[International expert consensus on gene therapy for hereditary hearing loss: based on clinical trials].
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Journal Article↗
Biomedical subjects
L Guo
Publications and source records attributed to L Guo.
Glutamate stimulates glutamate receptor interacting protein 1 degradation by ubiquitin-proteasome system to regulate surface expression of GluR2.
The glutamate receptor interacting protein 1 (GRIP1) is a scaffolding protein in postsynaptic density (PSD), tethering AMPA receptors to other signaling proteins. Here we report that glutamate stimulation caused a rapid reduction in protein levels of GRIP1, but not that of glutamate receptor (GluR) 1, GluR2 and protein interacting with C kinase 1 (PICK1) in rat primary cortical neuron cultures. Down-regulation of GRIP1 by glutamate was blocked by carbobenzoxyl-leucinyl-leucinyl-leucinal (MG132), a proteasome inhibitor and by expression of K48R-ubiquitin, a dominant negative form of ubiquitin. The GRIP1 reduction was inhibited by MK-801, an N-methyl-d-aspartate (NMDA) receptor antagonist, but not by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), an AMPA receptor antagonist. EGTA and 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetra acetic acid tetrakis (BAPTA), two Ca2+ chelators, but not nifedipine, an L-type Ca2+ channel blocker, prevented GRIP1 degradation. Furthermore, MG132 prevented glutamate-stimulated reduction in surface amount of GluR2, and knockdown of GRIP1 by RNAi against GRIP1 reduced surface GluR2 in neurons. Our results suggest that glutamate induces GRIP1 degradation by proteasome through an NMDA receptor-Ca2+ pathway and that GRIP1 degradation may play an important role in regulating GluR2 surface expression.
Animals↗
Measurement of the N-->Delta(+)(1232) transition at high-momentum transfer by pi(0) electroproduction.
We report a new measurement of the exclusive electroproduction reaction gamma(*)p-->pi(0)p to explore the evolution from soft nonperturbative physics to hard processes via the Q(2) dependence of the magnetic (M(1+)), electric (E(1+)), and scalar (S(1+)) multipoles in the N-->Delta transition. 9000 differential cross section data points cover W from threshold to 1.4 GeV/c(2), 4pi center-of-mass solid angle, and Q(2) from 3 to 6 GeV(2)/c(2), the highest yet achieved. It is found that the magnetic form factor G(M)(*) decreases with Q(2) more steeply than the proton magnetic form factor, the ratio E(1+)/M(1+) is small and negative, indicating strong helicity nonconservation, and the ratio S(1+)/M(1+) is negative, while its magnitude increases with Q(2).
Journal Article↗
Search for Theta++ pentaquarks in the exclusive reaction gammap-->K+K-p.
The reaction gammap --> pK+K- was studied at Jefferson Lab with photon energies from 1.8 to 3.8 GeV using a tagged photon beam. The goal was to search for a Theta++ pentaquark, a narrow, doubly charged baryon state having strangeness S=+1 and isospin I=1, in the pK+ invariant mass spectrum. No statistically significant evidence of a Theta++ was found. Upper limits on the total and differential cross section for the reaction gammap --> K-Theta++ were obtained in the mass range from 1.5 to 2.0 GeV/c2, with an upper limit for a narrow resonance with a mass M(Theta++) = 1.54 GeV/c2 of about 0.15 nb, 95% C.L.. This result places a stringent upper limit on the Theta++ width Gamma(Theta++) <0.1 MeV/c2.
Journal Article↗
Measurement of deeply virtual compton scattering with a polarized-proton target.
The longitudinal target-spin asymmetry AUL for the exclusive electroproduction of high-energy photons was measured for the first time in ep-->e;'pgamma. The data have been accumulated at JLab with the CLAS spectrometer using 5.7 GeV electrons and a longitudinally polarized NH3 target. A significant azimuthal angular dependence was observed, resulting from the interference of the deeply virtual Compton scattering and Bethe-Heitler processes. The amplitude of the sinvarphi moment is 0.252+/-0.042stat+/-0.020sys. Theoretical calculations are in good agreement with the magnitude and the kinematic dependence of the target-spin asymmetry, which is sensitive to the generalized parton distributions H and H.
