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. Author manuscript; available in PMC: 2015 Jun 21.
Published in final edited form as: Immunol Rev. 1991 Apr;120:35–50. doi: 10.1111/j.1600-065x.1991.tb00586.x

Alpha beta T-Lymphocyte Depleted Mice, a Model for γδ T-Lymphocyte Functional Studies

Amy Carbone §, Ronald Harbeck *, Angela Dallas *, David Nemazee **, Terri Finkel **, Rebecca O'Brien *, Ralph Kubo *,+, Willi Born *,+
PMCID: PMC4475640  NIHMSID: NIHMS513288  PMID: 1830862

INTRODUCTION

T cells of both the αβ and γδ lineage develop in the thymus. Whereas the γδ T-cell receptor (TCR)-CD3 complex appears to gain functional competence early in thymocyte development (Bucy et al. 1990, Coltey et al. 1989, Kyewski (in press)), at least three states of the αβ TCR-CD3 complex, associated with consecutive stages of thymocyte maturation, can be distinguished (Born et al. 1987, Finkel et al. 1989, McDuffie et al. 1986). During the earliest stage, represented by approximately one half of cortical thymocytes, the developing T cells express low levels of αβ TCR, mobilize only small amounts of Ca2+ after TCR engagement and are apparently insensitive to mechanisms that lead to negative selection. At the intermediate second stage, represented by the remaining cortical thymocytes, the developing T cells still express low levels of αβ TCR, but mobilize larger amounts of Ca2+ in response to TCR engagement and are tiow susceptible to negative selection. The third stage is represented by both medullary thymocytes and peripheral T lymphocytes. These mature T cells express high levels of αβ TCRs, can mobilize large amounts of Ca2+, and tend to perceive TCR engagement as an activating signal (Ellenhorn et al. 1988). Accordingly, efficient and selective depletion of mature peripheral αβ T lymphocytes might be achieved by interrupting thymocyte development during the sensitive cortical stage of development using anti-TCR antibodies, similar to earlier experiments with anti-major histocompatibility complex (MHC) monoclonal antibody (mAb) (Kruisbeek et al. 1985, 1983). This was first demonstrated with anti-CD3 (Owen et al. 1986, 1988) and with subset-specific anti-αβ TCR mAb (Born et al. 1987, McDuffie et al. 1986). Using framework-recognizing mAb that react with all mouse or rat αβ TCRs, virtually all lymphocytes bearing functional αβ TCRs can be depleted (Finkel et al. 1989, Hünig et al. 1989a, 1989b, Kubo et al. 1989). Although the possibility of secondary changes in other components of the immune system must be considered (Born and Ewijk, unpublished), αβ T-cell depletion provides an opportunity to conduct functional studies in vivo on lymphocytes expressing γδ TCRs, a population normally overshadowed by the predominant αβ T cells.

