Constituents of secondary organic carbon (SOC) in atmospheric aerosols are often mixed with inorganic components and compose a significant mass fraction of fine particulate matter in the atmosphere. Interactions between SOC and other condensed-phase species are not well understood. Here, we investigate the reactions of liquid-like and semi-solid SOC from ozonolysis of limonene (LSOC) and ?-pinene (PSOC) with NaCl using a set of complementary micro-spectroscopic analyses. These reactions result in chloride depletion in the condensed phase, release of gaseous HCl, and formation of organic salts. The reactions attributed to acid displacement by SOC acidic components are driven by the high volatility of HCl. Similar reactions can take place in SOC/NaNO3 particles. The results show that an increase in SOC mass fraction in the internally mixed SOC/NaCl particles leads to higher chloride depletion. Glass transition temperatures and viscosity of PSOC were estimated for atmospherically relevant conditions. Data show that the reaction extent depends on SOC composition, particle phase state and viscosity, mixing state, temperature, relative humidity (RH), and reaction time. LSOC shows slightly higher potential to deplete chloride than PSOC. Higher particle viscosity at low temperatures and RH can hinder these acid displacement reactions. Formation of organic salts from these overlooked reactions can alter particle physiochemical properties and may affect their reactivity and ability to act as cloud condensation and ice nuclei. The release and potential recycling of HCl and HNO3 from reacted aerosol particles may have important implications for atmospheric chemistry.
Complementary methods of high-resolution mass spectrometry and microspectroscopy were utilized for molecular analysis of secondary organic aerosol (SOA) generated from ozonolysis of two structural monoterpene isomers: d-limonene SOA (LSOA) and ?-pinene SOA (PSOA). The LSOA compounds readily formed adducts with Na(+) under electrospray ionization conditions, with only a small fraction of compounds detected in the protonated form. In contrast, a significant fraction of PSOA compounds appeared in the protonated form because of their increased molecular rigidity. Laboratory simulated aging of LSOA and PSOA, through conversion of carbonyls into imines mediated by NH3 vapors in humid air, resulted in selective browning of the LSOA sample, while the PSOA sample remained white. Comparative analysis of the reaction products in the aged LSOA and PSOA samples provided insights into chemistry relevant to formation of brown carbon chromophores. A significant fraction of carbonyl-imine conversion products with identical molecular formulas was detected in both samples. This reflects the high level of similarity in the molecular composition of these two closely related SOA materials. Several highly conjugated products were detected exclusively in the brown LSOA sample and were identified as potential chromophores responsible for the observed color change. The majority of the unique products in the aged LSOA sample with the highest number of double bonds contain two nitrogen atoms. We conclude that chromophores characteristic of the carbonyl-imine chemistry in LSOA are highly conjugated oligomers of secondary imines (Schiff bases) present at relatively low concentrations. Formation of this type of conjugated compounds in PSOA is hindered by the structural rigidity of the ?-pinene oxidation products. Our results suggest that the overall light-absorbing properties of SOA may be determined by trace amounts of strong brown carbon chromophores.
We evaluated the immunogenicity and efficacy of a candidate vaccine comprising the major outer membrane protein (MOMP) multi-epitope of Chlamydia trachomatis. A short gene of multi-epitope derived from MOMP containing multiple T- and B-cell epitopes was artificially synthesized. The recombinant plasmid pET32a(+) containing codon optimized MOMP multi-epitope gene was constructed. Expression of the fusion protein Trx-His-MOMP multi-epitope in Escherichia coli was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blot analysis. Balb/c mice were inoculated with the purified fusion protein subcutaneously three times with 2-week intervals. Results showed that the MOMP multi-epitope elicited not only strong humoral immune responses to C. trachomatis by generating significantly high levels of specific antibodies (IgG1 and IgG2a), but also a cellular immune response by inducing robust cytotoxic T lymphocyte responses in mice. Furthermore, the MOMP multi-epitope substantially primed secretion of IFN-?, revealing that this vaccine could induce a strong Th1 response. Finally, the mice vaccinated with the MOMP multi-epitope displayed a reduction of C. trachomatis shedding upon a chlamydial challenge and an accelerated clearance of the infected C. trachomatis. In conclusion, the MOMP multi-epitope vaccine may have the potentiality for the development of effective prophylactic and therapeutic vaccines against the C. trachomatis infection.
