DETECTION AND LOCALIZATION OF CYTOKINE IMMUNOREACTIVITY IN RETROOCULAR CONNECTIVE-TISSUE IN GRAVES OPHTHALMOPATHY

Paracrine interactions between fibroblasts residing in the retro-ocular space and infiltrating lymphocytes/macrophages are thought to be of central importance in the pathogenesis of Graves' opthalmopathy (GO). Although various roles have been suggested for interferon-gamma (IFNgamma), tumour necrosis factor-alpha (TNFalpha) and interleukin-1alpha (IL-1alpha) in GO, their actual presence in Graves' retro-ocular connective tissue has not been demonstrated. We examined surgical specimens obtained during orbital decompression from patients with severe GO (n = 6), and from normal individuals (n = 5), for the presence of IFNgamma, TNFalpha and IL-1alpha. We used immunohistochemical methods on frozen tissue sections and primary fibroblast cultures, and sodium dodecylsulfate polyacrylamide-gel electrophoresis of tissue extracts and tissue culture supernatants. In addition, immunohistochemical staining of tissues for characterization of the mononuclear cell infiltrates was performed. Aggregates of mononuclear cells in retro-ocular connective and fatty tissue were found in five of six GO tissue specimens, but in none of the control specimens. We detected immunoreactivity for the three cytokines (IFNgamma, TNFalpha and IL-1alpha) in the five GO tissue specimens that contained mononuclear cell aggregates. In addition, IL-1alpha immunoreactivity was demonstrable in primary and subsequent GO fibroblast cultures and in their supernatants. In contrast, no immunoreactivity for any of these cytokines was detected in tissue specimens, primary cultures or culture supernatants derived from normal individuals. The presence of mononuclear cell infiltrates and associated immunoreactivity for IFNgamma, TNFalpha, IL-1alpha in retro-ocular connective tissue derived from patients with GO suggests that the previously demonstrated in vitro functions of these cytokines may indeed be operative in vivo. Our results further support the concept that IFNgamma, TNFalpha and IL-1alpha may play a role in the pathogenesis of GO.