PARTIAL DEPLETION OR INACTIVATION OF αβ T LYMPHOCYTES

We first studied the effect of TCR engagement on thymocyte maturation in situ using the mouse Vβ8-specific mAb, KJ16 (Haskins et al. 1984, Roehm et al. 1984). In fetal thymus organ cultures (FTOC), small amounts of this antibody (a single dose of 500 nanogram/ml culture medium) were found to be sufficient to prevent the generation of mature Vβ8-bearing cells (Born et al. 1987). For in vivo depletion or inactivation of Vβ8-bearing cells, starting at birth, about 100-fold higher amounts of this antibody were required every 2nd day (McDuffie et al. 1986). The actual antibody concentrations in situ were not determined, however, and effective antibody concentrations might well be similar. By comparing immunofluorescent staining patterns with mAb KJ16 and secondary anti-Ig reagents, or secondary reagents alone, it was found that the anti-TCR mAb reached all available binding sites in the thymus, both in vivo and in vitro. The presence of the antibody resulted in a rapid and reversible modulation of receptors normally expressed on immature, low-density TCR-bearing thymocytes. Receptor modulation, however, did not seem to affect the appearance and development of immature, low-density TCR-bearing thymocytes, because withdrawal of the antibody was followed by a rapid reappearance of these cells, a phenomenon that can be best observed in the organ culture system. In contrast, T cells bearing high levels of Vβ8 failed to develop under the antibody treatment in either the thymus or the peripheral lymphoid organs. Pre-existing, mature T lymphocytes were much less susceptible to this effect. These data, together with ultrastructural evidence for receptor-mediated interactions between cortical thymocytes and epithelial cells (Farr et al. 1985), suggested that TCR modulation at an intermediate cortical stage of thymocyte development probably prevented further maturation, resulting instead in cell inactivation or death. In this regard, the effect of the antibody treatment resembles the physiological process of negative selection. An alternative possibility, namely that the antibody might opsonize cells and thus render them susceptible to Fc-recognizing, cytotoxic reticulocytes, was excluded because pepsin-treated or purified Fab′ KJ16 had a similar effect. Although our earlier organ culture experiments using mAb KJ16 had suggested that most cortical thymocytes are insensitive to receptor modulation, later organ culture studies using a framework-recognizing, pan-specific anti-mouse TCR mAb, H57-597 (see below), showed that cortical, TCR-bearing thymocytes are divided into an insensitive, probably less mature, and a sensitive, more mature population. As mentioned earlier, these cortical subsets appear to differ by the way their αβ TCR and CD3-complexes are functionally coupled (Finkel et al. 1989).

PAN-SPECIFIC ANTI-TCR ANTIBODIES

To generate a monoclonal antibody that would react with all mouse αβ TCRs, hamsters were immunized with affinity-purified receptor isolated from the T-cell hybridoma DO-11.10 (Kubo et al. 1989). DO-11.10 expresses Vβ8 and Vα13 (Yagüe et al. 1988). Sera from immunized animals were tested for anti-αβ TCR reactivity by immunoprecipitation of receptor from an unrelated T-cell hybridoma expressing different Vβ, Dβ and Jβ segments (2QK34.7), as well as for their ability to stimulate that hybridoma. Cells from an animal whose serum tested positive in these assays were fused to the mouse myeloma variant, X63-Ag8.653 (Kearney et al. 1979) to generate B-cell hybridomas. Two hybridomas from this fusion were reactive with the TCR of 2QK34.7, suggesting that they probably recognized framework TCR determinants. Monoclonal Ab H57-597 stimulated IL-2 production by all αβ T-cell hybridomas tested, whereas T-cell hybridomas bearing γδ TCR did not respond. Similarly, H57-597 precipitated αβ TCR but not γδ TCR from hybridoma lines. Under conditions that result in the dissociation of TCR αβ from CD3, the antibody precipitated αβ TCR alone, but not CD3. In addition, sequential immunoprecipitation of a digitonin lysate of neonatal thymocytes with H57-597 and subsequently with anti-CD3 mAb demonstrated that only γδ TCR-CD3 complex had remained after removal of the H57-597 precipitate. Finally, the staining pattern of H57-597 with murine thymocytes (Fig. 1A) and peripheral T lymphocytes (Fig. 3C) indicated that this mAb recognizes all murine αβ TCRs.

Figure 1.

Figure 1

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Effect of anti-TCR αβ treatment in vivo on adult C57Bl/6 thymocytes. Mice were injected with PBS or H57-597 as described in the text, and rested for 1 d before staining. Thymocytes were cultured in suspension for 3-5 h, then stained with biotin-conjugated H57-597 followed by phycoerythrin (PE)-streptavidin (A, B), unconjugated H57-597 followed by FITC-labeled rabbit anti-hamster Ig antiserum (E, F), or FITC-labeled rabbit anti-hamster Ig antiserum alone (C, D). Stained cells were analyzed on an EPICS C cytofluorograph.

