Delirium is very common in patients admitted to intensive care unit (ICU), and may worsen survival in these patients. Several meta-analyses have evaluated the antidelirium effects of dexmedetomidine in ICU patients, but their findings were inconsistent. Recently, several large multicentre randomised clinical trials (RCTs) were published, but they have not yet to be included in any meta-analysis. We will conduct a meta-analysis adding these data to evaluate the effects of dexmedetomidine on delirium and mortality in ICU patients, aiming to terminate controversy and provide robust evidence for guiding clinical practice.

Transcription factor Foxm1 plays a critical role during embryonic development and its expression is repressed during retinoic acid (RA)-induced differentiation of pluripotent P19 embryonal carcinoma cells at the early stage, correlated with downregulation of expression of pluripotency markers. To study whether Foxm1 participates in the maintenance of pluripotency of stem cells, we knock down Foxm1 expression in P19 cells and identify that Oct4 are regulated directly by Foxm1. Knockdown of Foxm1 also results in spontaneous differentiation of P19 cells to mesodermal derivatives, such as muscle and adipose tissues. Maintaining Foxm1 expression prevents the downregulation of pluripotency-related transcription factors such as Oct4 and Nanog during P19 cell differentiation. Furthermore, overexpression of FOXM1 alone in RA-differentiated P19 cells (4 days) or human newborn fibroblasts restarts the expression of pluripotent genes Oct4, Nanog and Sox2. Together, our results suggest a critical involvement of Foxm1 in maintenance of stem cell pluripotency.

Tissue regeneration relies on resident stem cells (SCs), whose activity and lineage choices are influenced by the microenvironment. Exploiting the synchronized, cyclical bouts of tissue regeneration in hair follicles (HFs), we investigate how microenvironment dynamics shape the emergence of stem cell lineages. Employing epigenetic and ChIP-seq profiling, we uncover how signal-dependent transcription factors couple spatiotemporal cues to chromatin dynamics, thereby choreographing stem cell lineages. Using enhancer-driven reporters, mutagenesis, and genetics, we show that simultaneous BMP-inhibitory and WNT signals set the stage for lineage choices by establishing chromatin platforms permissive for diversification. Mechanistically, when binding of BMP effector pSMAD1 is relieved, enhancers driving HF-stem cell master regulators are silenced. Concomitantly, multipotent, lineage-fated enhancers silent in HF-stem cells become activated by exchanging WNT effectors TCF3/4 for LEF1. Throughout regeneration, lineage enhancers continue reliance upon LEF1 but then achieve specificity by accommodating additional incoming signaling effectors. Barriers to progenitor plasticity increase when diverse, signal-sensitive transcription factors shape LEF1-regulated enhancer dynamics.

The genetic diversity and relationships among ethnic minority populations of southwest China were investigated using seven polymorphic restriction enzyme sites in the β-globin gene cluster. The haplotypes of 1392 chromosomes from ten ethnic populations living in southwest China were determined. Linkage equilibrium and recombination hotspot were found between the 5' sites and 3' sites of the β-globin gene cluster. 5' haplotypes 2 (+---), 6 (-++-+), 9 (-++++) and 3' haplotype FW3 (-+) were the predominant haplotypes. Notably, haplotype 9 frequency was significantly high in the southwest populations, indicating their difference with other Chinese. The interpopulation differentiation of southwest Chinese minority populations is less than those in populations of northern China and other continents. Phylogenetic analysis shows that populations sharing same ethnic origin or language clustered to each other, indicating current β-globin cluster diversity in the Chinese populations reflects their ethnic origin and linguistic affiliations to a great extent. This study characterizes β-globin gene cluster haplotypes in southwest Chinese minorities for the first time, and reveals the genetic variability and affinity of these populations using β-globin cluster haplotype frequencies. The results suggest that ethnic origin plays an important role in shaping variations of the β-globin gene cluster in the southwestern ethnic populations of China.

