhances RANKL → bone resorption and joint damage

Which cytokine drives Treg instability and plasticity?

IL-6 — promotes loss of FoxP3 and Th17 reprogramming

What transcription factors help stabilize Tregs?

FOXP3, HELIOS, BACH2, STAT5-CA — enhance suppressive stability and prevent reprogramming

What are the four key criteria for effective Treg-based therapies?

1. Suppression (IL-10, TGF-β, APC modulation)

2. Specificity (TCR/CAR engineering)

3. Stability (FOXP3 maintenance)

4. Survival (IL-2, TCR signals, metabolic support)

What is IPEX syndrome and its cause?

A severe autoimmune disease caused by mutations in FOXP3, leading to absence of functional Tregs.

What are Treg-based therapy targets in clinical immunology?

• GVHD prevention (post-HSCT)

• Solid organ transplantation

• Autoimmune diseases (e.g., T1D, Crohn’s disease)

What evidence supports the tolerogenic role of immature DCs in vivo?

Hawiger et al., 2001: Antigen targeting to immature DCs (via DEC-205) in mice induces tolerance, not immunity.

What happens when dendritic cells are depleted in mice?

Ohnmacht et al., 2009: Constitutive DC depletion causes fatal autoimmunity, confirming their central role in immune homeostasis.

How does DC maturity affect antigen presentation outcome in mice?

Probst et al., 2003:

• Immature DCs expressing viral antigen → tolerance

• Mature DCs → immunity

What human condition is associated with DC deficiency?

Bigley et al., 2011: Human DC deficiency leads to reduced FoxP3⁺ Treg numbers, showing DCs are essential for peripheral Treg maintenance.

Which apoptotic pathway is involved in deletion of autoreactive T cells by DCs?

Fas–FasL interaction from CD95⁺ “killer DCs” induces apoptosis of autoreactive T cells in peripheral tolerance.

What experimental findings show instability of CD25^low Tregs?

Komatsu et al.:

• CD25^low Tregs are prone to FoxP3 loss

• More likely to convert into IL-17–producing cells (ex-Tregs)

How does IL-6 promote autoimmunity through Treg instability?

IL-6 induces Treg-to-Th17 conversion → Th17 cells secrete IL-17 and RANKL → osteoclastogenesis and bone destruction in autoimmune arthritis.

What trial outcome supports combining tacrolimus with Treg grafts in HSCT?

• Treg graft alone: ↑ acute and chronic GVHD

• Treg + tacrolimus: ↓ GVHD, better Treg/CD4⁺ ratio, improved safety

  Tacrolimus enhances Treg-based transplant efficacy.

What are the major targets of Treg therapy in autoimmunity and transplantation?

• GVHD prevention post-HSCT

• Organ transplant tolerance

• Type 1 Diabetes, Crohn’s, RA, etc.

What tools enhance Treg specificity for clinical use?

• TCR engineering

• CAR-Treg development

• SynNotch receptors

• Lineage-specific TFs (e.g., T-bet, STAT3)

Which molecules promote FOXP3 stability in Tregs?

• HELIOS

• BACH2

• STAT5-CA
  Block degradation by suppressing CHIP, DBC1, PKCθ.

What molecular pathways support Treg survival?

• IL-2 (external)

• TCR and co-stimulation

• Metabolic signalling via PI3K–AKT and JNK1

What enzyme degrades extracellular ATP to suppress DC activation?

CD39, expressed on Tregs, hydrolyses ATP → reduces inflammation and DC activation.

How does Neuropilin-1 (Nrp-1) contribute to tolerance?

Stabilizes prolonged Treg–DC contact, blocking access of effector T cells to APCs and reducing antigen presentation.

What are examples of diseases caused by failure in tolerance mechanisms?

• IPEX (FOXP3 mutation)

• Autoimmune arthritis (Treg → Th17)

• IBD (gut Treg dysfunction)

• GVHD (lack of Treg-mediated suppression)

What are Treg "living drug" properties needed for therapy?

1. Suppressive capacity

2. Antigen specificity

3. Phenotypic stability (FoxP3⁺)

4. Survival and persistence

What type of antigens require mucosal tolerance to avoid chronic inflammation?

• Food antigens

• Environmental allergens

• Commensal microbiota

Which immune cells are key to tolerance toward commensals in mucosal tissues?

• Tregs (especially pTregs/iTregs)

• Immature dendritic cells

• Supported by cytokines like TGF-β and IL-10

What local metabolites promote Treg development in the gut?

• Short-chain fatty acids (SCFAs) from microbiota

• Retinoic acid (RA)

• Both enhance FoxP3 expression and mucosal Treg induction

What tissue plays a key role in balancing immunity and tolerance to gut microbiota?

The lamina propria — rich in immune cells and tolerogenic signals, balancing defense and tolerance

What is the role of CD39⁺ Tregs in mucosal tissues?

They degrade extracellular ATP, a danger signal, limiting DC activation and inflammation at mucosal surfaces.

How does the microbiota influence systemic immune tolerance?

Through metabolites (e.g., SCFAs), microbial antigen exposure, and crosstalk with DCs and epithelial cells → promotes peripheral Treg induction

Which chemokine receptors are involved in Treg homing to mucosal tissues?

• CCR9 (small intestine)

• CCR6, CX3CR1 (large intestine, lamina propria)

• Help Tregs localize to commensal-rich environments

What transcription factor promotes gut-homing and mucosal Treg specialization?

GATA-3 — important in intestinal tissue-resident Tregs and mucosal homeostasis

What feature distinguishes iTregs from tTregs in response to microbiota?

iTregs are induced peripherally from naïve T cells in response to microbial antigens, particularly in a TGF-β/RA-rich environment.

Which mucosal immune disease is associated with Treg dysfunction?

Inflammatory Bowel Disease (IBD) — loss of tolerance to commensals due to defective Treg function or balance with Th17 cells

How can mucosal Treg function be therapeutically enhanced?

• Probiotics or SCFA supplementation

• TGF-β mimetics

• RA analogs

• Microbiota-based therapies

Why is tissue-specific Treg programming important in immunotherapy?

It ensures local suppression at the site of inflammation or autoimmunity, minimizing systemic immunosuppression and enhancing therapeutic precision.

How does FOXP3+ Treg expression relate to gut homeostasis?

Maintains tolerance to microbiota, prevents overactive inflammation, and supports intestinal epithelial integrity

What are potential risks of mucosal Treg failure?

• Chronic inflammation (e.g., colitis, IBD)

• Loss of oral tolerance → food allergies

• Autoimmunity

How might engineered CAR-Tregs be used in mucosal disease?

CAR-Tregs can be designed to recognize microbial antigens or inflammatory markers in tissues like the gut → deliver targeted immunosuppression in IBD.