Epidermal growth factor receptor-mediated cell motility: phospholipase C activity is required,but mitogen-activated protein kinase activity is not sufficient for induced cell movement

We recently have demonstrated that EGF receptor (EGFR)-induced cell motility requires receptor kinase activity and autophosphorylation (P. Chen, K. Gupta, and A. Wells. 1994. J. Cell Biol. 124:547-555). This suggests that the immediate downstream effector molecule contains a src homology-2 domain. Phospholipase C gamma (PLC gamma) is among the candidate transducers of this signal because of its potential roles in modulating cytoskeletal dynamics. We utilized signaling-restricted EGFR mutants expressed in receptor devoid NR6 cells to determine if PLC activation is necessary for EGFR-mediated cell movement. Exposure to EGF (25 nM) augmented PLC activity in all five EGFR mutant cell lines which also responded by increased cell movement. Basal phosphoinositide turnover was not affected by EGF in the lines which do not present the enhanced motility response. The correlation between EGFR-mediated cell motility and PLC activity suggested, but did not prove, a causal link. A specific inhibitor of PLC, {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 (1 microM) diminished both the EGF- induced motility and PLC responses, while its inactive analogue {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343 had no effect on these responses. Both the PLC and motility responses were decreased by expression of a dominant-negative PLC gamma-1 fragment in EGF-responsive infectant lines. Lastly, anti-sense oligonucleotides (20 microM) to PLC gamma-1 reduced both responses in NR6 cells expressing wild-type EGFR. These findings strongly support PLC gamma as the immediate post receptor effector in this motogenic pathway. We have demonstrated previously that EGFR-mediated cell motility and mitogenic signaling pathways are separable. The point of divergence is undefined. All kinase-active EGFR mutants induced the mitogenic response while only those which are autophosphorylated induced PLC activity. {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 did not affect EGF-induced thymidine incorporation in these motility-responsive infectant cell lines. In addition, the dominant-negative PLC gamma-1 fragment did not diminish EGF-induced thymidine incorporation. All kinase active EGFR stimulated mitogen-activated protein (MAP) kinase activity, regardless of whether the receptors induced cell movement; this EGF-induced MAP kinase activity was not affected by {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 at concentrations that depressed the motility response. Thus, the signaling pathways which lead to motility and cell proliferation diverge at the immediate post-receptor stage, and we suggest that this is accomplished by differential activation of effector molecules.     

Diacylglycerol regulates acute hypoxic pulmonary vasoconstriction via TRPC6

Background

Hypoxic pulmonary vasoconstriction (HPV) is an essential mechanism of the lung that matches blood perfusion to alveolar ventilation to optimize gas exchange. Recently we have demonstrated that acute but not sustained HPV is critically dependent on the classical transient receptor potential 6 (TRPC6) channel. However, the mechanism of TRPC6 activation during acute HPV remains elusive. We hypothesize that a diacylglycerol (DAG)-dependent activation of TRPC6 regulates acute HPV.

Methods

We investigated the effect of the DAG analog 1-oleoyl-2-acetyl-sn-glycerol (OAG) on normoxic vascular tone in isolated perfused and ventilated mouse lungs from TRPC6-deficient and wild-type mice. Moreover, the effects of OAG, the DAG kinase inhibitor {"type":"entrez-nucleotide","attrs":{"text":"R59949","term_id":"830644","term_text":"R59949"}}R59949 and the phospholipase C inhibitor {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075"}}U73122 on the strength of HPV were investigated compared to those on non-hypoxia-induced vasoconstriction elicited by the thromboxane mimeticum U46619.

Results

OAG increased normoxic vascular tone in lungs from wild-type mice, but not in lungs from TRPC6-deficient mice. Under conditions of repetitive hypoxic ventilation, OAG as well as {"type":"entrez-nucleotide","attrs":{"text":"R59949","term_id":"830644","term_text":"R59949"}}R59949 dose-dependently attenuated the strength of acute HPV whereas U46619-induced vasoconstrictions were not reduced. Like OAG, {"type":"entrez-nucleotide","attrs":{"text":"R59949","term_id":"830644","term_text":"R59949"}}R59949 mimicked HPV, since it induced a dose-dependent vasoconstriction during normoxic ventilation. In contrast, {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075"}}U73122, a blocker of DAG synthesis, inhibited acute HPV whereas {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125"}}U73343, the inactive form of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075"}}U73122, had no effect on HPV.

Conclusion

These findings support the conclusion that the TRPC6-dependency of acute HPV is induced via DAG.     

Upregulation of the novel lncRNA U731166 is associated with migration,invasion and vemurafenib resistance in melanoma

Our previous work using a melanoma progression model composed of melanocytic cells (melanocytes, primary and metastatic melanoma samples) demonstrated various deregulated genes, including a few known lncRNAs. Further analysis was conducted to discover novel lncRNAs associated with melanoma, and candidates were prioritized for their potential association with invasiveness or other metastasis‐related processes. In this sense, we found the intergenic lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 (ENSG00000230454) and decided to explore its effects in melanoma. For that, we silenced the lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 expression using shRNAs in a melanoma cell line. Next, we experimentally investigated its functions and found that migration and invasion had significantly decreased in knockdown cells, indicating an essential association of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 for cancer processes. Additionally, using naïve and vemurafenib‐resistant cell lines and data from a patient before and after resistance, we found that vemurafenib‐resistant samples had a higher expression of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166. Also, we retrieved data from the literature that indicates lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 may act as a mediator of RNA processing and cell invasion, probably inducing a more aggressive phenotype. Therefore, our results suggest a relevant role of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 in metastasis development. We also pointed herein the lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 as a new possible biomarker or target to help overcome clinical vemurafenib resistance.     