Gata3 is a DNA-binding transcription factor involved in cellular differentiation in a variety of tissues including inner ear, hair follicle, kidney, mammary gland and T-cells. In a previous study in 2009, Maeda et al. (Dev. Dyn.238, 2280-2291; doi:10.1002/dvdy.22035) found that Gata3 mutants could be rescued from midgestational lethality by the expression of a Gata3 transgene in sympathoadrenal neuroendocrine cells. The rescued embryos clearly showed multiple defects in lens fibre cell differentiation. To determine whether these defects were truly due to the loss of Gata3 expression in the lens, we generated a lens-specific Gata3 loss-of-function model. Analogous to the previous findings, our Gata3 null embryos showed abnormal regulation of cell cycle exit during lens fibre cell differentiation, marked by reduction in the expression of the cyclin-dependent kinase inhibitors Cdkn1b/p27 and Cdkn1c/p57, and the retention of nuclei accompanied by downregulation of Dnase IIβ. Comparisons of transcriptomes between control and mutated lenses by RNA-Seq revealed dysregulation of lens-specific crystallin genes and intermediate filament protein Bfsp2. Both Cdkn1b/p27 and Cdkn1c/p57 loci are occupied in vivo by Gata3, as well as Prox1 and c-Jun, in lens chromatin. Collectively, our studies suggest that Gata3 regulates lens differentiation through the direct regulation of the Cdkn1b/p27and Cdkn1c/p57 expression, and the direct/or indirect transcriptional control of Bfsp2 and Dnase IIβ.

During brain development, volatile anesthetic can rapidly interfere with physiologic patterns of dendritic development and synaptogenesis and impair the formation of precise neuronal circuits. KCC2 plays vital roles in spine development and synaptogenesis through its Cl- transport function and structural interactions with the spine cytoskeleton protein 4.1 N. The aim of this study was to dissect the mechanism of volatile anesthetics, which impair dendritic development and synaptogenesis via mediation of KCC2 cleavage.

Biological control mechanisms of plant diseases have been intensively studied. However, how plant pathogens respond to and resist or alleviate biocontrol agents remains largely unknown. In this study, a comparative transcriptome analysis was performed to elucidate how the pathogen of sclerotinia stem rot, Sclerotinia sclerotiorum, responds and resists to the biocontrol agent, Bacillus amyloliquefaciens. Results revealed that a total of 2,373 genes were differentially expressed in S. sclerotiorum samples treated with B. amyloliquefaciens fermentation broth (TS) when compared to control samples (CS). Among these genes, 2,017 were upregulated and 356 were downregulated. Further analyses indicated that various genes related to fungal cell wall and cell membrane synthesis, antioxidants, and the autophagy pathway were significantly upregulated, including glucan synthesis, ergosterol biosynthesis pathway, fatty acid synthase, heme-binding peroxidase related to oxidative stress, glutathione S-transferase, ABC transporter, and autophagy-related genes. These results suggest that S. sclerotiorum recruits numerous genes to respond to or resist the biocontrol of B. amyloliquefaciens. Thus, this study serves as a valuable resource regarding the mechanisms of fungal pathogen resistance to biocontrol agents.

The tea green leafhopper Empoasca onukii Matsuda (Hemiptera: Cicadellidae), the orange spiny whitefly, Aleurocanthus spiniferus (Quaintanca) (Hemiptera: Aleyrodidae), and the green plant bugs Apolygus lucorum Meyer-Dür (Hemiptera: Miridae) are the important piercing-sucking herbivores in tea trees Camellia sinensis (L.) O. Kuntze (Theaceae). The goal of this study was to evaluate the laboratory toxicities and field control efficacies of botanical insecticides including matrine, azadirachtin, veratrine, and pyrethrin to three tea pests. Via leaf-dip bioassay, toxicity tests with botanical insecticides indicated that there were significant differences between the LC50 values for botanical insecticides within the same insect species. Matrine had the highest toxicity to E. onukii, A. spiniferus, and A. lucorum with the LC50 values of 2.35, 13.10, and 44.88 mg/liter, respectively. Field tests showed that, among four botanical insecticides, matrine at dose of 9 g a.i. ha-1 can significantly reduce the numbers of E. onukii and A. spiniferus and the infestation of A. lucorum on the tea plants. Furthermore, botanical insecticides matrine and azadirachtin had no obvious influence on the coccinellids, spiders, and parasitoids densities in tea plantations. The results of this study indicated that use of botanical insecticides, such as matrine, has the potential to manipulate the population of E. onukii, A. spiniferus, and A. lucorum and will be an effective and environmentally compatible strategy for the control of tea pests.

