Immunophenotypic attributes of benign peripheral blood [gamma][delta] T cells and conditions associated with their increase.
Abstract: * Context.--In comparison to [alpha][beta] T cells, little is known about the immunophenotype of healthy peripheral blood [gamma][delta] T cells or about conditions associated with expansion of this usually minor T-cell subset.

Objective.--To study the immunophenotype of increased nonneoplastic peripheral blood [gamma][delta] T cells and to determine clinical conditions associated with this laboratory finding.

Design.--Flow cytometric T-cell phenotyping studies performed on 352 consecutive peripheral blood specimens were reviewed, and 62 cases (18%) in which [gamma][delta] T cells comprised either more than 5% of the total lymphocytes or had an absolute count of more than 200 cells per [micro]L or both, were studied further. Clinical data were available from 36 cases.

Results.--The [gamma][delta] T cells often had an immunophenotype distinct from the [alpha][beta] T cells, with differences in CD5 expression as the most common (n = 17), followed by differences in CD3 (n = 6) and CD7 (n = 3). CD16 coexpression by the [gamma][delta] T cells was also frequent (n = 20). In 28 (78%) of 36 cases, there were one or more associated conditions: infection/inflammatory disease (n = 18), autoimmune disease (n = 9), lymphoproliferative disorder (n = 6), and splenectomy (n = 3).

Conclusions.--Circulating [gamma][delta] T cells are immunophenotypically distinct from ap T cells, and mild increases in these cells are not uncommon and may be associated with immune system activation and splenectomy. Recognition of this phenomenon is important because reactive [gamma][delta] T cells can exhibit distinctive immunophenotypic features that are also encountered in neoplastic conditions, such as T-cell large granular lymphocytic leukemia.
Article Type: Report
Subject: T cells (Genetic aspects)
Phenotype (Identification and classification)
Immunopathology (Research)
Clinical pathology (Research)
Flow cytometry (Methods)
Authors: Roden, Anja C.
Morice, William G.
Hanson, Curtis A.
Pub Date: 11/01/2008
Publication: Name: Archives of Pathology & Laboratory Medicine Publisher: College of American Pathologists Audience: Academic; Professional Format: Magazine/Journal Subject: Health Copyright: COPYRIGHT 2008 College of American Pathologists ISSN: 1543-2165
Issue: Date: Nov, 2008 Source Volume: 132 Source Issue: 11
Topic: Event Code: 310 Science & research
Geographic: Geographic Scope: United States Geographic Code: 1USA United States
Accession Number: 230246865
Full Text: Early in their ontogeny, T cells commit to express either [alpha][beta] or [gamma][delta] T-cell receptor (TCR) heterodimers, resulting in phenotypically and functionally distinct T-cell lineages. (1) In healthy adults, [alpha][beta] T cells predominate, with [gamma][delta] T cells constituting only a small proportion (1% to 5%) of the total T-lymphocyte pool. The distributions of [alpha][beta] and [gamma][delta] T cells are distinct, with the former comprising more than 95% of peripheral blood (PB) T cells, and the latter preferentially found in the spleen and epithelial-rich tissues, such as skin, intestine, and reproductive tract.2,3 In addition, [gamma][delta] T cells have functional capabilities that distinguish them from [alpha][beta] T cells, including rapid chemokine and cytokine secretion and the capacity for major histocompatibility complex-independent antigen recognition. (4-7) Although the precise function of [gamma][delta] T cells remains unclear, based on the distinctive properties of these cells, they are postulated to play a sentinel role in both innate and antigen-specific mucosal immune responses. (8,9) [gamma][delta] T cells have also been implicated in some autoimmune conditions because increased numbers of [gamma][delta] T cells have been found in the PB of patients with primary sclerosing cholangitis and autoimmune hepatitis. (10)

Because of the relative paucity of circulating [gamma][delta] T cells and the obscure nature of their immune function, few articles have been published regarding their immunophenotype or clinical conditions that may be associated with their increase. To address this issue, the results of flow cytometric immunophenotyping (FCIP) studies in a series of 62 cases with a reactive increase in PB [gamma][delta] T cells were reviewed. Clinical and additional laboratory data were available in 36 (58%) of these 62 cases, and that information was reviewed in detail to determine medical conditions associated with reactive PB [gamma][delta] T-cell lymphocytosis.

