Effect of lymphocytes on hormonal secretion by autologous thyrocytes cultured in monolayers

Abstract

We studied the lymphocyte-induced alterations in hormonal metabolism and the production of tumour necrosis factor alpha (TNF-alpha) during coculture of thyrocytes and autologous lymphocytes from 20 patients with Graves' disease and from five normal subjects. Thyroglobulin (Tg) mRNA was assessed by slot-blot analysis under TSH stimulation. Tg, tri-iodothyronine (T3) and cAMP secretion in the presence of TSH were measured by RIA after 3 or 5 days of coculture. TNF-alpha levels produced after 5 days incubation were also assayed in lymphocyte culture and coculture media. Lymphocytes isolated from peripheral blood (PBLs) altered the production of Tg, T3 and cAMP in autologous thyrocytes. Intrathyroidal lymphocytes (ITLs) decreased Tg and cAMP secretion but had no effect on T3 secretion. The reductions in Tg and cAMP levels obtained with mechanically isolated ITLs (M-ITLs) were generally higher than those obtained with ITLs isolated by dispase (D-ITLs). No difference was seen between Graves' disease and normal cocultures. PBLs secreted large concentrations of TNF-alpha, larger than those obtained with M-ITLs whereas D-ITLs produced low amounts of this cytokine. In coculture, TNF-alpha levels were lower than those observed in lymphocyte culture. Significant correlations were obtained between TNF-alpha levels and the decrease in Tg, T3 and cAMP concentrations. The percentage of T lymphocytes was higher in PBLs and D-ITLs than in M-ITLs. B lymphocytes levels were higher in ITLs, especially M-ITLs, than in PBLs. TNF-alpha production by B lymphocytes was maximal in M-ITLs. In conclusion, lymphocytes induced a decrease in hormonal thyroid metabolism when cocultured with autologous thyrocytes. These perturbations may be attributed, at least partly, to TNF-alpha secreted by lymphocytes. TNF-alpha interacts via the adenylate cyclase pathway of TSH signal transduction.

Expression of membrane and soluble intercellular adhesion molecule-1 in Graves' disease

“…We also observed that the concentration of FCS did not influence the levels of sICAM-1 which were identical with 10% FCS or 10% NU-serum, a synthetic product containing only 1% serum. These results corroborate those of Ciampolillo et al (1993) who tested only membrane ICAM-1 culture Several reports have been published on human thyrocytes cocultured in monolayers with autologous PBL or ITL lymphocytes (Davies et al 1985, Iwatani et al 1986, Matsunaga et al 1986, Otsubo et al 1988, Sato et al 1993, Massart et al 1995, 1996. However, our present work is the first demonstration of the action of PBL and ITL cocultured with autologous thyrocytes on the expression of membrane and soluble ICAM-1.…”

“…Non-adherent cells were then removed and purified on Lymphoprep as above. Control of the purity of the thyrocyte and lymphocyte populations Thyrocytes Purity of thyroid cells was tested by FCA as previously described (Massart et al 1996). Briefly, phycoerythrin-conjugated CD45 and CD14 mAbs (Immunotech), were used for direct staining of membrane molecules specific for leukocytes and monocytes/macrophages respectively, with murine IgG1 as negative control.…”

“…The presence of thyrocytes was evaluated in lymphocyte preparations as described (Massart et al 1996). Briefly, contaminating thyrocytes in the ITL preparations were recognized with anti-Tg mAb purchased from Realef Laboratories (Paris, France) and with a secondary Ab (FITC-coupled F(ab ) 2 antimouse IgG, Silenus, Hawthorn Laboratories, Victoria, Australia) diluted 1:400.…”

Increased expression of tumor necrosis factor-alpha and decreased expression of thyroglobulin and thyroid peroxidase mRNA levels in the thyroids of iodide-treated BB/Wor rats

“…Studies suggest that increases in endogenous TNFa production may be involved in the regulation of thyroid function in ATD. That is, TNFa and TNFa mRNA are found in human thyroid tissues from patients with Graves' disease and Hashimoto's thyroiditis (1)(2)(3)(4). In FRTL-5 cells, we (5,6) and others (7)(8)(9)(10) have observed that TNFa inhibits type I 5 0 -deiodinase activity and thyroglobulin (Tg), thyroid peroxidase (TPO) and type I 5 0 -deiodinase gene expression.…”

Action of peripheral or intrathyroidal lymphocytes on autologous thyrocytes cultured in follicles in collagen gel

“…INTRODUCTION Lymphocytes isolated from peripheral blood (PBLs) from patients with Graves' disease have been shown to have a cytotoxic effect in vitro when cocultured with autologous thyrocyte monolayers whereas intrathyroidal lymphocytes (ITLs) have been found to exert little or no cytotoxicity, depending on the isolation method (Massart et al 1995). We have also demonstrated that the different populations of lymphocytes cultured over thyroid monolayers decrease thyroglobulin (Tg), triiodothyronine (T3) and cAMP production under thyrotrophin (TSH) stimulation (Massart et al 1996). Moreover, we have shown that this inhibition of biochemical and hormonal metabolism in thyroid monolayers is correlated with lymphocyte tumour necrosis factor a (TNF-a) production.…”