Tet enzymes (Tet1/2/3) oxidize 5-methylcytosine to promote DNA demethylation and partner with chromatin modifiers to regulate gene expression. Tet1 is highly expressed in embryonic stem cells (ESCs), but its enzymatic and non-enzymatic roles in gene regulation are not dissected. We have generated Tet1 catalytically inactive (Tet1m/m) and knockout (Tet1-/-) ESCs and mice to study these functions. Loss of Tet1, but not loss of its catalytic activity, caused aberrant upregulation of bivalent (H3K4me3+; H3K27me3+) developmental genes, leading to defects in differentiation. Wild-type and catalytic-mutant Tet1 occupied similar genomic loci which overlapped with H3K27 tri-methyltransferase PRC2 and the deacetylase complex Sin3a at promoters of bivalent genes and with the helicase Chd4 at active genes. Loss of Tet1, but not loss of its catalytic activity, impaired enrichment of PRC2 and Sin3a at bivalent promoters leading to reduced H3K27 trimethylation and deacetylation, respectively, in absence of any changes in DNA methylation. Tet1-/-, but not Tet1m/m, embryos expressed higher levels of Gata6 and were developmentally delayed. Thus, the critical functions of Tet1 in ESCs and early development are mediated through its non-catalytic roles in regulating H3K27 modifications to silence developmental genes, and are more important than its catalytic functions in DNA demethylation.

Various lines of evidence suggest that neonatal exposure to general anesthetics, especially repeatedly, results in neuropathological brain changes and long-term cognitive impairment. Although progress has been made in experimental models, the exact mechanism of GA-induced neurotoxicity in the developing brain remains to be clarified. Sirtuin 1 (SIRT1) plays an important role in synaptic plasticity and cognitive performance, and its abnormal reduction is associated with cognitive dysfunction in neurodegenerative diseases. However, the role of SIRT1 in GA-induced neurotoxicity is unclear to date. In this study, we found that the protein level of SIRT1 was inhibited in the hippocampi of developing mice exposed to sevoflurane. Furthermore, the SIRT1 inhibition in hippocampi was associated with brain-derived neurotrophic factor (BDNF) downregulation modulated by methyl-cytosine-phosphate-guanine-binding protein 2 (MeCP2) and cAMP response element-binding protein (CREB). Pretreatment of neonatal mice with resveratrol nearly reversed the reduction in hippocampal SIRT1 expression, which increased the expression of BDNF in developing mice exposed to sevoflurane. Moreover, changes in the levels of CREB and MeCP2, which were considered to interact with BDNF promoter IV, were also rescued by resveratrol. Furthermore, resveratrol improved the cognitive performance in the Morris water maze test of the adult mice with exposure to sevoflurane in the neonatal stage, without changing motor function in the open field test. Taken together, our findings suggested that SIRT1 deficiency regulated BDNF signaling via regulation of the epigenetic activity of MeCP2 and CREB, and resveratrol might be a promising agent for mitigating sevoflurane-induced neurotoxicity in developing mice.

