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1.  Ti3+-self doped brookite TiO2 single-crystalline nanosheets with high solar absorption and excellent photocatalytic CO2 reduction 
Scientific Reports  2016;6:23684.
Black brookite TiO2 single-crystalline nanosheets with outstanding photocatalytic activity toward CO2 reduction is prepared by a facile oxidation-based hydrothermal reaction method combined with post-annealing treatment. Large amount of Ti3+ defects are introduced into the bulk of brookite nanoparticles, which increases the solar energy absorption and enhances the photocatalytic activity.
doi:10.1038/srep23684
PMCID: PMC4810378  PMID: 27021203
2.  Syntaxin Opening by the MUN Domain Underlies the Function of Munc13 in Synaptic Vesicle Priming 
UNC-13-Munc13s play a central function in synaptic vesicle priming through their MUN domains. However, it is unclear whether this function arises from the ability of the MUN domain to mediate the transition from the Munc18-1–closed syntaxin-1 complex to the SNARE complex in vitro. The crystal structure of rat Munc13-1 MUN domain now reveals an elongated, arch-shaped architecture formed by α-helical bundles, with a highly conserved hydrophobic pocket in the middle. Mutation of two residues (NF) in this pocket abolishes the stimulation caused by the Munc13-1 MUN domain on SNARE complex assembly and on SNARE-dependent proteoliposome fusion in vitro. Moreover, the same mutation in UNC-13 abrogates synaptic vesicle priming in C. elegans neuromuscular junctions. These results strongly support the notion that orchestration of syntaxin-1 opening and SNARE complex assembly underlies the central role of UNC-13-Munc13s in synaptic vesicle priming.
doi:10.1038/nsmb.3038
PMCID: PMC4809529  PMID: 26030875
3.  VAMP8 facilitates cellular proliferation and temozolomide resistance in human glioma cells 
Neuro-Oncology  2014;17(3):407-418.
Background
Malignant glioma is a common and lethal primary brain tumor in adults. Here we identified a novel oncoprotein, vesicle-associated membrane protein 8 (VAMP8), and investigated its roles in tumorigenisis and chemoresistance in glioma.
Methods
The expression of gene and protein were determined by quantitative PCR and Western blot, respectively. Histological analysis of 282 glioma samples and 12 normal controls was performed by Pearson's chi-squared test. Survival analysis was performed using the log-rank test and Cox proportional hazards regression. Cell proliferation and cytotoxicity assay were conducted using Cell Counting Kit-8. Autophagy was detected by confocal microscopy and Western blot.
Results
VAMP8 was significantly overexpressed in human glioma specimens and could become a potential novel prognostic and treatment-predictive marker for glioma patients. Overexpression of VAMP8 promoted cell proliferation in vitro and in vivo, whereas knockdown of VAMP8 attenuated glioma growth by arresting cell cycle in the G0/G1 phase. Moreover, VAMP8 contributed to temozolomide (TMZ) resistance by elevating the expression levels of autophagy proteins and the number of autophagosomes. Further inhibition of autophagy via siRNA-mediated knockdown of autophagy-related gene 5 (ATG5) or syntaxin 17 (STX17) reversed TMZ resistance in VAMP8-overexpressing cells, while silencing of VAMP8 impaired the autophagic flux and alleviated TMZ resistance in glioma cells.
Conclusion
Our findings identified VAMP8 as a novel oncogene by promoting cell proliferation and therapeutic resistance in glioma. Targeting VAMP8 may serve as a potential therapeutic regimen for the treatment of glioma.
doi:10.1093/neuonc/nou219
PMCID: PMC4483099  PMID: 25209430
autophagy; glioma; proliferation; TMZ resistance; VAMP8
4.  Overexpression of eIF4E in colorectal cancer patients is associated with liver metastasis 
OncoTargets and therapy  2016;9:815-822.
Purpose
Liver metastasis is one of the leading causes of death in colorectal cancer (CRC) patients. The present study aimed to evaluate the value of eIF4E as a prognostic marker of colorectal liver metastasis (CLM) and identify the functional role of eIF4E in CRC metastasis.
Patients and methods
The expression level of eIF4E in CRC tissues was analyzed by immunohistochemical staining and Western blot. Expression of eIF4E in CRC cell lines was evaluated by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot. Cell Counting Kit-8 (CCK-8) and Transwell assays were performed to assess the effects of eIF4E on cell proliferation, migration, and invasion. Western blot was further used to investigate the mechanism of eIF4E in tumor metastasis.
