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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 Gr ü tzkau 2 , Thomas H ä upl 1.
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DISSECTING DISEASE-SPECIFIC DIFFERENCES IN RA AND OA BY TRANSCRIPTOME ANALYSES OF SYNOVIAL TISSUE, BLOOD AND BONE MARROW MONOCYTES Biljana Smiljanovic1, Bruno Stuhlmüller1, Wlodzimierz Maslinski3, Gerd R. Burmester1, Andreas Radbruch2, Andreas Grützkau2, Thomas Häupl1 1Department of Rheumatology and Clinical Immunology, Charité University Hospital, Berlin, Germany 2DeutschesRheuma – Forschungszentrum, Berlin, Germany 3Department ofPathophysiologyandImmunology, Institute ofRheumatology, Warsaw, Poland Background: Rheumatoid arthritis (RA) is a chronicinflammatorydiseaseassociatedwithjointdestruction. Joint inflammationischaracterisedbyinfiltrationandactivationofvarious immune cells. Toavoiddifficulties in samplingsynovialtissueandtoavoidfluctuation in cellularcompositionofleukocytes, whichaccompaniesinflammationboth in synovialtissueandblood, thisstudy was focused on transcriptomeanalysesofsynovialtissues, bloodandbonemarrowmonocytes. The mainaim was toanalyse potential ofbloodmonocytes in dissectinginflammation in RA and in reflectinginflammationthatis evident in synovialtissue. Osteoarthritis (OA), a non-inflammatorydisease, was usedascontrol in thisstudy. Results: Transcriptomes generated from synovial tissue biopsies (A), blood monocytes (B) and bone marrow monocytes (C) discriminated RA from OA patients (A) TRANSCRIPTOME OF SYNOVIAL TISSUE (2019 AFFYMETRIX IDS) • 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 10 RA patients 10 OA patients (B) TRANSCRIPTOME OF BLOOD MONOCYTES (379 AFFYMETRIX IDS) • 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 6 RA patients 6 OA patients (C) TRANSCRIPTOME OF BONE MARROW MONOCYTES (221 AFFYMETRIX IDS) • 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 8 RA patients 8 OA patients ***p<0.0001 From monocyte transcriptomes to validation at the protein level in synovial fluid and serum from RA and OA patients 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. SYNOVIAL FLUID (SF) SERUM sCD163 sCD14 sCD14 sCD163 *p=0.0143 **p=0.0015 ***p<0.0001 ***p=0.0001 p=0.2706 *p=0.0438 p=0.3088 TNFAIP6 sIL1R2 TNFAIP6 sIL1R2 **p=0.0014 p=0.1083 Conclusion: The RA geneexpressionprofile was themostspecificand robust in synovialtissue, demonstratingthedominanceoftheinflammatoryprocess in thejoints. Nevertheless, thesystemicnatureof RA was also evident atthelevelofbloodandbonemarrowmonocytes. Concerningthatbloodis a favourableandeasilyaccessible material fordiagnosisandthatmonocytesareabletoexhibitdisease-specificalterations, understandingthemonocyteresponse in different rheumaticdiseasesseemstobean advantageousapproachforbiomarkerdiscovery. This approachshouldbe essential foridentifyingtheobjectivecriteria relevant fordiseaseandtherapeuticstratificationofpatientswith RA. Materials and Methods : Gene-expression profilesfromsynovialtissuesbiopsiesweregeneratedbyAffymetrixHG-U133A arrays. Gene-expression profilesofbloodandbonemarrowmonocytesweregeneratedbyAffymetrix HG-U133Plus arrays. The BioRetisdatabase was usedforarraydataanalyses. Biljana Smiljanovic Department ofRheumatologyandClinical Immunology, Charité University Hospital Charitéplatz 1 D-10117 Berlin Germany E-Mail: smiljanovic@drfz.de Funding: BTCurecontractno 115142-2 ArthroMarkgrantno 01EC1009A