Surgical pain is associated with delirium in patients, and acupuncture can treat pain. However, whether electroacupuncture can attenuate the surgical pain-associated delirium via the gut-brain axis remains unknown. Leveraging a mouse model of foot incision-induced surgical pain and delirium-like behavior, we found that electroacupuncture stimulation at specific acupoints (e.g., DU20+KI1) attenuated both surgical pain and delirium-like behavior in mice. Mechanistically, mice with incision-induced surgical pain and delirium-like behavior showed gut microbiota imbalance, microglia activation in the spinal cord, somatosensory cortex, and hippocampus, as well as an enhanced dendritic spine elimination in cortex revealed by two-photon imaging. The electroacupuncture regimen that alleviated surgical pain and delirium-like behavior in mice also effectively restored the gut microbiota balance, prevented the microglia activation, and reversed the dendritic spine elimination. These data demonstrated a potentially important gut-brain interactive mechanism underlying the surgical pain-induced delirium in mice. Pending further studies, these findings revealed a possible therapeutic approach in preventing and/or treating postoperative delirium by using perioperative electroacupuncture stimulation in patients.

Stroke is a major public health concern worldwide. The lack of effective therapies heightens the need for new therapeutic agents. Previous study identified sirtuin 5 (SIRT5) as a positive regulator of microglia-induced excessive neuroinflammation following ischemic stroke. Interventions targeting SIRT5 should therefore alleviate neuroinflammation and protect against ischemic stroke. Here, we synthesized a membrane-permeable peptide specifically bound to SIRT5 through a chaperone-mediated autophagy targeting motif (Tat-SIRT5-CTM) and examined its therapeutic effect in vitro and in vivo. First, in primary microglia, Tat-SIRT5-CTM suppressed the binding of SIRT5 with annexin-A1 (ANXA1), leading to SIRT5 degradation and thus inhibition of SIRT5-mediated desuccinylation of ANXA1, followed by increased membrane accumulation and secretion of ANXA1. These changes, in turn, alleviated microglia-induced neuroinflammation. Moreover, following intravenous injection, Tat-SIRT5-CTM could efficiently pass through the blood‒brain barrier. Importantly, systemic administration of Tat-SIRT5-CTM reduced the brain infarct area and neuronal loss, mitigated neurological deficit scores, and improved long-term neurological functions in a mouse model of ischemic stroke. Furthermore, no toxicity was observed when high doses Tat-SIRT5-CTM were injected into nonischemic mice. Collectively, our study reveals the promising efficacy of the peptide-directed lysosomal degradation of SIRT5 and suggests it as an effective therapeutic approach for the treatment of ischemic stroke.

The recognition of the scorpion toxin maurotoxin (MTX) by the voltage-gated potassium (Kv1) channels, Kv1.1, Kv1.2, and Kv1.3, has been studied by means of Brownian dynamics (BD) simulations. All of the 35 available structures of MTX in the Protein Data Bank (http://www.rcsb.org/pdb) determined by nuclear magnetic resonance were considered during the simulations, which indicated that the conformation of MTX significantly affected both the recognition and the binding between MTX and the Kv1 channels. Comparing the top five highest-frequency structures of MTX binding to the Kv1 channels, we found that the Kv1.2 channel, with the highest docking frequencies and the lowest electrostatic interaction energies, was the most favorable for MTX binding, whereas Kv1.1 was intermediate, and Kv1.3 was the least favorable one. Among the 35 structures of MTX, the 10th structure docked into the binding site of the Kv1.2 channel with the highest probability and the most favorable electrostatic interactions. From the MTX-Kv1.2 binding model, we identified the critical residues for the recognition of these two proteins through triplet contact analyses. MTX locates around the extracellular mouth of the Kv1 channels, making contacts with its beta-sheets. Lys23, a conserved amino acid in the scorpion toxins, protrudes into the pore of the Kv1.2 channel and forms two hydrogen bonds with the conserved residues Gly401(D) and Tyr400(C) and one hydrophobic contact with Gly401(C) of the Kv1.2 channel. The critical triplet contacts for recognition between MTX and the Kv1.2 channel are Lys23(MTX)-Asp402(C)(Kv1), Lys27(MTX)-Asp378(D)(Kv1), and Lys30(MTX)-Asp402(A)(Kv1). In addition, six hydrogen-bonding interactions are formed between residues Lys23, Lys27, Lys30, and Tyr32 of MTX and residues Gly401, Tyr400, Asp402, Asp378, and Thr406 of Kv1.2. Many of them are formed by side chains of residues of MTX and backbone atoms of the Kv1.2 channel. Five hydrophobic contacts exist between residues Pro20, Lys23, Lys30 and Tyr32 of MTX and residues Asp402, Val404, Gly401, and Arg377 of the Kv1.2 channel. The simulation results are in agreement with the previous molecular biology experiments and explain the binding phenomena between MTX and Kv1 channels at the molecular level. The consistency between the results of the BD simulations and the experimental data indicated that our three-dimensional model of the MTX-Kv1.2 channel complex is reasonable and can be used in additional biological studies, such as rational design of novel therapeutic agents blocking the voltage-gated channels and in mutagenesis studies in both the toxins and the Kv1 channels. In particular, both the BD simulations and the molecular mechanics refinements indicate that residue Asp378 of the Kv1.2 channel is critical for its recognition and binding functionality toward MTX. This phenomenon has not been appreciated in the previous mutagenesis experiments, indicating this might be a new clue for additional functional study of Kv1 channels.

