1. DISSECTING DISEASE-SPECIFIC DIFFERENCES IN RA AND OA BY TRANSCRIPTOME ANALYSES OF SYNOVIAL TISSUE, BLOOD AND BONE MARROW MONOCYTES Biljana Smiljanovic 1, Bruno Stuhlmüller 1, Wlodzimierz Maslinski 3, Gerd R. Burmester 1, Andreas Radbruch 2, Andreas Grützkau 2, Thomas Häupl 1 1 Department of Rheumatology and Clinical Immunology, Charité University Hospital, Berlin, Germany 2 Deutsches Rheuma – Forschungszentrum, Berlin, Germany 3 Department of Pathophysiology and Immunology, Institute of Rheumatology, Warsaw, Poland Rheumatoid arthritis (RA) is a chronic inflammatory disease associated with joint destruction. Joint inflammation is characterised by infiltration and activation of various immune cells. To avoid difficulties in sampling synovial tissue and to avoid fluctuation in cellular composition of leukocytes, which accompanies inflammation both in synovial tissue and blood, this study was focused on transcriptome analyses of synovial tissues, blood and bone marrow monocytes. The main aim was to analyse potential of blood monocytes in dissecting inflammation in RA and in reflecting inflammation that is evident in synovial tissue. Osteoarthritis (OA), a non-inflammatory disease, was used as control in this study. Conclusion: Materials and Methods : Background: 6 RA patients 6 OA patients 10 RA patients 10 OA patients sCD163 ***p<0.0001 *p=0.0143 *p=0.0438 ***p<0.0001 sCD163 p=0.3088 **p=0.0015 p=0.2706 ***p=0.0001 Results: 2.From monocyte transcriptomes to validation at the protein level in synovial fluid and serum from RA and OA patients 1.Transcriptomes generated from synovial tissue biopsies (A), blood monocytes (B) and bone marrow monocytes (C) discriminated RA from OA patients sCD14 SYNOVIAL FLUID (SF)SERUM sCD14, sCD163, TNFAIP6 and IL1R2 as potential markers that discriminated RA from OA were measured in Synovial fluid and Serum sCD14, sCD163 and IL1R2 measured in SF showed differences between RA and OA, while in serum only sCD14 exhibited difference. (A) TRANSCRIPTOME OF SYNOVIAL TISSUE (2019 AFFYMETRIX IDS) (B) TRANSCRIPTOME OF BLOOD MONOCYTES (379 AFFYMETRIX IDS) The RA gene expression profile was the most specific and robust in synovial tissue, demonstrating the dominance of the inflammatory process in the joints. Nevertheless, the systemic nature of RA was also evident at the level of blood and bone marrow monocytes. Concerning that blood is a favourable and easily accessible material for diagnosis and that monocytes are able to exhibit disease-specific alterations, understanding the monocyte response in different rheumatic diseases seems to be an advantageous approach for biomarker discovery. This approach should be essential for identifying the objective criteria relevant for disease and therapeutic stratification of patients with RA. Gene-expression profiles from synovial tissues biopsies were generated by Affymetrix HG-U133A arrays. Gene-expression profiles of blood and bone marrow monocytes were generated by Affymetrix HG-U133Plus arrays. The BioRetis database was used for array data analyses. Biljana Smiljanovic Department of Rheumatology and Clinical Immunology, Charité University Hospital Charitéplatz 1 D-10117 Berlin Germany E-Mail: smiljanovic@drfz.de (C) TRANSCRIPTOME OF BONE MARROW MONOCYTES (221 AFFYMETRIX IDS) 8 RA patients 8 OA patients Genes involved in: Inflammation (276 Affy IDs): CCL2, CCL5, CCL8, CCL13, CCL18, CXCL1, CXCL9, CXCL10, CXCL13, IL6, IL7, IL8, IL15 Cell adhesion (65 Affy IDs): ICAM1, ICAM3, ITGB2, ITGA4, SELE Extracellular matrix formation (31 Affy IDs): SDC1, SDC2, SDC3, CD44, FN1, LAMA2 Profile indicates infiltration of various cell types: Mf, T-, B-, NK- cells Genes involved in: Inflammation (40 Affy IDs): CCR2, CXCR4, IL1R2, TNFAIP6, CD163 Cell adhesion (22 Affy IDs): CD44, ICAM2, ICAM4, ITGAL Inflammatory profile in blood Mo was weaker than in synovial tissue p=0.1083 sIL1R2TNFAIP6 sIL1R2TNFAIP6 **p=0.0014 Genes involved in: Inflammation (18 Affy IDs): IL1R2, TNFAIP6, IL10, FAS Cell adhesion (13 Affy IDs): ITGA4, ITGA6, ITGB1, ALCAM Inflammatory profile in bone marrow Mo was far weaker than in peripheral blood Mo Funding:BTCure contract no 115142-2 ArthroMark grant no 01EC1009A