Small molecule sensitization to TRAIL is mediated via nuclear localization,phosphorylation and inhibition of chaperone activity of Hsp27

The small chaperone protein Hsp27 confers resistance to apoptosis, and therefore is an attractive anticancer drug target. We report here a novel mechanism underlying the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) sensitizing activity of the small molecule {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511, an inactive analog of the phosphoinositide 3-kinase inhibitor inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002, in HeLa cells that are refractory to TRAIL-induced apoptosis. On the basis of the fact that {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511 is derived from {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002, itself derived from quercetin, and earlier findings indicating that quercetin and {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 affected Hsp27 expression, we investigated whether {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511 sensitized cancer cells to TRAIL via a conserved inhibitory effect on Hsp27. We provide evidence that upon treatment with {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511, Hsp27 is progressively sequestered in the nucleus, thus reducing its protective effect in the cytosol during the apoptotic process. {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511-induced nuclear translocation of Hsp27 is linked to its sustained phosphorylation via activation of p38 kinase and MAPKAP kinase 2 and the inhibition of PP2A. Furthermore, Hsp27 phosphorylation leads to the subsequent dissociation of its large oligomers and a decrease in its chaperone activity, thereby further compromising the death inhibitory activity of Hsp27. Furthermore, genetic manipulation of Hsp27 expression significantly affected the TRAIL sensitizing activity of {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511, which corroborated the Hsp27 targeting activity of {"type":"entrez-nucleotide","attrs":{"text":"LY303511","term_id":"1257646067","term_text":"LY303511"}}LY303511. Taken together, these data indicate a novel mechanism of small molecule sensitization to TRAIL through targeting of Hsp27 functions, rather than its overall expression, leading to decreased cellular protection, which could have therapeutic implications for overcoming chemotherapy resistance in tumor cells.     

Calcium control of ciliary reversal in ionophore-treated and ATP- reactivated comb plates of ctenophores

Previous work showed that ctenophore larvae swim backwards in high-KCl seawater, due to a 180 degrees reversal in the direction of effective stroke of their ciliary comb plates (Tamm, S. L., and S. Tamm, 1981, J. Cell Biol., 89: 495-509). Ion substitution and blocking experiments indicated that this response is Ca2+ dependent, but comb plate cells are innervated and presumably under nervous control. To determine whether Ca2+ is directly involved in activating the ciliary reversal mechanism and/or is required for synaptic triggering of the response, we (a) determined the effects of ionophore {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187 and Ca2+ on the beat direction of isolated nerve-free comb plates dissociated from larvae by hypotonic, divalent cation-free medium, and (b) used permeabilized ATP- reactivated models of comb plates to test motile responses to known concentrations of free Ca2+. We found that 5 microM {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187 and 10 mM Ca2+ induced dissociated comb plate cells to beat in the reverse direction and to swim counterclockwise in circular paths instead of in the normal clockwise direction. Detergent/glycerol-extracted comb plates beat actively in the presence of ATP, and reactivation was reversibly inhibited by micromolar concentrations of vanadate. Free Ca2+ concentrations greater than 10(-6)M caused reversal in direction of the effective stroke but no significant increase in beat frequency. These results show that ciliary reversal in ctenophores, like that in protozoa, is activated by an increase in intracellular free Ca2+ ions. This allows the unique experimental advantages of ctenophore comb plate cilia to be used for future studies on the site and mechanism of action of Ca2+ in the regulation of ciliary motion.     

Extensive homologous recombination in classical swine fever virus: A re-evaluation of homologous recombination events in the strain AF407339

In this short report, the genome-wide homologous recombination events were re-evaluated for classical swine fever virus (CSFV) strain {"type":"entrez-nucleotide","attrs":{"text":"AF407339","term_id":"15488012","term_text":"AF407339"}}AF407339. We challenged a previous study which suggested only one recombination event in {"type":"entrez-nucleotide","attrs":{"text":"AF407339","term_id":"15488012","term_text":"AF407339"}}AF407339 based on 25 CSFV genomes. Through our re-analysis on the 25 genomes in the previous study and the 41 genomes used in the present study, we argued that there should be possibly at least two clear recombination events happening in {"type":"entrez-nucleotide","attrs":{"text":"AF407339","term_id":"15488012","term_text":"AF407339"}}AF407339 through genome-wide scanning. The reasons for identifying only one recombination event in the previous study might be due to the limited number of available CSFV genome sequences at that time and the limited usage of detection methods. In contrast, as identified by most detection methods using all available CSFV genome sequences, two major recombination events were found at the starting and ending zones of the genome {"type":"entrez-nucleotide","attrs":{"text":"AF407339","term_id":"15488012","term_text":"AF407339"}}AF407339, respectively. The first one has two parents {"type":"entrez-nucleotide","attrs":{"text":"AF333000","term_id":"13383931","term_text":"AF333000"}}AF333000 (minor) and {"type":"entrez-nucleotide","attrs":{"text":"AY554397","term_id":"49860681","term_text":"AY554397"}}AY554397 (major) with beginning and ending breakpoints located at 19 and 607 nt of the genome respectively. The second one has two parents {"type":"entrez-nucleotide","attrs":{"text":"AF531433","term_id":"22347661","term_text":"AF531433"}}AF531433 (minor) and {"type":"entrez-nucleotide","attrs":{"text":"GQ902941","term_id":"298113017","term_text":"GQ902941"}}GQ902941 (major) with beginning and ending breakpoints at 8397 and 11,078 nt of the genome respectively. Phylogenetic incongruence analysis using neighbor-joining algorithm with 1000 bootstrapping replicates further supported the existence of these two recombination events. In addition, we also identified additional 18 recombination events on the available CSFV strains. Some of them may be trivial and can be ignored. In conclusion, CSFV might have relatively high frequency of homologous recombination events. Genome-wide scanning of identifying recombination events should utilize multiple detection methods so as to reduce the risk of misidentification.     

Genome sequences published outside of Standards in Genomic Sciences, March-April 2012

The purpose of this table is to provide the community with a citable record of publications of ongoing genome sequencing projects that have led to a publication in the scientific literature. While our goal is to make the list complete, there is no guarantee that we may have omitted one or more publications appearing in this time frame. Readers and authors who wish to have publications added to subsequent versions of this list are invited to provide the bibliographic data for such references to the SIGS editorial office.

