Arstila et al. estimated an upper bound of 10⁸ different combinations (1). Pre-T cells having rearranged a  chain expand 1000-fold before the  chain is rearranged, and only 10% of these cells leave the thymus to enter the mature repertoire. Thus, it was argued that each  chain can maximally pair with any of about 100 different  chains.

This is indeed correct for all descendants of any particular pre-T cell having rearranged a particular  chain--but another pre-T cell rearranging the same  chain may bind to 100 different  chains. Thus, to calculate the upper bound on TCR diversity, one has to consider the frequency with which identical -chain rearrangements are expected. This frequency can be estimated from the turnover rate of the naïve T cell repertoire. In human adults, the total body production of naïve T cells has been estimated at about 10⁸ per day (2), a figure obtained from recovery rates following T cell depletion (2) and from an estimated 0.1% turnover (3) in a pool of 10¹¹ naïve T lymphocytes. Assuming that most of this production is of thymic origin (4) and that more than 90% of the cells die before leaving the thymus (1), this implies a daily production of at least 10⁹ pre-T cells. The 1000-fold expansion of the pre-T cells (1) before -chain rearrangement implies that approximately 10⁶  chains should be made every day. Because this is close to the Arstila et al. estimate of total -chain diversity, every  chain should be rearranged about every day.

Over the 1000-day expected life-span (2, 3) of the progeny of a pre-T cell expressing a single  chain, therefore, 1000 recurrences of the same -chain rearrangement might be expected. Hence, the upper bound for the total TCR diversity could easily be 1000-fold larger than calculated by Arstila et al. Such an upper bound, at 10¹¹, would allow almost every T cell in the naïve repertoire to have a unique TCR. The true TCR diversity may be several fold lower, however, owing to factors such as proliferation after the -chain rearrangement and possible restrictions in -chain pairing.

Can Kemir
José A.M. Borghans
Rob J. de Boer
Department of Theoretical Biology
Utrecht University
Padualaan 8
3584 CH Utrecht, The Netherlands
E-mail: R.J.deBoer{at}bio.uu.nl

REFERENCES

1. T. P. Arstila, et al., Science 286, 958 (1999) [Abstract/Free Full Text] .
2. D. R. Clark, R. J. De Boer, K. C. Wolthers, F. Miedema, Adv. Immunol. 73, 301 (1999) [Web of Science] [Medline] .
3. C. A. Michie, A. McLean, C. Alcock, P. C. Beverley, Nature 360, 264 (1992) [CrossRef] [Medline] .
4. B. F. Haynes, et al., J. Clin. Invest. 103, 453 (1999) [Web of Science] [Medline] .

Response: Kemir et al. argue that although any developing TCR  chain will be paired at most with 100 different  chains, the same  chain may appear repeatedly and garner other sets of 100  chains, increasing the total TCR diversity from the 10⁸ we estimated (1). We studied the diversity of the human TCR in the blood of healthy adult donors at a given moment, not over time. Also, we did not measure the upper limit of -to- pairing; our estimate was based on what is known of T cell development and TCR rearrangement. Thus, the comment of Kemir et al. actually goes beyond our data.

Because any expansion after -chain rearrangement will increase only clone size, not diversity, the argument of Kemir et al. hinges on the assumption that the estimated total turnover of naïve T cells equals thymic production of pre-T cells. That assumption is incorrect, however, and ignores the well-documented role of post-developmental division in the maintenance of the naïve T cell population, especially in adults. Murine T cells may go through up to six cell cycles after -chain rearrangement even before emigrating from the thymus (2). Haynes et al., cited by Kemir et al., specifically argued for "minimal contributions of the thymus to maintenance or reconstitution of the peripheral pool of T cells . . ." in humans [(3), p. 457], and showed that the presence or absence of thymic function and even the surgical removal of the thymus had no impact on the reconstitution of the T cell compartment, including the naïve CD4⁺ cells, in treated HIV-infected individuals. Naïve T cells, long after having completed TCR rearrangement, clearly have a considerable capacity for self-renewal.

The suggestion of Kemir et al. can also be viewed as a question of clone size. If the size of the repertoire is 10⁸ different TCRs, as we suggest, the average clone among 10¹¹ naïve T cells would consist of 1000 cells, the progeny of a single intrathymic -chain rearrangement after 10 cell cycles. These cycles should therefore be detectable in the naïve T cell population, and indeed this appears to be the case. Studying the disappearance of cells damaged by therapeutic irradiation, McLean and Michie (4) concluded that, on average, naïve T cells divide once every 3.5 years and die after 20 years, which suggests six post-thymic cell cycles in the life-span of an average naïve T cell. Other experimental approaches have suggested higher division rates. From age 25 to 70 years the mean telomere length in the naïve T cell population decreases from 9.5 kb to 8.0 kb, so an estimated loss of 50 to 100 base pairs (bp) per cell cycle translates to 7 to 13 divisions during the 20-year life-span of naïve cells (5). De Boer and Noest have argued that this estimate of telomere loss is too high; their estimate, 35 to 70 bp per cycle (6), would mean 10 to 19 cycles. At any given time the fraction of naïve T cells in cell cycle is 0.8% (7), which suggests a rate as high as one division per 125 days, or 60 cycles per life-span. The available data thus can easily accommodate 10 divisions producing the average naïve clone.

Studies on the frequency of antigen-specific T cell precursors provide an independent line of evidence that points to a diversity close to what we proposed. A conservative estimate of the frequency of such precursors in the naïve repertoire would be one per million; some studies have reported significantly higher frequencies (8, 9). Thus, a total repertoire of 10⁸ TCRs would predict an epitope-specific response to consist of 100 clones, while Kemir et al.'s repertoire of 10¹¹ TCRs predicts a composition of 100,000 responding clones. The existing literature is more compatible with our prediction (10-14). Thus, we submit that the phenomenon that Kemir et al. postulate, although in principle possible, has little impact on the total diversity.

T. Petteri Arstila
Department of Virology
Haartman Institute
University of Helsinki
00014 Helsinki, Finland
E-mail: petteri.arstila{at}helsinki.fi
Armanda Casrouge
Véronique Baron
Jos Even
Jean Kanellopoulos
Philippe Kourilsky
Unité de Biologie Moléculaire du Gène
INSERM U277
Institut Pasteur
Paris Cedex 15, France

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11. M. F. C. Callan et al., Nat. Med. 2, 906 (1996).
12. Y. N. Naumov, K. T. Hogan, E. N. Naumova, J. T. Pagel, J. Gorski, J. Immunol. 160, 2842 (1998) [Abstract/Free Full Text] .
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