Mpox virus (MPXV) primarily infects human skin to cause lesions. Currently, robust models that recapitulate skin infection by MPXV are lacking. Here we demonstrate that human induced pluripotent... Show moreMpox virus (MPXV) primarily infects human skin to cause lesions. Currently, robust models that recapitulate skin infection by MPXV are lacking. Here we demonstrate that human induced pluripotent stem cell-derived skin organoids are susceptible to MPXV infection and support infectious virus production. Keratinocytes, the predominant cell type of the skin epithelium, effectively support MPXV infection. Using transmission electron microscopy, we visualized the four stages of intracellular virus particle assembly: crescent formation, immature virions, mature virions and wrapped virions. Transcriptional analysis showed that MPXV infection rewires the host transcriptome and triggers abundant expression of viral transcripts. Early treatment with the antiviral drug tecovirimat effectively inhibits infectious virus production and prevents host transcriptome rewiring. Delayed treatment with tecovirimat also inhibits infectious MPXV particle production, albeit to a lesser extent. This study establishes human skin organoids as a robust experimental model for studying MPXV infection, mapping virus-host interactions and testing therapeutics.Mpox virus productively infects human induced pluripotent stem cell-derived skin organoids and is inhibited by tecovirimat, revealing a model system for analysis of mpox virus pathogenesis and drug screening. Show less
We have recently introduced multiple reaction monitoring (MRM) mass spectrometry as a novel tool for glycan biomarker research and discovery. Herein, we employ this technique to characterize the... Show moreWe have recently introduced multiple reaction monitoring (MRM) mass spectrometry as a novel tool for glycan biomarker research and discovery. Herein, we employ this technique to characterize the site-specific glycan alterations associated with primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). Glycopeptides associated with disease severity were also identified. Multinomial regression modelling was employed to construct and validate multi-analyte diagnostic models capable of accurately distinguishing PBC, PSC, and healthy controls from one another (AUC = 0.93 +/- 0.03). Finally, to investigate how disease-relevant environmental factors can influence glycosylation, we characterized the ability of bile acids known to be differentially expressed in PBC to alter glycosylation. We hypothesize that this could be a mechanism by which altered selfantigens are generated and become targets for immune attack. This work demonstrates the utility of the MRM method to identify diagnostic site-specific glycan classifiers capable of distinguishing even related autoimmune diseases from one another. Show less
Merleev, A.A.; Park, D.; Xie, Y.X.; Kailemia, M.J.; Xu, G.G.; Ruhaak, L.R.; ... ; Maverakis, E. 2020
Alterations in the human glycome have been associated with cancer and autoimmunity. Thus, constructing a site-specific map of the human glycome for biomarker research and discovery has been a... Show moreAlterations in the human glycome have been associated with cancer and autoimmunity. Thus, constructing a site-specific map of the human glycome for biomarker research and discovery has been a highly sought-after objective. However, due to analytical barriers, comprehensive site-specific glycoprofiling is difficult to perform. To develop a platform to detect easily quantifiable, site-specific, disease-associated glycan alterations for clinical applications, we have adapted the multiple reaction monitoring mass spectrometry method for use in glycan biomarker research. The adaptations allow for highly precise site-specific glycan monitoring with minimum sample prep. Using this technique, we successfully mapped out the relative abundances of the most common 159 glycopeptides in the plasma of 97 healthy volunteers. This plasma glycome map revealed 796 significant (FDR<0.05) site-specific inter-protein and intra-protein glycan associations, of which the vast majority were previously unknown. Since age and gender are relevant covariants in biomarker research, these variables were also characterized. 13 glycopeptides were found to be associated with gender and 41 to be associated with age. Using just five age-associated glycopeptides, a highly accurate age prediction model was constructed and validated (r(2)=0.62 +/- 0.12). The human plasma site-specific glycan map described herein has utility in applications ranging from glycan biomarker research and discovery to the development of novel glycan-altering interventions. Show less
