Sevoflurane impairs m6A-mediated mRNA translation and leads to fine motor and cognitive deficits.

Clinical surgical practices have found that children who undergo multiple anesthesia may have an increased risk of deficiencies in cognition and fine motor control. Here, we report that YT521-B homology domain family 1 (YTHDF1), a critical reader protein for N6-methyladenosine-modified mRNA, was significantly downregulated in the prefrontal cortex of young mice after multiple sevoflurane anesthesia exposures. Importantly, sevoflurane led to a decrease in protein synthesis in mouse cortical neurons that was fully rescued by YTHDF1, suggesting that anesthesia may affect early brain development by affecting m6A-dependent mRNA translation. Transcriptome-wide experiments showed that numerous mRNA targets related to synaptic functions in the prefrontal mouse cortex were associated with m6A methylation and YTHDF1. In particular, we found that synaptophysin, a critical presynaptic protein, was specifically modified by m6A methylation and associated with YTHDF1, and m6A methylation of synaptophysin decreased with multiple sevoflurane exposures. Importantly, we showed that fine motor control skills and cognitive functions were impaired in mice with multiple anesthesia exposures, and these effects were fully reversed by reintroducing YTHDF1 through a blood-brain barrier (BBB)-crossing viral delivery system. Finally, we found that the fine motor skills in children who underwent prolonged anesthesia were compromised 6 months after surgery. Our findings indicated that impairment in the translational regulation of mRNA via N6-methyladenosine methylation is a potential mechanism underlying the effects of anesthesia on neural development in the young brain. 1. N6-methyladenosine (m6A) modifications were involved in anesthesia-induced neurotoxicity. 2. Sevoflurane impairs m6A-mediated mRNA translation and leads to fine motor deficits in young mice. 3. YTHDF1, a m6A reader protein, rescued sevoflurane-induced protein synthesis inhibition and fine motor deficits in young mice.

Sequence-specific extracellular microRNAs activate TLR7 and induce cytokine secretion and leukocyte migration.

Toll-like receptors (TLRs) can contribute to central nervous system disease pathologies via recognition of microRNAs (miRNAs); however, it remains to be determined which miRNAs are able to activate this signaling. Here we report that numerous miRNAs induced the production of tumor necrosis factor alpha in multiple myeloid cell types, including microglia, and that this effect was abolished in cells deficient in TLR7. Examination of closely related miRNAs that differed in their ability to activate TLR7 resulted in the identification of a motif (UGCUUAU) in miR-20a-5p and specific nucleotides (all the uridines and surprisingly the cytosine as well) in a key area of miR-20a-5p and miR-148b-3p that were vital for the secretion of cytokines via TLR7 stimulation. A 10-nucleotide sequence including this motif was identified to be the shortest single-stranded RNA to signal via TLR7. An miRNA containing this motif induced the secretion of multiple proinflammatory molecules, which was dependent on the phosphoinositide 3-kinase, mitogen-activated protein kinase, and nuclear factor kappa-light-chain-enhancer of activated B cell signaling pathways. Wild-type mice administered miR-20a-5p, which contained this motif, demonstrated increased leukocyte migration. This effect was significantly ameliorated in TLR7-knockout mice, and mice administered miR-20b-5p, in which the motif was mutated, did not exhibit leukocyte migration. We provide a detailed analysis of miRNAs that activate endosomal TLR7 and identify key nucleotide features of a sequence motif recognized by TLR7.

Neobaicalein prevents isoflurane anesthesia-induced cognitive impairment in neonatal mice via regulating CREB1.

OBJECTIVES: Isoflurane (ISO) is widely used in the clinic and research. The authors aimed to explore whether Neobaicalein (Neob) could protect neonatal mice from ISO-induced cognitive damage. METHOD: The open field test, Morris water maze test, and tail suspension test was performed to assess the cognitive function in mice. Enzyme-linked immunosorbent assay was used to evaluate inflammatory-related protein concentrations. Immunohistochemistry was used to assess Ionized calcium-Binding Adapter molecule-1 (IBA-1) expression. Hippocampal neuron viability was detected using the Cell Counting Kit-8 assay. Double immunofluorescence staining was employed to confirm the interaction between proteins. Western blotting was used to assess protein expression levels. RESULTS: Neob notably improved cognitive function and exhibited anti-inflammatory effects; moreover, under iso-treatment, it exhibited neuroprotective effects. Furthermore, Neob suppressed interleukin-1ß, tumor necrosis factor-α, and interleukin-6 levels and upregulated interleukin-10 levels in ISO-treated mice. Neob significantly mitigated iso-induced increases in IBA-1-positive cell numbers of the hippocampus in neonatal mice. Furthermore, it inhibited ISO-induced neuronal apoptosis. Mechanistically, Neob was observed to upregulate cAMP Response Element Binding protein (CREB1) phosphorylation and protected hippocampal neurons from ISO-mediated apoptosis. Moreover, it rescued ISO-induced abnormalities of synaptic protein. CONCLUSIONS: Neob prevented ISO anesthesia-induced cognitive impairment by suppressing apoptosis and inflammation through upregulating CREB1.

