Lipopolysaccharides (LPS) of Gram-negative bacteria are critical for the defence against cytotoxic substances and must be transported from the inner membrane (IM) to the outer membrane (OM) through a bridge formed by seven membrane proteins (LptBFGCADE). The IM component LptB2FG powers the process through a yet unclarified mechanism. Here we report three high-resolution cryo-EM structures of LptB2FG alone and complexed with LptC (LptB2FGC), trapped in either the LPS- or AMP-PNP-bound state. The structures reveal conformational changes between these states and substrate binding with or without LptC. We identify two functional transmembrane arginine-containing loops interacting with the bound AMP-PNP and elucidate allosteric communications between the domains. AMP-PNP binding induces an inward rotation and shift of the transmembrane helices of LptFG and LptC to tighten the cavity, with the closure of two lateral gates, to eventually expel LPS into the bridge. Functional assays reveal the functionality of the LptF and LptG periplasmic domains. Our findings shed light on the LPS transport mechanism.

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Intracerebral haemorrhage (ICH) is a severe neurological disorder caused by bleeding within the brain tissue. Inflammation has been implicated in ICH pathogenesis and is a potential therapeutic target for ICH. Haemin, an activator of haem oxygenase-1 (HO-1), rapidly increases HO-1 protein expression and activity and has been shown to distinctly affect anti-inflammatory functions after central nervous system (CNS) injury. However, less is known about the mechanisms that underlie the antiinflammatory effects of haemin in aged rats post-ICH. Here, we performed microarray analysis to identify miRNAs that respond strongly to HO-1 regulation in ICH rats and found that miR-21-5p induced the most significant change. Using Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment and Gene Ontology (GO) analysis, we focused on dual-specificity phosphatase 8 (DUSP8) from the predicted miR-21-5p targets. Luciferase reporter assays confirmed that miR-21-5p bound directly to DUSP8. MiR-21-5p upregulation in vitro downregulated DUSP8 expression. Importantly, intracerebroventricularly injecting antagomir for miR-21-5p (A-miR-21-5p), which was used to inhibit miR-21-5p in aged ICH rats, significantly reduced the neurological defects, repaired cognitive impairment, alleviated blood-brain barrier (BBB) permeability, inhibited neuronal apoptosis posthaemorrhage and accelerated haematoma absorption. In addition, serum miR-21-5p levels were notably elevated in patients relative to healthy individuals and were correlated with National Institutes of Health Stroke Scale (NIHSS) scores and clinical outcomes. In summary, A-miR-21-5p increased HO-1 expression in cerebral haematomas, thus eliciting the

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Excessive production of amyloid β (Aβ) induced by familial mutations in amyloid precursor protein (APP) and presenilin 1 (PS1) results in neuronal oxidative insults, mitochondrial dysfunction, and apoptosis, which play an essential role in the pathological development of Alzheimer's disease (AD). Aloperine, a quinolizidine alkaloid derived from the leaves of the Sophora plant, has displayed multiple pharmacological functions in several chronic diseases. In the current study, we investigated the neuro-protective effects of aloperine against cytotoxicity in mouse Neuro2a (N2a) cells transfected with Swedish amyloid precursor protein (Swe-APP) mutant and presenilin 1 exon 9 deletion mutant (N2a/Swe.D9). We found that aloperine ameliorated oxidative stress patterns in N2a/Swe.D9 cells by reducing the production of reactive oxygen species (ROS) and 4-hydroxy-2-nonenal (4-HNE). Additionally, aloperine treatment led to elevated generation of ATP and increased mitochondrial membrane potential (MMP) in N2a/Swe.D9 cells. Importantly, we found that aloperine treatment reduced the vulnerability of N2a/Swe.D9 cells to HO. Aloperine also inhibited apoptosis of N2a/Swe.D9 cells via a mitochondria-dependent pathway. These findings suggest that aloperine may have pharmacological potential for the treatment of AD.

Growing evidence indicates that innate immune molecules regulate microglial activation in Alzheimer's disease (AD); however, their effects on amyloid pathology and neurodegeneration remain inconclusive. Here, we conditionally deleted one allele of myd88 gene specifically in microglia in APP/PS1‐transgenic mice by 6 months and analyzed AD‐associated pathologies by 9 months. We observed that heterozygous deletion of myd88 gene in microglia decreased cerebral amyloid β (Aβ) load and improved cognitive function of AD mice, which was correlated with reduced number of microglia in the brain and inhibited transcription of inflammatory genes, for example, tnf‐α and il‐1β, in both brain tissues and individual microglia. To investigate mechanisms underlying the pathological improvement, we observed that haploinsufficiency of MyD88 increased microglial recruitment toward Aβ deposits, which might facilitate Aβ clearance. Microglia with haploinsufficient expression of MyD88 also increased vasculature in the brain of APP/PS1‐transgenic mice, which was associated with up‐regulated transcription of osteopontin and insulin‐like growth factor genes in microglia. Moreover, MyD88‐haploinsufficient microglia elevated protein levels of LRP1 in cerebral capillaries of APP/PS1‐transgenic mice. Cell culture experiments further showed that treatments with interleukin‐1β decreased LRP1 expression in pericytes. In summary, haploinsufficiency of MyD88 in microglia at a late disease stage attenuates pro‐inflammatory activation and amyloid pathology, prevents the impairment of microvasculature and perhaps also protects LRP1‐mediated Aβ clearance in the brain of APP/PS1‐transgenic mice, all of which improves neuronal function of AD mice.

Alzheimer's disease (AD), the most common cause of dementia in the elderly, is pathologically characterized by extracellular deposition of amyloid‐β peptides (Aβ) and microglia‐dominated inflammatory activation in the brain. p38α‐MAPK is activated in both neurons and microglia. How p38α‐MAPK in microglia contributes to AD pathogenesis remains unclear. In this study, we conditionally knocked out p38α‐MAPK in all myeloid cells or specifically in microglia of APP‐transgenic mice, and examined animals for AD‐associated pathologies (i.e., cognitive deficits, Aβ pathology, and neuroinflammation) and individual microglia for their inflammatory activation and Aβ internalization at different disease stages (e.g., at 4 and 9 months of age). Our experiments showed that p38α‐MAPK‐deficient myeloid cells were more effective than p38α‐MAPK‐deficient microglia in reducing cerebral Aβ and neuronal impairment in APP‐transgenic mice. Deficiency of p38α‐MAPK in myeloid cells inhibited inflammatory activation of individual microglia at 4 months but enhanced it at 9 months. Inflammatory activation promoted microglial internalization of Aβ. Interestingly, p38α‐MAPK‐deficient myeloid cells reduced IL‐17a‐expressing CD4‐positive lymphocytes in 9 but not 4‐month‐old APP‐transgenic mice. By cross‐breeding APP‐transgenic mice with Il‐17a‐knockout mice, we observed that IL‐17a deficiency potentially activated microglia and reduced Aβ deposition in the brain as shown in 9‐month‐old myeloid p38α‐MAPK‐deficient AD mice. Thus, p38α‐MAPK deficiency in all myeloid cells, but not only in microglia, prevents AD progression. IL‐17a‐expressing lymphocytes may partially mediate the pathogenic role of p38α‐MAPK in peripheral myeloid cells. Our study supports p38α‐MAPK as a therapeutic target for AD patients.