Alginate is known to prevent elimination of Pseudomonas aeruginosa biofilms. Alginate lyase (AlgL) might therefore facilitate treatment of Pseudomonas aeruginosa-infected cystic fibrosis patients. However, the catalytic activity of wild-type AlgL is not sufficiently high. Therefore, molecular modeling and site-directed mutagenesis of AlgL might assist in enzyme engineering for therapeutic development. AlgL, isolated from Azotobacter vinelandii, catalyzes depolymerization of alginate via a β-elimination reaction. AlgL was modeled based on the crystal structure template of Sphingomonas AlgL species A1-III. Based on this computational analysis, AlgL was subjected to site-directed mutagenesis to improve its catalytic activity. The kcat/Km of the K194E mutant showed a nearly 5-fold increase against the acetylated alginate substrate, as compared to the wild-type. Double and triple mutants (K194E/K245D, K245D/K319A, K194E/K245D/E312D, and K194E/K245D/K319A) were also prepared. The most potent mutant was observed to be K194E/K245D/K319A, which has a 10-fold improved kcat value (against acetylated alginate) compared to the wild-type enzyme. The antibiofilm effect of both AlgL forms was identified in combination with piperacillin/tazobactam (PT) and the disruption effect was significantly higher in mutant AlgL combined with PT than wild-type AlgL. However, for both the wild-type and K194E/K245D/K319A mutant, the use of the AlgL enzyme alone did not show significant antibiofilm effect.

Truncated tissue factor (tTF) fusion protein targeting tumor vasculature can induce tumor vascular thrombosis and necrosis. Here, we generated (RGD)₃-tTF in which three arginine-glycine-aspartic (RGD) targeting integrin α(v)β₃ and tTF induce blood coagulation in tumor vessels.

Cardiac microvascular endothelial cells (CMECs) dysfunction induced by hypoxia is an important pathophysiological event in myocardium ischemic injury, whereas, the underlying mechanism is not fully clarified. FoxO transcription factors regulate target genes involved in apoptosis and cellular reactive oxygen species (ROS) production. Therefore, the present study was designed to elucidate the potential role of FoxOs on the hypoxia-induced ROS formation and apoptosis in CMECs. Exposure to low oxygen tension stimulated ROS accumulation and increased apoptosis in CMECs within 6-24 h. Hypoxia also significantly increased the expressions of HIF-1α and FoxO3a. However, hypoxia decreased the phosphorylation of Akt and FoxO3a, correlated with increased nuclear accumulation. Conversely, the expression of FoxO1 was not significantly altered by hypoxia. After inhibition of HIF-1α by siRNA, we observed that hypoxia-induced ROS accumulation and apoptosis of CMECs were decreased. Meanwhile, knockdown of HIF-1α also inhibited hypoxia induced FoxO3a expression in CMECs, but did not affect FoxO1 expression. Furthermore, hypoxia-induced ROS formation and apoptosis in CMECs were correlated with the disturbance of Bcl-2 family proteins, which were abolished by FoxO3a silencing with siRNA. In conclusion, our data provide evidence that FoxO3a leads to ROS accumulation in CMECs, and in parallel, induces the disturbance of Bcl-2 family proteins which results in apoptosis.

