Objective Acute-on-chronic liver failure (ACLF) has a high prevalence and short-term mortality. Monocytes play an important role in the development of ACLF. However, the monocyte subpopulations with unique features and functions in ACLF and associated with disease progression remain poorly understood. We investigated the specific monocyte subpopulations associated with ACLF progression and their roles in inflammatory responses using the single-cell RNA sequencing (scRNA-seq). Methods We performed scRNA-seq on 17,310 circulating monocytes from healthy controls and ACLF patients and genetically defined their subpopulations to characterize specific monocyte subpopulations associated with ACLF progression. Results Five monocyte subpopulations were obtained, including pro-inflammatory monocytes, CD16 monocytes, HLA monocytes, megakaryocyte-like monocytes, and NK-like monocytes. Comparisons of the monocytes between ACLF patients and healthy controls showed that the pro-inflammatory monocytes had the most significant gene changes, among which the expressions of genes related to inflammatory responses and cell metabolism were significantly increased while the genes related to cell cycle progression were significantly decreased. Furthermore, compared with the ACLF survival group, the ACLF death group had significantly higher expressions of pro-inflammatory cytokines (e.g., IL-6) and their receptors, chemokines (e.g., CCL4 and CCL5), and inflammation-inducing factors (e.g., HES4). Additionally, validation using scRNA-seq and flow cytometry revealed the presence of a cell type-specific transcriptional signature of pro-inflammatory monocytes THBS1, whose production might reflect the disease progression and poor prognosis. Conclusions We present the accurate classification, molecular markers, and signaling pathways of monocytes associated with ACLF progression. Therapies targeting pro-inflammatory monocytes may be a promising approach for blocking ACLF progression.

IntroductionPTGES3, also known as p23, is a molecule chaperone of Hsp90 that is involved in the pathogenesis of malignant tumors. Increasing studies have shown that PTGES3 plays a nonnegligible role in tumor development. However, analysis of PTGES3 in pan-cancer has not been performed yet.MethodsWe explored the role of PTGES3 in 33 types of tumors and depicted the potentialimmune-related pathways among them. Using multiple databases includingTCGA, LinkedOmics, GDSC, and TIMER, we made a comprehensive analysis to explore whether there was an interaction between PTGES3 and prognosis, DNA methylation, copy number variation (CNV), tumor mutational burden (TMB), microsatellite instability (MSI), and tumor immune microenvironment (TME).ResultsOur study revealed that PTGES3 expression level was upregulated in most cancers. PTGES3 was also associated with a positive or negative prognosis in a variety of cancers, which was mainly associated with DNA methylation, CNV, MSI, TMB, andmismatch repair-related genes. High PTGES3 expression was related to the infiltration of Th2 subsets of CD4+ T cells and immune checkpoint-related genes in most cancers, especially in hepatocellular carcinoma (HCC). Enrichment analysis demonstrated that PTGES3 was involved in cellular processes including DNA replication and spliceosome. The relationship between PTGES3 expression and HCC progression was verified at the protein level through immune histochemical analysis.ConclusionsOur research demonstrated theprognostic predictive value of PTGES3 in a wide range of cancers, which was alsoassociated with the process of tumor immune infiltration. As a result, it suggestedthat PTGES3 was a valuable prognostic biomarker in HCC treatment.

Background/Aims: The occurrence and development of hepatitis B virus-associated acute-onchronic liver failure (HBV-ACLF) is closely related to the immune pathway. We explored the heterogeneity of peripheral blood T cell subsets and the characteristics of exhausted T lymphocytes, in an attempt to identify potential therapeutic target molecules for immune dysfunction in ACLF patients.Methods: A total of 83,577 T cells from HBV-ACLF patients and healthy controls were screened for heterogeneity by single-cell RNA sequencing. In addition, exhausted T-lymphocyte subsets were screened to analyze their gene expression profiles, and their developmental trajectories were investigated. Subsequently, the expression of exhausted T cells and their capacity in secreting cytokines (interleukin 2, interferon γ, and tumor necrosis factor α) were validated by flow cytometry.Results: A total of eight stable clusters were identified, among which CD4 + TIGIT + subset and CD8 + LAG-3 + subset, with high expression of exhaust genes, were significantly higher in the HBV-ACLF patients than in normal controls. As shown by pseudotime analysis, T cells experienced a transition from naïve T cells to effector T cells and then exhausted T cells. Flow cytometry confirmed that the CD4 + TIGIT + subset and CD8 + LAG-3 + subset in the peripheral blood of the ACLF patients were significantly higher than those in the healthy controls. Moreover, in vitro cultured CD8 + LAG-3 + T cells were significantly fewer capable of secreting cytokines than CD8 + LAG-3subset.Conclusions: Peripheral blood T cells are heterogeneous in HBV-ACLF. The exhausted T cells markedly increase during the pathogenesis of ACLF, suggesting that T-cell exhaustion is involved in the immune dysfunction of HBV-ACLF patients. (Gut Liver 2024;18:520-530)