T cell memory Friday, February 10, 17

Transcription

1 T cell memory

2 expansion contraction memory recall EFFECTORS 10 6 pathogen load secondary challenge MEMORY 10 2 NAIVE >1 year days post-infection In mouse, ~100 naïve, CD8s become ~3x10 6 effectors in 7 days, then contract to ~10 5 by 14 days. Maybe 10x increase in TOTAL CD8 numbers! CD4 T cells response with ~same kinetics but ~10x lower numbers

3 Massive T cell response to acute viral or bacterial infection Clonal expansion of ag-specific T cells expansion contraction memory recall EFFECTORS 10 6 pathogen load secondary challenge MEMORY 10 2 NAIVE >1 year days post-infection In mouse, ~100 naïve, CD8s become ~3x10 6 effectors in 7 days, then contract to ~10 5 by 14 days. Maybe 10x increase in TOTAL CD8 numbers! CD4 T cells response with ~same kinetics but ~10x lower numbers N.B. EBV, infectious mononucleosis

4 Human CD8 T cell response to the yellow fever vaccine Tetramer HLA-A2+ peptide Blood

5 Dryvax (live Vaccinia) in Humans: antibody, CD4, CD8 CD8 antibody CD4

6 Human CD8 memory to smallpox vaccination CD8 memory Half-life = 8-15 yrs

7 Human CD4 memory to smallpox vaccination assessed by intracellular cytokine staining PBL stim with virus and ICCS SMALLPOX 1972 end vaccination 1978 eradicated CD4 memory Half-life 8-12 years Filled= >2 fold bg Open= <2 fold bg Slifka (2003) Nat Med 9, 1131

8 Serum antibody levels to smallpox vaccination Measured by ELISA and correlated with neutralization Not much boost with 2nd, 3rd vacc Correlation with neutralization

9 CD8 Effector cells are HETEROGENEOUS At the peak of response, a fraction of expanded CD8s express IL-7Rα (CD127) at high levels and these are enriched in memory precursors (MPEC). At the peak of response, most effectors are KLRG1 hi and these are destined to die by apoptosis, short-lived effectors (SLEC). How does effector cell heterogeneity come about?

10 Single OT-1 CD8 transfer and Listeria-Ova immunization ie naïve T cells are NOT pre-committed to dead-end Effector(SLEC) vs Memory(MTEC) Listeria-Ova OT-I cells are CD45.1 transferred into CD45.2 recip D.H. Busch, Immunity, vol 27, 2007

11 Single naïve OT-I CD8 transferred, Listeria-Ova infection makes heterogeneous effectors and makes memory cells % of spleen is about 10 4 cells Heterogeneous progeny. They also make memory ie at least 14 div

12 Do Memory cells arise from Effectors? Are memory precursors separate from effector precursors? Schoenberger, Science 323, 466 (2009)

13 Do Memory CD8s derive from cells that were once Effector CD8s? YES Sorted IFNγ producers make memory (Zink(CD8), Weaver(CD4)) CD8s which turn on Granzyme make memory (Fearon (2009) Science 323; ) gzmb promoter driving cre-er..(tamoxifen-inducible) cross to reporter mouse, ROSA-fl(stop)fl-YFP

14 Mice primed with flu and given tamoxifen on days 1-8. Assay response day 10 and 49 then boost with a 2nd flu and assay 7 days later. A fraction of effectors turn on cre (thus YFP+) They survive to memory They re-respond to 2nd hit ie they made memory cells.

15 Numbers of flu-specific (tetramer+) CD8+ T cells expressing granzyme reporter, same % become YFP+ in 2ndry as in 1ry Squares + = total CD8 ag-specific X = YFP+ CD8 ag-specific

16 Numbers of flu-specific (tetramer+) CD8+ T cells expressing granzyme reporter, same % become YFP+ in 2ndry as in 1ry Squares + = total CD8 ag-specific X = YFP+ CD8 ag-specific

17 Heterogeneity of CD8 Effectors and Memory Cells IL-7Rlo, KLRG1 hi, IL-2- SLEC N CD62L + CD44 - IL-7Rhi(CD127) KLRG1 lo MPEC IL-7Rhi,KLRG1 lo, IL-2+ CC62L lo,ccr7 lo,cd27 lo, kill, less turnover EM CM CD62L lo, CCR7 hi, IL-2+,higher turnover Longer-lived. Effectors Memory

18 Are SLECs really short-lived effectors? (Olson, Immunity, 2013)

19 Most protective population...

20 90-99% effectors die, while 1-10% live to memory. ie 90-99% will die by apoptosis, while 1-10% will survive contraction and become long-lived memory cells. What determines who dies?