MATERIALS AND METHODS

The results of T-cell FCIP studies from 352 consecutive PB specimens analyzed in the Mayo Clinic (Rochester, Minn) clinical flow cytometry laboratory during a 4-month period were reviewed. Cases in which [gamma][delta] T cells comprised either more than 5% of the lymphoid events (forward and side light scatter) or had a calculated absolute count of more than 200/[micro]L, or both, were selected for further study. These criteria for increased circulating [gamma][delta] T cells were generated through comparison to published reference ranges (2,3,11,12) and were chosen because they also allowed for accurate identification of this cell population in all histograms. Cases in which there was an established diagnosis of T-cell malignancy (n = 2) or clonal TCR [gamma]-chain gene rearrangements were detected (n = 2) were excluded. In all cases, a Wright-Giemsastained PB smear was reviewed to evaluate the morphology of the lymphoid cells.

Flow cytometric immunophenotyping was performed according to previously published techniques. (13) Briefly, nucleated cells were isolated through an isotonic lysis procedure, and 4-color FCIP was performed on the isolated cells using fluorochrome-conjugated antibodies to antigens associated with T cells and natural killer (NK) cells in the following combinations: tube 1, CD3-fluorescein isothiocyanate, CD16-phycoerythrin; tube 2, CD2fluorescein isothiocyanate, CD4-peridinin chlorophyll-[alpha] protein, CD5-allophycocyanin, CD8-phycoerythrin; and tube 3: CD3-allophycocyanin, CD4-peridinin chlorophyll protein, CD7-fluorescein isothiocyanate, and [gamma][delta] TCR-phycoerythrin. In 8 (13%) of the 62 cases, additional 3-color FCIP studies had been performed using antibodies to the cytotoxic T-cell-associated and NK-cell-associated antigens CD56, CD57, CD94, and CD161; and to the killing inhibitory receptors (KIRs) CD158a, CD158b, and CD158e (p70). (13) All antibody conjugates were from BD Biosciences (San Jose, Calif), with the exception of the anti-CD94 antibody conjugate (Ancell Corporation, Bayport, Minn) and the anti-CD158b antibody conjugate (BD Biosciences Pharmingen, San Diego, Calif). These data were collected on a FACSCalibur instrument (BD Biosciences) and analyzed using CellQuest Pro software (BD Biosciences). TCR [gamma]-chain rearrangements were evaluated by a polymerase chain reaction method previously described. (14)

Clinical information was obtained from review of the medical records, which were available in 36 of the study group cases. As a case control group, the flow cytometry data and medical records from 37 consecutively analyzed PB FCIP cases with normal T-cell phenotypes, which did not meet either criteria for increased [gamma][delta] T cells, were reviewed. All the patients consented to the use of their medical records for research, and the study was approved by the Mayo Clinic Institutional Review Board.

RESULTS

Sixty-two (18%) of 352 consecutively analyzed PB specimens had an increase in [gamma][delta] T cells sufficient to meet criteria for study inclusion (Table 1). The total lymphocyte count was elevated more than 2900/[micro]L in 29 (47%) of these 62 cases; in all cases, PB smear review failed to reveal either cytologically malignant lymphoid cells or a definite increase in large granular lymphocytes. Twenty-eight (45%) of the 62 study cases met both criteria for inclusion because their [gamma][delta] T cells comprised more than 5% of all lymphocytes and they had an absolute count greater than 200/[micro]L blood. Another 28 of the included cases met only the criterion of more than 5% [gamma][delta] T cells, and in 6 (10%) of the 62 cases, the absolute [gamma][delta] T cell count was more than 200 cells/[micro]L yet comprised less than 5% of all lymphocytes.

Among all 62 cases that met criteria for study inclusion, a relatively wide range was observed for both the percentage of [gamma][delta] T cells and the absolute [gamma][delta] T-cell counts (Table 1). In most cases (n = 43; 69%), the absolute [gamma][delta] T-cell count was less than 10% of the absolute total lymphocyte count; [gamma][delta] T cells comprised between 10% and 20% of the total lymphocytes in 14 (22%) of the 62 cases and more than 20% in 5 (8%) of the cases. Similarly, the absolute [gamma][delta] T-cell count was less than 500/[micro]L blood in most cases (n = 57; 92%). In the remaining cases (5/62; 8%), the absolute [gamma][delta] T-cell count was between 500 and 1000/[micro]L blood; none of the cases had more than 1000 78 T cells per [micro]L. The median values for both the percentage of [gamma][delta] T cells and the absolute [gamma][delta] T-cell counts were near the cutoffs established for determining an increase in these cells, further indicating that, in most cases, the increase in [gamma][delta] T cells was modest.

