THU0225 THE MOLECULAR PROFILING OF MONOCYTES FROM PATIENTS WITH PRIMARY ANTIPHOSPHOLIPID SYNDROME IDENTIFIES SEVERAL NETWORKS RELATED TO THEIR ATHEROTHROMBOTIC STATUS. ROLE OF ANTIPHOSPHOLIPID ANTIBODIES ON MONOCYTE MIRNA SECRETION

Abstract

Objectives 1. To characterize the mRNAs and microRNAs transcriptomes of monocytes, key immune cells in the atherothrombotic pathology of Antiphospholipid Syndrome patients (APS). 2. To evaluate the role of antiphospholipid antibodies (aPL) in the regulation of these processes. 3. To identify and characterize circulating microRNAs secreted by activated monocytes. Methods Monocytes from peripheral blood of 40 APS patients and 40 healthy donors were purified by negative immunomagnetic selection. Gene expression microarray (Agilent G4112F platform) and nCounter microRNA expression array (Nanostring) were performed on purified RNA. Functional categorization of altered genes and miRNAs was made using IPA software, and miRNA-mRNA interaction networks showing inverse correlation expression were identified. Genes and miRNAs integrating the networks were validated in the whole APS cohort by RT-PCR, as well as on a set of thrombotic non-autoimmune patients. Predicted miRNA-mRNA interactions were tested by microRNA transfection experiments. In parallel, a miRNA array in the plasma of those patients was performed, and common deregulated miRNAs between monocytes and plasma were identified. Monocytes isolated from HD were treated in vitro with purified aPLs and the altered gene/miRNA expression induced in monocytes was evaluated. Moreover, the effects of aPLs on the secretion of common deregulated microRNAs in the supernatant of cultured cells were assessed. Results Microarray identified 518 altered genes in APS monocytes. Relevant biofunctions on which these genes were involved included hematological and cardiovascular system development and function, inflammatory response, and embryonic development, among others. Gene alterations were validated in the whole cohort, demonstrated to be stable along the time, divergent of the gene profile in monocytes from non-autoimmune thrombotic patients, and associated to thrombotic recurrences and early atherosclerosis. Analysis of miRNA profiles showed altered expression of 22 miRNAs in APS monocytes. Fifty-four genes were inversely correlated and predicted as CVD-related target genes of 19 differentially expressed miRNAs in monocytes. Association of these genes and miRNAs with the occurrence and type of thrombotic events, obstetric complications and presence of atheroma plaques were demonstrated. Transfection studies further confirmed the relationship between specific miRNAs and their identified target genes. In vitro studies demonstrated the specific modulation of several genes/miRNAs by aPLs. Furthermore, those autoantibodies promoted the increase in several secreted miRNAs, found simultaneously altered in plasma and monocytes, and deeply involved in the development of inflammation and cardiovascular disease. Conclusion 1. Gene and microRNA expression profiles allowed the identification of relevant genes and pathways altered in monocytes and plasma of APS patients, associated with the pathogenesis of the disease and modulated, at least partially, by aPLs. 2. Specific microRNA-miRNA regulatory networks control the biological processes and factors related to the CV pathology in APS. 3. aPL antibodies induce the secretion by monocytes of specific microRNAs that might act as messengers in the bloodstream to propagate their deleterious effects. Acknowledgement Funded by ISCIII, PI15/01333, PI18/00837 and RIER RD16/0012/0015, co-funded with FEDER. Disclosure of Interests None declared