Results
The upregulation frequency of eIF4E in the CLM group (82.5%) was higher than that in the non-CLM group (65.0%). Of the 80 patients recruited for the follow-up study, 23 were in the low eIF4E group (ratio of tumor to nontumor tissue
Conclusion
The results of the present study indicated that high eIF4E levels in CRC patients predicted a high risk of liver metastasis. Knockdown of eIF4E inhibited CRC cell metastasis in part through regulating the expression of cyclin D1, VEGF, MMP-2, and MMP-9.
doi:10.2147/OTT.S98330
PMCID: PMC4767060  PMID: 26929650
eIF4E; colorectal cancer; liver metastasis; functional study
Biomedical Reports  2016;4(4):427-432.
Vasoreactivity is the most basic and direct indicator to reflect the artery vascular functional state in the body. The majority of previous studies have shown that a high-fat diet (HD) is often associated with a variety of cardiovascular diseases. However, the type of exercise that improves vasoreactivity, as is induced by a HD, remains to be elucidated. In the present study, the effects of aerobic moderate-intensity intermittent exercise through swimming were investigated on thoracic aorta vascular ring contraction and free radical metabolism using Sprague-Dawley rat models of common diet (CD; 23 g protein, 49 g carbohydrate, 4 g fat, 5 g fiber, 7 g bone meal and 6 g vitamins per 100 g), HD (peanuts, milk chocolate and sweet biscuits, in a weight ratio of 3:2:2:1), CD with intermittent exercise (CIE) and HD with intermittent exercise (HIE). The food utilization rate in the swimming group (CIE) decreased in comparison with the CD group. Lee's index in the CIE group decreased in comparison to that of CD after 8 weeks (P<0.05) and also HIE decreased compared to HD (P<0.05) after 8 weeks. Compared with the HD group, contractile response of the thoracic aortic rings to NA decreased in the HIE group, while high-density lipoprotein cholesterol content increased, total cholesterol, triglycerides and low-density lipoprotein cholesterol decreased (P<0.05); the malondialdehyde (MDA) concentration reduced in the myocardium, but the superoxide dismutase (SOD) level improved (P<0.05). In the HIE group, nitric oxide level was similar to the CD group. Compared with CD, contractile response of the thoracic aortic rings to NA increased in the CIE (P<0.05), the MDA concentration reduced in the myocardium, but the SOD level improved (P<0.05). Tunica media smooth muscle of the thoracic aortic rings in the CIE group arranged more regularly in comparison with the CD group (without swimming training). In conclusion, intermittent exercise improves the thoracic aorta vasoreactivity and function by enhanced antioxidant enzyme activity and reduced free radical generating.
doi:10.3892/br.2016.604
PMCID: PMC4812502  PMID: 27073626
intermittent exercise; vasoreactivity; high-fat diet
Cell stem cell  2014;14(2):228-236.
SUMMARY
Pancreatic β cells are of great interest for the treatment of type 1 diabetes. A number of strategies already exist for the generation of β cells, but a general approach for reprogramming nonendodermal cells into β cells could provide an attractive alternative in a variety of contexts. Here, we describe a stepwise method in which pluripotency reprogramming factors were transiently expressed in fibroblasts in conjunction with a unique combination of soluble molecules to generate definitive endoderm-like cells that did not pass through a pluripotent state. These endoderm-like cells were then directed toward pancreatic lineages using further combinations of small molecules in vitro. The resulting pancreatic progenitor-like cells could mature into cells of all three pancreatic lineages in vivo, including functional, insulin-secreting β-like cells that help to ameliorate hyperglycemia. Our findings may therefore provide a useful approach for generating large numbers of functional β cells for disease modeling and, ultimately, cell-based therapy.
doi:10.1016/j.stem.2014.01.006
PMCID: PMC4747235  PMID: 24506886
Scientific Reports  2016;6:19364.
A combination of the advanced chemometrics method with quantum mechanics calculation was for the first time applied to explore a facile yet efficient analysis strategy to thoroughly resolve femtosecond transient absorption spectroscopy of ortho-nitroaniline (ONA), served as a model compound of important nitroaromatics and explosives. The result revealed that the ONA molecule is primarily excited to S3 excited state from the ground state and then ultrafast relaxes to S2 state. The internal conversion from S2 to S1 occurs within 0.9 ps. One intermediate state S* was identified in the intersystem crossing (ISC) process, which is different from the specific upper triplet receiver state proposed in some other nitroaromatics systems. The S1 state decays to the S* one within 6.4 ps and then intersystem crossing to the lowest triplet state within 19.6 ps. T1 was estimated to have a lifetime up to 2 ns. The relatively long S* state and very long-lived T1 one should play a vital role as precursors to various nitroaromatic and explosive photoproducts.
doi:10.1038/srep19364
PMCID: PMC4726040  PMID: 26781083
Journal of proteome research  2014;10.1021/pr500325q.