The results of FCIP studies using antibodies to the antigens associated with T cells and NK cells are summarized in Table 2. In all cases, the [gamma][delta] T cells were CD3+ (Figure 1, A). These cells were uniformly CD4-; they also appeared CD8-, although, in some cases, the histograms suggested dim expression of this antigen. In most cases (n = 42; 68%), the [gamma][delta] T cells had unremarkable expression of T-cell-associated antigens. In the remaining 20 (32%) of 62 cases, the [gamma][delta] T cells showed distinctive patterns of T-cell-associated antigen expression when compared with the [alpha][beta] T cells in the sample. Among this latter group, differences in CD5 were most common with either partial absence (4/20; 20%) or complete absence (11/20; 55%) of expression of this antigen by the [gamma][delta] T cells seen (Figure 1, C and D; Table 2). In 6 (10%) of 62 cases, the [gamma][delta] T cells showed relatively bright CD3 expression (Figure 1, A through C). In most cases, alterations in T-cell antigen expression occurred singly; however, in minor subsets, the diminished CD5 expression was associated with either bright expression of CD3 (n = 3; 15%) or diminished expression of CD7 (n = 2; 10%). In a single case (5%), the [gamma][delta] T cells showed both bright CD3 expression and diminished expression of both CD5 and CD7. No correlation between the presence of altered expression of one or more T-associated antigens and the absolute [gamma][delta] T-cell count was observed.

[FIGURE 1 OMITTED]

A group of 37 cases, with essentially normal T-cell phenotypic findings and lacking an increase in [gamma][delta] T cells (median percentage of [gamma][delta] T cells in absolute lymphocyte count, 2.15; range, 0.2-4.9; median absolute [gamma][delta] T-cell count per [micro]L, 51.6; range, 3.4-182.2), was also obtained as control for clinical findings associated with increased circulating [gamma][delta] T cells. Review of the phenotyping analyses in this group revealed a single case with a slight increase in CD8+ T cells, which showed slightly diminished CD5 expression (as compared with the [CD4.sup.+] T cells) and normal CD2, CD3, CD7 expression. The [CD8.sup.+] T cells in this single case also showed partial expression of CD57 but lacked expression of other NK-cell-associated antigens, such as CD16, CD56, and CD94, and bone marrow examination failed to reveal evidence of a lymphoproliferative disorder. Another case in this group showed a slight increase in NK cells. None of these control cases with low numbers of [gamma][delta] T cells contained a phenotypically distinct cell population. Specifically, no cases with either definitive coexpression of CD16 or loss of CD5 by the few [gamma][delta] T cells present were identified in this group. Because of the lack of a distinct T-cell or NK-cell population in these control cases, KIR-expression studies were not performed.

Expression of the NK-cell-associated antigen CD16 was assessed in all study cases. In approximately one third of the cases (n = 20), the [gamma][delta] T cells were [CD16.sup.+] (Figures 1, B, and 2, A through D). Interestingly, the expression of CD16 was frequently associated with altered CD5 expression. In 10 (50%) of the 20 [CD16.sup.+] cases, the [gamma][delta] T cells showed complete absence of CD5, and in an additional 2 (10%) cases, a significant subset of the [gamma][delta] T cells were [CD5.sup.-]. Because both the diminished expression of CD5 and the coexpression of CD16 are associated with a diagnosis of T-cell large granular lymphocytic leukemia (T-LGL) in [alpha][beta] T cells, in 8 (13%) of the 62 cases in which the [gamma][delta] T cells exhibited one or both of these features, additional FCIP studies were performed using antibodies to an extended panel of NK-cell-associated antigens. (13) These studies are part of our standard laboratory evaluation of potential LGL cases and include antibodies to NK-associated major histocompatibility complex receptors, such as the KIRs. The results of these studies are summarized in Table 3. CD56 expression by the [gamma][delta] T cells was detected in only 3 (38%) of the 8 cases. In contrast, CD57 and CD94 expression were frequent; however, in most instances CD94 was only expressed by a subset of the cells (Figure 2, B). The KIRs, CD158a, CD158b, and CD158e (p70), were expressed in all cases; in each case, there appeared to be subsets of the [gamma][delta] T cells that were positive for different KIR antigens (Figure 2, C and D). This pattern of KIR expression has been described in reactive, nonclonal T-cell and NK-cell populations. (15) The uniform expression of one or more KIR antigens, which is associated with clonal T-cell and NK-cell expansions, was not seen in any case. (13,16) Further studies were not performed because the immunophenotyping results indicated the [gamma][delta] T cells were reactive in nature. In 7 (11%) of 62 of the cases studied, T-cell receptor gene rearrangement tests had been ordered by the requesting clinician; in all cases, no clonal TCR gene rearrangements were detected.

