Study On Immune Features Of Cytokine Storm In COVID-19 Patients By Single-cell Transcriptomics And Bioinformatics

Coronavirus disease 2019(COVID-19)named by the World Health Organization is a contagious disease leading to an ongoing global pandemic,caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).Dysfunction of immune response in patients with COVID-19 is an important factor affecting the symptoms and lethality.The death risk of patients is related to the cytokine storm caused by SARS-CoV-2 infection.More and more evidence showed that patients with COVID-19 may suffer from cytokine release syndrome with high expression of pro-inflammatory cytokines,which is also one of the main causes of severe illness and death in patients with COVID19.Based on the mechanism of cytokine storm in patients with COVID-19,immunosuppressive treatments,such as tocilizumab,targeting interleukin-6 receptor(IL-6R)have been applied to ameliorate lethal inflammatory symptoms in patients with COVID-19.However,the cellular and molecular mechanism involved in the cytokine storm in COVID-19 patients is still unclear,and immunopathogenic mechanism of COVID-19 remains to be studied.Single-cell genomics,especially single-cell transcriptomics,being a cutting-edge technology in life science and biomedicine over recent years,has been widely applied in several scientific fields such as in neuroscience,embryonic development,tumor immunity,etc.,especially in the studies of clinical diseases,which provides unique and in-depth insights of the pathogenesis.Since the outbreak of COVID-19 pandemic,single-cell omics technology has gradually deepened the understanding of COVID-19,which contributed to medical intervention in prevention,antibody vaccine development and other aspects.In many directions of COVID-19 study,understanding the interaction pattern between the virus and the host after SARS-CoV-2 infection,especially the immune response caused by virus infection,has established an immunological basis for explaining pathogenesis and developing treatment strategies for COVID-19 patients.Single-cell RNA sequencing(scRNA-seq),a high throughput,low cost and high sensitivity method in detecting gene expression,is used as the main technique in our studies to investigate the immunological response and characteristics of cytokine storm in patients with COVID-19.This thesis mainly includes two parts:in the first part,we mainly focused on the immunological characteristics of cytokine storm in patients with COVID-19 after SARS-CoV-2 infection;In the second part,we mainly studied the common and disease-specific immune signatures related to cytokine storm in patients with a variety of infectious diseases(including COVID-19,sepsis and human immunodeficiency virus(HIV)infection).In the first part of the work,our research can be divided into two main contents:the study of cytokine storm based on two patients with severe COVID-19 and the study of cytokine storm based on a large number of COVID-19 cases.In the first section,we analyzed the peripheral immune cells of two patients with severe COVID-19 before and after treatment with tocilizumab.By comparing patients in severe stage,remission stage and healthy controls,we identified a monocyte subset associated with inflammatory cytokine storm.By differential expression analysis and cytokine ligand-receptor interaction analysis,we found that these monocytes can secrete many pro-inflammatory cytokines,and through cytokine-mediated network interacts with other immune cells,leading to cytokine storm.Since the samples used in this section were only from two severe patients,we subsequently joined the single cell alliance to further explore the sources of the cytokine storm using the single cell transcriptome data of large-scale samples.In this section,we first extensively explored a variety of cell subtypes of different cell lineages in this big dataset,defined seven hyper-inflammatory cell subtypes using the newly developed computational pipeline.By cell proportion analysis in the large queue,we found a monocyte subset and some megakaryocytes relating to cytokine storm.We applied the cell-cell communication algorithm to construct the cytokine ligand-receptor interaction landscapes among peripheral,lung,and peripherallung hyper-inflammatory cell subtypes,providing a resourceful cellular and molecular analysis for the origins and interactions of cytokine storms in peripheral and lung of COVID-19 patients.In the second part,we used several bioinformatics methods to conduct integrative analysis on a large-scale single-cell transcriptome datasets of peripheral blood mononuclear cells(PBMC)from COVID-19,sepsis,and of HIV infected patients.We used the newly developed pipeline to identify ten hyper-inflammatory cell subtypes,finding that monocyte subsets are the main source of the differences on transcriptome levels of these three infectious diseases.Through differential expression analysis of monocytes,on the one hand,we found that pro-inflammatory monocytes in COVID-19 and HIV highly express the gene signatures relating to cytokine storm;on the other hand,monocyte subsets in COVID-19 patients share immunosuppressive features with sepsis.To further explain the differences and heterogeneity of inflammation in patients with COVID-19,we constructed a "three-stage" model of heterogeneity.We found the heterogeneity of COVID-19 patients profiled in this model to be related to the inflammatory and immunosuppressive characteristics in monocytes.Finally,we discussed the application of relevant bioinformatics methods in the study of recurrent abortion.We systematically studied the relative changes of decidual immune microenvironment in patients with recurrent abortion by using a variety of bioinformatics methods,revealed the abnormal development of NK cells and the underlying disorder mechanism,and constructed a disease specific interaction network between the main cell types of decidual immune microenvironment,providing new insights for decidual immune microenvironment in patients with recurrent abortion.In conclusion,this work provides insights into cytokine storm caused by SARSCoV-2 infection at cellular and molecular levels from different perspectives,and providing potential resources for the treatment of COVID-19 patients.