Purpose Neuroinflammation is an essential causative factor in the pathogenesis and progression of cognitive impairment. The present study aims to evaluate the critical role of microRNA-138-5p (miR-138-5p) in hippocampal neuroinflammation and cognitive impairment through the NLRP3/caspase-1 signaling pathway in rats. Material and Methods We established the cognitive impairment rat model and RM (Rat microglia) microglial cellular inflammation model by intracerebroventricular (icv) injection or stimulation of lipopolysaccharide (LPS). Morris water maze (MWM) and Y-maze tests were performed to assess the cognitive behaviors. Quantitative real-time polymerase chain reaction (qRT-PCR), Enzyme-linked immune-sorbent assay (ELISA) and Western blot analysis were utilized to evaluate mRNA or protein expression. Bioinformatic analysis and dual-luciferase reporter gene assay were performed to verify the targeting relationship between NLRP3 and miR-138-5p. Besides, Hematoxylin and eosin (H&E) staining and immunohistochemistry were applied to observe the neuronal morphology and detect the positive cells of the hippocampus, respectively. Results Compared to the control groups, LPS-treated rats exhibited significantly impaired learning and memory in MWM and Y-maze tests. The expression of NLRP3, caspase-1 and pro-inflammation cytokines (IL-1β and IL-18) were upregulated, while miR-138-5p was downregulated both in rat hippocampus and RM cells treated with LPS. MiR-138-5p is downregulated in microarray data of cognitive impairment animals and could directly target the 3ʹ-UTR of NLRP3. Furthermore, upregulation of miR-138-5p improved impaired cognitive functions, while inhibited hippocampal neuroinflammation demonstrated by decreased expression of NLRP3/caspase-1 axis, pro-inflammation cytokines and microglial activation. This study demonstrates for the first time that miR-138-5p suppresses the hippocampal NLRP3/caspase-1 signaling pathway activation in cognition impaired rats. Conclusion The low expression of miR-138-5p after LPS administration may contribute to the activation of the NLRP3/caspase-1 pathway, leading to hippocampal neuroinflammation and cognitive impairment in rat models. These findings indicate a promising therapeutic avenue for cognitive disorders.
Acute lung injury (ALI) is a common lung pathology that is accompanied by alveolar macrophage (AM) activation and inflammatory response. This study investigated the role of the long non-coding RNA NONRATT004344 (hereafter named lncRNA NLRP3) in regulating the Nod-like receptor protein 3 (NLRP3)-triggered inflammatory response in early ALI and the underlying mechanism as well. We established LPS-induced ALI models to explore their interactive mechanisms in vitro and in vivo. Luciferase reporter assays were performed to determine that miR-138-5p could bind to lncRNA NLRP3 and NLRP3. We observed increased lncRNA NLRP3 expression, decreased miR-138-5p expression, NLRP3 inflammasome activation, and upregulated caspase-1, IL-1β, and IL-18 expression in the LPS-induced ALI model. Furthermore, lncRNA NLRP3 overexpression activated the NLRP3 inflammasome and promoted IL-1β and IL-18 secretion; the miR-138-5p mimic abolished these effects in vivo and in vitro. Consistently, miR-138-5p inhibition reversed the effects of lncRNA NLRP3 silencing on the expression of NLRP3-related molecules and inhibition of the NLRP3/caspase-1/IL-1β signalling pathway. Mechanistically, lncRNA NLRP3 sponging miR-138-5p facilitated NLRP3 activation through a competitive endogenous RNA (ceRNA) mechanism. In summary, our results suggested that lncRNA NLRP3 binding miR-138-5p promotes NLRP3-triggered inflammatory response via lncRNA NLRP3/miR-138-5p/NLRP3 ceRNA network (ceRNET) and provides insights into the treatment of early ALI.
ObjectivesSepsis is a clinical disease that is typically treated in the intensive care unit, and the complex pathophysiology under this disease has not been thoroughly understood. While ferroptosis is involved in inflammation and infection, its effect in sepsis is still unknown. The study aimed to identify ferroptosis-related genes in sepsis, providing translational potential therapeutic targets.MethodsThe dataset GSE65682 was used to download the sample source from the Gene Expression Omnibus (GEO) database. Consensus weighted gene co-expression network analysis (WGCNA) was performed to find suspected modules of sepsis. The differentially expressed genes (DEGs) most significantly associated with mortality were intersected with those altered by lipopolysaccharide (LPS) treatment and were further analyzed for the identification of main pathways of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The related pathway markers were further verified by qPCR.ResultsA total of 802 blood samples with sepsis were included for WGCNA, which identified 21 modules. Intersected with ferroptosis databases and LPS treatment groups, we identified two ferroptosis-related genes: PEBP1 and LPIN1. Only LPIN1 contributes to a poor outcome. Then, 205 DEGs were further identified according to the high or low LPIN1 expression. Among them, we constructed a gene regulatory network with several transcriptional factors using the NetworkAnalyst online tool and identified that these genes mostly correlate with inflammation and immune response. The immune infiltration analysis showed that lower expression of LPIN1 was related to macrophage infiltration and could be an independent predictor factor of the survival status in sepsis patients. Meanwhile, the multivariate Cox analysis showed that LPIN1 had a significant correlation with survival that was further verified by in vitro and in vivo experiments.ConclusionIn conclusion, LPIN1 could become a reliable biomarker for patient survival in sepsis, which is associated with immune and inflammation status.
Purpose To explore the molecular mechanism and search for candidate lncRNA and mRNA associated with pyroptosis in the gene expression profile of LPS-induced acute lung injury (ALI). Methods We investigated lncRNA and mRNA expression in lipopolysaccharide (LPS)-induced ALI at an early stage. RNA sequencing (RNA-Seq) was carried out to analyze lncRNA and mRNA expression profiles between the LPS-induced and control groups. We used bioinformatics analysis to predict target genes of early differential lncRNAs among obtained the differential mRNAs. Results A total of 78 lncRNAs and 248 mRNAs were upregulated at 2 hours and downregulated at 9 hours, and 21 lncRNAs and 107 mRNAs were downregulated at 2 and upregulated at 9 hours in early ALI models. We predicted 7 cis-and trans-regulated target genes of the top 20 lncRNAs. Gene Ontology (GO) analysis indicated that the target genes for the screened lncRNAs were most enriched in three-terms: regulation of protein serine/threonine kinase activity, pertussis, and cellular response to LPS. Additionally, target genes of lncRNAs were the top three enriched in pertussis, osteoclast differentiation, and cAMP signaling pathways with Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. We also identified vital mRNAs and lncRNAs. Protein-protein interaction (PPI) network analysis suggested that Tnf, Jun, and Atf3 were the top three key genes. Hub lncRNA4344 (NONRATT004344.2) and cis-regulated target mRNA (NLRP3) were validated in vitro. Finally, luciferase assay results confirmed that lncRNA4344 sponged miR‐138-5p to promote pyroptosis in inflammatory responses to LPS‐induced acute lung injury by targeting NLRP3. Conclusion Based on analysis of lncRNA and mRNA expression profiles by RNA-Seq and experimental verification, this study is the first to reveal that lncRNA4344 sponged miR‐138-5p to promote pyroptosis in inflammatory responses of LPS‐induced acute lung injury by targeting NLRP3. These newly identified lncRNA, miRNA, and mRNA might be novel potential targets for early treatment and prevention in early ALI.
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