Poster - 31th German Conference on Bioinformatics 2016 ......Title: Poster - 31th German Conference...
Transcript of Poster - 31th German Conference on Bioinformatics 2016 ......Title: Poster - 31th German Conference...
Background and Objective:
Materials and Methods:
Results:
Conclusion:CD14+ cells purified from bone marrow and blood of RA and OA patients undergoinghip replacement surgery were profiled with Affymetrix HG-U133 Plus 2.0 arrays. TheBioRetis database was used for array analyses. For functional interpretation of arraydata Ingenuity Pathway Analysis and Gene Ontology were applied. 68 differentreference transcriptomes of healthy bone marrow progenitors and various stages ofactivated and differentiated monocytes were used for more detailed functionalinterpretation. Flow cytometry was applied for profiling of monocyte subsets:CD14+CD16-, CD14+CD16+ and CD14dimCD16+ in bone marrow, blood and synovialfluid samples. immunoClust algorithm was applied for automated analyses of FACSdata.
Alteration of RA monocytes was evident already in bone marrow and wascharacterized by increased monocytopoiesis and/or premature release intocirculation. Comprehensive analyses of Mo profiles by reference transcriptomesprovided a very detailed insight into gene patterns related to maturation,differentiation and activation of Mo. Flow cytometry analyses of Mo subsets in RAbone marrow, blood and synovial fluid depicted increased expression of CD16 on Moduring their maturation and differentiation from bone marrow into blood and duringtheir migration and activation from blood into synovium. The most obvious activationof Mo occurs in the joint and is depicted by a specific Mo subset that expresses thehighest level of CD14, CD16 & CD163.
Most biologics for rheumatoid arthritis (RA) target processes involved in monocyte activation. To determine when, where and how monocytes become involved in pathogenesisof RA, we analysed monocytes from bone marrow, blood and synovial fluid by gene-expression profiling and cytometry, and compared their activation patterns withosteoarthritis (OA).
ArthroMarkgrantno01EC1009A
Geneexpressionprofilingandcytometryanalysisfrombonemarrowmonocytes,blood
andsynovialfluidfromrheumatoidandosteoarthritispatients
BiljanaSmiljanovic1,TillSorensen1,MarcBonin-Andresen1,BrunoStuhlmuller1,GerdR.Burmester1,AndreasGrutzkau2,ThomasHaupl1
1DepartmentofRheumatologyandClinicalImmunology,CharitéUniversityHospital,Berlin,Germany,2GermanArthritisResearchCenter,Berlin,Germany
MarcBoninDepartmentofRheumatologyandClinicalImmunologyCharitéUniversityHospitalCharitéplatz1D-10117BerlinGermany
Tel:+49(0)30450513296Fax:+49(0)30450513968E-Mail: [email protected]:www.charite-bioinformatik.de
Contacts:
www.charite-bioinformatik.de
1.TranscriptomesofRAbonemarrowandbloodmonocytesshowedaminoroverlapandmoreprominentalterationsinblood
2.FunctionalanalysisofRA-BM&RA-PBtranscriptomesemphasisedgenesinvolvedininflammationandhemopoiesis