Medicines for the treatment of most human pathologies are encumbered by unwanted side effects that arise from the deposition of an effective drug into the wrong tissues. The logical remedy for these undesirable properties involves selective targeting of the therapeutic agent to pathologic cells, thereby avoiding collateral toxicity to healthy cells. Since significant advantages can also accrue by incorporating a therapeutic or imaging agent into a nanoparticle, many laboratories are now combining both benefits into a single formulation. This review will focus on the major guiding principles in the design of ligand-targeted nanoparticles, including optimization of their chemical and physical properties, selection of the ideal targeting ligand, engineering of the appropriate surface passivation and linker strategies to achieve selective delivery of the entrapped cargo to the desired diseased cell.
So far, nonsequential double ionization (NSDI) of atoms can be well understood within a semiclassical or even classical picture. No quantum effect appears to be required to explain the data observed. We theoretically study electron correlation resulting from NSDI of argon in a low-intensity laser field using a quantum-mechanical S-matrix theory. We show that quantum interference between the contributions of different intermediate excited states of the singly charged argon ion produces a transition from back-to-back to side-by-side emission with increasing laser intensity, which is in close agreement with the experimental data. For higher intensities, this transition is enhanced by the consequences of depletion of the excited states.
Atmospheric aging of naturally emitted marine aerosol often leads to formation of internally mixed particles composed of sea salts and water-soluble organic compounds of anthropogenic origin. Mixing of sea salt and organic components has profound effects on the evolving chemical composition and hygroscopic properties of the resulted particles, which are poorly understood. Here, we have studied chemical composition and hygroscopic properties of laboratory generated NaCl particles mixed with malonic acid (MA) and glutaric acid (GA) at different molar ratios using micro-FTIR spectroscopy, atomic force microscopy, and X-ray elemental microanalysis. Hygroscopic properties of internally mixed NaCl and organic acid particles were distinctly different from pure components and varied significantly with the type and amount of organic compound present. Experimental results were in a good agreement with the AIM modeling calculations of gas/liquid/solid partitioning in studied systems. X-ray elemental microanalysis of particles showed that Cl/Na ratio decreased with increasing organic acid component in the particles with MA yielding lower ratios relative to GA. We attribute the depletion of chloride to the formation of sodium malonate and sodium glutarate salts resulted by HCl evaporation from dehydrating particles.
Context:Progesterone contributes to maintenance of human pregnancy, in part by inhibiting activity of the human pro-labor genes corticotropin-releasing hormone (CRH) and cyclooxygenase-2 (COX-2). However, the molecular mechanisms underlying action of progesterone remain poorly defined. We have shown that in human placenta, constitutively activated non-canonical NF-?B pathway positively regulates CRH and COX-2, which is further stimulated by glucocorticoid receptor signaling.Objective:We investigated the role of progesterone receptor (PR) in regulation of nuclear activity of RelB/NF-?B2, and in turn, expression of placental CRH and COX-2.Methods:We used a variety of techniques including gene silencing, ectopic expression, chromatin immunoprecipitation (ChIP), Western blot, RT-qPCR, and immunohistochemical staining assays in human placental tissues and primary culture of human cytotrophoblast.Results:We identified PR isoform-A (PR-A) as the only isoform of PR produced in human placenta. PR-A levels were lower in term placenta than in mid-term placenta. Depletion of PR-A by short interfering RNA derepressed inhibition of CRH and COX-2 by progesterone (P4) and the synthetic progestin, 17?-hydroxyprogesterone caproate (17-OHP). Overexpression of PR-A inhibited transcription of CRH and COX-2, which was further down-regulated by treatment of P4 or 17-OHP. Such an inhibition was mediated by negative functional interaction of PR-A with activity of RelB/NF-?B2.Conclusion:Progesterone inhibits the pro-labor genes CRH and COX- 2 via PR-A repression of the non-canonical NF-?B signaling in human placenta. Characterization of these pathways may identify potential drug targets for prevention of preterm birth.