Figure 3.

Figure 3

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Anti-TCR αβ treatment eliminates peripheral TCR αβ-bearing lymphocytes. Whole (A, B) or nylon wool purified (C) lymph node cells of PBS- or H57-597-treated mice were cultured in suspension for 3–5 h, then stained with biotinylated H57-597 and fluoresceinated 145-2C11 (FITC anti-CD3) or HO13-4-6 (FITC-anti-Thy-1) followed by PE-streptavidin.

For in vivo treatments, mice were injected intraperitoneally (i.p.) with 100 μg of purified H57-597 in 50 μl of phosphate-buffered saline (PBS) on the day of birth and the following day. Thereafter, mice received 200 μg of antibody i.p. every other day until the animals were sacrificed for analysis. Control littermates received 50 μl of PBS. During the 1st week, injections were performed by tunneling a 30 gauge needle into the peritoneum over the left anterior thorax to minimize trauma and loss of injected material.

Initially, mice were monitored over periods of up to 6 months for possible non-specific effects of the treatment (unpublished). Although not undernourished, H57-597-treated mice were consistently smaller than age-matched, PBS-treated controls, suggesting a developmental delay. Yields of thymocytes, spleen cells, and lymph node cells were reduced to various degrees (up to 20-fold), in comparison to PBS-treated mice and normal untreated littermates. Mortality was increased in the groups that received frequent injections but there was no difference between antibody- and PBS-treated mice.

Hünig et al. (1989a) generated a monoclonal antibody reactive with all rat TCRs by immunizing BALB/c mice with rat T-cell blasts. Immune spleen cells were fused to the nonproducer cell line X63-Ag8.653. Supernatants of hybridomas were first screened for antibody binding to rat T-cell blasts and subsequently for the induction of DNA synthesis in rat splenocytes. One mAb (R73) was obtained that apparently detected a framework determinant on rat αβ TCRs.

EFFECT ON ADULT THYMOCYTES

Treatment of mice with H57-597 resulted in complete loss of high-density αβ TCR-positive cells in the thymus (Fig. 1, A-F). Failure to detect surface receptor was not due to masking of TCR determinants by injected antibody since staining with a secondary rabbit anti-hamster Ig antibody alone (Fig. 1, C and D) or in conjunction with unconjugated H57-597 (Fig. 1, E and F) did not reveal high-density TCR-positive thymocytes. Thymocytes from H57-597-treated mice and from PBS-treated controls were routinely incubated in suspension cultures for 3–5 hours in order to better visualize and quantitate low density TCR-positive cells, because this manipulation results in increased receptor density (A.C., unpublished). Low-density TCR-bearing thymocytes were only slightly diminished. To determine whether the absence of high-density TCR-positive thymocytes in H57-597-treated mice reflected merely receptor modulation or irreversible loss of these cells, thymocytes were incubated in suspension cultures for 24 or 72 h. Both the percentage and the intensity of low-density TCR-positive cells increased among cultured thymocytes from control and H57-597-treated animals after 24 h of incubation (Fig. 2, B and D, compare with Fig. 1, A and B). Nevertheless, no high-density TCR-positive cells (Fig. 2, D) were detected in experimental mice. The number of low-density TCR-positive thymocytes was estimated by calculating their proportion among TCR-negative and low-density TCR-positive cells only, so that cells from H57-597-treated mice could be directly compared to controls (Finkel et al. 1989). The proportion of these cells was significantly decreased in thymocytes that had not been cultured in suspension (approx. 40% of control cells, not shown) but did increase after 3–5 h of culture (81% of control) and 24 h of culture (84% of control). It seems likely that this was mainly due to receptor modulation followed by re-expression during the culture period, although other possibilities such as de novo expression of αβ TCRs in vitro (Owen et al. 1986) have not been excluded. In FTOC experiments, more low-density cells seemed to be eliminated by the antibody treatment (Finkel et al. 1989); this difference could be due to lower effective antibody concentrations or a greater supply of precursor cells in vivo.