Phylum Euryarchaeota

• Halococcus hamelinensis, sequence accession PRJNA80845 [1]
• “Methanocella conradii” HZ254, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003243","term_id":"379319823","term_text":"CP003243"}}CP003243 [2]
• Thermococcus litoralis NS-C, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHVB00000000","term_id":"374743520","term_text":"AHVB00000000"}}AHVB00000000 [3]

Phylum Crenarchaeota

• Candidatus Nitrosopumilus salaria” BD31, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AEXL00000000","term_id":"386807666","term_text":"AEXL00000000"}}AEXL00000000 [4]
• Candidatus Nitrosoarchaeum limnia, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHJG00000000","term_id":"367466343","term_text":"AHJG00000000"}}AHJG00000000 [5]

Phylum Deinococcus-Thermus

• Deinococcus gobiensis, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP002536","term_id":"324314064","term_text":"CP002536"}}CP002536 [6]

Phylum Proteobacteria

• Aggregatibacter actinomycetemcomitans strain ANH9381, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003099","term_id":"365745154","term_text":"CP003099"}}CP003099 [7]
• Alishewanella jeotgali, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHTH00000000","term_id":"374571619","term_text":"AHTH00000000"}}AHTH00000000 [8]
• Enterobacter aerogenes KCTC 2190, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP002824","term_id":"334732565","term_text":"CP002824"}}CP002824 [9]
• Escherichia coli O104:H4, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AFOB02000092","term_id":"340739517","term_text":"AFOB02000092"}}AFOB02000092 [10]
• Helicobacter pylori strains 17874, sequence accession PRJNA76569 [11]
• Helicobacter pylori strains P79, sequence accession PRJNA76567 [11]
• Janthinobacterium sp. Strain PAMC 25724, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHHB00000000","term_id":"372292151","term_text":"AHHB00000000"}}AHHB00000000 [12]
• Klebsiella oxytoca KCTC 1686, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003218","term_id":"365906294","term_text":"CP003218"}}CP003218 [13]
• Klebsiella pneumoniae subsp. pneumoniae HS11286, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003200","term_id":"364515570","term_text":"CP003200"}}CP003200 (chromosome), {"type":"entrez-nucleotide","attrs":{"text":"CP003223","term_id":"365803686","term_text":"CP003223"}}CP003223 (plasmid pKPHS1), {"type":"entrez-nucleotide","attrs":{"text":"CP003224","term_id":"365803828","term_text":"CP003224"}}CP003224 (plasmid pKPHS2), {"type":"entrez-nucleotide","attrs":{"text":"CP003225","term_id":"365803989","term_text":"CP003225"}}CP003225 (plasmid pKPHS3), {"type":"entrez-nucleotide","attrs":{"text":"CP003226","term_id":"365804142","term_text":"CP003226"}}CP003226 (plasmid pKPHS4), {"type":"entrez-nucleotide","attrs":{"text":"CP003227","term_id":"365804147","term_text":"CP003227"}}CP003227 (plasmid pKPHS5), {"type":"entrez-nucleotide","attrs":{"text":"CP003228","term_id":"365804153","term_text":"CP003228"}}CP003228 (plasmid pKPHS6) [14]
• Oceanimonas sp. GK1, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003171","term_id":"372983507","term_text":"CP003171"}}CP003171 [15]
• “Pseudogulbenkiania ferrooxidans” Strain 2002, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"NZ_ACIS01000000","term_id":"224827334","term_text":"ref||NZ_ACIS01000000"}}NZ_ACIS01000000 [16]
• Pseudomonas extremaustralis 14-3b, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHIP00000000","term_id":"371721769","term_text":"AHIP00000000"}}AHIP00000000 [17]
• Pseudomonas sp. Strain PAMC 25886, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHHC00000000","term_id":"372002201","term_text":"AHHC00000000"}}AHHC00000000 [18]
• Psychrobacter, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHVZ00000000","term_id":"375126956","term_text":"AHVZ00000000"}}AHVZ00000000 [19]
• Rahnella sp. Strain Y9602, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP002505","term_id":"321165934","term_text":"CP002505"}}CP002505 [20]
• Rhizobium sp. Strain PDO1-076, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHZC00000000","term_id":"375058045","term_text":"AHZC00000000"}}AHZC00000000 [21]
• Rhodospirillum photometricum DSM122, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"HE663493","term_id":"378401447","term_text":"HE663493"}}HE663493 [22]
• “Rickettsia sibirica sibirica”, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHIZ00000000","term_id":"374669031","term_text":"AHIZ00000000"}}AHIZ00000000 [23]
• Rickettsia sibirica subsp. mongolitimonae strain HA-91, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHZB00000000","term_id":"375318263","term_text":"AHZB00000000"}}AHZB00000000 [24]
• Salmonella enterica subsp. enterica Serotype Enteritidis Strain LA5, sequence accession [25]
• Salmonella enterica subsp. enterica Serotype Senftenberg Strain SS209, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CAGQ00000000","term_id":"379987603","term_text":"CAGQ00000000"}}CAGQ00000000 [26]
• Salmonella enterica subsp. enterica Serovar Typhi P-stx-12, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003278","term_id":"374352002","term_text":"CP003278"}}CP003278 (chromosome) and {"type":"entrez-nucleotide","attrs":{"text":"CP003279","term_id":"374356693","term_text":"CP003279"}}CP003279 (plasmid) [27]
• Sphingomonas echinoides ATCC 14820, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHIR00000000","term_id":"366040205","term_text":"AHIR00000000"}}AHIR00000000 [28]
• Strain HIMB55, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AGIF00000000","term_id":"374304583","term_text":"AGIF00000000"}}AGIF00000000 [29]
• Vibrio harveyi CAIM 1792, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHHQ00000000","term_id":"374671508","term_text":"AHHQ00000000"}}AHHQ00000000 [30]
• Wolbachia Strain wAlbB, sequence accession {"type":"entrez-nucleotide-range","attrs":{"text":"CAGB01000001 to CAGB01000165","start_term":"CAGB01000001","end_term":"CAGB01000165","start_term_id":"371933004","end_term_id":"371931774"}}CAGB01000001 to CAGB01000165 [31]
• Xanthomonas axonopodis pv. punicae Strain LMG 859, sequence accession {"type":"entrez-nucleotide-range","attrs":{"text":"CAGJ01000001 to CAGJ01000217","start_term":"CAGJ01000001","end_term":"CAGJ01000217","start_term_id":"372556075","end_term_id":"372551622"}}CAGJ01000001 to CAGJ01000217 [32]