We have designed, fabricated, and tested a compact gas-phase reactor for performing in situ soft x-ray scanning transmission x-ray microscopy (STXM) measurements. The reactor mounts directly to the existing sample holder used in the majority of STXM instruments around the world and installs with minimal instrument reconfiguration. The reactor accommodates many gas atmospheres, but was designed specifically to address the needs of measurements under water vapor. An on-board sensor measures the relative humidity and temperature inside the reactor, minimizing uncertainties associated with measuring these quantities outside the instrument. The reactor reduces x-ray absorption from the process gas by over 85% compared to analogous experiments with the entire STXM instrument filled with process gas. Reduced absorption by the process gas allows data collection at full instrumental resolution, minimizes radiation dose to the sample, and results in much more stable imaging conditions. The reactor is in use at the STXM instruments at beamlines 11.0.2 and 184.108.40.206 at the Advanced Light Source.
A high-throughput RNA interference (RNAi) screen targeting 542 genes of the human kinome was used to discover regulators of RNAi. Here we report that the proto-oncogene Akt-3/PKB? (Akt3) phosphorylates Argonaute 2 (Ago2) at S387, which downregulates cleavage and upregulates translational repression of endogenous microRNA (miRNA)-targeted messenger RNAs (mRNAs). We further demonstrate that Akt3 coimmunoprecipitates with Ago2 and phosphorylation of Ago2 at S387 facilitates its interaction with GW182 and localization to cytoplasmic processing bodies (P bodies), where miRNA-targeted mRNAs are thought to be stored and degraded. Therefore, Akt3-mediated phosphorylation of Ago2 is a molecular switch between target mRNA cleavage and translational repression activities of Ago2.
We have developed a rapid and high throughput lipase-ANS (8-Anilino-l-naphthalenesulfonic acid) assay to evaluate the thermo-stability of lipases based on the ANS fluorescence signals increasing and shifting when this small fluorescence probes binds to lipase. The testing lipase samples were incubated at a temperature range of 25 degrees C to 65 degrees C for 30 min before mixed with ANS solution (0.20 mg/mL lipase and 0.05 mmol/L ANS in the buffer of 20 mmol/L Tris-HCl, 100 mmol/L NaCl, pH 7.2) in a cuvette or microplate. Fluorescence signals of the samples were measured at EX 378 nm, EM 465 nm with a fluorescence photometer or a plate reader, and Tm was calculated with the software of GraphPad Prism5.0. The Tm values of several mutants of Penicillium expansum lipase (PEL) were measured with this ANS assay and conventional method simultaneously and the results show that Tm values are comparative and consistent between these methods, suggesting that the lipase-ANS assay is a reliable, rapid and high throughput method for lipase thermo-stability measurement.
The discovery of potent N-hydroxyl caprolactam matrix metalloproteinase (MMP) inhibitors (6) based on the natural product Cobactin-T (2) is described. The synthetic method, which utilizes the ring closing metathesis reaction, is compatible to provide complementary (R) and (S) enantiomers. These compounds tested against MMP-2 and 9, show that the R stereochemistry (i.e., 16), which is opposite for that found in the natural product Cobactin-T is >1000-fold more active with IC(50) values of 0.2-0.6nM against both enzymes. The variation in the incorporation of the sulfonamide enzyme recognition element (Ar(2)XAr(1)SO(2)N(R(1)), 6), along with alterations in the RCM/double bond chemistry (R(2)) provided a series of sub nanomolar MMP inhibitors. For example, compounds 16 and 34 were found to be the most potent with IC(50) values against MMP-2 and MMP-9 found to be between 0.2 and 0.6nM with 34 being the most potent compound discovered (MMP-2 IC(50)=0.39nM and MMP-9 IC(50)=0.22nM). Compounds 16 and 34 showed acceptable drug-like properties in vivo with compound 34 showing oral bioavailability.
Essential hypertension has been recognized as a disease resulting from a combination of environmental and genetic factors. Recent studies demonstrated that microRNAs (miRNAs) are involved in cardiac hypertrophy and heart failure. However, little is known about the roles of miRNAs in essential hypertension.