[FIGURE 2 OMITTED]

The median age of the 62 subjects was 56 years (range, 1-89 years), and most (56/62; 90%) subjects were either between 19 and 50 years old (28/62; 45%) or between 51 and 75 years old (28/62; 45%). Additional clinical information was available for 36 subjects; the data are summarized in Table 4. These case history reviews were comprehensive; however particular emphasis was placed on conditions associated with immune system activation and prior splenectomy because these conditions have been described (2,13,17-20) as altering both the number and phenotype of PB [gamma][delta] T cells. In most cases (28/36, 78%), an associated clinical condition was present, and in 7 (25%) of these 28 cases, more than one condition was identified. Most often the increase in PB [gamma][delta] T cells was associated with either infectious/inflammatory disorders (18/28; 64%) or autoimmune diseases (9/28; 32%). The infectious/inflammatory disorders were found almost equally in women (10/ 18; 56%) and men (8/18; 44%). Most subjects with autoimmune disorders (8/9; 89%) were women. In 6 (21%) of the 28 cases, there was a history of a hematolymphoid disorder. Prior splenectomy was documented in 3 (11%) of the 28 cases. In 8 (22%) of 36 cases, no clinical features associated with the observed increase in PB [gamma][delta] T cells could be gleaned from the records. In this latter group, FCIP studies were ordered for follow-up of leukocytosis (n = 2), pancytopenia (n = 2), microhematuria (n = 1), chronic anemia (n = 1), and mild leukopenia (n = 1). In 1 case, there was no indication for FCIP in our records. No correlation was identified between the phenotype of [gamma][delta] T cells and clinical findings, and no statistically significant differences in number or percentage of circulating [gamma][delta] T cells were identified between cases with infectious/ inflammatory disorders, autoimmune diseases, or history of splenectomy and cases without any of these clinical findings.

To determine whether the observed clinical conditions were specifically associated with increased PB [gamma][delta] T cells, the clinical records from the control group of 37 consecutively analyzed PB cases with T-cell phenotyping studies (those that did not meet the criterion for increased [gamma][delta] T cells) were reviewed. The median age and the distribution of men to women in the control group were similar to those of the study cases. In contrast to the cases with increased [gamma][delta] T cells, review of the records revealed identifiable clinical conditions in relatively few (n = 13; 35%) of the control cases. Similar to the study group, however, the associated conditions most often identified in the control group were infectious/inflammatory disorders (8/13; 62%). These disorders were composed of a single case each of the following: urinary tract infection, upper respiratory infection, sinusitis, pancolitis, bacterial infection, pancreatitis, sacroiliitis of undetermined etiology, and hepatitis C. Other conditions identified in the controls included lymphoproliferative/myeloproliferative disease (n = 3), autoimmune disease (n = 2) and prior splenectomy (n = 1). In contrast to the study group, none of the controls had more than one associated condition. The medical records of the remaining 24 control cases did not contain any of these conditions or other associated findings.

