Impact of SARS-CoV-2 infection on the recovery of peripheral blood mononuclear cells by density gradient

Maria D I Manunta,Guido Baselli,Daniele Prati,Cristiana Bianco,Alessandra Bandera,Elisa Erba,Francesca Ferrari,Manuela Brasca,Mario Tirone,Alessandra Cattaneo,Mahnoosh Ostadreza,Luigi Santoro,Andrea Gori,Laura Porretti,Giuseppe Lamorte,Luca V C Valenti,Elena Trombetta

doi:10.1038/s41598-021-83950-2

Abstract

SARS-CoV-2 virus infection is responsible for coronavirus disease (COVID-19), which is characterised by a hyperinflammatory response that plays a major role in determining the respiratory and immune-mediated complications of this condition. While isolating peripheral blood mononuclear cells (PBMCs) from whole blood of COVID-19 patients by density gradient centrifugation, we noticed some changes in the floating properties and in the sedimentation of the cells on density medium. Investigating this further, we found that in early phase COVID-19 patients, characterised by reduced circulating lymphocytes and monocytes, the PBMC fraction contained surprisingly high levels of neutrophils. Furthermore, the neutrophil population exhibited alterations in the cell size and in the internal complexity, consistent with the presence of low density neutrophils (LDNs) and immature forms, which may explain the shift seen in the floating abilities and that may be predictive of the severity of the disease. The percentage of this subset of neutrophils found in the PBMC band was rather spread (35.4 ± 27.2%, with a median 28.8% and IQR 11.6–56.1, Welch’s t-test early phase COVID-19 versus blood donor healthy controls P < 0.0001). Results confirm the presence of an increased number of LDNs in patients with early stage COVID-19, which correlates with disease severity and may be recovered by centrifugation on a density gradient together with PBMCs.

Highlights

SARS-CoV-2 virus infection is responsible for coronavirus disease (COVID-19), which is characterised by a hyperinflammatory response that plays a major role in determining the respiratory and immunemediated complications of this condition
Patients were stratified according to their clinical status and the specimens were grouped in accordance with the time of sampling i.e. within 24 h from admission or 14 days after discharge
Magnification of a 14 d post-discharge sample and a sample taken within 24 h from admission are displayed in Fig. 1E, while Fig. 1F is a closer view of two samples taken at the time of admission into hospital

Summary

Introduction

SARS-CoV-2 virus infection is responsible for coronavirus disease (COVID-19), which is characterised by a hyperinflammatory response that plays a major role in determining the respiratory and immunemediated complications of this condition. The neutrophil population exhibited alterations in the cell size and in the internal complexity, consistent with the presence of low density neutrophils (LDNs) and immature forms, which may explain the shift seen in the floating abilities and that may be predictive of the severity of the disease The percentage of this subset of neutrophils found in the PBMC band was rather spread (35.4 ± 27.2%, with a median 28.8% and IQR 11.6–56.1, Welch’s t-test early phase COVID-19 versus blood donor healthy controls P < 0.0001). Results confirm the presence of an increased number of LDNs in patients with early stage COVID-19, which correlates with disease severity and may be recovered by centrifugation on a density gradient together with PBMCs. The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an enveloped single-stranded RNA virus belonging to the subfamily Coronavirinae in the family of Coronaviridae of the order N idovirales[1]. Prolonged activation of neutrophils can have detrimental effects contributing to lung injuries[61], the NET formation may play a crucial role in the exacerbation of the lung disease and in other complications associated with COVID-19 as well as in the mortality of COVID-19 p atients[62]

Methods

Results

Conclusion