Journal Article↗
Search for the Theta+ Pentaquark in the gammad--> DeltanK+ reaction measured with the CLAS spectrometer.
For the first time, the reaction gammad-->DeltanK+ has been analyzed in order to search for the exotic pentaquark baryon Theta+(1540). The data were taken at Jefferson Laboratory, using the Hall-B tagged-photon beam of energy between 0.8 and 3.6 GeV and the CEBAF Large Acceptance Spectrometer (CLAS). No statistically significant structures were observed in the nK+ invariant-mass distribution. The upper limit on the gammad-->DeltaTheta+ integrated cross section has been calculated and found to be between 5 and 25 nb, depending on the production model assumed. The upper limit on the differential cross section is also reported.
Journal Article↗
Surfactant copolymers prevent aggregation of heat denatured lysozyme.
We investigated the ability of certain triblock copolymer surfactant poloxamers of the form polyethylene oxide-polypropylene oxide-polyethylene oxide (PEO-PPO-PEO), to prevent formation of stable aggregates of heat denatured hen egg lysozyme. Differential scanning calorimetry (DSC) and synchrotron small angle x-ray scattering (SAXS) experiments were performed to study the thermodynamics and solution structures of lysozyme at temperatures between 20 and 90 degrees C in the presence and absence of poloxamers with various molecular weights (8.4-14.3 kDa), but similar hydrophile/hydrophobe (PEO:PPO) ratio of 80%. Poloxmer 188 was found to be very effective in preventing aggregation of heat denatured lysozyme and those functioned as a synthetic surfactant, thus enabling them to refold when the conditions become optimal. For comparison, we measured the ability of 8 kDa polyethylene glycol (PEG) to prevent lysozyme aggregation under same conditions. The results of these studies suggest that poloxamers are more efficient than PEG in preventing aggregation of heat denaturated lysozyme. To achieve equivalence, more than an order of magnitude higher concentration of PEG concentration was needed. Apparently, the presence of a hydrophobic segment in the poloxamers increases their ability to target the hydrophobic region of the unfolded proteins and protect them from self association. Given their biocompatibility and the low concentrations at which they effectively facilitate refolding of denatured proteins, they may be useful in the treatment of burns and other conditions resulting in the denaturation of proteins.
Hydrophobic and Hydrophilic Interactions↗
Search for the Theta+ pentaquark in the reaction gammad --> pK-K+n.
A search for the Theta+ in the reaction gammad --> pK-K+n was completed using the CLAS detector at Jefferson Lab. A study of the same reaction, published earlier, reported the observation of a narrow Theta+ resonance. The present experiment, with more than 30 times the integrated luminosity of our earlier measurement, does not show any evidence for a narrow pentaquark resonance. The angle-integrated upper limit on Theta+ production in the mass range of 1.52-1.56 GeV/c2 for the gammad --> pK-Theta+ reaction is 0.3 nb (95% C.L.). This upper limit depends on assumptions made for the mass and angular distribution of Theta+ production. Using Lambda(1520) production as an empirical measure of rescattering in the deuteron, the cross section upper limit for the elementary gamman --> K-Theta+ reaction is estimated to be a factor of 10 higher, i.e., approximately 3 nb (95% C.L.).
Journal Article↗
Measurement of two- and three-nucleon short-range correlation probabilities in nuclei.
