881 Antisynthetase syndrome and dermatomyositis immunophenotyping

Abstract

Antisynthetase syndrome (AS) and Dermatomyositis (DM) are rare inflammatory autoimmune diseases that some consider as separate entities. AS is poorly defined and some patients have skin lesions consistent with DM. To investigate differences between AS and DM relative to control (HC), we identified immunophenotypes. 3 AS, 3 DM, and 3 HC formalin-fixed, paraffin-embedded (FFPE) samples obtained from back, arm, or leg were stained via immunofluorescence for MxA and IFNβ. Three 20x regions were acquired on Nikon Eclipse. A panel of 35 metal conjugated antibodies separately was stained and areas of 500x800 μm (ROI) were ablated at 200Hz on the Hyperion Imaging System(Fluidigm). Cell segmentation was performed in Visiopharm. Mean pixel intensities (MPI) per cell were analyzed using histoCAT and ImageJ. One-way ANOVA and Dunn’s Multiple comparison test was performed with all values reported as AS/DM/HC mean± SEM. Skin lesions of AS and DM patients MPI did not differ significantly in MxA (15.6±1.0/12.7±2.7/1.4±0.5; p<0.05 with AS vs DM p>0.05) or IFNβ expression (18.4± 2.4/21,20.9± 1.2/1.3±0.2; p<0.05 with AS vs DM p>0.05). AS and DM lesions did not differ in the following cells/ROI except CD4: CD4 (70± 42/178± 28/11±2; AS vs DM p<0.001), CD8 (49± 37/55±21/20±12), MAC387 (6± 3/22± 10/4±2), pDC (4± 1/14± 7/5±1), CD11C+ (25± 8/44± 11/11±6), mast (14± 5/21± 12/25±11), and FOXP3+ CD4(18±14/64± 32/5±2); all p<0.001 with AS vs DM p>0.05 except CD4). AS and DM lesions also did not differ in MPI of key inflammatory pathways: pSTING (7.1± 1.4/8.2± 1.3/1.5±0.6), IL31 (0.87± 0.2/1.0± 0.1/0.1±0.02), IFNγ (1.5± 0.38/2.8± 0.2/0.3±0.08), IL4 (4.4± 0.7/4.6± 0.5/0.8±0.1), and IL17 (1.0± 0.2/1.2± 0.2/0.2±0.01); p<0.0001 with AS vs DM p>0.05. Patients with both AS and DM lesional skin did not differ from DM alone in immunophenotyped or type I IFN protein. AS with DM skin lesions were not differentiated from DM, as the pathogenic cellular characteristics between the two were the same.