DISSECTING DISEASE-SPECIFIC DIFFERENCES IN RA AND OA BY TRANSCRIPTOME ANALYSES OF SYNOVIAL TISSUE, BLOOD AND BONE MARROW MONOCYTES Biljana Smiljanovic1, Bruno Stuhlmüller1, Wlodzimierz Maslinski3, Gerd R. Burmester1, Andreas Radbruch2, Andreas Grützkau2, Thomas Häupl1 1Department of Rheumatology and Clinical Immunology, Charité University Hospital, Berlin, Germany 2DeutschesRheuma – Forschungszentrum, Berlin, Germany 3Department ofPathophysiologyandImmunology, Institute ofRheumatology, Warsaw, Poland Background: Rheumatoid arthritis (RA) is a chronicinflammatorydiseaseassociatedwithjointdestruction. Joint inflammationischaracterisedbyinfiltrationandactivationofvarious immune cells. Toavoiddifficulties in samplingsynovialtissueandtoavoidfluctuation in cellularcompositionofleukocytes, whichaccompaniesinflammationboth in synovialtissueandblood, thisstudy was focused on transcriptomeanalysesofsynovialtissues, bloodandbonemarrowmonocytes. The mainaim was toanalyse potential ofbloodmonocytes in dissectinginflammation in RA and in reflectinginflammationthatis evident in synovialtissue. Osteoarthritis (OA), a non-inflammatorydisease, was usedascontrol in thisstudy. Results: Transcriptomes generated from synovial tissue biopsies (A), blood monocytes (B) and bone marrow monocytes (C) discriminated RA from OA patients (A) TRANSCRIPTOME OF SYNOVIAL TISSUE (2019 AFFYMETRIX IDS) • 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 10 RA patients 10 OA patients (B) TRANSCRIPTOME OF BLOOD MONOCYTES (379 AFFYMETRIX IDS) • 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 6 RA patients 6 OA patients (C) TRANSCRIPTOME OF BONE MARROW MONOCYTES (221 AFFYMETRIX IDS) • 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 8 RA patients 8 OA patients ***p<0.0001 From monocyte transcriptomes to validation at the protein level in synovial fluid and serum from RA and OA patients 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. SYNOVIAL FLUID (SF) SERUM sCD163 sCD14 sCD14 sCD163 *p=0.0143 **p=0.0015 ***p<0.0001 ***p=0.0001 p=0.2706 *p=0.0438 p=0.3088 TNFAIP6 sIL1R2 TNFAIP6 sIL1R2 **p=0.0014 p=0.1083 Conclusion: The RA geneexpressionprofile was themostspecificand robust in synovialtissue, demonstratingthedominanceoftheinflammatoryprocess in thejoints. Nevertheless, thesystemicnatureof RA was also evident atthelevelofbloodandbonemarrowmonocytes. Concerningthatbloodis a favourableandeasilyaccessible material fordiagnosisandthatmonocytesareabletoexhibitdisease-specificalterations, understandingthemonocyteresponse in different rheumaticdiseasesseemstobean advantageousapproachforbiomarkerdiscovery. This approachshouldbe essential foridentifyingtheobjectivecriteria relevant fordiseaseandtherapeuticstratificationofpatientswith RA. Materials and Methods : Gene-expression profilesfromsynovialtissuesbiopsiesweregeneratedbyAffymetrixHG-U133A arrays. Gene-expression profilesofbloodandbonemarrowmonocytesweregeneratedbyAffymetrix HG-U133Plus arrays. The BioRetisdatabase was usedforarraydataanalyses. Biljana Smiljanovic Department ofRheumatologyandClinical Immunology, Charité University Hospital Charitéplatz 1 D-10117 Berlin Germany E-Mail: smiljanovic@drfz.de Funding: BTCurecontractno 115142-2 ArthroMarkgrantno 01EC1009A
