Dendritic Cell Maturation, T Cell Homing, Activation, and Checkpoint Inhibition

Course Administration and Deadlines

• Peer Review Assignments:

  • Following the prior group evaluation submission, students are assigned $2$ peer review videos to evaluate.

  • Submission Deadline: Friday at $11:59\text{ PM}$.

  • Troubleshooting Protocol: If assigned peer review videos are not visible in the portal, students must seek assistance from peers first (as student views differ from instructor views) or contact the instructor directly. No extensions or exceptions are granted for failing to complete peer reviews.

Overview of Lymphocyte Development and Immune Response Initiation

• Lymphocyte Origin and Maturation Sites:

  • B Cells: Early development and complete maturation occur within the bone marrow.

  • T Cells: Originating progenitor cells arise in the bone marrow and subsequently migrate to the thymus for T cell receptor gene rearrangement, selection, and maturation.

• Big Picture Dynamics of Peripheral Infection:

  • Pathogen Entry: Pathogens (e.g., bacteria) breach mechanical skin barriers at local injury sites (e.g., a foot wound).

  • Antigen Uptake: Resident immature dendritic cells in infected tissue phagocytose pathogens, degrade pathogen proteins, and load derived peptides onto Major Histocompatibility Complex ($\text{MHC}$) molecules.

  • Migration to Draining Lymph Node:

  • Immature dendritic cells undergo maturation and transition from an antigen-capturing phenotype to an antigen-presenting phenotype.

  • Dendritic cells cease phagocytosis, downregulate antigen uptake machinery, and upregulate surface expression of peptide-MHC complexes and co-stimulatory molecules.

  • Maturing dendritic cells enter lymphatic vessels and travel to the nearest lymph node, termed the draining lymph node.

  • High-density surface expression of $\text{MHC Class II}$ molecules (visualized as green fluorescence) marks activated dendritic cells migrating through tissue and lymph to present antigens to naive T cells.

Dendritic Cell Antigen Processing and Presentation Pathways

• Pathogen Recognition Mechanisms:

  • Dendritic cells express an array of Pattern Recognition Receptors ($\text{PRRs}$) capable of binding conserved Pathogen-Associated Molecular Patterns ($\text{PAMPs}$) found on bacterial and fungal pathogens.

  • Activation via $\text{PRRs}$ triggers phagocytosis, internal signaling, and initiates dendritic cell maturation and lymph node migration via afferent lymphatics.

• Toll-Like Receptor ($\text{TLR}$) Profile:

  • Humans express a total of $10$ functional Toll-Like Receptors ($\text{TLR1}$ through $\text{TLR10}$).

  • Dendritic cells express $9$ out of the $10$ human $\text{TLRs}$ (expressing all except $1$).

  • $\text{TLR}$ ligation activates dendritic cells, significantly enhancing their capabilities for antigen uptake, processing, and presentation.

• Pathways of Antigen Acquisition:

  • Receptor-Mediated Endocytosis: Cell-surface receptors bind specific extracellular pathogen components or intact pathogens. Binding induces endocytosis and lysosomal degradation, leading to peptide presentation on $\text{MHC Class II}$ molecules to $\text{CD4}^+$ T cells.

  • Macropinocytosis: Dendritic cells non-selectively engulf extracellular fluid and soluble antigens in large vesicles. Soluble antigens are processed and displayed on $\text{MHC Class II}$ molecules for presentation to $\text{CD4}^+$ T cells.

  • Direct Viral Infection and Endogenous Processing: Specific viruses bind surface entry receptors on dendritic cells and gain entry into the cytoplasm. Viral replication leads to synthesis of endogenous viral proteins within the dendritic cell cytosol. Cytosolic proteins are degraded by the proteasome and presented on $\text{MHC Class I}$ molecules to $\text{CD8}^+$ cytotoxic T cells.

Chemokine Signaling and Lymph Node Homing

• Chemokine Receptor $\text{CCR7}$ Expression:

  • Upon activation and maturation, dendritic cells upregulate the chemokine receptor $\text{CCR7}$.

  • Naive B cells, naive T cells, and activated dendritic cells all express $\text{CCR7}$.

• Chemokine Gradient ($\text{CCL21}$ and $\text{CCL19}$):

  • Stromal cells and High Endothelial Venules ($\text{HEVs}$) in the T cell zone of lymph nodes produce and secrete the chemokines $\text{CCL21}$ and $\text{CCL19}$.

  • $\text{CCR7}$ binding to $\text{CCL21}$ and $\text{CCL19}$ directs the migration of circulating naive T cells and B cells across $\text{HEVs}$ into the T cell region.

  • Expressed $\text{CCR7}$ allows maturing dendritic cells migrating in lymph to enter and position themselves within the T cell cortex of the lymph node.

Naive T Cell Homing and Extravasation Across High Endothelial Venules

• Entry Routes into Lymph Nodes:

  • Naive T cells enter lymph nodes via two primary routes: directly from arterial blood through High Endothelial Venules ($\text{HEVs}$), or from upstream lymph nodes via afferent lymphatic vessels.

• Endothelial and T Cell Adhesion Molecule Pairs:

  • High Endothelial Venules ($\text{HEVs}$) express specific vascular adhesion molecules on their luminal surface: $\text{GlyCAM-1}$, $\text{CD34}$, and $\text{ICAM-1}$.

  • Naive T cells express surface adhesion molecules including $\text{L-selectin}$ and $\text{LFA-1}$ (Lymphocyte Function-associated Antigen 1).

