Frequency analysis of autoreactive T-helper 1 and 2 cells in bullous pemphigoid and pemphigus vulgaris by enzyme-linked immunospot assay

Background Bullous pemphigoid (BP) and pemphigus vulgaris (PV) are autoimmu ne bullous skin diseases mediated by autoantibodies against adhesion molecu les of the skin. Previous studies have identified autoreactive T cells in p atients with BP and PV, which may be critical in providing B-cell help for autoantibody production. Objectives To evaluate the frequency of autoreacti ve T-helper (Th) 1 and Th2 cells in patients with BP (n = 7) or PV (n = 1) and in healthy controls (n = 11). Methods In an enzyme-linked immunospot (ELISPOT) assay, microtitre plates w ere coated with antihuman interleukin (IL)-5 IgG or antihuman interferon (I FN)-gamma IgG prior to culturing human peripheral blood lymphocytes (PBL) w ith BP180 or desmoglein (Dsg) 3 proteins for 7 days, Cytokine-producing aut oreactive T cells were visualized as spot-forming units, Results One BP patient with extensive blisters had 5.1 +/- 1.5 (mean +/- SD ) BP180-reactive Th1 cells and 2.9 +/- 1.5 Th2 cells per 10(5) PBL, In cont rast, PBL from six BP patients in remission or on immunosuppressive therapy did not form IFN-gamma- or IL-5-producing spots per less than or equal to 5 x 10(5) PBL. The patient with oral PV had 4.7 +/- 2.4 Th1 cells and 3.0 /- 0.4 Th2 cells per 10(5) PBL and a vigorous PBL response to Dsg3, In addi tion, three of 10 controls had BP180-reactive Th1 (2.7-13.8 per 10(5) PBL) and Th2 (0.3-1.8 per 10(5) PBL) cells and one control had 9.0 + 0.7 Th1 cel ls and 1.1 +/- 0.8 Th2 cells per 10(5) PBL, with reactivity to Dsg3. ELISPO T reactivity correlated with H-3-thymidine incorporation in six of seven pa tients and controls with autoreactive T-cell responses. Conclusions The ELISPOT assay seems to be promising for the quantitative an d qualitative analysis of autoreactive T-cell responses in BP and PV.