Phylum Tenericutes

• Mycoplasma hyorhinis Strain GDL-1, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003231","term_id":"367460291","term_text":"CP003231"}}CP003231 [33]

Phylum Firmicutes

• Bacillus subtilis, sequence accession BGSCID 3A27 through BGSCID 28A4 [34]
• Clostridium difficile Strain CD37, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHJJ00000000","term_id":"371930118","term_text":"AHJJ00000000"}}AHJJ00000000 [35]
• Clostridium perfringens, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AFES00000000","term_id":"380306484","term_text":"AFES00000000"}}AFES00000000 [36]
• Lactobacillus fructivorans KCTC 3543, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AEQY00000000","term_id":"316986063","term_text":"AEQY00000000"}}AEQY00000000 [37]
• Lactococcus lactis IO-1, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AP012281","term_id":"374672113","term_text":"AP012281"}}AP012281 [38]
• Lactobacillus plantarum strain NC8, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AGRI00000000","term_id":"376011245","term_text":"AGRI00000000"}}AGRI00000000 [39]
• Paenibacillus dendritiformis C454, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHKH00000000","term_id":"374393106","term_text":"AHKH00000000"}}AHKH00000000 [40]
• Paenibacillus sp. Strain Aloe-11, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AGFI00000000","term_id":"374477730","term_text":"AGFI00000000"}}AGFI00000000 [41]
• “Peptoniphilus rhinitidis” 1-13^T, sequence accession {"type":"entrez-nucleotide-range","attrs":{"text":"BAEW01000001 to BAEW01000056","start_term":"BAEW01000001","end_term":"BAEW01000056","start_term_id":"374722129","end_term_id":"374722074"}}BAEW01000001 to BAEW01000056 [42]
• Streptococcus macedonicus ACA-DC 198, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"HE613569","term_id":"372283141","term_text":"HE613569"}}HE613569 and {"type":"entrez-nucleotide","attrs":{"text":"HE613570","term_id":"372285142","term_text":"HE613570"}}HE613570 [43]
• Staphylococcus aureus VC40, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003033","term_id":"374362062","term_text":"CP003033"}}CP003033 [44]
• Streptococcus infantarius subsp. infantarius Strain CJ18, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003295","term_id":"374681091","term_text":"CP003295"}}CP003295 (chromosome), {"type":"entrez-nucleotide","attrs":{"text":"CP003296","term_id":"374682963","term_text":"CP003296"}}CP003296 (plasmid) [45]
• Streptococcus macedonicus ACA-DC 198, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"HE613569","term_id":"372283141","term_text":"HE613569"}}HE613569 (chromosome), {"type":"entrez-nucleotide","attrs":{"text":"HE613570","term_id":"372285142","term_text":"HE613570"}}HE613570 (plasmid pSMA198) [46]

Phylum Actinobacteria

• Actinoplanes sp. SE50/110, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"CP003170","term_id":"359832573","term_text":"CP003170"}}CP003170 [47]
• Amycolatopsis sp. Strain ATCC 39116, sequence accession [48]
• Nocardia cyriacigeorgica GUH-2, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"FO082843","term_id":"374843763","term_text":"FO082843"}}FO082843 [49]
• Salinibacterium sp., sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHWA00000000","term_id":"375323651","term_text":"AHWA00000000"}}AHWA00000000 [50]
• Streptomyces acidiscabies 84-104, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AHBF00000000","term_id":"372137622","term_text":"AHBF00000000"}}AHBF00000000 [51]