For more than 30 years the study of learning and memory in Drosophila melanogaster (fruit fly) has used an olfactory learning paradigm and has resulted in the discovery of many genes involved in memory formation. By varying learning programs, the creation of different memory types can be achieved, from short-term memory formation to long-term. Previous studies in the fruit fly used gene mutation methods to identify genes involved in memory formation. Presumably, memory creation involves a combination of genes, pathways and neural circuits. To examine memory formation at the protein level, a quantitative proteomic analysis was performed using olfactory learning and 15N labeled fruit flies. Differences were observed in protein expression and relevant pathways between different learning programs. Our data showed major protein expression changes occurred between short-term memory (STM) and long-lasting memory, and only minor changes were found between long-term memory (LTM) and anesthesia-resistant memory (ARM).
doi:10.1021/pr500325q
PMCID: PMC4412804  PMID: 24983411
Olfactory learning; Memory; Drosophila melanogaster; Quantitative proteomics; Metabolic labeling; Mass spectrometry
Nature Communications  2016;7:10080.
Pancreatic beta cells are of great interest for biomedical research and regenerative medicine. Here we show the conversion of human fibroblasts towards an endodermal cell fate by employing non-integrative episomal reprogramming factors in combination with specific growth factors and chemical compounds. On initial culture, converted definitive endodermal progenitor cells (cDE cells) are specified into posterior foregut-like progenitor cells (cPF cells). The cPF cells and their derivatives, pancreatic endodermal progenitor cells (cPE cells), can be greatly expanded. A screening approach identified chemical compounds that promote the differentiation and maturation of cPE cells into functional pancreatic beta-like cells (cPB cells) in vitro. Transplanted cPB cells exhibit glucose-stimulated insulin secretion in vivo and protect mice from chemically induced diabetes. In summary, our study has important implications for future strategies aimed at generating high numbers of functional beta cells, which may help restoring normoglycemia in patients suffering from diabetes.
Insulin-producing pancreatic beta cells, generated in vitro, could lead to new anti-diabetic therapies. Here, Zhu et al. convert human fibroblasts into endodermal progenitors that differentiate in vitro into glucose-responsive beta-like cells that, following transplantation in mice, protect from diabetes.
doi:10.1038/ncomms10080
PMCID: PMC4729817  PMID: 26733021
Applied and Environmental Microbiology  2015;81(13):4423-4431.
The CRISPR-Cas9 system is a powerful and revolutionary genome-editing tool for eukaryotic genomes, but its use in bacterial genomes is very limited. Here, we investigated the use of the Streptococcus pyogenes CRISPR-Cas9 system in editing the genome of Clostridium cellulolyticum, a model microorganism for bioenergy research. Wild-type Cas9-induced double-strand breaks were lethal to C. cellulolyticum due to the minimal expression of nonhomologous end joining (NHEJ) components in this strain. To circumvent this lethality, Cas9 nickase was applied to develop a single-nick-triggered homologous recombination strategy, which allows precise one-step editing at intended genomic loci by transforming a single vector. This strategy has a high editing efficiency (>95%) even using short homologous arms (0.2 kb), is able to deliver foreign genes into the genome in a single step without a marker, enables precise editing even at two very similar target sites differing by two bases preceding the seed region, and has a very high target site density (median interval distance of 9 bp and 95.7% gene coverage in C. cellulolyticum). Together, these results establish a simple and robust methodology for genome editing in NHEJ-ineffective prokaryotes.
doi:10.1128/AEM.00873-15
PMCID: PMC4475897  PMID: 25911483
PLoS ONE  2015;10(12):e0145031.
