Fig. 3

Scl70 antigen and anti-Scl70 autoantibodies form soluble immune complexes (sICs) that bind and activate CD16 on NK cells. A Bioactivity of sICs in serum from the described CAR-T cell patient, SLE patients (n = 3) and healthy controls (n = 4) in the presence of 0.05 µg/ml Scl70 antigen. Data are shown as mean ± SEM and are presented as fold over CD16 bioactivity in the absence of Scl70 antigen. For the CAR-T cell patient, serum samples before (blue) and after (orange) CAR-T cell treatment are shown. B Longitudinal measurement of CD16 activation by sICs (squares) in the presence of 0.05 µg/ml Scl70 antigen before (blue) and after (orange) CAR-T cell treatment and serum anti-Scl70 autoantibody concentration (triangles). Anti-Scl70 values below 5 U/ml are considered negative. C PBMC were stimulated with serum from 5 different Scl70 + SSc patients in the presence of the indicated concentrations of Scl70 antigen. Upregulation of the activation marker CD69 of NK cells, B cells and T cells was determined by flow cytometry. D left and middle: In the same experiment shown in (C), CD56^dim and CD56^bright NK cells were gated separately. Shown is the frequency of CD69-positive cells as a technical confirmation of activation of Fc-receptor-bearing CD56^dim NK cells by Scl70-containing immune complexes. Right; from the same PBMCs, NK cells were in parallel isolated and their degranulation without (0) and with (5 µg/ml) Scl70 antigen in the same five sera from Scl70 + SSc patients as shown in C and D left/middle was determined. CD107a + NK cells represent degranulated NK cells, expressed as percentage of all NK cells. These data confirm functionally relevant activation of NK cells by Scl70-containing immune complexes