Purpose To determine the phenotype and functional characteristics of type I collagen (CI)-specific T cells in scleroderma (SSc).
Methods Peripheral blood mononuclear cells (PBMC) were collected from SSc patients and normal controls. CFSE labeling and flow cytometry were used to follow antigen-specific T cell proliferation in response to bovine CI and β1,2 chain of CI. Phenotypic analysis and intracellular cytokine expression of proliferating cells was performed by multicolor flow cytometry and CI-specific T cell lines were generated by repeated stimulation of PBMC with CI and autologous PBMC. T cell line supernatants were assayed for cytokines using a multiplex cytokine bead array system. Normal human fibroblasts were then cultured with CI-specific T cell supernatants and CI (α2) gene transcription in fibroblasts was determined by real-time PCR.
Results 25 patients with SSc and 18 normal controls were studied. A T-cell proliferative response to CI was detected in 8 (32%) patients and in none of the controls (p = .01). Among responders, the proliferating T cell population was invariably CD3+CD4+. The CI-specific, CD4+ T cells expressed an activated, memory phenotype (CD25+CD45RO+) but did not stain for CD212. To confirm our findings, we generated CI-specific T cell lines that had the following phenotype: CD3+CD4+CD8-CD28dimCD49a+. Following activation of CI-specific T cells we detected intracellular interferon-γ but not IL-4 by flow cytometry. Supernatants from the T cell lines contained IL-2, IFN-γ, GM-CSF and TNF-α, but little or no IL-4 and IL-10, suggesting that CI-specific cells express a Th1 cytokine pattern. Moreover, supernatant from CI-specific T cells was able to suppress CI gene transcription in normal human fibroblasts.
Conclusion Circulating CI-specific memory CD4 T cells that have a Th1 cytokine profile are present in a subset of patients with SSc. These T cells may play a role in limiting fibrosis. Therefore, in vivo expansion of this T cell subset may be of therapeutic benefit in SSc. Supported by a grant from the Scleroderma Foundation and the Research Center of Excellence for Connective Tissue Disease, UTHSC.