Genetic Landscape And Immune Mechanism Of Monocytes In Developing Acute-on-Chronic Liver Failure

Objective:The unique function and subpopulation heterogeneity of monocytes in immunity in Acute-on-Chronic Liver Failure(ACLF)has not been fully elucidated,and this study aims to use single-cell RNA sequencing,sc RNA-seq)studies specific subpopulations of monocytes in ACLF disease and their immune mechanisms in inflammatory responses.Methods:We performed sc RNA-seq on peripheral blood mononuclear cells(PBMCs)of 3healthy controls(n=3)and 6 ACLF patients,including 3 ACLF survival ACLF and 3ACLF death groups.Cluster and annotation analysis were used to identify characteristics of healthy controls,ACLF survivors,ACF dead monocyte subsets,and ACLF disease progression.Results:Five monocytes subsets were obtained,namely pro-inflammatory monocytes,CD16 monocytes,HLA monocytes,megakaryocyte-like monocytes,and NK cell-like monocytes Monocytes)。 Pro-inflammatory Monocytes were the most significant in comparing monocytes from ACLF patients and healthy controls,respectively,with significantly increased expression of genes associated with inflammatory response,complement cascade,and cell metabolism,while genes associated with cell cycle progression were significantly reduced.In the comparison of ACLF disease groups,pro-inflammatory cytokines and their receptors(such as IL-6 and IL1B),chemokines(such as CCL4,CCL3,CXCL2)and inflammatory inducible factors(HES4)were further elevated in Pro-inflammatory Monocytes in the ACLF death group compared with the ACLF survival group.Through sc RNA-seq and flow cytometry analysis,it was found that the expression of THBS1 in Pro-inflammatory Monocytes was high,and the expression of pro-inflammatory factors such as TNF-α,IL-6,and IL-1β was increased.Mechanically,THBS1 activates the TGF-β/NK-κB signaling pathway,resulting in protein phosphorylation of signal transduction factors SMAD2 and SMAD3,which in turn stimulates the secretion of pro-inflammatory factors by immune cells and promotes the development of systemic inflammatory responses,indicating that intensified inflammation may be an influencing factor for the progression of ACLF disease and poor prognosis.Conclusion:Through the ACLF-derived PBMCs map,it was revealed that the pro-inflammatory monocytes subsets with unique pro-inflammatory functions were closely related to the progression of the disease,and the specific transcriptional characteristics of thrombin sensitive protein 1(Thrombospondin-1(THBS1)were discovered,which confirmed the specificity and potential of THBS1 as a biomarker of ACLF disease progression,and provided a new diagnostic and therapeutic target for reducing the mortality of ACLF.