COMMENT

This study was undertaken to determine how frequently increased PB [gamma][delta] T cells are encountered in a clinical flow cytometry laboratory, to characterize the immunophenotypic attributes of these cell populations, and to identify clinical conditions that may be associated with this laboratory result. Prior studies11,12 in healthy adults have indicated that [gamma][delta] T cells comprise less than 5% of the total PB T-cell pool, with an absolute range of 55 to 120 [gamma][delta] T cells per [micro]L. Based on these data, thresholds of [gamma][delta] T cells comprising either more than 5% of the total events in the lymphocyte gate (forward and side light scatter) or 200 cells per [micro]L were established for inclusion in this study. These levels also represented the threshold at which the [gamma][delta] T cells could be identified with confidence in all the data histograms. Using these criteria, [gamma][delta] T cells were increased in 62 (18%) of 352 PB specimens studied. Although this proportion of cases was somewhat higher than expected, the included study cases were considered valid because the inclusion criteria were derived from available reference range control data.

The detection of altered or atypical patterns of antigen expression is an important element in determining whether a lymphocyte population detected by flow cytometry is found to be abnormal and, therefore, possibly indicative of a lymphoproliferative disorder. [gamma][delta] T cells have previously been demonstrated (15,20-22) to have unique phenotypic attributes, which could make them appear abnormal in comparison to [alpha][beta] T cells. Indeed, the [gamma][delta] T cells in approximately one third of the cases in this study had patterns of T-cell-associated antigen expression that distinguished them from the [alpha][beta] T cells present. These features included distinctively bright CD3 staining of the [gamma][delta] T cells (n = 6), a finding not unexpected because [gamma][delta] T cells usually have a 2-fold higher surface density of TCR-CD3 expression than [alpha][beta] T cells. (21,22)

The most commonly encountered distinguishing immunophenotypic feature of the [gamma][delta] T cells was diminished or absent expression of CD5. This pattern of CD5 expression (or lack thereof), which has been described as an immunophenotypic aberrancy in [gamma][delta] T-cell neoplasms such as hepatosplenic T-cell lymphoma, (23) is also present in nonneoplastic [gamma][delta] T-cell populations, particularly those expressing the V[delta]1 variable region gene. (20) All [gamma][delta] T cells express either V[delta]1 or V[delta]2; in healthy PB, the V[delta]2 cells, which immunophenotypically closely resemble [alpha][beta] T cells, predominate. However, an increase in PB [CD5.sup.-], V[[delta]1.sup.+] [gamma][delta] T cells has been described (20) in reactive states, including human immunodeficiency virus infection. As V[delta]1 or V[delta]2 expression was not assessed in this study, it is unknown whether the observed alterations in CD5 expression were attributable to an increase in V[delta]1 cells. Furthermore, this phenotype did not correlate with any of the associated clinical conditions, although no cases of human immunodeficiency virus were included in the cadre of studied patients.

Coexpression of the NK-cell-associated antigen CD16 by the [gamma][delta] T cells was detected in 20 (32%) of the 62 study cases. An early study by Groh et al (24) described CD16 coexpression by a subset of healthy [gamma][delta] T cells in 1 (20%) of 5 analyzed cases. This slightly lower frequency of CD16 expression described in this prior work may be attributable to the small number of cases analyzed as well as to the use of older methodology, which may have had lower antigen-detection sensitivity. Interestingly, we noted an association between [gamma][delta] T-cell CD16 positivity and both CD57 coexpression and diminished expression of CD5. As noted above, diminished expression of CD5 by PB [gamma][delta] T cells is more commonly seen in the V[delta]1 subset. This [gamma][delta] T-cell subset has also been described as frequently being [CD57.sup.+] . Hence, the association of diminished expression of CD5 and coexpression of CD16 and CD57 observed in our cases with a reactive increase in [gamma][delta] T cells provides additional, albeit indirect, evidence to suggest that the observed changes may be attributable to an increase in V[[delta]1.sup.+] cells.