• Step-by-Step Extravasation (Diapedesis) Cascade:

  • Step 1: Rolling: $\text{L-selectin}$ on the naive T cell binds reversibly to carbohydrate moieties on $\text{GlyCAM-1}$ and $\text{CD34}$ on HEV endothelial cells, causing the fast-moving T cell to slow down and roll along the vessel wall (analogous to neutrophil rolling in innate inflammation).

  • Step 2: Chemokine Activation: Chemokines ($\text{CCL21}$ and $\text{CCL19}$) displayed on the endothelial lumen bind $\text{CCR7}$ on the rolling T cell.

  • Step 3: Conformational Change and High-Affinity Binding: Intracellular signaling downstream of $\text{CCR7}$ induces a conformational change in T cell $\text{LFA-1}$, converting it from a low-affinity state to a high-affinity state. High-affinity $\text{LFA-1}$ binds tightly to $\text{ICAM-1}$ on the HEV endothelium, completely arresting T cell movement.

  • Step 4: Diapedesis / Extravasation: The arrested T cell extravasates by squeezing between adjacent endothelial cells to migrate out of the blood vessel lumen and into the T cell cortex of the lymph node.

T Cell Inspection, Exit, and Retention Mechanisms

• Continuous Antigen Inspection:

  • Naive T cells migrating through the T cell zone dynamically interact with stationary dendritic cells, scanning displayed peptide-MHC complexes.

  • Adhesion interactions (such as $\text{LFA-1}$ binding to $\text{ICAM-1}$ or $\text{ICAM-2}$ on dendritic cells) facilitate transient cell-to-cell contact during inspection.

• Exit Signal ($\text{S1P}$ Gradient):

  • Sphingosine-1-Phosphate ($\text{S1P}$): A lipid mediator with chemokine-like chemoattractant properties present at higher concentrations in circulating blood and lymph than within lymph node tissue.

  • Naive T cells expressing the $\text{S1P}$ receptor ($\text{S1P}_1$) follow the gradient of $\text{S1P}$ to leave the lymph node via efferent lymphatic vessels if no specific antigen is recognized.

  • Unstimulated T cells re-enter the bloodstream to continue circulating through secondary lymphoid organs.

• Retention Signal ($\text{CD69}$ Expression):

  • Antigen Recognition: Binding of the T Cell Receptor ($\text{TCR}$) to a specific peptide-MHC complex on a dendritic cell triggers intracellular activation signaling.

  • Induction of $\text{CD69}$: T cell activation induces expression of the transmembrane protein $\text{CD69}$.

  • $\text{S1P}$ Receptor Downregulation: $\text{CD69}$ physically interacts with and internalizes/downregulates the $\text{S1P}$ receptor from the cell surface.

  • Outcome: Loss of surface $\text{S1P}$ receptor prevents the T cell from sensing the egress-promoting $\text{S1P}$ gradient, retaining the activated T cell within the lymph node to undergo clonal expansion and proliferation over several days.

  • Post-Proliferation Release: Following clonal expansion and differentiation into effector T cells, $\text{CD69}$ expression subsides, restoring surface $\text{S1P}$ receptor expression and enabling effector T cells to exit via efferent lymphatics.

T Cell Activation, Co-Stimulation, and Down-Regulation

• T Cell Receptor Complex Structure:

  • Signal-transducing subunits associated with the antigen-binding $\text{TCR}$ subunits include the $\text{CD3}$ complex (consisting of $\gamma$, $\delta$, and $\epsilon$ chains) and the $\zeta$ (zeta) chain homodimer.

• Two-Signal Requirement for T Cell Activation:

  • Signal 1: $\text{TCR}$ binding to specific peptide-MHC complex along with coreceptor ($\text{CD4}$ or $\text{CD8}$) engagement.

  • Signal 2 (Co-stimulation): Engagement of the co-stimulatory receptor $\text{CD28}$ on the T cell by $\text{B7}$ co-stimulatory molecules ($\text{B7.1/CD80}$ or $\text{B7.2/CD86}$) expressed exclusively on activated antigen-presenting cells (dendritic cells).

• Negative Regulation via $\text{CTLA-4}$:

  • To prevent over-activation and maintain immune homeostasis, T cells express $\text{CTLA-4}$ (Cytotoxic T-Lymphocyte-Associated Protein 4) following activation.

  • Mechanism: $\text{CTLA-4}$ binds to $\text{B7}$ molecules with significantly higher affinity and avidity than $\text{CD28}$.

  • Function: Acting as the immunological "brakes," $\text{CTLA-4}$ engagement delivers inhibitory signals that suppress T cell proliferation and cytokine production.

Immune Checkpoint Inhibitors in Cancer Immunotherapy

• Accelerator vs. Brakes Paradigm:

  • $\text{CD28}$: Immune "accelerator" (delivers positive co-stimulatory activation signals).

  • $\text{CTLA-4}$: Immune "brakes" (delivers negative co-inhibitory signals).

• Tumor Immune Evasion:

  • Malignant tumors exploit inhibitory immune checkpoint pathways (such as $\text{CTLA-4}$ binding to $\text{B7}$) to induce T cell exhaustion or inactivity, evading destruction by tumor-antigen-specific T cells.

• Therapeutic Checkpoint Inhibition:

  • Anti-$\text{CTLA-4}$ monoclonal antibodies are administered as therapeutic agents.

  • These antibodies specifically bind $\text{CTLA-4}$, sterically blocking its interaction with $\text{B7}$ on antigen-presenting cells.

  • By preventing the "brakes" from engaging, the anti-$\text{CTLA-4}$ antibody maintains active $\text{CD28}$-$\text{B7}$ co-stimulation, sustaining T cell activation, proliferation, and effector functions to target and eliminate cancer cells.