A comprehensive study to delineate the role of an extracellular vesicle-associated microRNA-29a in chronic methamphetamine use disorder.

Extracellular vesicles (EVs), which express a repertoire of cargo molecules (cf. proteins, microRNA, lipids, etc.), have been garnering a prominent role in the modulation of several cellular processes. Here, using both non-human primate and rodent model systems, we provide evidence that brain-derived EV (BDE) miRNA, miR-29a-3p (mir-29a), is significantly increased during chronic methamphetamine (MA) exposure. Further, miR-29a levels show significant increase both with drug-seeking and reinstatement in a rat MA self-administration model. We also show that EV-associated miR-29a is enriched in EV pool comprising of small EVs and exomeres and further plays a critical role in MA-induced inflammation and synaptodendritic damage. Furthermore, treatment with the anti-inflammatory drug ibudilast (AV411), which is known to reduce MA relapse, decreased the expression of miR-29a and subsequently attenuated inflammation and rescued synaptodendritic injury. Finally, using plasma from MUD subjects, we provide translational evidence that EV-miR29a could potentially serve as a biomarker to detect neuronal damage in humans diagnosed with MA use disorder (MUD). In summary, our work suggests that EV-associated miR-29a-3p plays a crucial role in MUD and might be used as a potential blood-based biomarker for detecting chronic inflammation and synaptic damage.

Multiple sevoflurane exposures don't disturb the T-cell receptor repertoire in infant rhesus monkeys' thymus.

AIMS: Multiple surgical procedures and anesthesia increase the risk of the development in children. However, the influence of such exposures on the developing childhood immunity organs is rarely reported. MATERIALS AND METHODS: High-throughput sequencing of T-cell receptor (TCR) repertoires (TCRseq) from rhesus monkeys' thymus was performed to investigate whether anesthetics could induce de novo antigen recognition via TCR or TCR development impairments. KEY FINDINGS: No significant difference between sevoflurane and control groups regarding VJ gene combinations and diversity of V and J gene was seen, nor was there an obvious change in similar average number of Complementarity Determining Region 3 (CDR3) aa clonotypes. Our analysis of Rank abundance, Gini coefficient, Simpson index, Normalized Shannon Diversity Entropy (NSDE), Morisita-Horn Similarity Index (MHSI) and Bhattacharyya Distance (BD) indicated there is no difference in TCR diversity and similarity. SIGNIFICANCE: These results suggest early events in thymic T cell development and repertoire generation are not abnormality after multiple sevoflurane exposure during childhood. The stabilization of the immune repertoires suggested the safety of sevoflurane in host immune response in children.

Neobaicalein prevents isoflurane anesthesia-induced cognitive impairment in neonatal mice via regulating CREB1

Abstract Objectives: Isoflurane (ISO) is widely used in the clinic and research. The authors aimed to explore whether Neobaicalein (Neob) could protect neonatal mice from ISO-induced cognitive damage. Method: The open field test, Morris water maze test, and tail suspension test was performed to assess the cognitive function in mice. Enzyme-linked immunosorbent assay was used to evaluate inflammatory-related protein concentrations. Immunohistochemistry was used to assess Ionized calcium-Binding Adapter molecule-1 (IBA-1) expression. Hippocampal neuron viability was detected using the Cell Counting Kit-8 assay. Double immunofluorescence staining was employed to confirm the interaction between proteins. Western blotting was used to assess protein expression levels. Results: Neob notably improved cognitive function and exhibited anti-inflammatory effects; moreover, under isotreatment, it exhibited neuroprotective effects. Furthermore, Neob suppressed interleukin-1β, tumor necrosis factor-α, and interleukin-6 levels and upregulated interleukin-10 levels in ISO-treated mice. Neob significantly mitigated iso-induced increases in IBA-1 - positive cell numbers of the hippocampus in neonatal mice. Furthermore, it inhibited ISO-induced neuronal apoptosis. Mechanistically, Neob was observed to upregulate cAMP Response Element Binding protein (CREB1) phosphorylation and protected hippocampal neurons from ISO-mediated apoptosis. Moreover, it rescued ISO-induced abnormalities of synaptic protein. Conclusions: Neob prevented ISO anesthesia-induced cognitive impairment by suppressing apoptosis and inflammation through upregulating CREB1.