No abstract

Background: Sepsis is a highly heterogeneous syndrome with diverse immune status and varied bioprocesses among individuals. The heterogeneity of sepsis could be associated with N6-methyladenosine (m6A) RNA methylation, due to m6A as a common and reversible posttranscriptional RNA modification involved in the regulation of whole bioprocesses. Therefore, we aim to identify m6A induced molecular subtypes of sepsis and furthermore explore the probable mechanism.Methods: Gene expression datasets with 479 consecutive patients admitted for sepsis to the intensive care unit (ICU) in the Amsterdam Academic Medical Center were included in present study at first. Secondly, twelve m6A methylation regulatory genes were determined via systematic review in published researches. Furthermore, we utilized unsupervised clustering (consensus k means clustering) to identify m6A induced molecular subtypes in sepsis based on m6A prognostic molecular, and assess the association of these subtypes with clinical traits and survival outcomes. Moreover, the probable mechanism and regulatory relationship of m6A in sepsis was also explored through Gene Set Enrichment Analysis (GSEA), Weighted gene coexpression network analysis (WGCNA), Gene Ontology (GO) analysis and Co-expression analysis.Results: Three m6A subtypes with different outcome were identified in sepsis cohort through unsupervised clustering on m6A prognostic molecular, designated Cluster 1/2/3 (log-rank P=0.004). The best outcome was found for patients classified as having cluster 3, and at 28 days, 21 of 144 people with cluster 3 had died [hazard ratio (HR) vs. all other clusters 5.42 (95% CI: 0.359-0.819); P=0.011], compared with 57 of 224 people with cluster 1 (HR 0.579, 95% CI: 0.364-0.920; P=0.037), and 36 of 112 people with cluster 2 (HR 0.477, 95% CI: 0.272-0.833; P=0.003). For exploration of the relationship between m6A subtypes and immunity, the GSEA found that patients in cluster 1 suffered from hyper-activated immunocompetent status; patients in cluster 2 indicated immunosuppressive status; and patients in cluster 3 showed the moderate immune activity (P<0.05). Co-expression analysis furthermore identified 82 immune molecules and 40 autophagy-related molecules could be regulated by prognostic m6A RNA methylation regulators (P<0.05) and correlation coefficient >0.6. In addition, WGCNA and GO analysis indicated that autophagy was significantly and widely activated in patients with cluster 3 (P<0.05).Conclusions: According to the heterogeneity in m6A methylation regulatory genes, three distinct subtypes in sepsis were identified with different RNA epigenetics, immune status, biological processes and outcomes, which initially uncovered that heterogeneity of sepsis may be largely caused by m6A RNA methylation.

Modulating immunoregulation of CD4 CD25 Treg is a critical mechanism for nicotinic anti-inflammatory pathway and it may be feasible to use selective agonists as an immunotherapy for SAP.

Accumulated evidence has demonstrated that the macrophage phenotypic switch from M0 to M1 is crucial in the initiation of the inflammatory process of acute respiratory distress syndrome (ARDS). Better insight into the molecular control of M1 macrophages in ARDS may identify potential therapeutic targets. In the current study, 36 candidate genes associated with the severity of ARDS and simultaneously involved in M1-polarized macrophages were first screened through a weighted network algorithm on all gene expression profiles from the 26 ARDS patients and empirical Bayes analysis on the gene expression profiles of macrophages. STAT1, IFIH1, GBP1, IFIT3, and IRF1 were subsequently identified as hub genes according to connectivity degree analysis and multiple external validations. Among these candidate genes, IFIH1 had the strongest connection with ARDS through the RobustRankAggreg algorithm. It was selected as a crucial gene for further investigation. For in vitro validation, the RAW264.7 cell line and BMDMs were transfected with shIFIH1 lentivirus and plasmid expression vectors of IFIH1. Cellular experimental studies further confirmed that IFIH1 was a novel regulator for promoting M1 macrophage polarization. Moreover, gene set enrichment analysis (GSEA) and in vitro validations indicated that IFIH1 regulated M1 polarization by activating IRF3. In addition, previous studies demonstrated that activation of IFIH1-IRF3 was stimulated by viral RNAs or RNA mimics. Surprisingly, the current study found that LPS could also induce IFIH1-IRF3 activation via a MyD88-dependent mechanism. We also found that only IFIH1 expression without LPS or RNA mimic stimulation could not affect IRF3 activation and M1 macrophage polarization. These findings were validated on two types of macrophages, RAW264.7 cells and BMDMs, which expanded the knowledge on the inflammatory roles of IFIH1 and IRF3, suggesting IFIH1 as a potential target for ARDS treatment.

Hyperbaric oxygen treatment (HBOT) has been used to reduce neuropathic pain. Melatonin and opioid receptors are involved in neuropathic pain, but it is not known if HBOT works through these pathways to achieve its antinociceptive effect. We divided anesthetized rats into two treatment and three sham groups. The two treatment groups received third-degree burns on their right hind paws, one treated in a hyperbaric chamber for a week and the other for two weeks. We evaluated the mechanical paw-withdrawal threshold (MWT) and expression of melatonin receptor 1 (MT1), melatonin receptor 2 (MT2), μ (MOR) and κ (KOR) opioid receptor, brain-derived neurotrophic factor (BDNF), Substance P, and calcitonin gene-related peptide (CGRP) in cuneate nucleus, dorsal horn, and hind paw skin by immunohistochemical, immunofluorescence assays and real-time quantitative polymerase chain reaction (RT-PCR). The group receiving one-week HBOT had increased expressions of MT1, MT2, MOR and KOR and decreased expressions of BDNF, Substance P, and CGRP. Their mechanically measured pain levels returned to normal within a week and lasted three weeks. This anti-allodynia effect lasted twice as long in those treated for two weeks. Our findings suggest that increasing the duration of HBOT can reduce burn-induced mechanical allodynia for an extended period of time in rats. The upregulation of melatonin and opioid receptors observed after one week of HBOT suggests they may be partly involved in attenuation of the mechanical allodynia. Downregulation of BDNF, substance P and CGRP may have also contributed to the overall beneficial effect of HBOT.