[Gd@C82(OH)22]n, a fullerene-based nanoparticle, exhibits potent anti-tumor effects in mouse tumor-bearing models without detectable toxicity and the pathological studies revealed a massive infiltration of leukocytes in the residual tumors of [Gd@C82(OH)22]n-treated mice. We report here that [Gd@C82(OH)22]n promotes macrophages secreting pro-inflammatory cytokines IL-6 and TNF-alpha, enhances the expression of MHC-II and costimulatory molecules such as CD40 and CD54, increases endocytosis and cell adhesion. Furthermore, [Gd@C82(OH)22]n-treated macrophages became functionally activated as illustrated by their capacity to activate allogeneic T cells. Taken together, our results indicate that [Gd@C82(OH)22]n nanoparticle is a potent activator of macrophages, which may in part account for its potent anti-tumor effect.
Malignant gliomas have a tendency to invade diffusely into surrounding healthy brain tissues, thereby precluding their successful surgical removal. Intersectin1 (ITSN1) as a molecular linker in the central nervous system is well known as an important regulator of endocytosis and exocytosis. ITSN1 has two isoforms: ITSN1-l and ITSN1-s. In this study, we show that siRNA-mediated down regulation of ITSN1-s inhibited migration and invasion of glioma cells. In addition, we demonstrate the possible mechanisms by which ITSN1-s functions in migration and invasion. Several key proteins, including cofilin, LIMK, PAK, FAK, integrin ?1, and MMP-9, which are critical for cells migration and invasion, were probably involved in ITSN1-s signaling pathways. These results suggest that ITSN1-s contributes to glioma cells migration and invasion by regulating the formation of cytoskeleton, influencing adhesion and increasing expression of MMP-9. Our results indicate that ITSN1-s is a critical factor in gliomas invasion and identify that ITSN1-s is a new potentially antiinvasion target for therapeutic intervention in gliomas.
The cellobiase gene from Aspergillus niger was cloned and connected with the strong promoter Pcbh1 from Trichoderma reesei to construct a recombinant plasmid pHB9 with the hygromycin B resistance marker. The plasmid was transformed into conidia of T. reesei using the modified PEG-CaCl(2) method. Main factors effecting the transformation were discussed and about 99-113 transformants/?g DNA could be obtained under optimal conditions. It was found that the molecular mass of the recombinant cellobiase was about 120 kDa by SDS-PAGE analysis. The activity of cellobiase could reach 5.3 IU/ml after 48 h fermentation, which was as high as 106 times compared with that of the host strain. Meanwhile, the filter paper activity of recombinant T. reesei was 1.44-fold of the host strain. Saccharification of corncob residue with the crude enzyme showed that the hydrolysis yield (84.2%) of recombinant T. reesei was 21% higher than that (69.5%) of the host strain.
NPRL2, one of the tumor suppressor genes residing in a 120-kb homozygous deletion region of human chromosome band 3p21.3, has a high degree of amino acid sequence homology with the nitrogen permease regulator 2 (NPR2) yeast gene, and mutations of NPRL2 in yeast cells are associated with resistance to cisplatin-mediated cell killing. Previously, we showed that restoration of NPRL2 in NPRL2-negative and cisplatin-resistant cells resensitize lung cancer cells to cisplatin treatment in vitro and in vivo. In this study, we show that sensitization of non-small cell lung cancer (NSCLC) cells to cisplatin by NPRL2 is accomplished through the regulation of key components in the DNA-damage checkpoint pathway. NPRL2 can phosphorylate ataxia telangiectasia mutated (ATM) kinase activated by cisplatin and promote downstream gamma-H2AX formation in vitro and in vivo, which occurs during apoptosis concurrently with the initial appearance of high-molecular-weight DNA fragments. Moreover, this combination treatment results in higher Chk1 and Chk2 kinase activity than does treatment with cisplatin alone and can activate Chk2 in pleural metastases tumor xenograft in mice. Activated Chk1 and Chk2 increase the expression of cell cycle checkpoint proteins, including Cdc25A and Cdc25C, leading to higher levels of G2/M arrest in tumor cells treated with NPRL2 and cisplatin than in tumor cells treated with cisplatin only. Our results therefore suggest that ectopic expression of NPRL2 activates the DNA damage checkpoint pathway in cisplatin-resistant and NPRL2-negative cells; hence, the combination of NPRL2 and cisplatin can resensitize cisplatin nonresponders to cisplatin treatment through the activation of the DNA damage checkpoint pathway, leading to cell arrest in the G2/M phase and induction of apoptosis. The direct implication of this study is that combination treatment with NPRL2 and cisplatin may overcome cisplatin resistance and enhance therapeutic efficacy.