Forest health issues are on the rise in the United States, resulting from introduction of alien pests and diseases, coupled with abiotic stresses related to climate change. Increasingly, forest scientists are finding genetic/genomic resources valuable in addressing forest health issues. For a set of ten ecologically and economically important native hardwood tree species representing a broad phylogenetic spectrum, we used low coverage whole genome sequencing from multiplex Illumina paired ends to economically profile their genomic content. For six species, the genome content was further analyzed by flow cytometry in order to determine the nuclear genome size. Sequencing yielded a depth of 0.8X to 7.5X, from which in silico analysis yielded preliminary estimates of gene and repetitive sequence content in the genome for each species. Thousands of genomic SSRs were identified, with a clear predisposition toward dinucleotide repeats and AT-rich repeat motifs. Flanking primers were designed for SSR loci for all ten species, ranging from 891 loci in sugar maple to 18,167 in redbay. In summary, we have demonstrated that useful preliminary genome information including repeat content, gene content and useful SSR markers can be obtained at low cost and time input from a single lane of Illumina multiplex sequence.
doi:10.1371/journal.pone.0145031
PMCID: PMC4689444  PMID: 26698853
Cancer Medicine  2015;4(11):1639-1650.
Frey’s syndrome (FS) is an unavoidable sequela following the surgery of the parotid gland. Although several treatment methods are available, their efficacy is short term or accompanied by unacceptable complications. In the past two decades, botulinum toxin type A (BTXA) has been widely used to treat FS. Although several systematic reviews have been reported recently, they were conflicting and with obvious deficiencies. Thus, we performed an objectively systematic review to determine whether BTXA is an effective and safe treatment for FS. A literature retrieval covering PubMed, Web of Science, Ovid, Embase and Cochrane library was performed on 16 January, 2015. Proportion meta-analysis and corresponding 95% confidence interval (CI) were performed to evaluate the efficacy and safety of BXTA in treatment of FS. A total of 499 records were retrieved and 22 articles with 23 studies were included after scrutiny by two independent authors. Statistical analyses regarding the effective rate, incidence of complications were used to estimate the efficacy and safety of BTXA. Our results suggested that the effective rate of BTXA for treatment of FS is 98.5% (95% CI = 0.971–0.994) and the incidence of complication is 3.6% (95% CI = 0.017–0.061). In conclusion, our study supports that BTXA produces meaningful benefits on the treatment of patients with FS. However, owing to lack of strong evidence, future studies with well-designed inclusion criteria and multicenter randomized controlled trials are needed to give more credible evidence, if possible.
doi:10.1002/cam4.504
PMCID: PMC4673990  PMID: 26310612
Botulinum toxin type A; efficacy; Frey’s syndrome; proportion meta-analysis; safety
Biophysics Reports  2015;1:71-80.
The absolute quantification of target proteins in proteomics involves stable isotope dilution coupled with multiple reactions monitoring mass spectrometry (SID-MRM-MS). The successful preparation of stable isotope-labeled internal standard peptides is an important prerequisite for the SID-MRM absolute quantification methods. Dimethyl labeling has been widely used in relative quantitative proteomics and it is fast, simple, reliable, cost-effective, and applicable to any protein sample, making it an ideal candidate method for the preparation of stable isotope-labeled internal standards. MRM mass spectrometry is of high sensitivity, specificity, and throughput characteristics and can quantify multiple proteins simultaneously, including low-abundance proteins in precious samples such as pancreatic islets. In this study, a new method for the absolute quantification of three proteases involved in insulin maturation, namely PC1/3, PC2 and CPE, was developed by coupling a stable isotope dimethyl labeling strategy for internal standard peptide preparation with SID-MRM-MS quantitative technology. This method offers a new and effective approach for deep understanding of the functional status of pancreatic β cells and pathogenesis in diabetes.
doi:10.1007/s41048-015-0012-1
PMCID: PMC4762191  PMID: 26942221
Dimethyl labeling; Stable isotope dilution-multiple reaction monitoring (SID-MRM); Mass spectrometry; Insulin
Molecular cell  2014;56(2):323-332.
Summary
Tyrosyl-tRNA synthetase (TyrRS) is known for its essential aminoacylation function in protein synthesis. Here we report a new function for TyrRS in DNA damage protection. We found that oxidative stress, which often down-regulates protein synthesis, induces TyrRS to rapidly translocate from the cytosol to the nucleus. We also found that angiogenin mediates or potentiates this stress-induced translocalization. The nuclear-localized TyrRS activates transcription factor E2F1 to up-regulate the expression of DNA damage repair genes such as BRCA1 and RAD51. The activation is achieved through direct interaction of TyrRS with TRIM28 to sequester this vertebrate-specific epigenetic repressors and its associated HDAC1 from deacetylating and suppressing E2F1. Remarkably, overexpression of TyrRS strongly protects against UV-induced DNA double-strand breaks in zebrafish, while restricting TyrRS nuclear entry completely abolishes the protection. Therefore, oxidative stress triggers an essential cytoplasmic enzyme used for protein synthesis to translocate to the nucleus to protect against DNA damage.