Non-Bacterial genomes

• Bluetongue Virus Serotype 2, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AJ783905","term_id":"70907482","term_text":"AJ783905"}}AJ783905 (Seg-6) and {"type":"entrez-nucleotide","attrs":{"text":"JQ681257","term_id":"383932043","term_text":"JQ681257"}}JQ681257 (Seg-1), {"type":"entrez-nucleotide","attrs":{"text":"JQ681257","term_id":"383932043","term_text":"JQ681257"}}JQ681257 (Seg-1), {"type":"entrez-nucleotide","attrs":{"text":"JQ681258","term_id":"383932045","term_text":"JQ681258"}}JQ681258 (Seg-2), {"type":"entrez-nucleotide","attrs":{"text":"JQ681259","term_id":"383932047","term_text":"JQ681259"}}JQ681259 (Seg-3), {"type":"entrez-nucleotide","attrs":{"text":"JQ681260","term_id":"383932049","term_text":"JQ681260"}}JQ681260 (Seg-4), {"type":"entrez-nucleotide","attrs":{"text":"JQ681261","term_id":"383932051","term_text":"JQ681261"}}JQ681261 (Seg-5), {"type":"entrez-nucleotide","attrs":{"text":"JQ681262","term_id":"383932053","term_text":"JQ681262"}}JQ681262 (Seg-7), JQ6812563 (Seg-8), JQ6812564 (Seg-9), to {"type":"entrez-nucleotide","attrs":{"text":"JQ681265","term_id":"383932059","term_text":"JQ681265"}}JQ681265 (Seg-10) [52]
• Virus Serotype 1, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"AJ585111","term_id":"118420282","term_text":"AJ585111"}}AJ585111 (Seg-2), {"type":"entrez-nucleotide","attrs":{"text":"AJ586659","term_id":"47826214","term_text":"AJ586659"}}AJ586659 (Seg-6), {"type":"entrez-nucleotide","attrs":{"text":"JQ282770","term_id":"383215071","term_text":"JQ282770"}}JQ282770 (Seg-1), {"type":"entrez-nucleotide","attrs":{"text":"JQ282771","term_id":"383215073","term_text":"JQ282771"}}JQ282771 (Seg-3), {"type":"entrez-nucleotide","attrs":{"text":"JQ282772","term_id":"383215075","term_text":"JQ282772"}}JQ282772 (Seg-4), {"type":"entrez-nucleotide","attrs":{"text":"JQ282773","term_id":"383215077","term_text":"JQ282773"}}JQ282773 (Seg-5), {"type":"entrez-nucleotide","attrs":{"text":"JQ282774","term_id":"383215079","term_text":"JQ282774"}}JQ282774 (Seg-7), {"type":"entrez-nucleotide","attrs":{"text":"JQ282775","term_id":"383215081","term_text":"JQ282775"}}JQ282775 (Seg-8), {"type":"entrez-nucleotide","attrs":{"text":"JQ282776","term_id":"383215083","term_text":"JQ282776"}}JQ282776 (Seg-9), and {"type":"entrez-nucleotide","attrs":{"text":"JQ282777","term_id":"383215085","term_text":"JQ282777"}}JQ282777 (Seg-10) [52]
• Chloroplast genome of Erycina pusilla, sequence accession JF_746994 [53]
• Danio rerio, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ434101","term_id":"384872525","term_text":"JQ434101"}}JQ434101 [54]
• Enterococcal Bacteriophage SAP6, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JF731128","term_id":"343411857","term_text":"JF731128"}}JF731128 [55]
• Eubenangee virus, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ070376","term_id":"383513837","term_text":"JQ070376"}}JQ070376 through {"type":"entrez-nucleotide","attrs":{"text":"JQ070385","term_id":"383513855","term_text":"JQ070385"}}JQ070385 [56]
• Fujian/411-like viruses, sequence accession {"type":"entrez-nucleotide-range","attrs":{"text":"CY087969 to CY088568","start_term":"CY087969","end_term":"CY088568","start_term_id":"380506411","end_term_id":"380507803"}}CY087969 to CY088568 [57]
• Hantavirus Variant of Rio Mamoré Virus, Maripa Virus, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ611712","term_id":"384382977","term_text":"JQ611712"}}JQ611712 (segment S), {"type":"entrez-nucleotide","attrs":{"text":"JQ611713","term_id":"384382979","term_text":"JQ611713"}}JQ611713 (segment M), and {"type":"entrez-nucleotide","attrs":{"text":"JQ611714","term_id":"384382981","term_text":"JQ611714"}}JQ611714 (segment L) [58]
• Pata virus, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ070386","term_id":"383513857","term_text":"JQ070386"}}JQ070386 through {"type":"entrez-nucleotide","attrs":{"text":"JQ070395","term_id":"383513875","term_text":"JQ070395"}}JQ070395 [59]
• Porcine Circovirus 2, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ413808","term_id":"383215164","term_text":"JQ413808"}}JQ413808 [60]
• Porcine Reproductive and Respiratory Syndrome Virus, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ326271","term_id":"382929416","term_text":"JQ326271"}}JQ326271 [61]
• Streptococcus mutans Phage M102AD, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"DQ386162","term_id":"88698119","term_text":"DQ386162"}}DQ386162 [62]
• Tilligery virus, sequence accession {"type":"entrez-nucleotide","attrs":{"text":"JQ070366","term_id":"383513817","term_text":"JQ070366"}}JQ070366 through {"type":"entrez-nucleotide","attrs":{"text":"JQ070375","term_id":"383513835","term_text":"JQ070375"}}JQ070375 [63]

     

Elevation of Extracellular Ca2+ Induces Store-Operated Calcium Entry via Calcium-Sensing Receptors: A Pathway Contributes to the Proliferation of Osteoblasts

Aims

The local concentration of extracellular Ca²⁺ ([Ca²⁺]_o) in bone microenvironment is accumulated during bone remodeling. In the present study we investigated whether elevating [Ca²⁺]_o induced store-operated calcium entry (SOCE) in primary rat calvarial osteoblasts and further examined the contribution of elevating [Ca²⁺]_o to osteoblastic proliferation.

Methods

Cytosolic Ca²⁺ concentration ([Ca²⁺]_c) of primary cultured rat osteoblasts was detected by fluorescence imaging using calcium-sensitive probe fura-2/AM. Osteoblastic proliferation was estimated by cell counting, MTS assay and ATP assay. Agonists and antagonists of calcium-sensing receptors (CaSR) as well as inhibitors of phospholipase C (PLC), SOCE and voltage-gated calcium (Cav) channels were applied to study the mechanism in detail.

Results

Our data showed that elevating [Ca²⁺]_o evoked a sustained increase of [Ca²⁺]_c in a dose-dependent manner. This [Ca²⁺]_c increase was blocked by TMB-8 (Ca²⁺ release inhibitor), 2-APB and BTP-2 (both SOCE blockers), respectively, whereas not affected by Cav channels blockers nifedipine and verapamil. Furthermore, NPS2143 (a CaSR antagonist) or {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 (a PLC inhibitor) strongly reduced the [Ca²⁺]_o-induced [Ca²⁺]_c increase. The similar responses were observed when cells were stimulated with CaSR agonist spermine. These data indicated that elevating [Ca²⁺]_o resulted in SOCE depending on the activation of CaSR and PLC in osteoblasts. In addition, high [Ca²⁺]_o significantly promoted osteoblastic proliferation, which was notably reversed by BAPTA-AM (an intracellular calcium chelator), 2-APB, BTP-2, TMB-8, NPS2143 and {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122, respectively, but not affected by Cav channels antagonists.

Conclusions

Elevating [Ca²⁺]_o induced SOCE by triggering the activation of CaSR and PLC. This process was involved in osteoblastic proliferation induced by high level of extracellular Ca²⁺ concentration.     

