1. Evaluation of Human Thymic Function during Health and HIV-1 Infection
Ping Ye and Denise Kirschner
DIMACS Workshop
9/23/2001

2. Outline Thymic function in healthy people
Thymopoiesis
Markers to represent recent thymic emigrants (RTE)
Thymopoiesis model
TREC model
Thymic function during pediatric HIV-1 infection
HIV-1 infection
Clinical and experimental studies
Thymic infection model
Evaluation of TREC

3. The Thymus Janeway CA et al. Immunobiology 5th ed. 2001
Haynes BF. Clin Immunol. 92(1):3-5,1999

4. Thymopoiesis
Berkowitz RD et al. J Immunol. 161(7): , 1998

5. Recent Thymic Emigrants (RTE)
Lack of phenotypic markers for human RTE
Human RTE generally refer to T cells that have undergone only a few cellular divisions after leaving the thymus
T cell receptor excision circles (TREC) have recently been used as a measure of the number of RTE

6. T Cell Receptor Excision Circles (TREC)
Pieces of DNA fragments that are generated during TCR gene rearrangement in the thymus and then exported from the thymus to periphery within T cells episomally
Haynes BF et al. Annu Rev Immunol. 18: , 2000
Janeway CA et al. Immunobiology 5th ed. 2001

7. TREC as A Measure of RTE

8. Parameters Estimating RTE
TREC concentration
Is affected by both RTE and peripheral T cell proliferation and death
Naïve T cells (CD45RA+, CD62L, CD95+)
May have long lifespan
May proliferate in an antigen-independent manner
May rapidly convert to memory cells
May be converted from memory cells
Thymic volume
Assuming thymic volume is proportional to RTE levels

9. Two Questions to Address
1st Can TREC concentration be properly used as measure of RTE?
2nd Whether thymic infection with different
HIV-1 strains contribute to differences in disease progression?

10. Thymopoiesis Model Cell Source THYMUS TN ITTP DP SP4 SP8 BLOOD/
TES(t) TN
ITTP DP
SP4
SP8
BLOOD/
LYMPHOID TISSUES
CD4+ RTE
CD8+ RTE

11. Simulation Results for Thymopoiesis Model
Thymocytes
RTE/day

12. Model Simulation Values Compared with Experimental Data

13. TREC Model THYMUS SP4 SP8 CD4+ RTE CD8+ RTE CD4+ T cell CD8+ T cell
BLOOD/
LYMPHOID TISSUES

14. Simulation Results for TREC Model
Douek DC et al. Nature. 369(6712):690-5, 1998.
Zhang L et al. J. Exp. Med. 187(11): , 1998

15. Four Events Affecting TREC Concentration
THYMUS
SP4
SP8 1
CD4+ RTE
CD8+ RTE 2 3
CD4+
T cell
CD8+
T cell 4
TREC
TREC
BLOOD/
LYMPHOID TISSUES
1 Thymic output
2 T cell division
3 T cell death
4 TREC degradation
TREC concentration is affected by

16. TREC Can Be Equally Affected by Thymic Output and T Cell Division (at Any Age)#

17. TREC Concentration during Aging
Healthy Volunteers
sjTREC
cjTREC
Age (years)
Douek DC et al. Nature. 396(6712): 690-5, 1998

18. Thymic Involution Induce TREC Decline (Over Entire Lifespan)
TREC concentration is a good marker for RTE in healthy people

19. Summary Thymic Function in Healthy people
Our model quantifies the number of RTE and TREC concentration over an 80-year lifespan
TREC concentration can be equally affected by thymic output and T cell division at any age
Thymic involution induces TREC concentration decline over the entire lifespan
TREC concentration is a good marker for both CD4+ and CD8+ RTE in healthy people

20. Outline Thymic function in healthy people
Thymopoiesis
Markers to represent recent thymic emigrants (RTE)
Thymopoiesis model
TREC model
Thymic function during pediatric HIV-1 infection
HIV-1 infection
Clinical and experimental studies
Thymic infection model
Evaluation of TREC

21. HIV-1 Structure Associated with AIDS Retrovirus family
Two identical ss RNA
Enveloped virus
Antigenic variation
Infects CD4+ cells
Entry requires CD4 and coreceptor

22. HIV-1 Entry

23. Classification of HIV-1 Strains
R5 strain
Uses CCR5 as coreceptor
Replicates slowly and is minimally cytopathic
Is primarily transmitted
Is prevalent in the early stage of disease
X4 strain
Uses CXCR4 as coreceptor
Replicates fast and is highly cytopathic
Is rarely transmitted
Appears in approximately 50% of patients in the late stage of disease
R5X4 strain
Uses either CCR5 or CXCR4 as coreceptor
Has similar properties as the X4 strain

24. HIV-1 Disease Progression
Hypotheses for the decline of CD4+ T cells in the blood
Changes in cell migration patterns
Changes in cell life-spans
Changes in cell production by the thymus

25. Clinical Studies of Thymic Infection with HIV-1
TREC levels in CD4+ and CD8+ T cells decline
Naïve CD4+ and CD8+ T cell numbers decline
Thymic volume decreases
The thymus undergoes morphological changes
“Thymic dysfunction” (TD+) is described in a subset of vertically-infected infants

26. “Thymic dysfunction” (TD+)
Two distinct modes of pediatric HIV-1 disease progression
Normal progressors 10 years to AIDS (85%)
Fast progressors 2-3 years to AIDS (15%)
lower CD4+ and CD8+ counts (DiGeorge Syndrome)

27. Experimental Studies of Thymic Infection with HIV-1
Coreceptors
CCR5 Expressed in low levels on DP, SP4, SP8 cells
CXCR4 Expressed in high levels on ITTP, DP cells,
low levels on SP4 and SP8 cells
HIV-1 strains
R5 strain Infects DP and SP4 cells
Minimally cytopathic
X4 strain Infects ITTP, DP and SP4 cells
Highly cytopathic
Thymocyte maturation stages
SP4,SP8 Support active viral replication
ITTP,DP Support less viral replication

28. Two Questions to Address
1st Can TREC concentration be properly used as measure of RTE?
2nd Whether thymic infection with different
HIV-1 strains contribute to differences in disease progression?

