DOI: 10.7272/q6ms3r18
title: Plasma cytokine measurements using Luminex in COVID-19 patients

description: Dexamethasone is the standard of care for critically ill patients with
 COVID-19, but its immunological effects in this setting and the mechanisms
 by which it decreases mortality are not understood. We performed bulk and
 single-cell RNA sequencing of the lower respiratory tract and blood, and
 plasma cytokine profiling to study the effect of dexamethasone on systemic
 and pulmonary immune cells. We find signatures of decreased viral injury,
 antigen presentation, and T cell recruitment in patients treated with
 dexamethasone. We identify compartment- and cell- specific differences in
 the effect of dexamethasone in patients with severe COVID that are
 reproducible in publicly available datasets. Our results highlight the
 importance of studying compartmentalized inflammation in critically ill
 patients.
description: The soluble plasma cytokines were quantified using the Luminex
 multiplex platform with custom-developed reagents (R&D Systems,
 Minneapolis, MN) or single-plex ELISA (R&D Systems, Minneapolis,
 MN). The quantified analytes were read on MAGPIX instrument and the raw
 data was analyzed using the xPONENT software. Analytes quantified using
 single-plex ELISA were read using optical density. The measurements
 outside of the limit of detection were imputed using 1/3 of the lower
 limit of the standard curve for analytes quantified by Luminex and 1/2 of
 the lower limit of the standard curve for analytes quantified by ELISA.
 The NA values correspond to measurements that were not performed for the
 corresponding cytokine in the patient. This data was
 represented using principal component analysis. For this analysis,
 variables with more than 10% missing values across the dataset were
 excluded. Patients with one or more remaining missing values were filtered
 out. Values were then <em>log2</em>-transformed and scaled. A
 PERMANOVA test was performed using Euclidean distances to estimate
 separation of the treatment groups. To compare circulating cytokine
 levels, Wilcoxon rank-sum tests on cytokine concentrations, including
 those with more than 10% missing values, were employed. Significant
 differences were selected using a 0.1 threshold on BH-adjusted p-values.
 The results of this analysis can be found in our manuscript which is
 accepted in Nature Communications, and the preprint is available in
 Research Square repository which can be accessed at https://www.researchsquare.com/article/rs-3168149/v1.
description: The data was analyzed using R programming.
description: We have uploaded Plasma cytokine measurements from Luminex in severe
 COVID-19 patients who did or did not receive dexamethasone in a
 tab-separated file called **dex_cytokine.tsv** file. **dex_cytokine: the
 plasma cytokine measurements in severe COVID-19 patients** The soluble
 plasma cytokines were quantified using the Luminex multiplex platform with
 custom-developed reagents (R&D Systems, Minneapolis, MN) or
 single-plex ELISA (R&D Systems, Minneapolis, MN). The quantified
 analytes were read on MAGPIX instrument and the raw data was analyzed
 using the xPONENT software. Analytes quantified using single-plex ELISA
 were read using optical density. The measurements outside of the limit of
 detection were imputed using 1/3 of the lower limit of the standard curve
 for analytes quantified by Luminex and 1/2 of the lower limit of the
 standard curve for analytes quantified by ELISA. The NA values correspond
 to measurements that were not performed for the corresponding cytokine in
 the patient. The table depicts expression measurements
 (picograms/microliter - pg/mL) for the following cytokines in plasma of
 severe COVID-19 patients who did or did not receive dexamethasone, which
 was collected on the day of patient's enrollment in the study: Ang-1,
 Ang-2, ICAM-1, IFN-gamma, IL-10, IL-18, IL-6, IL-8, IP-10, MMP-8, PAI-1,
 Protein C, RAGE, SP-D, Thrombomodulin, TNF R1, TREM-1, VEGF. The values
 represent concentrations of cytokines in plasma in picograms/microliter
 unit. The following patients did not receive dexamethasone: MVIR1-HS1,
 MVIR1-HS107, MVIR1-HS2, MVIR1-HS23, MVIR1-HS33, MVIR1-HS43, MVIR1-HS50,
 MVIR1-HS53, MVIR1-HS59, MVIR1-HS72, MVIR1-HS77, MVIR1-HS78, MVIR1-HS89,
 MVIR1-HS91, MVIR1-HS94. The rest of the cohort received dexamethasone. 
 This data was represented using principal component analysis. For this
 analysis only, variables with more than 10% missing values across the
 dataset were excluded. Patients with one or more remaining missing values
 were filtered out. Values were then *log2*-transformed and scaled. A
 PERMANOVA test was performed using Euclidean distances to estimate
 separation of the treatment groups. To compare circulating cytokine
 levels, Wilcoxon rank-sum tests on cytokine concentrations, including
 those with more than 10% missing values, were employed. Significant
 differences were selected using a 0.1 threshold on BH-adjusted p-values.
 The results of this analysis can be found in our manuscript which is
 accepted in Nature Communications, and the preprint is available in
 Research Square repository which can be accessed
 at [https://www.researchsquare.com/article/rs-3168149/v1](https://www.researchsquare.com/article/rs-3168149/v1).