Construction of immune-related LncRNAs classifier to predict prognosis and immunotherapy response in thymic epithelial tumors

The primary objective of this study was to construct an immune-related long noncoding RNAs (IRLs) classifier to precisely predict the prognosis and immunotherapy response of patients with thymic epithelial tumors (TET). Based on univariable Cox regression analysis and Lasso regression, six prognosis-related IRLs (AC004466.3, {"type":"entrez-nucleotide","attrs":{"text":"AC138207.2","term_id":"29165571","term_text":"AC138207.2"}}AC138207.2, {"type":"entrez-nucleotide","attrs":{"text":"AC148477.2","term_id":"45504179","term_text":"AC148477.2"}}AC148477.2, {"type":"entrez-nucleotide","attrs":{"text":"AL450270.1","term_id":"11244554","term_text":"AL450270.1"}}AL450270.1, HOXB-AS1 and SNHG8) were selected to build an IRL classifier. Importantly, results of qRT-PCR validated that higher expression levels of {"type":"entrez-nucleotide","attrs":{"text":"AC138207.2","term_id":"29165571","term_text":"AC138207.2"}}AC138207.2, {"type":"entrez-nucleotide","attrs":{"text":"AC148477.2","term_id":"45504179","term_text":"AC148477.2"}}AC148477.2, {"type":"entrez-nucleotide","attrs":{"text":"AL450270.1","term_id":"11244554","term_text":"AL450270.1"}}AL450270.1 and SNHG8 as well as lower expression levels of AC004466.3, and HOXB-AS1 in TETs samples compared with normal controls. The IRL classifier could effectively classify patients into the low-risk and high-risk groups based on the different survival parameters. In terms of predictive ability and clinical utility, the IRL classifier was superior to Masaoka staging system. Additionally, IRL classifier is significantly associated with immune cells infiltration (dendritic cells, activated CD4 memory T cells and tumor-infiltrating lymphocyte (TIL), T cell subsets in particular), immune microenvironment (immune score and immune checkpoint inhibitors) and immunogenicity (TMB) in TETs, which hints that IRL classifier is tightly correlated with immune characteristics and might guide more effective immunotherapy strategies for TETs patients. Encouragingly, according to TIDE algorithm, there were more immunotherapy responders in the low-risk IRL subgroup and the IRL score was robustly negatively linked to the immunotherapeutic response. To sum up, the IRL classifier was established, which can be used to predict the prognosis, immune infiltration status, immunotherapy response in TETs patients, and may facilitate personalized counseling for immunotherapy.     

Genomic Basis of a Polyagglutinating Isolate of Neisseria meningitidis

Containment strategies for outbreaks of invasive Neisseria meningitidis disease are informed by serogroup assays that characterize the polysaccharide capsule. We sought to uncover the genomic basis of conflicting serogroup assay results for an isolate ({"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917) from a patient with acute meningococcal disease. To this end, we characterized the complete genome sequence of the {"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917 isolate and performed a variety of comparative sequence analyses against N. meningitidis reference genome sequences of known serogroups. Multilocus sequence typing and whole-genome sequence comparison revealed that {"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917 is a member of the ST-11 sequence group, which is most often associated with serogroup C. However, sequence similarity comparisons and phylogenetic analysis showed that the serogroup diagnostic capsule polymerase gene (synD) of {"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917 belongs to serogroup B. These results suggest that a capsule-switching event occurred based on homologous recombination at or around the capsule locus of {"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917. Detailed analysis of this locus uncovered the locations of recombination breakpoints in the {"type":"entrez-nucleotide","attrs":{"text":"M16917","term_id":"167250","term_text":"M16917"}}M16917 genome sequence, which led to the introduction of an ∼2-kb serogroup B sequence cassette into the serogroup C genomic background. Since there is no currently available vaccine for serogroup B strains of N. meningitidis, this kind capsule-switching event could have public health relevance as a vaccine escape mutant.     

Angiopoietin-Like 4 Mediates PPAR Delta Effect on Lipoprotein Lipase-Dependent Fatty Acid Uptake but Not on Beta-Oxidation in Myotubes

Peroxisome proliferator-activated receptor (PPAR) delta is an important regulator of fatty acid (FA) metabolism. Angiopoietin-like 4 (Angptl4), a multifunctional protein, is one of the major targets of PPAR delta in skeletal muscle cells. Here we investigated the regulation of Angptl4 and its role in mediating PPAR delta functions using human, rat and mouse myotubes. Expression of Angptl4 was upregulated during myotubes differentiation and by oleic acid, insulin and PPAR delta agonist {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516. Treatment with {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 or Angptl4 overexpression inhibited both lipoprotein lipase (LPL) activity and LPL-dependent uptake of FAs whereas uptake of BSA-bound FAs was not affected by either treatment. Activation of retinoic X receptor (RXR), PPAR delta functional partner, using bexarotene upregulated Angptl4 expression and inhibited LPL activity in a PPAR delta dependent fashion. Silencing of Angptl4 blocked the effect of {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 and bexarotene on LPL activity. Treatment with {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 but not Angptl4 overexpression significantly increased palmitate oxidation. Furthermore, Angptl4 overexpression did not affect the capacity of {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 to increase palmitate oxidation. Basal and insulin stimulated glucose uptake, glycogen synthesis and glucose oxidation were not significantly modulated by Angptl4 overexpression. Our findings suggest that FAs-PPARdelta/RXR-Angptl4 axis controls the LPL-dependent uptake of FAs in myotubes, whereas the effect of PPAR delta activation on beta-oxidation is independent of Angptl4.     

Intracellular ca2+ stores could participate to abscisic acid-induced depolarization and stomatal closure in Arabidopsis thaliana

In Arabidopsis thaliana cell suspension,abscisic acid (aBa) induces changes in cytosolic calcium concentration ([Ca²⁺]_cyt) which are the trigger for aBa-induced plasma membrane anion current activation, H⁺-aTPase inhibition, and subsequent plasma membrane depolarization. In the present study, we took advantage of this model to analyze the implication of intracellular Ca²⁺ stores in aBa signal transduction through electrophysiological current measurements, cytosolic Ca²⁺ activity measurements with the apoaequorin Ca²⁺ reporter protein and external pH measurement. Intracellular Ca²⁺ stores involvement was determined by using specific inhibitors of CICR channels: the cADP-ribose/ryanodine receptor (Br-cADPR and dantrolene) and of the inositol trisphosphate receptor ({"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122). In addition experiments were performed on epidermal strips of A. thaliana leaves to monitor stomatal closure in response to ABA in presence of the same pharmacology. Our data provide evidence that ryanodine receptor and inositol trisphosphate receptor could be involved in ABA-induced (1) Ca²⁺ release in the cytosol, (2) anion channel activation and H⁺-ATPase inhibition leading to plasma membrane depolarization and (3) stomatal closure. Intracellular Ca²⁺ release could thus contribute to the control of early events in the ABA signal transduction pathway in A. thaliana.     

