Abstract

BackgroundPulmonary arterial hypertension (PAH) is one of the most important complications that seriously threatens the prognosis of patients with systemic lupus erythematosus (SLE), with complicated and unclear pathogenesis.ObjectivesBased on genomic studies and functional experiments, we aim to investigate candidate biomarkers and targeted therapy for the early diagnosis and timely treatment of SLE-PAH patients.Methods:1) In order to screen susceptible genes of SLE-PAH, a number of 150 peripheral blood from SLE-PAH patients were subject to whole-exome sequencing (WES), and genome-wide association study (GWAS) was performed by comparing with 934 healthy controls.2) The transcriptional expression levels on peripheral blood of SLE-PAH patients were examined by RT-qPCR to further evaluate the possible pathogenesis of the above screened genes.3) Intervention experiments on human pulmonary artery endothelial cells (hPAEC) were performed to figure out the potential pathogenesis of the selected gene in vitro. RNA-seq and gene ontology were applied to identify the downstream pathways.4) Established by pristane injection and hypoxia induction, SLE-PAH mice model was used to evaluate the pathogenicity and therapeutic value of selected gene. Pulmonary arterial pressure (PAP) was measured by right heart catheterization after tail-intravenous injection of therapeutic vectors.Results:1) The tumor necrosis factor receptor-associated factor 5 (TRAF5) was identified as a susceptible gene of SLE-PAH based on WES and GWAS.2) The significant reductions of TRAF5 on transcriptional level in peripheral blood of SLE-PAH patients were identified, indicating clinical diagnosis values.3) Knockdown of TRAF5 significantly increased early apoptosis of hPAEC and triggered the pathogenesis of PAH through distinct pathways.4) SLE-PAH mouse model was successfully established since they showed lupus phenotype and the mean PAPs were measured as over 40mmHg. Tail-intravenous injection of TRAF5-overexpression vector attenuated PAH.ConclusionLack of TRAF5 triggers the pathogenesis of PAH in SLE patients through inducing hPAEC abnormality. It is a susceptible gene of SLE-PAH and could be a candidate marker for diagnosis and therapy for SLE-PAH patients.Figure 1.A) Genomic and protein simulation structure TRAF5 (a susceptible gene of SLE-PAH). Red dots represent mutation sites that screened from SLE-PAH patients. P.G468R mutation causes dysfunction of protein. B) TRAF5 mRNA expression levels in PBMC of healthy-controls, SLE-PAH and SLE-nPAH patients. *p<0.05. C) shTRAF5 transfected human PAEC. The transfection efficiency reaches 80% (The percentage of EGFP positive cells in all cells of bright field) when MOI=20. All three knockdown vectors of shTRAF5 showed significantly down regulation of TRAF5. *p<0.05, **p<0.01. D) FACS was performed to detect early apoptotic cells labeled with Annexin V-APC (Apoptosis Detection Kit). The group of shTRAF5 showed significantly increased apoptosis compared with groups of control/shScramble. *p<0.05 E) Wound healing experiments were performed in different groups, and the distances of scratches at the same area in each group were measured at the time point of 0h, 6h, 24h, 48h. Abnormal migration was observed in shTRAF5 transfected PAECs.REFERENCES:NIL.Acknowledgements:NIL.Disclosure of InterestsNone Declared.

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