Figure 2.

Figure 2

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Anti-TCR αβ treatment eliminates mature TCR αβ-bearing thymocytes. Thymocytes of PBS- or H57-597-treated mice were cultured in suspension for 24 or 72 h, then stained and analyzed as described in the legend to Fig. 1.

The absence of high-density TCR-positive cells after 24 h of suspension culture probably indicates their complete elimination from the thymus of antibody-treated animals. Thymocytes were also cultured for 72 h. In thymocytes of PBS-treated control mice, a 5-fold relative increase in high-density TCR-positive cells and a decrease in TCR-negative thymocytes was found (Fig. 2, F). These proportional changes could be caused by in vitro maturation or selective growth of certain thymocytes. In thymocytes of antibody-treated mice, high-density TCR-positive cells were still not present after 72 h of culture, although TCR-negative cells were reduced as in the control cultures (Fig. 2, H). These data confirm that mature αβ TCR-bearing thymocytes are irreversibly depleted due to the antibody treatment.

Similar to the studies in mice, the generation of αβ TCR-positive cells in rats could be suppressed by injections of purified mAb R73 from birth (Hünig et al. 1989b). As with mice, αβ TCR-suppressed rats were found to lack mature thymocytes expressing high levels of the αβ TCR. No effect was observed on low-density TCR-expressing thymocytes, however. This difference of observations concerning low-density TCR-bearing cells could be due to anatomical dissimilarities between rats and mice, or to differences in the affinity or dosage of the two antibodies, since reportedly only a partial saturation of available binding sites on the rat cells was achieved (Hünig et al. 1989b).

EFFECT ON PERIPHERAL αβ T LYMPHOCYTES

Peripheral lymphocytes of antibody-treated mice were analyzed by two-color flow cytometry. In unseparated lymph node cells, no TCR αβ-bearing cells were found, whereas 49% of PBS-treated control lymph node cells were TCR αβ-positive (Fig. 3, A). Similar results were obtained with spleen cells from antibody-treated mice, and lymph node and spleen cells incubated in suspension cultures for 24 h to permit re-expression of TCRs (data not shown). Thus, as with anti-CD3 mAb (Rueff-Juy et al. 1989), neonatal injections resulted in a total depletion of mature αβ T lymphocytes. In contrast, it has been reported that adult injections were only temporarily immunosuppressive (Hirsch et al. 1988). Staining of lymph node cells with anti-Thy-1 mAb and H57-597 revealed in the experimental mice the presence of a residual population of Thy-1-bearing cells, approximately 5% of Thy-1-positive cells in controls (Fig. 3, B). These cells expressed Thy-1 at slightly higher levels than normal lymph node-derived T lymphocytes. Enrichment by passage over nylon wool showed that most of these cells were TCR αβ- and γδ-negative. Only a very small fraction expressed TCR αβ, and at 5–10 times lower levels than normal T lymphocytes (Fig. 3, C). The residual Thy-1-bearing cells also differed from normal T-cell populations with regard to CD4 and CD8 expression. Most were CD8-positive, 10% bore CD4 and 14% were CD4/8 double-negative (Fig. 4A, B). Among Thy-1-positive lymph node cells of controls, most bore CD4, fewer cells were CD8-positive and only a very small fraction was CD4/8 double-negative.

Figure 4.

Figure 4

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Anti-TCR αβ treatment reveals a population of Thy-1-positive, TCR αβ/γδ-negative peripheral T lymphocytes with altered CD4/8 surface phenotype. Nylon wool purified lymph node cells of PBS- or H57-597-treated mice were cultured in suspension for 3–5 h, then stained with biotinylated GK1.5 (biotin anti-CD4) or biotinylated 53-6.72 (biotin-anti-CD8) and FITC-anti-Thy 1 followed by PE-streptavidin.

