Plasma cytokine measurements using Luminex in COVID-19 patients
- 1. University of California, San Francisco
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.
Notes
The data was analyzed using R programming.
Funding provided by: National Institutes of Health
ROR ID: https://ror.org/01cwqze88
Award Number:
Methods
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 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.
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- https://www.researchsquare.com/article/rs-3168149/v1 (URL)