Profilin-1 is expressed in human atherosclerotic plaques and induces atherogenic effects on vascular smooth muscle cells

Background

Profilin-1 is an ubiquitous actin binding protein. Under pathological conditions such as diabetes, profilin-1 levels are increased in the vascular endothelium. We recently demonstrated that profilin-1 overexpression triggers indicators of endothelial dysfunction downstream of LDL signaling, and that attenuated expression of profilin-1 confers protection from atherosclerosis in vivo.

Methodology

Here we monitored profilin-1 expression in human atherosclerotic plaques by immunofluorescent staining. The effects of recombinant profilin-1 on atherogenic signaling pathways and cellular responses such as DNA synthesis (BrdU-incorporation) and chemotaxis (modified Boyden-chamber) were evaluated in cultured rat aortic and human coronary vascular smooth muscle cells (VSMCs). Furthermore, the correlation between profilin-1 serum levels and the degree of atherosclerosis was assessed in humans.

Principal Findings

In coronary arteries from patients with coronary heart disease, we found markedly enhanced profilin expression in atherosclerotic plaques compared to the normal vessel wall. Stimulation of rat aortic and human coronary VSMCs with recombinant profilin-1 (10⁻⁶ M) in vitro led to activation of intracellular signaling cascades such as phosphorylation of Erk1/2, p70^S6 kinase and PI3K/Akt within 10 minutes. Furthermore, profilin-1 concentration-dependently induced DNA-synthesis and migration of both rat and human VSMCs, respectively. Inhibition of PI3K (Wortmannin, {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002) or Src-family kinases (SU6656, PP2), but not PLCγ ({"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122), completely abolished profilin-induced cell cycle progression, whereas PI3K inhibition partially reduced the chemotactic response. Finally, we found that profilin-1 serum levels were significantly elevated in patients with severe atherosclerosis in humans (p<0.001 vs. no atherosclerosis or control group).

Conclusions

Profilin-1 expression is significantly enhanced in human atherosclerotic plaques compared to the normal vessel wall, and the serum levels of profilin-1 correlate with the degree of atherosclerosis in humans. The atherogenic effects exerted by profilin-1 on VSMCs suggest an auto-/paracrine role within the plaque. These data indicate that profilin-1 might critically contribute to atherogenesis and may represent a novel therapeutic target.     

Aberrantly Expressed lncRNAs in Primary Varicose Great Saphenous Veins

Long non-coding RNAs (lncRNAs) are key regulatory molecules involved in a variety of biological processes and human diseases. However, the pathological effects of lncRNAs on primary varicose great saphenous veins (GSVs) remain unclear. The purpose of the present study was to identify aberrantly expressed lncRNAs involved in the prevalence of GSV varicosities and predict their potential functions. Using microarray with 33,045 lncRNA and 30,215 mRNA probes, 557 lncRNAs and 980 mRNAs that differed significantly in expression between the varicose great saphenous veins and control veins were identified in six pairs of samples. These lncRNAs were sub-grouped and mRNAs expressed at different levels were clustered into several pathways with six focused on metabolic pathways. Quantitative real-time PCR replication of nine lncRNAs was performed in 32 subjects, validating six lncRNAs ({"type":"entrez-nucleotide","attrs":{"text":"AF119885","term_id":"7770206","term_text":"AF119885"}}AF119885, {"type":"entrez-nucleotide","attrs":{"text":"AK021444","term_id":"10432630","term_text":"AK021444"}}AK021444, {"type":"entrez-nucleotide","attrs":{"text":"NR_027830","term_id":"240255595","term_text":"NR_027830"}}NR_027830, {"type":"entrez-nucleotide","attrs":{"text":"G36810","term_id":"2734477","term_text":"G36810"}}G36810, {"type":"entrez-nucleotide","attrs":{"text":"NR_027927","term_id":"242246950","term_text":"NR_027927"}}NR_027927, uc.345-). A coding-non-coding gene co-expression network revealed that four of these six lncRNAs may be correlated with 11 mRNAs and pathway analysis revealed that they may be correlated with another 8 mRNAs associated with metabolic pathways. In conclusion, aberrantly expressed lncRNAs for GSV varicosities were here systematically screened and validated and their functions were predicted. These findings provide novel insight into the physiology of lncRNAs and the pathogenesis of varicose veins for further investigation. These aberrantly expressed lncRNAs may serve as new therapeutic targets for varicose veins. The Human Ethnics Committee of Shanghai East Hospital, Tongji University School of Medicine approved the study (NO.: 2011-DF-53).     

Virtual Screening as a Strategy for the Identification of Xenobiotics Disrupting Corticosteroid Action

Background

Impaired corticosteroid action caused by genetic and environmental influence, including exposure to hazardous xenobiotics, contributes to the development and progression of metabolic diseases, cardiovascular complications and immune disorders. Novel strategies are thus needed for identifying xenobiotics that interfere with corticosteroid homeostasis. 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2) and mineralocorticoid receptors (MR) are major regulators of corticosteroid action. 11β-HSD2 converts the active glucocorticoid cortisol to the inactive cortisone and protects MR from activation by glucocorticoids. 11β-HSD2 has also an essential role in the placenta to protect the fetus from high maternal cortisol concentrations.