Again, similar findings have been reported in αβ-suppressed rats (Hünig et al. 1989b). Peripheral lymphoid organs were found to be devoid of αβ TCR-bearing cells. Consistently, however, a small population of lymphocytes in spleen and lymph nodes expressed αβ TCR at roughly 5-fold lower levels than control cells. As in mice, it is not known whether this phenotype is an experimental artefact or typical of certain immature cells that manage to escape thymic selection.

γδ T LYMPHOCYTES

For one- and two-color fluorescent analyses of γδ cells, the pan-specific anti-γδ TCR antibody 4O3.A10, kindly provided by Dr. Osami Kanagawa, was used (Itohara et al. 1989). Sizes of γδ cell populations in thymus and peripheral lymphoid organs were not significantly changed by the depletion of αβ TCR-bearing lymphocytes. This is in agreement with findings in FTOC (Kyewski (in press)). In the normal adult thymus, γδ TCR-bearing cells were infrequent (less than 1%) and the same was true for thymocytes of H57-597-treated mice (Fig. 5). Similarly, γδ-positive cells accounted for only about 1% of whole, unseparated normal lymph node cells. In αβ-suppressed mice, their relative frequency increased (Fig. 6, A, B). Because of the T-cell depletion in H57-597-treated mice, nylon wool passage could be used efficiently to enrich and quantitate γδ cells (Fig. 6B). Total numbers of γδ cells in lymph nodes (Fig. 6) and spleen (not shown), however, were found to be largely unaltered. These data do not address the possibility of other, qualitative changes in the γδ cell population. In αβ-suppressed rats, the frequency of CD5-positive, TCR αβ-negative cells was increased, suggesting that γδ cells in these animals might be elevated above normal levels. Alternatively, this CD5-bearing population, similarly to the residual Thy-1-bearing population in mice, may be TCR-negative (Hünig et al. 1989b).

Figure 5.

Figure 5

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TCR αβ-positive thymocytes are not replaced by TCR γδ-bearing thymocytes. Thymocytes of 10-wk-old, PBS- or H57-597-treated mice were cultured in suspension for 3–5 h, then stained with fluoresceinated 145-2C11 (FITC-anti-CD3) (A, B) or unconjugated 403.A10 (pan-specific anti-TCR γδ; a gift of Dr. O. Kanagawa), followed by FITC-labeled rabbit anti-hamster antiserum (C, D).

Figure 6.

Figure 6

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Anti-TCR αβ treatment increases relative frequencies but not absolute numbers of peripheral TCR γδ-bearing lymphocytes. Whole (A) or nylon wool purified (B) lymph node cells of PBS- or H57-597-treated mice were cultured in suspension for 5 h, then stained with biotinylated H57-597 or 403.A10 (biotin-anti TCR γδ), and fluoresceinated 145-2C11 (FITC-anti CD3) followed by PE-streptavidin.

ANTIGEN RESPONSES

The virtual absence of mature αβ T cells in αβ-suppressed mice, together with the ohservation that γδ T cells are at least grossly unaffected, suggests that αβ-suppressed mice might be a sensitive vehicle for testing antigen specificity and functions of γδ T cells. Since alloantigen-reactive γδ clones have been isolated by others (Matis et al. 1987, 1989, Rivas et al. 1989), we have attempted to stimulate spleen cells of αβ-suppressed mice with alloantigen. Alloreactivity, as measured in bulk mixed lymphocyte cultures, was not detectable (unpublished observation). This is perhaps not surprising since frequencies of alloantigen-reactive clones among γδ cells appear to be considerably lower than among αβ cells (O'Brien et al. 1989).

It has also been reported that αβ-suppressed rats were essentially devoid of alloreactive cells, even when IL-2 was added to facilitate the responses (Hünig et al. 1989b). This is in line with unpublished data by the same authors showing that αβ-suppressed rats accept skin grafts that are rejected by normal controls.