doi:10.1016/j.molcel.2014.09.006
PMCID: PMC4224670  PMID: 25284223
Tyrosyl-tRNA synthetase; oxidative stress; nuclear localization; E2F1; transcription; DNA damage; angiogenin; TRIM28; KAP1; HDAC1
Background
Chemokine-like factor (CKLF)-like MARVEL transmembrane domain-containing family (CMTM) is a gene family involved in multiple malignancies. CMTM4 is a member of this family and is located at chromosome 16q22.1, a locus that harbours a number of tumour suppressor genes. It has been defined as a regulator of cell cycle and division in HeLa cells; however, its roles in tumourigenesis remain poorly studied.
Methods
An integrated bioinformatics analysis based on the array data from the GEO database was conducted to view the differential expression of CMTM4 across multiple cancers and their corresponding control tissues. Primary clear cell renal cell carcinoma (ccRCC) and the paired adjacent non-tumour tissues were then collected to examine the expression of CMTM4 by western blotting, immunohistochemistry, and quantitative RT-PCR. The ccRCC cell lines A498 and 786-O and the normal renal tubular epithelial cell line HK-2 were also tested for CMTM4 expression by western blotting. Cell Counting Kit-8 (CCK-8) and viable cell counting assays were used to delineate the growth curves of 786-O cells after CMTM4 overexpression or knockdown. Wound healing and transwell assays were performed to assess the cells’ ability to migrate. The effects of CMTM4 on cellular apoptosis and cell cycle progression were analysed by flow cytometry, and cell cycle hallmarks were detected by western blotting and RT-PCR. The xenograft model in nude mice was used to elucidate the function of CMTM4 in tumourigenesis ex vivo.
Results
By omic data analysis, we found a substantial downregulation of CMTM4 in ccRCC. Western blotting then confirmed that CMTM4 was dramatically reduced in 86.9 % (53/61) of ccRCC tissues compared with the paired adjacent non-tumour tissues, as well as in the 786-O and A498 ccRCC cell lines. Restoration of CMTM4 significantly suppressed 786-O cell growth by inducing G2/M cell cycle arrest and p21 upregulation, and cell migration was also inhibited. However, knockdown of CMTM4 led to a completely opposite effect on these cell behaviours. Overexpression of CMTM4 also markedly inhibited the tumour xenograft growth in nude mice.
Conclusions
CMTM4 is downregulated and exhibits tumour-suppressor activities in ccRCC, and could be exploited as a target for ccRCC treatment.
doi:10.1186/s13046-015-0236-4
PMCID: PMC4609138  PMID: 26474560
CMTM4; Clear cell renal cell carcinoma; Brain cancer; Tumour suppressor gene; G2/M cell cycle arrest; p21
PLoS ONE  2015;10(10):e0140300.
The brain active patterns were organized differently under resting states of eyes open (EO) and eyes closed (EC). The altered voxel-wise and regional-wise resting state active patterns under EO/EC were found by static analysis. More importantly, dynamical spontaneous functional connectivity has been observed in the resting brain. To the best of our knowledge, the dynamical mechanisms of intrinsic connectivity networks (ICNs) under EO/EC remain largely unexplored. The goals of this paper were twofold: 1) investigating the dynamical intra-ICN and inter-ICN temporal patterns during resting state; 2) analyzing the altered dynamical temporal patterns of ICNs under EO/EC. To this end, a cohort of healthy subjects with scan conditions of EO/EC were recruited from 1000 Functional Connectomes Project. Through Hilbert transform, time-varying phase synchronization (PS) was applied to evaluate the inter-ICN synchrony. Meanwhile, time-varying amplitude was analyzed as dynamical intra-ICN temporal patterns. The results found six micro-states of inter-ICN synchrony. The medial visual network (MVN) showed decreased intra-ICN amplitude during EC relative to EO. The sensory-motor network (SMN) and auditory network (AN) exhibited enhanced intra-ICN amplitude during EC relative to EO. Altered inter-ICN PS was found between certain ICNs. Particularly, the SMN and AN exhibited enhanced PS to other ICNs during EC relative to EO. In addition, the intra-ICN amplitude might influence the inter-ICN synchrony. Moreover, default mode network (DMN) might play an important role in information processing during EO/EC. Together, the dynamical temporal patterns within and between ICNs were altered during different scan conditions of EO/EC. Overall, the dynamical intra-ICN and inter-ICN temporal patterns could benefit resting state fMRI-related research, and could be potential biomarkers for human functional connectome.
doi:10.1371/journal.pone.0140300
PMCID: PMC4607488  PMID: 26469182
Scientific Reports  2015;5:13017.
