Background: Chronic kidney disease (CKD) increases cardiovascular risk. Underlying mechanisms, however, remain obscure. The uremic toxin indoxyl sulfate is an independent cardiovascular risk factor in CKD. We explored the potential impact of indoxyl sulfate on pro-inflammatory activation of macrophages and its underlying mechanisms. Methods: We examined in viro the effects of clinically relevant concentrations of indoxyl sulfate on pro-inflammatory responses of macrophages and the roles of organic anion transporters (OATs) and organic anion transporting polypeptides (OATPs). A systems approach, involving unbiased global proteomics, bioinformatics, and network analysis, then explored potential key pathways. To address the role of Dll4 in indoxyl sulfate-induced macrophage activation and atherogenesis in CKD in vivo, we used 5/6 nephrectomy and Delta-like 4 (Dll4) antibody in LDL receptor-deficient (Ldlr−/−) mice. To further determine the relative contribution of OATP2B1 or Dll4 to pro-inflammatory activation of macrophages and atherogenesis in vivo, we used siRNA delivered by macrophage-targeted lipid nanoparticles in mice. Results: We found that indoxyl sulfate-induced pro-inflammatory macrophage activation is mediated by its uptake through transporters, including OATP2B1, encoded by the SLCO2B1 gene. The global proteomics identified potential mechanisms, including Notch signaling and the ubiquitin-proteasome pathway, that mediate indoxyl sulfate-triggered pro-inflammatory macrophage activation. We chose the Notch pathway as an example of key candidates for validation of our target discovery platform and for further mechanistic studies. As predicted computationally, indoxyl sulfate triggered Notch signaling, which was preceded by the rapid induction of Dll4 protein. Dll4 induction may result from inhibition of the ubiquitin-proteasome pathway, via the deubiquitinating enzyme USP5. In mice, macrophage-targeted OATP2B1/Slco2b1 silencing and Dll4 antibody inhibited pro-inflammatory activation of peritoneal macrophages induced by indoxyl sulfate. In Ldlr−/− mice, Dll4 antibody abolished atherosclerotic lesion development accelerated in CKD mice. Moreover, co-administration of indoxyl sulfate and OATP2B1/Slco2b1 or Dll4 siRNA encapsulated in macrophage-targeted lipid nanoparticles in Ldlr−/− mice suppressed lesion development. Conclusion: These results suggest that novel crosstalk between OATP2B1 and Dll4-Notch signaling in macrophages mediates indoxyl sulfate-induced vascular inflammation in CKD.

The current pandemic of COVID-19 caused thousands of deaths and healthcare professionals struggle to properly manage infected patients. This review summarizes information about SARS-CoV-2 receptor binding dynamics and intricacies, lung autopsy findings, immune response patterns, evidence-based explanations for the immune response, and COVID-19-associated hypercoagulability.

ADP-ribosylation is a post-translational modification that is catalyzed by the ADP-ribosyltransferase enzyme family. Major emphasis to date has been ADP-ribosylation's role in cancer; however, there is growing interest in its role in inflammation and cardiovascular disease. Despite a recent boom in ADP-ribosylation mass spectrometry-based proteomics, there are limited computational resources to evaluate the quality of reported ADP-ribosylated (ADPr) proteins. We recently developed a novel mass spectral annotation strategy (RiboMaP) that facilitates identification and reporting of ADPr peptides and proteins. This strategy can monitor the fragmentation properties of ADPr peptide-unique fragment ions, termed m-ions and p-ions, that in turn provide spectral quality scores for candidate ADP-ribosyl peptides. In this study, we leveraged the availability of publicly available ADP-ribosylome data, acquired on various mass spectrometers, to evaluate the broader applicability of RiboMaP. We observed that fragmentation spectra of ADPr peptides vary considerably across datasets; nonetheless, RiboMaP improves ADPr peptide spectral annotation across all studies. We then reanalyzed our own previously published in vitro ADP-ribosylome data to determine common responses to the pro-inflammatory cytokine, IFN-γ. We conclude that despite these recent advances in the field of ADPr proteomics, studies in the context of inflammation and cardiovascular disease still require further bench-to-informatics workflow development in order to capture ADPr signaling events related to inflammatory pathways.