Background/Purpose: Antiphospholipid syndrome is an autoimmune thrombophillia characterized by recurrent thromboembolism and or pregrancy morbidity in the presence of antiphospholipid antibodies (aPL), which recognize either a serum apolipoprotein, known as β2glycoprotein I (β2GPI), or other PL-binding proteins such as prothrombin. Complexes of anti-β2GPI/β2GPI activate platelets, monocytes and endothelial cells, probably by activating TLR4 and TLR6 leading eventually to NFκB and MAPK activation resulting to Tissue Factor (TF) and proinflammatory cytokine expression. Therefore, we assessed the whole transcriptome of endothelial cells that have been stimulated with anti-β2GPI/β2GPI.

Methods: Human umbilical Vein Endothelial cells (HUVECs) were isolated from 4 Healthy control women upon delivery.  Healthy donor HUVEC were stimulated with IgG isolated from APS patients with high anti-β2GPI titers or IgG from healthy individuals in the presence of β2GPI. Consequently, total mRNA was isolated, cDNA libraries were created and whole transcriptome sequencing (RNASeq) was performed. Gene expression data were validated in protein levels with immunocytochemistry and immunohistochemistry in placenta tissues from APS patients and healthy individuals. Data underwent KEGG Pathway Database and Gene Ontology analysis.

Results: Whole transcriptome analysis of HUVECs stimulated with anti-β2GPI/β2GPI complexes revealed 680 differentially expressed genes, among which 377 were upregulated and 303 downregulated in the aPL treated endothelial cells. Characteristic examples of the upregulated genes are IL-6, IL-8, VCAM1, SELE and TGFB2 and TGFBR1. Bioinformatics analysis revealed that the upregulated genes belong mainly to the cytokine-cytokine receptor interaction (hsa053323), MAPK signaling pathway (hsa04010), TNF signaling pathway (hsa04668) and NOD-like receptor pathway (hsa04621). Characteristic examples of the downregulated genes include the CBX4, CBX8, BCOR and HDAC7 genes. Several proteins encoded by these genes play role in the epigenetic modification of DNA. Immunohistochemical staining on placenta biopsies from APS patients and healthy individuals for IL-6, IL-8, IL-18, NFkB, TF, TNF-a, E-SELECTIN, MAPK8, TGFB2 and TGFBR1 showed increased intensity in the signal of endothelial cells on APS specimens validating thus the RNASeq results in the tissues.

Conclusion: RNASeq of endothelial cells treated with anti-β2GPI/β2GPI reveals a thoroughly analysed proinflamatory and procoagulant phenotype. Moreover, differential expression of DNA modifying proteins suggests the possible epigenetic regulation of gene expression on endothelial cells in APS syndrome. Ongoing experiments aim to analyze histone acetylation and methylation status of the promoters of the selected genes that were shown to be differentially expressed.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/whole-transcriptome-analysis-maps-proinflammatory-and-procoagulant-pathways-in-apl-treated-huvecs/