The ratios of inclusive electron scattering cross sections of 4He, 12C, and 56Fe to 3He have been measured at 1 < xB <. At Q2 > 1.4 GeV2, the ratios exhibit two separate plateaus, at 1.5 < xB < 2 and at xB > 2.25. This pattern is predicted by models that include 2- and 3-nucleon short-range correlations (SRC). Relative to A = 3, the per-nucleon probabilities of 3-nucleon SRC are 2.3, 3.1, and 4.4 times larger for A = 4, 12, and 56. This is the first measurement of 3-nucleon SRC probabilities in nuclei.
Journal Article↗
Eta' photoproduction on the proton for photon energies from 1.527 to 2.227 GeV.
Differential cross sections for the reaction gamma p --> eta' p have been measured with the CLAS spectrometer and a tagged photon beam with energies from 1.527 to 2.227 GeV. The results reported here possess much greater accuracy than previous measurements. Analyses of these data suggest for the first time the coupling of the eta'N channel to both the S11(1535) and P11(1710) resonances, known to couple strongly to the etaN channel in photoproduction on the proton, and the importance of J = 3/2 resonances in the process.
Journal Article↗
Long splice variant N type calcium channels are clustered at presynaptic transmitter release sites without modular adaptor proteins.
The presynaptic N type Ca channel (CaV2.2) is associated with the transmitter release site apparatus and plays a critical role in the gating of transmitter release. It has been suggested that a distinct CaV2.2 long C terminal splice variant is targeted to the nerve terminal and is anchored at the release face by calcium/calmodulin-dependent serine protein kinase (CASK) and Munc-18-interacting protein (MINT), two modular adaptor proteins. We used the isolated chick ciliary ganglion calyx terminal together with two new antibodies (L4569, L4570) selective for CaV2.2 long C terminal splice variant to test these hypotheses. CaV2.2 long C terminal splice variant was present at the presynaptic transmitter release sites, as identified by Rab3a-interacting molecule (RIM) co-staining and quantitative immunocytochemistry. CASK was also present at the terminal both in conjunction with, and independent of its binding partner, MINT. Immunoprecipitation of CaV2.2 long C terminal splice variant from brain lysate coprecipitated CASK, confirming that these two proteins can form a complex. However, CASK was not colocalized either with CaV2.2 long C terminal splice variant or the transmitter release site marker RIM at the calyx terminal release face. Neither was MINT colocalized with CaV2.2 long C terminal splice variant. Our results show that native CaV2.2 long C terminal splice variant is targeted to the transmitter release sites at an intact presynaptic terminal. However, the lack of enrichment of CASK at the release site combined with the failure of this protein or its partner MINT to colocalize with CaV2.2 argues against the idea that these modular adaptor proteins anchor CaV2.2 at presynaptic nerve terminals.
Adaptor Proteins, Signal Transducing↗
Search for Theta+ (1540) Pentaquark in High-Statistics Measurement of gammap-->K0K+n at CLAS.
The exclusive reaction gammap-->K0K+n was studied in the photon energy range between 1.6 and 3.8 GeV searching for evidence of the exotic baryon Theta+ (1540)-->nK+. The decay to nK+requires the assignment of strangeness S=+1 to any observed resonance. Data were collected with the CLAS detector at the Thomas Jefferson National Accelerator Facility corresponding to an integrated luminosity of 70 pb-1. No evidence for the Theta+ pentaquark was found. Upper limits were set on the production cross section as function of center-of-mass angle and nK+ mass. The 95% C.L. upper limit on the total cross section for a narrow resonance at 1540 MeV was found to be 0.8 nb.
Journal Article↗
Establishment of a cell-based drug screening system for identifying selective down-regulators of mPGES-1.
