Characteristics of T-cell large granular lymphocyte proliferations associated with neutropenia and inflammatory arthropathy

Abstract

Introduction: The purpose of this study was to analyze the data of patients with T-cell large granular lymphocyte (T-LGL) lymphocytosis associated with inflammatory arthropathy or with no arthritis symptoms.

Methods: Clinical, serological as well as histopathological, immunohistochemical, and flow cytometric evaluations of blood/bone marrow of 21 patients with T-LGL lymphocytosis were performed. The bone marrow samples were also investigated for T-cell receptor (TCR) and immunoglobulin (IG) gene rearrangements by polymerase chain reaction with heteroduplex analysis.

Results: Neutropenia was observed in 21 patients, splenomegaly in 10, autoimmune diseases such as rheumatoid arthritis (RA) in 9, unclassified arthritis resembling RA in 2, and autoimmune thyroiditis in 5 patients. T-LGL leukemia was recognized in 19 cases. Features of Felty syndrome were observed in all RA patients, representing a spectrum of T-LGL proliferations from reactive polyclonal through transitional between reactive and monoclonal to T-LGL leukemia. Bone marrow trephines from T-LGL leukemia patients showed interstitial clusters and intrasinusoidal linear infiltrations of CD3+/CD8+/CD57+/granzyme B+ lymphocytes, reactive lymphoid nodules, and decreased or normal granulocyte precursor count with left-shifted maturation. In three-color flow cytometry (FCM), T-LGL leukemia cells demonstrated CD2, CD3, and CD8 expression as well as a combination of CD16, CD56, or CD57. Abnormalities of other T-cell antigen expressions (especially CD5, CD7, and CD43) were also detected. In patients with polyclonal T-LGL lymphocytosis, T cells were dispersed in the bone marrow and the expression of pan-T-cell antigens in FCM was normal. Molecular studies revealed TCRB and TCRG gene rearrangements in 13 patients and TCRB, TCRG, and TCRD in 4 patients. The most frequently rearranged regions of variable genes were Vbeta-Jbeta1, Jbeta2 and Vgamma If Vgamma10-Jgamma. Moreover, in 4 patients, additional rearrangements of IG kappa and lambda variable genes of B cells were also observed.

Conclusion: RA and neutropenia patients represented a continuous spectrum of T-LGL proliferations, although monoclonal expansions were most frequently observed. The histopathological pattern and immunophenotype of bone marrow infiltration as well as molecular characteristics were similar in T-LGL leukemia patients with and without arthritis.

Figure 1

Histopathological features of bone marrow in patients with arthritis and T-cell large granular lymphocyte (T-LGL) lymphocytosis. (a) Patient 1 with rheumatoid arthritis (RA) and T-LGL leukemia. Staining for CD57 demonstrates intrasinusoidal linear arrays and interstitial clusters of T cells (EnVision stain, ×100). (b) Granzyme B highlights cytotoxic granules in these cells (EnVision stain, ×200). (c) Patient 10 with polyclonal T-LGL lymphocytosis. Staining for CD8 shows dispersed T cells (EnVision stain, ×200). (d) Patient 9 with unclassified arthritis, T-LGL leukemia, and IGKV and IGLV gene rearrangements. CD3 staining shows interstitial and nodular infiltration of T cells (EnVision stain, ×100). (e) Patient 9. The lymphoid nodule contains few CD20⁺ B cells (EnVision stain, ×200). (f) Patient 7 with RA and T-LGL leukemia. A decreased count of granulocytic precursors (myeloperoxydase⁺) is shown (EnVision stain, ×200). IGKV, immunoglobulin kappa variable; IGLV, immunoglobulin lambda variable

Figure 2

Flow cytometric analyses of patient 15 with T-cell large granular lymphocyte leukemia. (a) CD8⁺CD4^- leukemic cells. (b) CD3⁺/CD45RA⁺ leukemic cells. (c) CD2⁺/CD7^- leukemic cells and double-stained population in the region R2 consistent with normal T lymphocytes. (d) CD5^-/CD25^- leukemic cells and CD5⁺/CD25^-/+ expression on normal T lymphocytes in the region R3. (e) CD7^- leukemia cells express slightly weaker levels of CD43 compared with normal CD43^+higherCD7⁺ cells consistent with normal T lymphocytes in the region R2. (f) CD3⁺ population with coexistence of CD16 antigens. (g) CD3⁺CD56^- leukemic cells. (h) CD2⁺CD57^- leukemic cells. CD2⁺CD57⁺-reactive T lymphocytes in the region R2. (i) Leukemic cells are positive for CD3 and TCRαβ. FITC, fluorescein isothiocyanate; PE, phycoerythrin; TCR, T-cell receptor.

Figure 3

Ethidium bromide-stained polyacrylamide gel showing polymerase chain reaction products derived from TCR gene rearrangements in patients with rheumatoid arthritis and T-cell large granular lymphocyte (T-LGL) proliferations. (a) Polyclonal expansion of T-LGLs in patient 10. Lane 1: TCRB gene rearrangement–negative, polyclonal smear (tube A); lane 2: TCRB gene rearrangement-negative, polyclonal smear (tube B); lane 3: TCRB gene rearrangement-negative, polyclonal smear (tube C); lane 4: standard 50 base pairs (bp); lane 5: TCRG gene rearrangement-negative, polyclonal smear (tube A); lane 6: TCRG gene rearrangement-negative, polyclonal smear (tube B); and lane 7: TCRD gene rearrangement-negative, polyclonal smear. (b) Monoclonal expansion in polyclonal background in patient 7. Lane 1: TCRG gene rearrangement: monoclonal product 180 bp (i) in tube A; lane 2: TCRG gene rearrangement: monoclonal product 210 bp (ii) in polyclonal background (tube B); lane 3: TCRD gene rearrangement: monoclonal product 160 bp (iii); lane 4: TCRB (tube A) gene rearrangement-negative, polyclonal smear; lane 5: standard 50 bp; lane 6: TCRB (tube B) gene rearrangement-negative, polyclonal smear; and lane 7: TCRB (tube C) gene rearrangement-negative, polyclonal smear. (c) Monoclonal gene rearrangements in patient 1 with T-LGL leukemia. Lane 1: TCRB gene rearrangement-negative (tube A); lane 2: TCRB gene rearrangement-positive, monoclonal product 250 bp (iv) in tube B; lane 3: TCRB (tube C): gene rearrangement-negative, polyclonal smear; lane 4: standard 50 bp; lane 5: TCRG gene rearrangement-positive, monoclonal product 230 bp (v) in tube A; lane 6: TCRG gene rearrangement-positive, monoclonal product 180 bp (vi) in tube B; and lane 7: TCRD gene rearrangement-negative, polyclonal smear. TCR, T-cell receptor.