Both coexpression of NK-associated antigens, such as CD16 and CD57, and diminished expression of CD5 are features also associated with T-LGL. (13) For this reason in 8 of the cases with increased [gamma][delta] T cells in which one or both of these features was present, additional flow cytometry studies were performed using antibodies to NK-cell-associated antigens, including the NK-cell receptors to further pursue this possibility. The NK-cell receptors are a group of receptors for class I major histocompatibility complex and related antigens, which include CD94/NKG2 complexes, as well as the KIRs CD158a, CD158b, and CD158e. Analysis of these antigens is helpful in diagnosing both T-LGL and hepatosplenic T-cell lymphoma (13,25,26) because, in T-LGL, uniform expression of a single KIR antigen is detected in approximately one half of cases, and in hepatosplenic T-cell lymphoma, uniform expression of multiple KIR antigens is often seen. In contrast to these malignancy-associated KIR expression patterns, in the 8 study cases in which NK-cell receptor FCIP studies were performed, only minor subsets of the reactive [gamma][delta] T cells were positive for each of the KIR antigen. Also, CD94 was often expressed by the reactive [gamma][delta] T cells; that expression and the polytypic KIR expression pattern by a minor cell subset are consistent with previously reported (27) expression patterns of these antigens by healthy PB [gamma][delta] T cells. These FCIP results, in conjunction with the absence of associated cytopenias or demonstrable clonal TCR gene rearrangements, essentially excluded a diagnosis of T-LGL in all instances.

Although the precise role of [gamma][delta] T cells in global immune function remains unclear, they have been implicated in a variety of immune responses, such as responses to mycobacterial pathogens and the pathogenic immune responses of autoimmune diseases, (13,17,18,28) celiac disease, (29) and multiple sclerosis.30 In keeping with these known stimulators of the [gamma][delta] T-cell compartment, review of the clinical records revealed that the observed increase in [gamma][delta] T cells was frequently associated with activation of the immune system, by either infection (50% of cases) or autoimmune disease (25% of cases). The number of study cases with these associated conditions was 2-fold and 5-fold greater, respectively, than in the matched control group. [gamma][delta] T cells have also been implicated in tumor immunity, which may account for the association of an increase in these cells with a previous diagnosis of a nonT-cell hematolymphoid neoplasm identified in 6 study cases, almost twice the number identified in the control group. (31,32) Also, prior splenectomy was present in 3 of the study cases; in contrast, only a single splenectomized patient was found in the control group. This difference was not unexpected given that up to 30% of splenic lymphocytes are [gamma][delta] T cells, and an increase of PB [gamma][delta] T cells following splenectomy has been previously described. (13)

CONCLUSIONS

An increase in PB [gamma][delta] T cells is not an uncommon finding in FCIP studies. These increases are typically slight and can be seen in response to a wide variety of conditions, most of which are associated with acute or chronic activation of the immune system. Although these cells typically remain a minor component of the total T-cell pool, it is important to recognize that reactive increases in [gamma][delta] T cells can occur because these cells may exhibit patterns of T-cell antigen expression that mimic those seen in T-cell lymphoproliferative disorders, such as T-LGL or other T-cell malignancies. For this reason, determining the pathologic significance of a detectable increase in PB [gamma][delta] T cells requires correlation with other clinical and laboratory features.

Accepted for publication April 2, 2008.

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Anja C. Roden, MD; William G. Morice, MD, PhD; Curtis A. Hanson, MD

From the Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minn.

The authors have no relevant financial interest in the products or companies described in this article.

Reprints: Anja C. Roden, MD, Department of Laboratory Medicine and Pathology, Mayo Clinic, Hilton 11, 200 First St SW, Rochester, MN 55905 (e-mail: roden.anja@mayo.edu).
Table 1. Lymphocyte Counts in Patients With Increased Circulating
[gamma][delta] T Cells

78 T-Cell Inclusion Criteria Met (n = 62) *

                            >5% of ALC and
Test Result                 >200/[micro]L          >5% of ALC Only

No. (%) of patients            28 (45)               28 (45)
in group
% Lymphocyte, median         41.1 (9.5-96.0)       34.5 (4.2-63.0)
(range) ([dagger])
ALC (cells/[micro]L),        3500 (800-8200)       1350 (300-3600)
median (range)
% T cells (of ALC),          78.4 (56.6-97.2)      78.7 (57.3-92.5)
median (range)
Absolute T-cell count        2600 (800-5900)       1100 (200-2600)
(cells/[micro]L),
median (range)
% [gamma][delta] T           12.1 (5.3-32.3)        8.4 (5.5-42.2)
cells (of absolute
T-cell count), median
(range)
Absolute [gamma][delta]     345.8 (203.0-641.9)   115.1 (37.5-198)
T-cell count (cells/
[micro]L), median (range)