2. DISSECTING DISEASE-SPECIFIC DIFFERENCES IN RA AND OA BY TRANSCRIPTOME ANALYSES OF SYNOVIAL TISSUE, BLOOD AND BONE MARROW MONOCYTES Biljana Smiljanovic 1, Bruno Stuhlmüller 1, Wlodzimierz Maslinski 3, Gerd R. Burmester 1, Andreas Radbruch 2, Andreas Grützkau 2, Thomas Häupl 1 1 Department of Rheumatology and Clinical Immunology, Charité University Hospital, Berlin, Germany 2 Deutsches Rheuma – Forschungszentrum, Berlin, Germany 3 Department of Pathophysiology and Immunology, Institute of Rheumatology, Warsaw, Poland Rheumatoid arthritis (RA) is a chronic inflammatory disease associated with joint destruction. Joint inflammation is characterised by infiltration and activation of various immune cells. To avoid difficulties in sampling synovial tissue and to avoid fluctuation in cellular composition of leukocytes, which accompanies inflammation both in synovial tissue and blood, this study was focused on transcriptome analyses of synovial tissues, blood and bone marrow monocytes. The main aim was to analyse potential of blood monocytes in dissecting inflammation in RA and in reflecting inflammation that is evident in synovial tissue. Osteoarthritis (OA), a non-inflammatory disease, was used as control in this study. Conclusion: Materials and Methods : Background: 6 RA patients 6 OA patients 10 RA patients 10 OA patients sCD163 ***p<0.0001 *p=0.0143 *p=0.0438 ***p<0.0001 sCD163 p=0.3088 **p=0.0015 p=0.2706 ***p=0.0001 Results: 2.From monocyte transcriptomes to validation at the protein level in synovial fluid and serum from RA and OA patients 1.Transcriptomes generated from synovial tissue biopsies (A), blood monocytes (B) and bone marrow monocytes (C) discriminated RA from OA patients sCD14 SYNOVIAL FLUID (SF)SERUM sCD14, sCD163, TNFAIP6 and IL1R2 as potential markers that discriminated RA from OA were measured in Synovial fluid and Serum sCD14, sCD163 and IL1R2 measured in SF showed differences between RA and OA, while in serum only sCD14 exhibited difference. (A) TRANSCRIPTOME OF SYNOVIAL TISSUE (2019 AFFYMETRIX IDS) (B) TRANSCRIPTOME OF BLOOD MONOCYTES (379 AFFYMETRIX IDS) The RA gene expression profile was the most specific and robust in synovial tissue, demonstrating the dominance of the inflammatory process in the joints. Nevertheless, the systemic nature of RA was also evident at the level of blood and bone marrow monocytes. Concerning that blood is a favourable and easily accessible material for diagnosis and that monocytes are able to exhibit disease-specific alterations, understanding the monocyte response in different rheumatic diseases seems to be an advantageous approach for biomarker discovery. This approach should be essential for identifying the objective criteria relevant for disease and therapeutic stratification of patients with RA. Gene-expression profiles from synovial tissues biopsies were generated by Affymetrix HG-U133A arrays. Gene-expression profiles of blood and bone marrow monocytes were generated by Affymetrix HG-U133Plus arrays. The BioRetis database was used for array data analyses. Biljana Smiljanovic Department of Rheumatology and Clinical Immunology, Charité University Hospital Charitéplatz 1 D-10117 Berlin Germany E-Mail: smiljanovic@drfz.de (C) TRANSCRIPTOME OF BONE MARROW MONOCYTES (221 AFFYMETRIX IDS) 8 RA patients 8 OA patients Genes involved in: Inflammation (276 Affy IDs): CCL2, CCL5, CCL8, CCL13, CCL18, CXCL1, CXCL9, CXCL10, CXCL13, IL6, IL7, IL8, IL15 Cell adhesion (65 Affy IDs): ICAM1, ICAM3, ITGB2, ITGA4, SELE Extracellular matrix formation (31 Affy IDs): SDC1, SDC2, SDC3, CD44, FN1, LAMA2 Profile indicates infiltration of various cell types: Mf, T-, B-, NK- cells Genes involved in: Inflammation (40 Affy IDs): CCR2, CXCR4, IL1R2, TNFAIP6, CD163 Cell adhesion (22 Affy IDs): CD44, ICAM2, ICAM4, ITGAL Inflammatory profile in blood Mo was weaker than in synovial tissue p=0.1083 sIL1R2TNFAIP6 sIL1R2TNFAIP6 **p=0.0014 Genes involved in: Inflammation (18 Affy IDs): IL1R2, TNFAIP6, IL10, FAS Cell adhesion (13 Affy IDs): ITGA4, ITGA6, ITGB1, ALCAM Inflammatory profile in bone marrow Mo was far weaker than in peripheral blood Mo Funding:BTCure contract no 115142-2 ArthroMark grant no 01EC1009A