29. Thymic Model With HIV-1 Infection
Cell Source
THYMUS
TES(t) TN X4
ITTP
ITTPX R5 X4
DPR DP
DPX R5 X4
SP8R
SP4R
SP4
SP4X
SP8X
SP8 R5 X4 R5 X4
CD4+ RTE
CD8+ RTE
BLOOD/LYMPHOID TISSUES

30. Virtual Pediatric Thymic Infection

31. Simulations of CD4+ RTE Predict Bimodal Pattern of Blood CD4+ T Cell Counts

32. Simulation Values at Year Two of Virtual Infection Compared with Experimental Data

33. Bifurcation Parameters Implications for Treatment
Viral influx from blood into the thymus
Virulence of strains (viral infection rate and production rate)
HIV-1 induced death of uninfected thymocytes
Progenitor cells from bone marrow
Carrying capacity of the thymus

34. TREC Concentration during HIV-1 Infection
HIV-1 Infected Subjects
sjTREC
Age
Age (years)
Douek DC et al. Nature. 396(6712): 690-5, 1998

35. Two Questions to Address
1st Can TREC concentration be properly used as measure of RTE?
2nd Whether thymic infection with different
HIV-1 strains contribute to differences in disease progression?

36. T Cell Kinetics during HIV-1 Infection
We include these effects in the model to represent HIV-1 infection

37. T Cell Counts and TREC Concentrations in HIV-1 Infected Subjects (Age 30)
T cells
TREC
Margolick JB et al. J Acquir Immune Defic Syndr. 6(2): , 1993
Zhang L et al. J. Exp. Med. 187(11): , 1998

38. Model Prediction for Thymic Output during HIV-1 Infection
TREC concentration is a good marker for CD4+ RTE
TREC concentration is NOT a good marker for CD8+ RTE

39. Summary Thymic Function during Pediatric HIV-1 Infection
Infection with R5 and X4 strains induces different degrees of thymic dysfunction, which is related to CD4+ T cell counts and disease progression as seen in pediatric patients
Thymic infection in pediatric patients is more severe than in adult patients, likely due to higher viral loads and a more active thymus
Suppressing viral load in blood, high drug efficacy within the thymus, and improving inherent thymic function are necessary for reconstitution of RTE levels
Protease inhibitors have high levels of efficacy directly suppressing viral replication within the thymus, while reverse transcriptase inhibitors have low efficacy
TREC concentration is a reliable marker for CD4+ RTE, but not for CD8+ RTE in HIV-1 infected subjects

40. Conclusions
Peripheral T cell turnover should be examined together with TREC concentration as a measure of RTE
Infection with different HIV-1 strains induces different degrees of thymic dysfunction, contributing to CD4+ T cell depletion and disease progression
With adequate suppression of viral replication within both the blood and the thymus during HAART, thymic function recovery can reconstitute immune cell numbers

41. Publications
Ping Ye and Denise Kirschner (2002). Reevaluation of T cell receptor excision circles as a measure of human recent thymic emigrants. Journal of Immunology, 168(10),
Ping Ye, Athena Kourtis, and Denise Kirschner (2002). The effects of different HIV-1 strains on human thymic function. AIDS Research and Human Retroviruses, 18(17) (In press).
Ping Ye, Athena Kourtis, and Denise Kirschner . Reconstitution of thymic function in HIV-1 patients treated with highly active anti-retroviral therapy (submitted).

42. Acknowledgments Dr. Denise Kirschner Collaborator: Dr. Athena Kourtis
Dr. Ramit Mehr
Kirschner lab members
Funding
NIH and Whitaker Foundation to DEK
Rackham Graduate School and Elizabeth Glaser Pediatric AIDS Foundation to PY

43. Clinical Studies of Thymic Function during HAART
TREC levels recover
Naïve CD4+ and CD8+ T cell numbers increase
Thymic volume increases
Children with “thymic dysfunction” response well to HAART

44. Model of Thymic Function Reconstitution during HAART
Cell Source
THYMUS
TES(t) TN
ITTP X4
ITTPX
DPR R5 X4
DPX DP R5 X4
SP8R
SP4R
SP4
SP4X
SP8X
SP8 R5 X4 R5 X4
CD4+ RTE
CD8+ RTE
BLOOD/LYMPHOID TISSUES
Reduce viral influx into the thymus
Decrease viral infection and production (if thymic drug efficacy > 0%)
Decrease HIV-1 induced death of uninfected thymocytes
Increase progenitor cells from bone marrow
Increase carrying capacity of the thymus

45. Simulations of pediatric thymocyte dynamics during HAART

46. Simulations of CD4+ RTE Predict HAART Agent Thymic Efficacy
RTI
HAART