Test Result                 >200/[micro]L Only

No. (%) of patients             6 (10)
in group
% Lymphocyte, median         60.2 (49.0-85.0)
(range) ([dagger])
ALC (cells/[micro]L),        8350 (4700-10 100)
median (range)
% T cells (of ALC),          81.5 (62.5-90.1)
median (range)
Absolute T-cell count        6100 (4000-9100)
(cells/[micro]L),
median (range)
% [gamma][delta] T            4.6 (2.7-5.7)
cells (of absolute
T-cell count), median
(range)
Absolute [gamma][delta]     234.1 (203.5-369.6)
T-cell count (cells/
[micro]L), median (range)

* ALC indicates total absolute lymphocyte count; >5%, T cells
comprised >5% of ALC; >200/[micro]L, absolute T-cell count was >200 X
106 cells/[micro]L.

([dagger]) Percentage of lymphocytes in total white blood cell count.

Table 2. Alterations of T-Cell--Associated Antigen
Expression in Reactive [gamma][delta] T Cells (n = 62)

                                           Diminished   Bright
          Complete Loss,   Partial Loss,   Intensity,   Intensity,
Antigen      No. (%)          No. (%)       No. (%)     No. (%)

CD2           0                0              0           0
CD3           0                0              0           6 (10)
CD5          11 (18)           4 (6)          1 (2)       1 (2)
CD7           0                0              3 (5)       0

Table 3. Patterns of CD5 and Natural Killer Cell-Associated Antigen
Expression in Reactive, Circulating [gamma][delta] T Cells (n = 8) *

Case   CD5       CD16   CD57      CD94       KIR (%)

 1     Pos       Neg    Pos       Pos        Partial (17), polytypic
 2     Partial   Neg    Pos       Partial    Partial (7), polytypic
 3     Partial   Pos    Pos       Pos        Partial (16), polytypic
 4     Pos       Neg    Neg       Partial    Partial (11), polytypci
 5     Neg       Pos    Partial   Partial    Partial (3), polytypic
 6     Neg       Neg    Pos       Neg        Partial (4), polytypic
 7     Pos       Neg    Partial   Partial    Partial (13), polytypic
 8     Pos       Neg    Partial   Partial    Partial (15), polytypic

       [gamma]][delta]T-Cell Count,
Case      No. of Cells/[micro]L

 1                 183.6
 2                 641.9
 3                 334.9
 4                 518.3
 5                 232.3
 6                 580.5
 7                 395.0
 8                 478.9

* KIR (%) indicates percentage of [gamma][delta] T cells expressing
killing inhibitory receptors; [gamma][delta] T-cell count, absolute
number; pos, positive expression; neg, not expressed; polytypic,
subset of cells positive for CD158a, CD158b, and CD158e; and partial,
subset is positive.

Table 4. Associated Clinical Features in Subjects
With Increased Circulating [gamma][delta] T Lymphocytes * and
Control Subjects ([dagger])
                                                     Study     Control
                                                     Group,    Group,
Clinical Conditions                                  No. (%)   No. (%)

Infection/inflammatory disorders ([double dagger])   18 (50)    8 (21)
Autoimmune disease ([section])                        9 (25)    2 (5)
Lymphoproliferative disorder ([parallel])             6 (17)    4 (10)
Splenectomy                                           3 (8)     1 (2)
None identified                                       8 (22)   24 (62)

* n = 36; in 7 study subjects multiple associated clinical findings
were present, so column total > 36.

([dagger]) n = 39.

([double dagger]) Includes parasitic infestations (positive serology
or culture; n = 3), fungal infection (positive culture; n = 2),
presumed viral infections (n = 2), upper respiratory tract
infections (n = 4), and chronic sinusitis, granulomatous
panniculitis, pancreatitis, chronic hepatitis C, hepatitis
B, fever of unknown origin, and psoriasis (n = 1 each).

([section]) Includes rheumatoid arthritis (n = 2) and immune
thrombocytopenic purpura, ankylosing spondylitis, Sjogren
syndrome, celiac disease, Graves disease, multiple sclerosis,
pernicious anemia, and autoimmune-induced pancytopenia (n = 1 each).

([parallel]) Includes Hodgkin lymphoma, diffuse large B-cell
lymphoma, lymphomatoid papulosis, myelodysplastic syndrome,
monoclonal gammopathy of uncertain significance, and orbital
inflammatory pseudotumor (n = 1 each).
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