We demonstrate the use of cryogenic super-resolution correlative light and electron microscopy (csCLEM) to precisely determine the spatial relationship between proteins and their native cellular structures. Several fluorescent proteins (FPs) were found to be photoswitchable and emitted far more photons under our cryogenic imaging condition, resulting in higher localization precision which is comparable to ambient super-resolution imaging. Vitrified specimens were prepared by high pressure freezing and cryo-sectioning to maintain a near-native state with better fluorescence preservation. A 2-3-fold improvement of resolution over the recent reports was achieved due to the photon budget performance of screening out Dronpa and optimized imaging conditions, even with thin sections which is at a disadvantage when calculate the structure resolution from label density. We extended csCLEM to mammalian cells by introducing cryo-sectioning and observed good correlation of a mitochondrial protein with the mitochondrial outer membrane at nanometer resolution in three dimensions.
doi:10.1038/srep13017
PMCID: PMC4604464  PMID: 26462878
Traditional Chinese medicine injections (TCMIs) have played an irreplaceable role for treating some clinical emergency, severe illness, and infectious diseases in China. In recent years, the incidence rates of adverse drug reactions (ADRs) of TCMIs have increased year by year. Danhong injection (DHI) is one representative TCMI comprised of Danshen and Honghua for treating cardiovascular and cerebrovascular diseases in clinic. In present study, the postmarketing safety surveillance and reevaluation of DHI were reported. Total 30888 patients in 37 hospitals from 6 provinces participated in the study. The results showed that the ADR incidence rate of DHI was 3.50‰. Seventeen kinds of new adverse reactions of DHI were found. The main type of ADRs of DHI was type A (including sweating, dizziness, headache, flushing, vasodilation, eye hemorrhage, faintness, chest pain, palpitations, breathlessness, anxious, nausea, flatulence, vomiting, hypotension, hypertension, local numbness, dyspnea, joint disease, and tinnitus) accounting for 57.75%. The severities of most ADRs of DHI were mild and moderate reactions accounting for 25.93% and 66.67%, respectively. The main disposition of ADRs of DHI was drug withdrawal and without any treatments. The results can provide basis for amendment and improvement of the instructions of DHI, as well as demonstration and reference for the postmarketing safety surveillance and reevaluation of other TCMIs. And the rationality, scientificity, and safety of clinical applications of TCMIs could be improved.
doi:10.1155/2015/610846
PMCID: PMC4609862  PMID: 26508981
Despite the identification of many genes and pathways involved in the persistence phenomenon of bacteria, the relative importance of these genes in a single organism remains unclear. Here, using Escherichia coli as a model, we generated mutants of 21 known candidate persister genes and compared the relative importance of these mutants in persistence to various antibiotics (ampicillin, gentamicin, norfloxacin, and trimethoprim) at different times. We found that oxyR, dnaK, sucB, relA, rpoS, clpB, mqsR, and recA were prominent persister genes involved in persistence to multiple antibiotics. These genes map to the following pathways: antioxidative defense pathway (oxyR), global regulators (dnaK, clpB, and rpoS), energy production (sucB), stringent response (relA), toxin–antitoxin (TA) module (mqsR), and SOS response (recA). Among the TA modules, the ranking order was mqsR, lon, relE, tisAB, hipA, and dinJ. Intriguingly, rpoS deletion caused a defect in persistence to gentamicin but increased persistence to ampicillin and norfloxacin. Mutants demonstrated dramatic differences in persistence to different antibiotics at different time points: some mutants (oxyR, dnaK, phoU, lon, recA, mqsR, and tisAB) displayed defect in persistence from early time points, while other mutants (relE, smpB, glpD, umuD, and tnaA) showed defect only at later time points. These results indicate that varying hierarchy and importance of persister genes exist and that persister genes can be divided into those involved in shallow persistence and those involved in deep persistence. Our findings suggest that the persistence phenomenon is a dynamic process with different persister genes playing roles of variable significance at different times. These findings have implications for improved understanding of persistence phenomenon and developing new drugs targeting persisters for more effective cure of persistent infections.
doi:10.3389/fmicb.2015.01003
PMCID: PMC4588708  PMID: 26483762
Escherichia coli; persistence; persister gene; knockout mutant; antibiotics; ranking
World Journal of Gastroenterology : WJG  2015;21(36):10375-10384.