Cocaine binds and inhibits dopamine transporter (DAT), norepinephrine transporter (NET) and serotonin transporter. The residues forming cocaine binding sites are unknown. RTI-113, a cocaine analog, is 100× more potent at inhibiting DAT than inhibiting NET. Here we show that removing the hydroxyl group from residue Tyr151 in NET by replacing it with Phe, the corresponding residue in DAT, increased the sensitivity of NET to RTI-113, while the reverse mutation in DAT decreased the sensitivity of DAT to RTI-113. In contrast, RTI-31, another cocaine analog having the same structure as RTI-113 but with the phenyl group at the 2β position replaced by a methyl group, inhibits the transporter mutants equally well whether a hydroxyl group is present at the residue or not. The data suggest that this residue contributes to cocaine binding site and is close to the 2β position of cocaine analogs. These results are consistent with our previously proposed cocaine-DAT binding model where cocaine initially binds to a site that does not overlap with, but is close to, the dopamine-binding site. Computational modeling and molecular docking yielded a binding model that explains the observed changes in RTI-113 inhibition potencies.

Population migrations in Southwest and South China have played an important role in the formation of East Asian populations and led to a high degree of cultural diversity among ethnic minorities living in these areas. To explore the genetic relationships of these ethnic minorities, we systematically surveyed the variation of 10 autosomal STR markers of 1,538 individuals from 30 populations of 25 ethnic minorities, of which the majority were chosen from Southwest China, especially Yunnan Province. With genotyped data of the markers, we constructed phylogenies of these populations with both D(A) and D(C) measures and performed a principal component analysis, as well as a clustering analysis by structure. Results showed that we successfully recovered the genetic structure of analyzed populations formed by historical migrations. Aggregation patterns of these populations accord well with their linguistic affiliations, suggesting that deciphering of genetic relationships does in fact offer clues for study of ethnic differentiation.

Transcription factor FOXM1 is involved in stimulating cell proliferation, enhancing DNA damage repair, promoting metastasis of cancer cells, and the inhibition of FOXM1 has been shown to prevent the initiation and progression of multiple cancers and FOXM1 is considered to be an effective target for tumor therapeutic drug development. The N-terminus of FOXM1 has been found to prevent transcriptional activities of FOXM1 and to mediate the interaction between FOXM1 and SMAD3.

Many essential oils (EOs) regulate the quorum-sensing (QS) system of pathogens and inhibit the virulence expression. Interference with QS can potentially reduce bacterial multidrug resistance and aid the biological control of bacterial disease. In the present work, the antibacterial and anti-QS activities of Cinnamomum camphora leaf EO were investigated. A total of 23 chemical components with relative levels ≥0.11%, including a large number of terpene compounds, were identified in C. camphora leaf EO by gas chromatography-mass spectrometry (GC-MS). The principal component was linalool, followed by eucalyptol, with relative levels of 51.57% and 22.07%, respectively. The minimum inhibitory concentration (MIC) and antibacterial activity of C. camphora EO were examined, and P. aeruginosa and E. coli ATCC25922 showed the highest and lowest sensitivity to C. camphora EO, respectively. Tests of QS inhibitory activity revealed that C. camphora EO significantly decreased the production of violacein and biofilm biomass in C. violaceum, with the maximum inhibition rates of 63% and 77.64%, respectively, and inhibited the biofilm formation and swarming movement, independent of affecting the growth of C. violaceum. Addition of C. camphora EO also resulted in downregulation of the expression of the acyl-homoserine lactones (AHL) synthesis gene (cviI) and transcription regulator (cviR), and inhibited the expression of QS-regulated virulence genes, including vioA, vioB, vioC, vioD, vioE, lasA, lasB, pilE3, and hmsHNFR. Collectively, the prominent antibacterial activity and anti-QS activities clearly support that C. camphora EO acts as a potential antibacterial agent and QS inhibitor in the prevention of bacterial contamination.