3.FunctionalanalysesofRA-BM&RA-PBprofilesby68referencetranscriptomesshowedtheshiftstowardimmatureprofiles(shifttotheleft)
4.MonocytematurationduringmigrationfrombonemarrowviabloodtosynovialfluidinRApatientsanalyzedbyFACS
RAbonemarrowMoprofileanalysedbyMolecularnetwork- Ingenuity(IPA)&Geneontology(GO)
RAbloodMoprofileanalysedbyMolecularnetwork- Ingenuity(IPA)&Geneontology(GO)
(1)RAtranscriptomeofbonemarrow(BM)Mo (2)RAtranscriptomeofblood(PB)Mo (3)Principalcomponentanalysis(PCA)ofBM&PBprofilesfromRA&OAMo
221differentiallyexpressedprobe-setsbetweenRA&OAbonemarrowMo
379differentiallyexpressedprobe-setsbetweenRA&OAbloodMo
571differentiallyexpressedprobe-setsinBM&PBMofromRA&OApatients
OA-BMOA-PBRA_BMRA_PB
RA
OA
RA
OA
Biologicalprocesses Genesanti-apoptosis Up-reg:CLU,FAS,IL10,SOCS3,YWHAZ
Down-reg:ANXA1,NOTCH2NL,VNN1hemopoiesis Up-reg:FLT3,IL10,PICALM,ZBTB16
Down-reg:NOTCH2NLInflammatory Up-reg:FPR2,IL10,IL8,TNFAIP6,TPST1
Down-reg:ANXA1,C3AR1,VNN1responsecelladhesion Up-reg:ALCAM,ARF6,FPR2,ITGA4,ITGA6,ITGB1
Down-reg:CD36
Biologicalprocesses Genesinflammatoryresponse Up-reg:CCR2,CD163,FPR2,LTB4R,NLRC4,PXK
Down-reg:CAMK1D,CCL2,CSF1R,ITGAL,MIF,TNFanti-apoptosis Up-reg:BAG4,CLEC5A,NAIP,SERPINB2,THBS1,VNN1
Down-reg:CCL2,FOXO1,RIPK2,TCF7L2,TNFcellcyclearrest Up-reg:CDKN2B,GADD45A,MAP2K6,MYC,THBS1
Down-reg:CDKN1C,GAS2L1,TCF7L2hemopoiesis Up-reg:PICALM,RUNX1
Down-reg:BCL11A,CSF1R,IKZF1,ROGDI
• RA-BMprofileshowedamoreimmatureprofilethanOA-BMandischaracterisedbyearlymyelopoieticgenesandgenesup-regulatedbyG-CSFthatindicatedfasteregressofMofromBM
• RA-PBprofileexhibitedalsoamoreimmatureprofilethanOA-PB,characterizedbylatemyelopoieticgenesfromBM,pronouncedG-CSFeffectsandabsenceofgenesspecificforCD16+Mo
- Mofrombonemarrow,bloodandsynovialfluidwereanalysedbyFACS
- Mosubsets:CD14+CD16- &CD14~CD16+- Frequency,CD14,CD16&CD163expressionofMowereanalysed
- immunoClustsoftwareforautomatedanalysisofFACSdata- CD14+CD16- MoarethedominantsubsetinBMfromRA&OApatients.
- FrequencyofCD14~CD16+Mo,whichisthemoredifferentiatedMosubset,increasedinPBandexpressionofCD14decreasedwhileexpressionofCD16increasedonthissubset.
- CD14++CD16+subsetappearsonlyinSFandrepresentsthemostdifferentiated(thehighestCD16expression)andthemostactivated(thehighestlevelofCD14&CD163)Mosubset.
AnalysisofRA-BMprofilebyreferencetranscriptomes68Referencetranscritpomesofbonemarrow&bloodsamples AnalysisofRA-PBprofilebyreferencetranscriptomes
Bonemarrow(RA&OA) Blood(RA&OA) SF&Blood(RA)Mosubsets
CD14+CD16- &CD14~CD16+BM&PBfromRA&OA
MosubsetsCD14+CD16+&CD14~CD16+
&CD14++CD16+fromPB&SFofRA
RA
OA
RA
OA
RA-BMprofileUp-reginRA-BM
Down-reginRA-BM
Frequency CD14expression CD16expression CD163expression
1.Latemyelopoiesis
2.WeakTNF/LPSeffect
1.Earlymyelopoiesis
2.WeakTNF/LPSeffect
3. G-CSF effect
1.Myelopoiesis
2.TNF/LPSeffect
1. Early&Latemyelopoiesis
2.TNF/LPSeffect
3. G-CSF effect
RA-PBprofile
Up-reginRA-PB
Down-reginRA-PB
3.CD16+pattern