OBJECTIVE: To establish a drug screening system based on transcriptional regulation of microsomal PGE(2) synthase 1 (mPGES-1), cyclooxygenase 1 (COX-1) and 2 (COX-2) for discovering selective down-regulators of mPGES-1. METHODS: The upstream regulatory sequences of mPGES-1, COX-1, COX-2 were respectively cloned into pGL3B-neo vector containing luciferase gene and neomycin resistance gene (the pGL3B-neo vector had been previously constructed by cloning the neomycin resistance gene into the Sal I site of the pGL3-Basic vector). After that, the recombinant reporter gene vectors pGL3B-neo/mPGES-1, pGL3B-neo/COX-1, pGL3B-neo/COX-2 were respectively transfected into A549 cells and therefore stable cell lines, namely M 1, M 2 and M 3, were obtained. Samples were detected then by testing luciferase activity of M 1, M 2 and M 3 cells in microtiter wells to identify compounds that can selectively down-regulate mPGES-1 expression. Through luciferase activity testing, the compounds which had more than 40 % inhibition ratio on M 1 and less than 20 % inhibition ratio on M 2 and M 3 cells could be regarded as hits. RESULTS: Using the cell-based reporter gene assay, we screened compounds for selectively down-regulation of mPGES-1 expression and several compounds were discovered. CONCLUSION: A cell-based drug screening system was established to screen selective down-regulators of mPGES-1 expression, and compound CM188 was identified, which might become a lead compound for novel anti-arthritic drugs.
Acetates↗
Mitochondria and calpains mediate caspase-dependent apoptosis induced by doxycycline in HeLa cells.
Doxycycline (Dc) has been demonstrated to inhibit cell growth and induce apoptosis in tumor cells, although its mechanism of action is not fully understood. The present study demonstrates that apoptosis can be induced in HeLa cells. Western blot data demonstrated that cytochrome c (Cyt c), Smac (the second mitochondria-derived activator of caspase), calpain I, caspase-9, -3 and -8 were involved in the apoptotic process, while the pan caspase inhibitor zVAD-fmk almost completely inhibited Dc-induced apoptosis. We further demonstrated that the release of mitochondrial proteins and the activation of calpains occurred upstream of the caspase cascade, in which caspase-9 was activated in response to the release of Cyt c, that caspase-8 activation was caspase and calpain dependent, and that caspase-3 was activated mainly by caspase-8 and -9. Caspase-8 played important roles in the activation of caspase-3 and induction of apoptosis, whereas the role of the caspase-9 was limited.
Apoptosis↗
Cytokeratin-positive subserosal myofibroblasts in gastroduodenal ulcer; another type of myofibroblasts.
To investigate the distribution and origin of alpha-smooth muscle actin (ASMA)-positive stromal cells in the perforation of human gastroduodenal ulcers. Perforative lesions of 24 surgically resected gastroduodenal ulcers were examined immunohistochemically for ASMA, HCD, CD34, CD31, CAM5.2 and HMW-CK, and double staining of ASMA and CAM5.2 was also performed. In addition, to determine the cell source of collagen, in situ hybridization of collagen I mRNA was performed. In the normal gastroduodenal wall, the reticular network of CD34-positive stromal cells was identified in the muscularis mucosa, submucosa, muscular propria, and subserosa. In the subepithelial area, many myofibroblasts were observed, whereas no CD34-positive stromal cells were seen. In areas neighboring ulcerative lesions, no CD34-positive stromal cells were observed, but a significant number of myofibroblasts were present there. In the deep layer of ulceration, numerous fusiform or stellate stromal cells strongly positive for ASMA and CAM5.2 were observed in the subserosal area around the perforation. In the same site, many cells co-expressing ASMA and CAM5.2 were identified by double staining. In contrast, in the surface layer of ulceration, stromal cells expressing only ASMA were observed. The cytokeratin-positive subserosal myofibroblastic cell in human gastroduodenal ulcer is a novel type of myofibroblast.
Actins↗
Effect of copper and thiomolybdates on bovine theca cell differentiation in vitro.
