T-cell clonality in synovial fluid from rheumatoid joints before and after culture in interleukin-2.

T-cell receptor (TCR) gamma gene rearrangements which have been amplified in polymerase chain reactions (PCRs) and analysed by high resolution polyacrylamide gel electrophoresis have been used to investigate the clonal diversity of T-cells in joint effusions from 16 patients with rheumatoid arthritis, one with systemic lupus erythematosus and one with psoriatic arthropathy. Polyclonality was found in every case but an oligoclonal subset of dominant rearrangements was also demonstrated in all but the patient with psoriasis. Marked changes in the relative preponderance of the various clonotypes were observed in 29 of 48 paired tests from 12 cases before and after culture in media containing interleukin-2 (IL-2) showing that SF mononuclear cells cultured in vitro with IL-2 are not representative of those present in vivo.

An immunohistological study of cutaneous lymphocytes in vitiligo.

In a study of 49 biopsies from the margins of depigmented cutaneous lesions in 18 patients with vitiligo, highly significant overall increases were found in CD3+, CD4+, and CD8+ T cells, though cell numbers in individual cases were often within the normal range. Many of the T cells were activated (MHC class II+, interferon gamma+), of CD45RO (UCHL1+) memory subset, and many expressed the cutaneous lymphocyte-associated antigen (HECA-452+) typical of skin-homing T cells. Immunohistologically, the most intense epidermal T-cell infiltration was present within 0.6 mm of the edge of the lesion in 10 of 13 double-stained sections with a clearly defined zone of melanocyte depletion. In 40 lesions from 17 patients seen 11-64 weeks after biopsy, no apparent association was found between T-cell numbers and disease activity as assessed by Köbnerization of biopsy wounds or spread of depigmentation. These findings are consistent with the hypothesis that lesional T cells rather than circulating antimelanocytic antibody may be responsible for the supposedly autoimmune but characteristically patchy destruction of cutaneous melanocytes in vitiligo. Nevertheless, many of the infiltrating T cells are probably innocent bystanders attracted by upregulated cell adhesion molecules near sites of melanocyte damage.

Abnormal expression of MHC class II and ICAM-1 by melanocytes in vitiligo.

The relationship between damage to cutaneous melanocytes and antimelanocyte autoimmunity in vitiligo is unclear. We have demonstrated abnormal expression of MHC class II molecules by perilesional melanocytes in 13/21 patients with vitiligo and a six-fold increase in the number expressing the intercellular adhesion molecule ICAM-1. These molecules have important roles in normal antigen presentation and activation of helper T lymphocytes, and their expression by melanocytes may contribute to the abnormal immune response in vitiligo. MHC class II is not expressed by melanocytes in psoriasis and is unlikely to be induced in vitiligo by cytokines released from activated non-melanocyte-specific T lymphocytes.

A sensitive method of screening for dominant T cell clones by amplification of T cell gamma gene rearrangements with the polymerase chain reaction.

A sensitive method of screening for dominant T cell clones in small samples of DNA has been developed using the polymerase chain reaction to amplify and identify T cell gamma receptor gene rearrangements. It can detect such rearrangements in nanogram quantities of DNA from cultured T cell clones, even in the presence of 20-100 parts of polyclonal lymph node DNA, and works with DNA extracted from paraffin sections of cloned T cells which have been fixed in formalin. Presumptive clonal reactions have been obtained in preliminary tests on 8 of 10 unfixed T cell lymphomas but in 0 of 10 reactive lymph nodes.

Hypothesis: symmetrical cutaneous lymphoma.

Patients with malignant lymphoma may have cutaneous and subcutaneous involvement that exhibits a striking symmetry about the coronal axis. The symmetry of these lesions may be caused by site-specific migration from the circulation, preferential proliferation by lymphocytes of the neoplastic clones at defined anatomical sites, or both mechanisms. Similar behaviour by benign lymphocytes may explain the symmetry and selective anatomical distribution of lesions in other skin diseases.

A strategy for demonstrating the clonal origin of small numbers of T lymphocytes in histopathological specimens.

A strategy is described for detecting large T-cell clones among small numbers of lymphoid cells in fresh or formalin-fixed tissue samples using the polymerase chain reaction (PCR) to amplify and identify rearrangements of the V and J genes of the T-cell gamma receptor. Following hybridization of primers with gene-specific sequences at judiciously selected locations on each of the eight potentially active V and five potentially active J genes, the PCR can theoretically amplify DNA segments which span the join between rearranged V and J genes and are of approximately 384 different sizes, each segment size reflecting different gamma gene clonal rearrangements. Large monoclonal populations of T lymphocytes, indicated by excessive amounts of particular PCR segment sizes, can be further characterized by direct nucleotide sequencing of the hypervariable N regions of these segments, and the presence of such clones can be confirmed directly in tissue sections by in situ hybridization with N region-specific oligonucleotide probes.

Epithelial antigens in malignant mixed Müllerian tumours of endometrium.

Immunohistochemical staining of formalin fixed paraffin sections with the mouse monoclonal antibodies E29 and CAM 5.2 effectively distinguish epithelium (antigen positive) from stroma (antigen negative) in normal endometrium. These antibodies were used as epithelial markers in the study of eight malignant mixed Müllerian tumours (MMT) of endometrium and gave normal reactions with well differentiated neoplastic glands; in contrast, negative or very weak staining was observed in poorly differentiated epithelial cells, present in large numbers in three cases. Abnormal antigen-containing cells were observed in the stroma of seven MMT. In some cases these are probably due to stromal invasion by malignant epithelium but in others they may represent an early stage of epithelial differentiation in mesenchymal cells of the malignant stroma.

A family study of vitiligo patterns.

The anatomical distribution of vitiligo has been studied in families with evidence of organ-specific autoimmune disease. No examples of similar pattern inheritance were found in first degree relatives in contrast to published reports of similar vitiligo patterns in identical twins. The genetic predisposition to develop vitiligo apparently allows for a diversity of anatomical pattern. A similar mechanism may be responsible for the occurrence of different organ-specific autoimmune diseases in members of the same family.