Because γδ cells in normal mice have been shown to respond to mycobacterial antigen administered in complete Freund's adjuvant (IFA) (Janis et al. 1989), we wished to examine γδ cell reactivity in the absence of αβ T lymphocytes. Table I shows that γδ cells rapidly accumulated in the draining lymph nodes of αβ-suppressed mice that had been immunized subcutaneously with purified protein derivative (PPD) in IFA. Responses in H57-597-treated animals were almost as strong and rapid as in PBS-treated controls. The data suggest that αβ T lymphocytes are not required for this γδ cell response. Whether γδ cells accumulate under these conditions because they recognize mycobacterial antigens or perhaps inflammatory autoantigens (Born et al. 1990a, 1990b), and whether they proliferate in situ or merely home to the draining lymph nodes remains open.

TABLE I.

Expansion of γδ cells in response to PPD/IFA

# days after PPD injection Ab Treatment Uninjected (Left side)* PPD-Injected (Right side)* Stimulation Index
2 PBS 1.2 9.1 7.6
H57-597 8.8 51.0 5.8
4 PBS 0.4 11.9 31.3
H57.597 1.6 23.5 14.7
8 PBS 3.8 31.5 8.3
H57-597 4.6 15.4 3.3
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Adult H57-597-treated, or PBS-treated control mice were injected in the right front and rear footpads with 50 μg PPD (M. tuberculosis) in IFA. Draining lymph nodes of injected and non-injected control sides, and experimental and control mice are compared. Absolute numbers of γδ cells at different time points are not comparable because of changes in the experimental protocol.

*

Number of cells × 104.

Finally, we examined T cell-dependent and -independent B-cell responses in αβ-suppressed mice. For this purpose, α/β-suppressed and PBS-injected age- and sex-matched control mice were subcutaneously immunized with arsenate-keyhole limpet hemocyanin (ars-KLH) in CFA. and with TNP-Ficoll i.p. One week later mice were sacrificed, the sera analyzed by ELISA for serum antibody production and Ig levels, and the spleens analyzed by flow cytometry with a panel of monoclonal antibodies specific for various cell surface markers. The αβ-suppressed mice tested had considerably fewer spleen cells than controls. They also showed subnormal percentages of B cells as defined by the following surface markers: IgM, IgD, kappa, and B220 (Table II). This suggests that B-cell development, homing, or clonal expansion is impaired in αβ-suppressed mice. The large reduction of Ly-1-positive cells in the H57-597-treated animals was an indicator of their T-lymphocyte depletion. The proportions of B cells bearing high levels of IgD (approximately 85%) were similar in suppressed and normal splenocytes (not shown).

TABLE II.

Cell surface immunofluorescence analysis of αβ suppressed, immunized mice: B lymphocyte markers

% positive
Spleen cells IgD IgM Kappa Ly-1* Iab B220
H57-597-treated
    1 25.9 32.0 27.8 9.5 47.1 35.0
    2 29.6 32.2 32.3 5.4 45.7 35.5
PBS-treated
    3 37.5 45.8 46.6 34.7 55.4 49.1
    4 41.2 48.2 49.5 35.2 55.2 51.6
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Percentages of IgD, IgM, kappa, Ly-1, Iab and B220 positive cells among splenocytes of 2 H57-597-treated and 2 control mice. Eight-week-old male C57Bl/6 mice were injected intraperitoneally with 100 μg TNP-Ficoll (Biosearch, San Rafael, CA), and in the front and rear footpads with 50 μg KLH-arsenate in CFA. Eight days later, spleen cells were stained with mAb specific for the incicated surface markers and analyzed using an EPICS profile cytofluorograph.

*

High-density Ly-1. The sensitivity of this staining did not allow quantitative measurement of B cells of the Ly-1 lineage that bear this antigen at a very low surface density.