Methods and Principal Findings

We employed a previously constructed 3D-structural library of chemicals with proven and suspected endocrine disrupting effects for virtual screening using a chemical feature-based 11β-HSD pharmacophore. We tested several in silico predicted chemicals in a 11β-HSD2 bioassay. The identified antibiotic lasalocid and the silane-coupling agent {"type":"entrez-nucleotide","attrs":{"text":"AB110873","term_id":"44885399","term_text":"AB110873"}}AB110873 were found to concentration-dependently inhibit 11β-HSD2. Moreover, the silane {"type":"entrez-nucleotide","attrs":{"text":"AB110873","term_id":"44885399","term_text":"AB110873"}}AB110873 was shown to activate MR and stimulate mitochondrial ROS generation and the production of the proinflammatory cytokine interleukin-6 (IL-6). Finally, we constructed a MR pharmacophore, which successfully identified the silane {"type":"entrez-nucleotide","attrs":{"text":"AB110873","term_id":"44885399","term_text":"AB110873"}}AB110873.

Conclusions

Screening of virtual chemical structure libraries can facilitate the identification of xenobiotics inhibiting 11β-HSD2 and/or activating MR. Lasalocid and {"type":"entrez-nucleotide","attrs":{"text":"AB110873","term_id":"44885399","term_text":"AB110873"}}AB110873 belong to new classes of 11β-HSD2 inhibitors. The silane {"type":"entrez-nucleotide","attrs":{"text":"AB110873","term_id":"44885399","term_text":"AB110873"}}AB110873 represents to the best of our knowledge the first industrial chemical shown to activate MR. Furthermore, the MR pharmacophore can now be used for future screening purposes.     

High Prevalence and Significant Association of ESBL and QNR Genes in Pathogenic Klebsiella pneumoniae Isolates of Patients from Kolkata, India

Pathogenic Klebsiella pneumoniae, resistant to beta-lactam and quinolone drugs, is widely recognized as important bacteria causing array of diseases. The resistance property is obtained by acquisition of plasmid encoded bla_TEM, bla_SHV, bla_CTX-M, QNRA, QNRB and QNRS genes. The aim of this study was to document the prevalence and association of these resistant genes in K. pneumoniae infecting patients in India. Approximately 97 and 76.7 % of the 73 K. pneumoniae isolates showed resistance towards beta-lactam and quinolone drugs respectively. Bla genes were detected in 74 % of K. pneumoniae isolates; with prevalence in the following order: bla_TEM > bla_SHV > bla_CTXM. QNR genes were detected in 67 % samples. Chi-square analysis revealed significant association between presence of bla and qnr genes in our study (P value = 0.000125). Sequence analysis of some bla_TEM, bla_SHV, bla_CTX-M and QNRB PCR products revealed presence of bla_TEM1 (GenBank accession: {"type":"entrez-nucleotide","attrs":{"text":"JN193522","term_id":"343796760","term_text":"JN193522"}}JN193522), bla_TEM116 ({"type":"entrez-nucleotide","attrs":{"text":"JN193523","term_id":"343796762","term_text":"JN193523"}}JN193523 and {"type":"entrez-nucleotide","attrs":{"text":"JN193524","term_id":"343796764","term_text":"JN193524"}}JN193524), bla_SHV11, bla_CTXM72 variants ({"type":"entrez-nucleotide","attrs":{"text":"JF523199","term_id":"329024835","term_text":"JF523199"}}JF523199) and QNRB1 ({"type":"entrez-nucleotide","attrs":{"text":"JN193526","term_id":"343796802","term_text":"JN193526"}}JN193526 and {"type":"entrez-nucleotide","attrs":{"text":"JN193527","term_id":"343796804","term_text":"JN193527"}}JN193527) in our samples.     

Prevalence,antimicrobial resistance profile,and characterization of multi-drug resistant bacteria from various infected wounds in North Egypt

Multi-drug resistant (MDR) bacteria associated with wounds are extremely escalating. This study aims to survey different wounds in Alexandria hospitals, North Egypt, to explore the prevalence and characteristics of MDR bacteria for future utilization in antibacterial wound dressing designs. Among various bacterial isolates, we determined 22 MDR bacteria could resist different classes of antibiotics. The collected samples exhibited the prevalence of mono-bacterial infections (60%), while 40% included poly-bacterial species due to previous antibiotic administration. Moreover, Gram-negative bacteria showed dominance with a ratio of 63.6%, while Gram-positive bacteria reported 36.4%. Subsequently, the five most virulent bacteria were identified following the molecular approach by 16S rRNA and physiological properties using the VITEK 2 automated system. They were deposited in GenBank as Staphylococcus haemolyticus MST1 ({"type":"entrez-nucleotide","attrs":{"text":"KY550377","term_id":"1304487819","term_text":"KY550377"}}KY550377), Pseudomonas aeruginosa MST2 ({"type":"entrez-nucleotide","attrs":{"text":"KY550378","term_id":"1304487820","term_text":"KY550378"}}KY550378), Klebsiella pneumoniae MST3 ({"type":"entrez-nucleotide","attrs":{"text":"KY550379","term_id":"1304487821","term_text":"KY550379"}}KY550379), Escherichia coli MST4 ({"type":"entrez-nucleotide","attrs":{"text":"KY550380","term_id":"1304487822","term_text":"KY550380"}}KY550380), and Escherichia coli MST5 ({"type":"entrez-nucleotide","attrs":{"text":"KY550381","term_id":"1304487823","term_text":"KY550381"}}KY550381). In terms of isolation source, S. haemolyticus MST1 was isolated from a traumatic wound, while P. aeruginosa MST2 and E. coli MST4 were procured from hernia surgical wounds, and K. pneumoniae MST3 and E. coli MST5 were obtained from diabetic foot ulcers. Antibiotic sensitivity tests exposed that K. pneumoniae MST3, E. coli MST4, and E. coli MST5 are extended-spectrum β-lactamases (ESBLs) bacteria. Moreover, S. haemolyticus MST1 belongs to the methicillin-resistant coagulase-negative staphylococcus (MRCoNS), whereas P. aeruginosa MST2 exhibited resistance to common empirical bactericidal antibiotics. Overall, the study provides new insights into the prevalent MDR bacteria in Egypt for further use as specific models in formulating antibacterial wound dressings.