Malignant gliomas have a high proliferation ability and high tendency to invade diffusely into surrounding healthy brain tissues, thereby precluding their successful surgical removal. Intersectin1 (also called ITSN1) as a molecular linker in the central nervous system is well known as an important regulator of endocytosis and exocytosis. ITSN1 has two isoforms: ITSN1-l and ITSN1-s. In this study, we show that siRNA-mediated down regulation of ITSN1-s induced glioma cells apoptosis. In addition, we demonstrate the possible mechanisms by which ITSN1-s functions in glioma cells apoptosis. Our data demonstrate that several key proteins, including FAK, Akt, Bcl-2, BAD which are critical for cells apoptosis were probably involved in ITSN1-s signaling pathways. Our results indicate that ITSN1-s is an effecter in regulation of gliomas cells apoptosis, and identify that ITSN1-s may be a new potentially anti-apoptosis target for therapeutic of gliomas.
Top-selective surface modification has been widely used for the synthesis of Janus nanoparticles (NPs). Herein we demonstrate that polymer single crystals can serve as generic substrates to immobilize NPs and the resultant NPs are Janus in nature. This technique is generic because various NPs as well as polymer single crystal substrates can be used. Single crystals of poly(ethylene oxide), polycaprolactone, and polyethylene-block-poly(ethylene oxide) have been successfully used to immobilize gold, magnetic, and semiconducting NPs. Subsequent dissolution of the single crystals led to various types of Janus NPs and NP clusters with different polymer brushes.
Argonaute (AGO) proteins bind to small RNAs and mediate small RNA-induced silencing in eukaryotes. Using a minimal in vitro system, we show that bacterially expressed human AGO1 and AGO2 but not AGO3 and AGO4 possess strand-dissociating activity of microRNA (miRNA) duplexes. Both AGO1 and AGO2 function as RNA chaperones, capable of performing multiple rounds of strand dissociation. Unexpectedly, both AGO1 and AGO2 demonstrate passenger strand cleavage activity of a small interfering RNA (siRNA) duplex, but only AGO2 has target RNA cleavage activity. These observations indicate that passenger strand and mRNA endonuclease activities are mechanistically distinct. We further validate these observations in mammalian extracts and cultured mammalian cells, in which we demonstrate that AGO1 uses only miRNA duplexes when assembling translational repression-competent complexes, whereas AGO2 can use both miRNA and siRNA duplexes. We show that passenger strand cleavage and RNA chaperone activities that are intrinsic to both AGO1 and AGO2 are sufficient for RNA-induced silencing complex (RISC) loading.
Scientific and technological interest in one-dimensional nanomaterials, in particular carbon nanotubes, is a result of their fascinating properties and their ability to serve as templates for directed assembly. For applications in nanoelectronics it is necessary to create ordered arrays of nanotubes for large-scale integrated circuits, an area in which there has been significant progress, and to produce controllable patterns on individual nanotubes so that multiple transistors can be fabricated on them, an area where progress has been slower. Here, we show that judiciously selected crystalline block copolymers can be periodically decorated along carbon nanotubes, leading to amphiphilic, alternating patterns with a period of approximately 12 nm. In addition, end-functionalization of the block copolymers allowed gold nanoparticles to be periodically attached to the nanotubes. This approach provides a facile technique for the periodic patterning of one-dimensional nanomaterials.
The synthesis and biological evaluation of JAK3 based staurosporine compounds is described. The compounds are constructed completely de novo, and a ring closing metathesis strategy is used to assemble the sugar mimetic portion. These analogs show potent JAK3 activity against isolated enzyme and in T-cells. One analog (32) showed unique biological effects during in vitro and in vivo tests including inhibition of STAT5 phosphorylation, blockade of mast cell responses, and reduction of JAK3 based effects in mice models of allergic disease.