AIM: To investigate the effects of Recql5 deficiency on liver injury induced by lipopolysaccharide/D-galactosamine (LPS/D-Gal).
METHODS: Liver injury was induced in wild type (WT) or Recql5-deficient mice using LPS/D-Gal, and assessed by histological, serum transaminases, and mortality analyses. Hepatocellular apoptosis was quantified by transferase dUTP nick end labeling assay and Western blot analysis of cleaved caspase-3. Liver inflammatory chemokine and cytochrome P450 expression was analyzed by quantitative reverse transcription-PCR. Neutrophil infiltration was evaluated by myeloperoxidase activity. Expression and phosphorylation of ERK, JNK, p65, and H2A.X was determined by Western blot. Oxidative stress was evaluated by measuring malondialdehyde production and nitric oxide synthase, superoxide dismutase, glutathione peroxidase, catalase, and glutathione reductase activity.
RESULTS: Following LPS/D-Gal exposure, Recql5-deficient mice exhibited enhanced liver injury, as evidenced by more severe hepatic hemorrhage, higher serum aspartate transaminase and alanine transaminase levels, and lower survival rate. As compared to WT mice, Recql5-deficient mice showed an increased number of apoptotic hepatocytes and higher cleaved caspase-3 levels. Recql5-deficient mice exhibited increased DNA damage, as evidenced by increased γ-H2A.X levels. Inflammatory cytokine levels, neutrophil infiltration, and ERK phosphorylation were also significantly increased in the knockout mice. Additionally, Recql5-deficicent mice exhibited increased malondialdehyde production and elevated inducible nitric oxide synthase, superoxide dismutase, glutathione peroxidase, catalase, and glutathione reductase activity, indicative of enhanced oxidative stress. Moreover, CYP450 expression was significantly downregulated in Recql5-deficient mice after LPS/D-Gal treatment.
CONCLUSION: Recql5 protects the liver against LPS/D-Gal-induced injury through suppression of hepatocyte apoptosis and oxidative stress and modulation of CYP450 expression.
doi:10.3748/wjg.v21.i36.10375
PMCID: PMC4579884  PMID: 26420964
Recql5; Liver injury; Apoptosis; Oxidative stress; CYP450
The first total synthesis of the proposed structure of cycloinumakiol (1) and its C5-epimer (18) are achieved in a concise and efficient fashion: 9 and 5 steps from known 3-hydroxybenzocyclobutenone with overall yields of 15% and 33%, respectively. A key for the success of this approach is use of a catalytic C–C activation strategy for constructing the tetracyclic core of 1 through carboacylation of a sterically hindered trisubstituted olefin with benzocyclobutenone. In addition, the structure of the natural cycloinumakiol was reassigned to 19-hydroxyltotarol (7) through X-ray diffraction analysis. This work demonstrates the potential of C–C activation for streamlining complex natural-product synthesis.
doi:10.1002/anie.201404802
PMCID: PMC4214140  PMID: 25138969
total synthesis; C–C activation; cycloinumakiol; structural revision; Rh catalysis
Objective: The Cycle of Willis unites the internal carotid and vertebrobasilar system and maintains the stability of blood supply to the brain. This present study aims to analyze the anatomic variations in the anterior part of the Cycle of Willis. Methods: Forty five formalin-preserved human brains (90 cerebral hemispheres) with intact dura mater from Chinese adults were dissected under the surgical microscope. The anterior components of the Circle of Willis were observed and measured. Anatomic variations of this segment were observed and photographed. The data collected in this investigation was statistically analyzed. Results: In approximately 13.3% (6/45) of all cerebral samples, a caliber difference of ≥ 0.5 mm was noted between the left A1 and the right segments. The A2 segments of left and right hemispheres were derived from the contralateral A1 segment in 6.7% (3/45) of the brain specimens. Simple anterior communicating artery (ACoA) was observed in 37.8% (17/45) whereas complex ACoA was noted in 60% (27/45) of cerebral samples. Conclusion: As it demonstrates the anatomic variations of ACAC, all surgical approaches should be performed after angiographic demonstrations. Understanding of these anatomic variations plays a pivotal role in neurovascular procedures.
PMCID: PMC4658873  PMID: 26628984
Circle of Willis; anterior circulation; microanatomy; variation
Scientific Reports  2015;5:13931.
