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Abnormal coagulation and increased risk of thrombosis are some of the symptoms associated with COVID-19 severity. Anti-phospholipid antibodies (aPLs) present in critically ill COVID-19 patients contribute to systemic thrombosis. The aim of this study was to identify key common genes to characterize genetic crosstalk between COVID-19 and antiphospholipid syndrome (APS) using bioinformatics analysis and explore novel mechanisms of immune-mediated thrombosis in critically ill COVID-19 patients. The transcriptome data of mononuclear cells from severe COVID-19 patients and APS patients were evaluated to obtain the common genes. The protein-protein interaction network and cytoHubba module analysis in Cytoscape software were used to find the associated hinge genes and hub genes. Among the common differentially expressed genes, TIMELESS depletion was identified only in patients with severe COVID-19 and not in patients with mild COVID-19, and it was validated with the GSE159678 dataset. Functional analyses using gene ontology terms and the Kyoto Encyclopedia of Genes and Genomes pathway suggested that TIMELESS might contribute to the production of antiphospholipid antibody and thrombosis in both COVID-19 and APS patients. The potential role of TIMELESS and autophagy genes in momonuclear cells were further investigated, and GSK3B was found to be associated with TIMELESS. Autophagy targeting agents have a therapeutic potential against COVID-19 and thrombogenesis in APS, which may be related to the role of autophagy genes in the modification of circadian clock proteins. Interference with TIMELESS and other genes associated with it to regulate autoantibody expression may be a potential strategy for immunotherapy against thrombogenesis in severe COVID-19 patients.

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Anti-ß2 glycoprotein I (anti-ß2GPI) antibodies are important contributors to the development of thrombosis. Anti-ß2GPI antibody complexes with ß2GPI are well known to activate monocytes and endothelial cells via the intracellular NF-kB pathway with prothrombotic implications. By contrast, the interaction of anti- ß2GPI/ß2GPI complexes with platelets has not been extensively studied. The p38 mitogen-activated protein kinase (MAPK) pathway has been recognized to be an important intracellular signaling pathway in the coagulation cascade and an integral component of arterial and venous thrombosis. The present study reveals that levels of anti- ß2GPI/ß2GPI complexes in sera are positively associated with p38MAPK phosphorylation of platelets in thrombotic patients. Furthermore, SB203580 inhibits anti-ß2GPI/ß2GPI complex-induced platelet activation. Thrombus formation decreased in p38MAPK-/- mice after treatment with anti-ß2GPI/ß2GPI complexes. In conclusion, p38MAPK may be a treatment target for anti-ß2GPI antibody-associated thrombotic events.

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Renal fibrosis is glomerulosclerosis and renal tubulointerstitial fibrosis caused by the increase of interstitial cells and intercellular substances and the accumulation of extracellular matrix, and is a common pathological manifestation of renal disease progressing to end-stage renal failure. It has proved that Astragalus polysaccharide (AP) has curative effect on renal disease; however, its therapeutic mechanism on renal fibrosis is still unclear. Metabolomics approach provides an opportunity to identify novel molecular biomarkers. The purpose of this study is to study the changes of serum metabolic profile of rats with unilateral tubal ligation and replication of renal fibrosis model and the therapeutic effect of AP on it. The blood samples of rats in the control group, renal fibrosis model group, and AP treatment group collected on the 21st day were analyzed by metabolomics method based on UPLC-Q-TOF-MS. Principal component analysis (PCA) showed that clustering was obvious and significantly separated, and paired partial least squares discriminant analysis (OPLS-DA) was used for further analysis. Combined with the network databases such as HMDB and KEGG and a large number of literatures, 32 potential biomarkers related to renal fibrosis were preliminarily screened out and further verified by MS/MS secondary debris information. After pretreatment with AP, 20 biomarkers were significantly regulated, and correlated with phenylalanine, tyrosine, and tryptophan biosynthesis, phenylalanine metabolism, arachidonic acid metabolism, etc. It also revealed the metabolic changes of renal fibrosis and intervention effect of AP. These data uncover a link between metabolism and the molecular mechanism with potential implications in the understanding of the intervention effect of AP. Conclusively, UPLC-Q-TOF-MS-based metabolomics can be valuable and promising strategy to understand the disease mechanism and natural drug pretreatment.

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Hyperhomocysteinemia induces the proliferation of vascular smooth muscle cells (VSMCs). Hydrogen sulfide (H2S) inhibits the phenotype switch of VSMCs and calcium-sensing receptor (CaSR) regulated the production of endogenous H2S. However, whether CaSR inhibits the proliferation of VSMCs by regulating the endogenous cystathionine-gamma-lyase (CSE, a major enzyme that produces H2S) pathway in high homocysteine (HHcy) has not been previously investigated. The intracellular calcium concentration, the concentration of H2S, the cell viability, the proliferation and the expression of proteins of cultured VSMCs from rat thoracic aortas were measured, respectively. The results showed that the [Ca(2+)]i and the expression of p-CaMK and CSE increased upon treatment with CaSR agonist. In HHcy, the H2S concentration decrease, the proliferation and migration rate increased, the expression of Cyclin D1, PCNA, Osteopontin and p-Erk1/2 increased while the α-SM actin, P21(Cip/WAK-1) and Calponin decreased. The CaSR agonist or exogenous H2S significantly reversed the changes of VSMCs caused by HHcy. In conclusion, our results demonstrated that CaSR regulate the endogenous CSE/H2S is related to the PLC-IP3 receptor and CaM signal pathways which inhibit the proliferation of VSMCs, and the latter is involved in the Erk1/2 dependent signal pathway in high homocysteine.

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Anti-ß2 glycoprotein I (anti-ß2GPI ) antibodies are important contributors to thrombosis, especially in patients with antiphospholipid syndrome (APS). However, the mechanism by which anti-ß2GPI antibodies are involved in the pathogenesis of thrombosis is not fully understood. In this report, we investigated the role of anti- ß2GPI antibodies in complexes with ß2GPI as mediators of platelet activation, which can serve as a potential source contributing to thrombosis. We examined the involvement of the apolipoprotein E receptor 2' (apoER2') and glycoprotein I ba (GP I ba) in platelet activation induced by the anti-ß2GPI /ß2GPI complex. The interaction between the anti-ß2GPI /ß2GPI complex and platelets was examined using in vitro methods, in which the Fc portion of the antibody was immobilized using protein A coated onto a microtiter plate. Platelet activation was assessed by measuring GPII b/III a activation and P-selectin expression and thromboxane B2 production as well as p38 mitogen-activated protein kinase phosphorylation. Our results revealed that the anti-ß2GPI /ß2GPI complex was able to activate platelets, and this activation was inhibited by either the anti-GP I bα antibody or the apoER2' inhibitor. Results showed that the anti-ß2GPI /ß2GPI complex induced platelet activation via GPI ba and apoER2', which may then contribute to the prothrombotic tendency in APS patients.

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