Sera taken from immunized and pre-immune mice that were either αβ-suppressed or non-suppressed were analyzed for specific antibody and IgM levels (Fig. 7). Suppressed mice responded to the T cell-independent antigen TNP-Ficoll but did not respond to the T cell-dependent antigen ars-KLH. Control non-suppressed mice responded to both antigens. Total serum IgM levels in all mice were similar, suggesting that the “normal” serum IgM is essentially unaffected by the chronic anti-αβ treatment.

Figure 7.

Figure 7

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Specific and nonspecific serum IgM in immunized, αβ TCR-suppressed mice. Eight days after immunization (see Table II), diluted sera of normal and H57-597-suppressed male C57Bl/6 mice were incubated in polyvinyl chloride microtiter wells coated with TNP-BSA, ars-BSA, or BSA alone to detect specific antibodies, or with anti-mouse IgM (M41), rat anti λ (L22.18), or normal rat Ig to detect total or λ-bearing IgMs. Wells were then washed, incubated with horseradish peroxidase-conjugated rat anti-mouse IgM and the bound peroxidase was measured using a standard colorimetric assay. 1: Pool of sera from 5 non-immunized, αβ TCR-suppressed mice. 2: Pool of two normal controls. 3,4: Individual non-suppressed, TNP-Ficoll- and KLH-ars-immunized mice. 5,6: Individual H57-597-treated, TNP-Ficoll- and KLH-ars-immunized mice.

CONCLUSIONS

The data presented in this paper show that, starting at birth, mice can be efficiently depleted of all mature αβ TCR-bearing lymphocytes by in vivo treatment with the framework-recognizing, pan-specific anti-TCR αβ mAb H57-597. It is possible that residual Thy-1-positive cells in lymph nodes and spleen are progeny of immature thymocytes that escaped normal selectional processes. It remains to be seen whether these cells represent a population that is, perhaps in smaller numbers, also present under nonnal conditions. It is also unclear whether these cells have retained function; for example, natural killer-like cytotoxicity. B lymphocytes in immune, αβ-suppressed mice were reduced in numbers, but no changes were detected in their overall composition. Also, T cell-independent B-cell responses were not significantly changed, suggesting that the B-lymphocyte compartment remains functionally competent in the absence of αβ T lymphocytes. Most interestingly, perhaps, thymic and peripheral populations of TCR γδ-bearing lymphocytes appeared to be largely unaffected by the depletion of αβ cells. Since γδ cells also showed nearly normal responses after immunization, the data suggest that both the development and the functional responses of γδ cells may be regulated independent of αβ T lymphocytes. These results support the view that γδ cells are a distinct functional lymphocyte subset rather than “αβ T cells in disguise.” Observations suggesting that most γδ clones may be unable to recognize alloantigens, and that γδ cells may also be unable to substitute for αβ-positive T-helper cells in T cell-dependent B-cell responses, further stress the distinction between the two T-cell subsets. Neonatal αβ suppression may provide a useful model for studies on γδ cell function.

SUMMARY

Adult mice can be depleted of essentially all mature αβ T lymphocytes by chronic treatment with the framework-recognizing, pan-specific anti-TCR αβ mAb, H57-597. Similar findings have been reported in rats. γδ cell populations remain essentially unaltered in size and reactivity. Suppression of αβ T-cell development results in the loss of alloantigen reactivity and of B-cell help, suggesting that γδ and αβ populations differ in their functional capabilities. Indirect effects of the antibody treatment include quantitative changes in splenic B cells, as well as reduced sizes and weights of experimental animals, αβ-suppressed mice and rats may provide model systems for studies on γδ cell function in vivo.

ACKNOWLEDGMENTS

This work was supported by NIH grant AI27903-01. W. Born is a recipient of a Cancer Research Institute Investigator Award. The authors wish to thank William Townend for help with cytofluorimetric analysis, Deirdre Gay for the maintenance of αβ-suppressed mice, and Margaret Hammond for expert secretarial assistance.

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