ARIH1 inhibits influenza A virus replication and facilitates RIG-I dependent immune signaling by interacting with SQSTM1/p62.

BACKGROUND: Multiple host factors are involved in modulating type I interferon expression induced by viruses; however, the mechanism is not fully elucidated. Influenza A virus infection causes severe respiratory symptoms and triggers a series of signaling cascades and host innate immune responses, including interferon production. The co-IP/MS technology was used to screen several antiviral factors in the early stage. Among these factors, ariadne-1 homolog (ARIH1) caught our attention. METHODS: Western blot assay was performed to detect the level of proteins and software ImageJ was used to analyze the band intensities. Polymerase activity assay was conducted to evaluate the polymerase activity of influenza A virus. Tissue culture infective dose (TCID50) assay was performed to measure influenza A virus titers, and quantitative RT-PCR assay was applied to test the mRNA level of IFN-ß, ISG56, and CXCL10. Luciferase reporter assay was used to confirm the target of ARIH1 in RIG-I signaling. Immunoprecipitation assay was performed to detect the interaction and the ubiquitination of the proteins. All data were analyzed by biostatistical methods and presented as means ± standard deviation from three independent experiments. Statistical significance was determined using two-tailed student's t test. A P value of less than 0.05 was considered statistically significant, and a P value of less than 0.01 was considered highly significant (ns, P ≥ 0.05; *, P < 0.05; and **, P < 0.01). RESULTS: We found that ARIH1, a member of E3 ubiquitin ligases, enhanced cellular antiviral responses. Subsequent study showed that ARIH1 was up-regulated during influenza A virus infection. Further analysis showed that ARIH1 enhanced IFN-ß and downstream gene expression by affecting the degradation of RIG-I through the SQSTM1/p62 signaling pathway. CONCLUSION: This newly revealed mechanism shows that cellular response increases of ARIH1 and promotes IFN-ß expression to boost host survival during viral infection.

Transcriptomic analysis and laboratory experiments reveal potential critical genes and regulatory mechanisms in sepsis-associated acute kidney injury.

Background: Sepsis-associated acute kidney injury (SA-AKI) is one of the most frequent and serious complications of sepsis. However, the transcriptional regulatory network of the pathophysiological mechanism of the kidney has not been revealed. This study identified new mechanisms in SA-AKI using bioinformatics analyses and laboratory-based experiments. Methods: We performed transcriptomic profiling of mouse kidneys after cecal ligation and puncture (CLP) to mimic clinical sepsis. RNA from kidney samples from the CLP and control groups was isolated and analyzed using bulk messenger RNA (mRNA)-seq. Differentially expressed genes (DEGs) between the two groups were identified, and GO, KEGG and GSEA pathway enrichment analyses were performed. The protein-protein interaction (PPI) network of DEGs and hub genes was analyzed. The hub genes were verified using quantitative real-time polymerase chain reaction (qPCR) or Western blotting. The interaction network, targeted microRNAs (miRNAs) and long noncoding RNAs (lncRNAs) of hub genes were predicted, and the critical miRNA-hub gene regulatory axis was verified using qPCR, Western blotting, malondialdehyde (MDA) determination and flow cytometry. Correlation analyses of N6-adenosine methylation (m6A) RNA methylation regulators and hub genes and m6A modification analysis were performed. Results: A total of 4,754 DEGs were identified between the two groups using high-throughput sequencing. The pathways in which DEGs were enriched included ferroptosis (the highest enrichment score), apoptosis, and the PI3K-Akt, NF-kappa B and IL-17 signaling pathways. Seven (Hmox1, Spp1, Socs3, Mapk14, Lcn2, Cxcl1 and Cxcl12) of the 15 hub genes were involved in the KEGG pathway. mmu-miR-7212-5p-Hmox1 was a key RNA regulatory axis in ferroptosis. m6A RNA methylation modifications were involved in SA-AKI. The correlation analyses showed the close interactions among the m6A RNA methylation regulators and important hub genes. Conclusions: The findings of this study provide new insights into the mechanism regulating the occurrence and progression of SA-AKI. The mmu-miR-7212-5p-Hmox1 axis in ferroptosis and m6A RNA methylation regulators may have potential clinical significance for the future treatment of SA-AKI. The datasets generated for this study can be found in the repository of the GEO database (Series number: GSE186822).

Methylsulfonylmethane protects against lethal dose MRSA-induced sepsis through promoting M2 macrophage polarization.

BACKGROUND: Multi-drug-resistant bacterial infections, which have become a global threat, lack effective treatments. The discoveries of non-antibiotics with different modes of antibacterial action, such as methylsulfonylmethane (MSM), are a promising new treatment for multi-drug-resistant pathogens. METHODS: We constructed a mouse peritonitis infection model to evaluate the effects of MSM against methicillin-resistant Staphylococcus aureus (MRSA) infection. The time-kill kinetics of MSM against MRSA and the effect of MSM on the integrity of bacterial cell membrane were measured. Viability effects of MSM on THP1 cells were performed by CCK-8 cytotoxicity assay. Systematic inflammatory factor levels of mice were detected using ELISA. The immune response of peritoneal macrophages during MRSA-infection was evaluated using RNA sequencing. Gene Ontology function, Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses, and correlation analyses were applied to analysis RNA sequencing data. RT-qPCR, western blotting and flow cytometry were performed to analysis the gene and protein expression levels of macrophages. RESULTS: In in vitro experiments, MSM did not show significant killing effects on the growth of MRSA directly and did not destroy bacterial membrane integrity. MSM also displayed no significant effects on the proliferative capacity of THP1 cells. However, MSM treatment protected mice against a lethal dose MRSA-infection and decreased systemic inflammation. MSM upregulated metabolic pathway in peritoneal macrophages, especial glycolysis, during MRSA infection. MSM increased the expression of M2 markers (such as Arg1), promoted phosphorylation of STAT3 (which regulates M2 polarization), and decreased the expression of M1 markers in peritoneal macrophages. Additionally, MSM treatment increased the expression of H3K18 lactylation specific target genes, including Arg1. GNE-140, the LDHA-specific inhibitor of glycolysis, blocked the MSM-induced Arg1 expression in this disease model. CONCLUSIONS: MSM protects against MRSA infection through immunomodulation. MSM promotes the expression of Arg1 by lactate-H3K18la pathway to control macrophage to M2 polarization; it firstly provides therapeutic potential for drug-resistant infections and sepsis.