Corticosteroids have been proved to be ineffective for Guillain-Barré syndrome, but the mechanism remains unknown. In a rabbit model of axonal Guillain-Barré syndrome, treatment with corticosteroids significantly reduced macrophage infiltration in the spinal ventral roots and the survival rate as well as clinical improvement. On 30th day after onset, there was significantly higher frequency of axonal degeneration in the corticosteroids-treated rabbits than saline-treated rabbits. Corticosteroids may reduce the scavengers that play a crucial role for nerve regeneration, thus delay the recovery of this disease.
doi:10.1038/srep13931
PMCID: PMC4565078  PMID: 26355080
Interventional Neuroradiology  null;20(4):442-447.
Summary
This study evaluated the results of endovascular embolization of multiple intracranial aneurysms. A retrospective hospital chart and radiograph review were made of all patients with multiple intracranial aneurysms seen between March 2010 and January 2011.
Ten patients presented with subarachnoid hemorrhage, four with mass effect, two with brain ischemia and twenty were incidental. These 36 patients harbored 84 aneurysms, 63 of which were treated with endovascular techniques, two by surgical clipping, and 19 were left untreated. Of the coil-treated lesions, a complete endovascular occlusion was achieved in 54 aneurysms (85.7%), and eight (12.7%) presented neck remnants with one (1.6%) stented only. Twenty-six patients (72.2%) underwent coil embolization of more than one aneurysm in the first session. Follow-up angiographic studies in 31 patients demonstrated an unchanged or improved result in 93.0% of the aneurysms (53 lesions) and coil compaction in 7.0% (four lesions). The overall clinical outcome was excellent in 33 patients (91.7%), good in one (2.8%) and fair in two (5.5%).
Endovascular techniques may be a particularly suitable method for treating multiple intracranial aneurysms.
doi:10.15274/INR-2014-10037
PMCID: PMC4187440  PMID: 25207907
multiple; aneurysm; embolization; intracranial; coils
Acta Pharmacologica Sinica  2015;36(9):1137-1144.
Aim:
Ca2+-release-activated Ca2+ (CRAC) channel, a subfamily of store-operated channels, is formed by calcium release-activated calcium modulator 1 (ORAI1), and gated by stromal interaction molecule 1 (STIM1). CRAC channel may be a novel target for the treatment of immune disorders and allergy. The aim of this study was to identify novel small molecule CRAC channel inhibitors.
Methods:
HEK293 cells stably co-expressing both ORAI1 and STIM1 were used for high-throughput screening. A hit, 1-phenyl-3-(1-phenylethyl)urea, was identified that inhibited CRAC channels by targeting ORAI1. Five series of its derivatives were designed and synthesized, and their primary structure-activity relationships (SARs) were analyzed. All derivatives were assessed for their effects on Ca2+ influx through CRAC channels on HEK293 cells, cytotoxicity in Jurkat cells, and IL-2 production in Jurkat cells expressing ORAI1-SS-eGFP.
Results:
A total of 19 hits were discovered in libraries containing 32 000 compounds using the high-throughput screening. 1-Phenyl-3-(1-phenylethyl)urea inhibited Ca2+ influx with IC50 of 3.25±0.17 μmol/L. SAR study on its derivatives showed that the alkyl substituent on the α-position of the left-side benzylic amine (R1) was essential for Ca2+ influx inhibition and that the S-configuration was better than the R-configuration. The derivatives in which the right-side R3 was substituted by an electron-donating group showed more potent inhibitory activity than those that were substituted by electron-withdrawing groups. Furthermore, the free N–H of urea was not necessary to maintain the high potency of Ca2+ influx inhibition. The N,N′-disubstituted or N′-substituted derivatives showed relatively low cytotoxicity but maintained the ability to inhibit IL-2 production. Among them, compound 5b showed an improved inhibition of IL-2 production and low cytotoxicity.
Conclusion:
1-Phenyl-3-(1-phenylethyl)urea is a novel CRAC channel inhibitor that specifically targets ORAI1. This study provides a new chemical scaffold for design and development of CRAC channel inhibitors with improved Ca2+ influx inhibition, immune inhibition and low cytotoxicity.
doi:10.1038/aps.2015.52
PMCID: PMC4561970  PMID: 26256403
CRAC channel; ORAI1; 1-phenyl-3-(1-phenylethyl)urea; YM58483; high-throughput screening; IL-2 production; structure modification; SAR

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