21 How is the extent of contraction regulated? by competition between effector cells after the peak of the primary response versus contraction is programmed or cell-intrinsic not regulated by competition

22 bim-/- CD8s do not contract Bim a BH3-only member of Bcl2 family, required for contraction (Strasser) wt OT-I bim -/- OT-I Day 7 CD CD Spleen CD45.2 CD45.2 Day 14 CD CD CD45.2 CD45.2 Day 7 to day14. acute contraction: wt numbers decrease 10-fold bim-/- numbers decrease less than 2-fold

23 Is contraction regulated by competition for resources? Naïve T cells Effector T cells Memory T cell pool WT contraction Use bim-/- as non-dying competitors versus WT contraction?? bim -/- Competitors Martin Prlic

24 DAY 7 wt and bim -/- OT-I T cells expand equally Endog CD8s wt alone bim -/- alone 10 4 Mix 1:1 CD CD OT wt OT-1 MIX = 5,000 WT + 5,000 bim-/- Thy CD45.2 ko OT-1

25 Memory Contraction of wt cells is not influenced by bim -/- cells wt alone bim -/- alone Mix 1: CD CD Thy wt OT bim-/- OT-1 CD % of 5.2% = about 0.3% ie same decrease as wt alone

26 Factors influencing the differentiation of T cells after activation Asymmetric division? TCR affinity

27 Transcription factor Blimp-1 is involved in SLEC versus MPEC decision during the CD8 response

28 Mutual antagonism of Bcl-6 and Blimp-1, transcriptional repressors that regulate terminal effector versus memory differentiation in B cells and CD4s and CD8s 2 opposing players in differentiation Blimp-1 promotes Effector differentiation

29 CD8 T cell differentiation regulation by Blimp-1 and BCl-6 IL-2, IL-12

30 CD4 T cell differentiation

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32 What determines upreg of Bcl6(Tfh) versus Blimp-1(Th1)?? Level of TCR-ag interaction? DC vs B cells as APC? Amount of IL-2 / CD25 signals (promote Blimp-1)?

33 CD4 differentiation into Th1, Tfh or GC Tfh varies by clone (ie TCR repertoire) The whole mouse CD4 response Single ag-specific naïve CD4 T cells transferred, immunized with Listeria. 6 days later, ag-specific cells phenotyped for effector cell differentiation. Propose that TCR-MHC/pep dwell time, related to TCR affinity and ag dose, regulates CD25/Il-2 signals Jenkins (2013) Cell 153, 785 Single CD4 cells

34 CD4 Differentiation IL-2/IL-12à Blimp-1+ Tbet(Th1), vs Bcl6(Tfh) Based on (2011) Pepper et al Immunity 35, 553 (attenuated Listeria)

35 Models of CD4 effector and central memory Th1, Th2 or Th17 effectors can become EFFECTOR Mem, Treg, Tfh die/change Tfh or Tcm interact with B cells And become CENTRAL memory

36 Lineage committed Tfh and Th1 Memory cells following acute LCMV Infection Hale et al (2013) Immunity 38, 805

37 A third type of memory T cell Tcm.circulate like naïve T cells, LN homing, CCR7, great potential for expansion. Tem.circulate blood to non-lymphoid tissues to LN. Faster effector function, less expansion. Trm.non-recirculating, stay in tissues eg skin, ganglia, mucosa, brain

38 Tissue-Resident Memory Scheme showing Trm in dorsal root Sensory Ganglia which are latent for herpes simplex 1 (HSV-1) Transplant under kidney capsule Shave, abrade. HSV-1 flank skin day 0 HSV infects epithelia and sensory nerve Endings, then to ganglia Sensory DRG innervating site > day 20 Sensory neurons are latently infected HSV reactivate due to stress Assay + 5 days Carbone (2008) Science 319; Cold sores. Herpes zoster(shingles)

39 Local re-expansion of resident memory CD8+ T cells in DRG Put 2 DRG grafts under same kidney, one from mouse which has marked CD8s, one from a mouse that does not. Expansion occurs in the DRG, not in LN or spleen Science 319, (2008)

40 Tissue Resident T cell Memory Evidence for tissue-resident memory (Trm) CD8s in skin, DRG, IEL and LP in gut, brain. CD103+, CD69+, S1PR1lo, KLF2lo Non-Recirculating, separate from Tcm and Tem

41 Mechanisms promoting Trm residence in tissues. CD69 inhibits S1PR1; KLF2 regulates S1PR1 CD103 binds E-cadherin Immunity (2014) 41, 886

42 Issues studying Trm: Difficult to discern cells in vasculature (perfusion doesn t work) Difficult to study in humans Flow cytometry underestimates the true size of the population 42

43 Flow cytometry underestimates Trm 43

44 CD8 Trm in Mouse epidermis Female reproductive tract

45 Trm are the dominant memory population (Thome, Cell, 2014)

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47 Prospect for HIV vaccine? Antibody sites are variable T cell response too slow? Need response at site of entry? A vaccine for SIV generates EFFECTOR CD4 and CD8 T cell memory Use of a Rhesus CMV vaccine vector.

48 Cytomegalovirus (CMV) targets multiple class I pathways and becomes latent

49 Recombinant Rhesus-CMV vectors for SIV vaccine Picker, L et al Nat Med (2009) 15, 293 and Nature (2011) 473, 523

50 Recombinant Rhesus-CMV vectors for SIV vaccine Vaccine subcutaneous, SIV challenge multiple low dose rectal 13/24 protected

51 T em phenotype of CMV-SIV vector vaccinated Rhesus monkeys T em provide faster response to mucosal infection than T cm

52 Different types of memory cells respond differently...

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