Our recent study demonstrated that constitutively activated RelB/NF-?B2 positively regulates the CRH in the human placenta. In the current study, we explored the role of the glucocorticoid receptor (GR) signaling in constitutive activation of the noncanonical NF-?B pathway. A glucocorticoid response element (GRE) motif search suggests that both NF-?B inducing kinase (NIK) and RelB genes, which are key regulators of the noncanonical NF-?B pathway, have a putative GRE within their promoter, approximately 1 kb upstream from the transcription start site. By using chromatin immunoprecipitation assay we identified that the GR and phosphorylated GR at Ser211 were associated with the GREs of both NIK and RelB. Dexamethasone stimulated expression of NIK, RelB, NF-?B2 as well as CRH and cyclooxygenase-2 (COX-2). Repression of GR by short interfering RNA resulted in inhibition of NIK, RelB, NF-?B2, CRH, and COX-2. In addition, depletion of GR attenuated glucocorticoid-mediated up-regulation of NIK, RelB, NF-?B2, CRH, and COX-2. Furthermore, siRNA specifically targeting NIK down-regulated CRH and COX-2. Taken together, these results suggest that constitutive activation of the noncanonical NF-?B pathway in term human placenta is driven by the GR signaling, which in turn up-regulates placental CRH and other NF-?B-responsive genes.
MicroRNAs (miRNAs) are small noncoding RNAs that post-transcriptionally regulate protein output from the majority of human mRNAs. In contrast to the consensus view that all miRNAs are associated with Argonaute (Ago) proteins, we determine that miRNAs are expressed in a 13-fold excess relative to Agos in HeLa cells and that miRNAs are bound to mRNAs in a sevenfold excess relative to Agos, implying the existence of miRNA-mRNA duplexes not stoichiometrically bound by Agos. We show that all four human Agos can repress miRNA-mRNA duplexes, but only Ago2 can cleave small interfering RNA-mRNA duplexes in vitro. We visualize direct Ago binding to miRNA-mRNA duplexes in live cells using fluorescence lifetime imaging microscopy. In contrast to the consensus view that Agos bind miRNA duplexes, these data demonstrate that Agos can bind and repress miRNA-mRNA duplexes and support a model of catalytic Ago function in translational repression.
Placental CRH may be part of a clock that governs the length of human gestation. The mechanism underlying differential regulation of CRH in the human placenta is poorly understood. We report here that constitutively activated RelB/nuclear factor-?B2 (NF-?B)-2 (p100/p52) acts as an endogenous stimulatory signal to regulate CRH by binding to an NF-?B enhancer of CRH gene promoter in the human placenta. Nuclear staining of NF-?B2 and RelB in villous syncytiotrophoblasts and cytotrophoblasts was coupled with cytoplasmic CRH in syncytial knots of cytotrophoblasts. Chromatin immunoprecipitation identified that CRH gene associated with both RelB and NF-?B2 (p52). Dexamethasone increased synthesis and nuclear translocation of RelB and NF-?B2 (p52) and their association with the CRH gene. In contrast, progesterone, a down-regulator of placental CRH, repressed NF-?B2 (p100) processing, nuclear translocation of RelB and NF-?B2 (p52), and their association with the CRH gene. Luciferase reporter assay determined that the NF-?B enhancer of CRH was sufficient to regulate transcriptional activity of a heterologous promoter in primary cytotrophoblasts. RNA interference-mediated repression of RelB or NF-?B2 resulted in significant inhibition of CRH at both transcriptional and translational levels and prevented the dexamethasone-mediated up-regulation of CRH transcription and translation. These results suggest that the noncanonical NF-?B pathway regulates CRH production in the human placenta and is responsible for the positive regulation of CRH by glucocorticoids.
Based on fractal theory and damage mechanics, the aim of this paper is to describe the monofractal and multifractal characteristics of corrosion morphology and develop a new approach to characterize the nonuniform corrosion degree of reinforcing bars. The relationship between fractal parameters and tensile strength of reinforcing bars are discussed. The results showed that corrosion mass loss ratio of a bar cannot accurately reflect the damage degree of the bar. The corrosion morphology of reinforcing bars exhibits both monofractal and multifractal features. The fractal dimension and the tensile strength of corroded steel bars exhibit a power function relationship, while the width of multifractal spectrum and tensile strength of corroded steel bars exhibit a linear relationship. By comparison, using width of multifractal spectrum as multifractal damage variable not only reflects the distribution of corrosion damage in reinforcing bars, but also reveals the influence of nonuniform corrosion on the mechanical properties of reinforcing bars. The present research provides a new approach for the establishment of corrosion damage constitutive models of reinforcing bars.
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