Detection of clonally rearranged T-cell-receptor gamma chain genes from T-cell malignancies and acute inflammatory rheumatic disease using PCR amplification, PAGE, and automated analysis.
Clonal expansions of T cells carrying identical T-cell-receptor (TCR) genes are the hallmark of T-cell malignancies, but they can also result from a strong immune reaction to a dominant epitope. The basis for the molecular detection of clonal T cells is amplification of the V-(D)-N-J region of the TCR gene. We evaluated PCR amplification of the rearranged gamma TCR from genomic DNA extracted from peripheral blood and subsequent polyacrylamide gel electrophoresis (PAGE) in an automated DNA sequencer. We determined the sensitivity for the detection of clonal T cells and propose a standardized evaluation procedure for the electrophoretic profiles generated by the DNA sequencer. The sensitivity of our method was 0.6-1.25% of clonal T cells within a polyclonal background. Sixteen patients with T-cell malignancies, ten with acute inflammatory rheumatic diseases, and twelve healthy controls were examined. Among the systemic T-cell malignancies, all but one patient with T-PLL (8/ 9) revealed a clonal PCR signal. No clonal signal was detectable in any patient in clinical complete remission (5/5) or in either of the two patients with lymphomas limited to cutaneous sites. However, clonal T cells were detected in one patient with polymyalgia rheumatica and in one with reactive arthritis. A polyclonal signal was found in the remaining eight patients with acute inflammatory rheumatic diseases and in 12 healthy controls. Taking our results together, the PCR/PAGE assay is able to reliably distinguish clonal from polyclonal T-cell populations. However, although the sensitivity is limited to approximately 1%, clonal T cells can be found in the peripheral blood of some patients with autoimmune diseases and not only in T-cell malignancies.
Witzens
M
M
Department of Internal Medicine V (Hematology, Oncology, Rheumatology), University of Heidelberg, Germany.
Möhler
T
T
Willhauck
M
M
Scheibenbogen
C
C
Lee
K H
KH
Keilholz
U
U
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D016918:Arthritis, Reactive / Q000097:blood / Q000235:genetics; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005784:Gene Amplification; D015334:Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor; D006801:Humans; D007119:Immunoblastic Lymphadenopathy / Q000097:blood / Q000235:genetics; D019169:Jurkat Cells; D015458:Leukemia, T-Cell / Q000097:blood* / Q000235:genetics*; D008180:Lupus Erythematosus, Systemic / Q000097:blood / Q000235:genetics; D016399:Lymphoma, T-Cell / Q000097:blood* / Q000235:genetics*; D008875:Middle Aged; D009182:Mycosis Fungoides / Q000097:blood / Q000235:genetics; D015434:Panniculitis / Q000097:blood / Q000235:genetics; D016133:Polymerase Chain Reaction; D011111:Polymyalgia Rheumatica / Q000097:blood / Q000235:genetics; D016692:Receptors, Antigen, T-Cell, gamma-delta / Q000235:genetics*; D012216:Rheumatic Diseases / Q000097:blood* / Q000235:genetics*; D012751:Sezary Syndrome / Q000097:blood / Q000235:genetics
D016692:Receptors, Antigen, T-Cell, gamma-delta