AMI - Lecture 4 - T cell priming in relation to Covid

explain the 3 signals by which the DC activates the naive T cell

• ag presentation → ag-MHC/TCR

• co-stimulation → upregulation by PRR, CD80/86 molecules are upregulated. these bind to CD28. essential for proliferation and survival

• cytokines → these determine the effector cell subset

What are the different T cell subsets and which cytokines are involved

Explain T cell exhaustion

• happens when there is chronic exposure to ag (viral infection, cancer)

• exhausted T cell express high levels of PD1 (programmed cell death protein 1)

  • PD1 → inhibitory receptor that dampen immune response and T cell activity

• T cell exhaustion results in T cells that;

  • can’t proliferate well

  • can’t produce cytokines

  • can’t kill as much

  • not responsive to cytokines

• Solution → immune checkpoint therapy. they block the checkpoints and inhibitory signals

Characteristics of naïve, effector and memory T cells

• Naive cells recirculate between lymphoid
  organs and have NO effector functions.
  Numbers too low to detect.

• Effector cells migrate to tissues
  and have strong/fast effector functions (already differentiated).
  Numbers high, can be detected ex vivo

• Memory cells migrate to lymphoid organs or
  tissues and have effector functions
  Numbers low, can be detected with sensitive assays

Difference central and effector memory t cells

Tissue resident memory cell

stay in the tissue and provide local protection.

• First responder in tissue > local memory responses.

• Attraction of other immune cells

• Up- and downregulate surface molecule receptors (EXPRESSION OF DIFFERENT RECEPTORS by which they stay in the tissue)

• Down: what enters the blood circulation/avoid exit cues.

• The presence of Trm has been shown in mice and humans via multiple methods

  • sew 2 mice together, looked at where the cells went to through blood stream, stayed at the same place.

Explain MHC polymorphism + peptide binding groove

• the peptide binding groove is polymorphic for MHC 1+2

  • 6 different for MHC1

  • 6 different for MHC2 → peptide binding groove is bigger here

• polymorphism important → recognize wide range of pathogens, defense against evolving pathogens

• MHC molecules has revealed peptide motifs for each MHC molecule. The peptide motifs enables prediction of MHC binding peptides from proteins

• Every HLA binds multiple peptides.

• Different MHC alleles bind different peptide sets

Techniques to measure ag specific T cells; CFSE/CTV

• label T cells with dyes that don’t kill the cell > track division with flow cytometry = count cell divisions.

• Can combine with markers

Techniques to measure ag specific T cells; elispot assay

• sensitive kind of ELISA. intracellular cytokine staining),

• ELIspot: coat the well with an antibody against the cytokine. → Then you put the cells on top of it with the antigen. → Only the cells that respond to the antigen will start making Interferon gamma. → The t cells will be washed away and then the ELISA will be done and spots will show the responding cells.

Techniques to measure ag specific T cells; intracellular cytokine assay

• stimulate T cells with antigen → the T cells will start to produce cytokines and then a drug is given so the secretion of the cytokines is blocked. → With a detergent the cell can be permeabilized → Then the antibodies inside the cell will be stained. This determines the number of cytokine-producing cells

• analyzed by flow cytometry

Activation induced marker assay (AIM)

• stain for surface markers!!

• the cells that are activated express cytokines and surface markers. Cells are incubated with antigen, then they start to express surface markers. These can be stained.

MHC/ag tetramer staining

• quantification of antigen specific T cell responses.

• Use the MHC to stain. However, the affinity of TCR and MHC is not as high as antibodies. So tetramer is used;→ The 4 MHC molecules can bind, 4 because the affinity is higher than 1.

  • Determines the number of antigen, specific T cell

  • Additional markers by flow cytometry

  • Quantification directly ex vivo

  • No culture necessary

  • Peptide needs to be identified

  • Works best for MHC-I

Why is covid more serious than normal endemic virus

• have spike proteins, are critical to enter host cells

• have nucleocapsids (surround + protects viral RNA)

• infect respiratory cells, binds ace2 receptors

Explain the persistence of covid in B ant T cells

• after 6-8 months following cells still persistent;

  • memory B and plasma cells → low levels if ab response = secretion

  • memory B cells + ab are used to target the spike protein + nucleocapsid proteins

  • CD4/8 T cells are also still in the body after 6-8 months

  • CD4/8 cells are specific to multiple proteins on virus → spike protein + non structural proteins

Explain the 2 different covid vaccines

• cross reactivity → people who have never been infected with Covid still recognize parts of protein. Regions of the virus share homology between common cold and Covid. → may explain why some people get less sick.

• RNA (moderna) → very expensive, stored at -18

  • israel developed this, death rates dropped after 50% was vaccinated

• viral vector vaccines (astrazeneca) → modified, non replicating virus, easier to produce and distribute.