Subfertility that will respond to appropriate copper supplementation is a widespread problem in the national dairy herd. The aims of this study were to determine the effect of copper and/or copper chelating thiomolybdates on LH-induced differentiation by looking at the effects on androgen production, steroidogenic enzymes (cytochrome P450 17alpha-hydroxylase and cytochrome P450 side-chain cleavage) and lysyl oxidase mRNA expression in cultured theca cells maintained under serum-free conditions. The effect of thiomolybdates and copper on LH differentiation was investigated by supplementing (ammonium) tetrathiomolybdate to optimum theca cell culture media at 0-100 microg/ml, copper (chloride) at equimolar concentrations (0-51.6 microg/ml) or equimolar combinations of both media. Lysyl oxidase mRNA expression was investigated with semi-quantitative RT-PCR, whilst expression of cytochrome P450 17alpha-hydroxylase and cytochrome P450 side-chain cleavage mRNA was examined using real time PCR. Both PCRs used bovine specific primers and cell lysates obtained from bovine theca cells cultured for 6 days and in the presence or absence of the 100 microg/ml dose of thiomolybdate and equimolar copper thiomolybdate. Thiomolybdate depressed androstenedione production in a dose-dependent manner at doses greater than 1 microg/ml at 96 h (P<0.05); doses above 20 microg/ml were all greatly reduced at all time points and at 192 h when related to numbers of cells (P<0.001). Copper alone had no effect at physiological doses, but the use of the equimolar copper thiomolybdate reversed the effect of tetrathiomolybdates on androstenedione production at the 20 microg/ml dose. Thiomolybdate supplementation, with and without copper, had no significant effect on the level of lysyl oxidase or cytochrome P450 side-chain cleavage expression. However, cytochrome P450 17alpha-hydroxylase expression was significantly increased (P<0.05) by tetrathiomolybdate, possibly due to a local regulatory system. In conclusion, these results demonstrate that thiomolybdates can prevent LH-induced differentiation of bovine theca cells in vitro and that these effects can be ameliorated by copper supplementation. Our results also indicate that it is unlikely that the effects of thiomolybdate are mediated at the transcriptional level and further work is required to determine if thiomolybdate exerts its effects through post-translation processing or some other unrelated mechanism. Overall, these data support the hypothesis that copper responsive subfertility results from perturbation of the normal pattern of ovarian steroidogenesis.
Animals↗
Beam-helicity asymmetries in double-charged-pion photoproduction on the proton.
Beam-helicity asymmetries for the two-pion-photoproduction reaction gammap-->ppi(+)pi(-) have been studied for the first time in the resonance region for center-of-mass energies between 1.35 and 2.30 GeV. The experiment was performed at Jefferson Lab with the CEBAF Large Acceptance Spectrometer using circularly polarized tagged photons incident on an unpolarized hydrogen target. Beam-helicity-dependent angular distributions of the final-state particles were measured. The large cross-section asymmetries exhibit strong sensitivity to the kinematics and dynamics of the reaction. The data are compared with the results of various phenomenological model calculations, and show that these models currently do not provide an adequate description for the behavior of this new observable.
Journal Article↗
Human papillomavirus infection in women who develop high-grade cervical intraepithelial neoplasia or cervical cancer: a case-control study in the UK.
Human papillomavirus (HPV) testing might identify older women who could be withdrawn from the cervical screening programme, or require less frequent screening. A case-control study using the United Kingdom cervical screening population was set up to help address this issue. Cases comprised 575 women who developed cervical intraepithelial neoplasia (CIN) grade 2 or worse over a 13-year period following a cytologically normal baseline smear, and were stratified by age group ('under 20', '20-39' and 40 years or over). Controls (n=601) were women who remained disease free over this interval and were the same age on average as cases. DNA was extracted from the baseline smears and tested for HPV by PCR using GP5+/6+ consensus primers. HPV+ samples were tested for HPV types 16 and 18 using specific PCR primers. In all, 27.0% of cases tested positive for HPV at baseline, compared with 15.4% of controls (odds ratio (OR)=2.00; 95% confidence interval (CI), 1.50-2.68). Among women aged 40 years or over, the OR for HPV 16 was 8.95 (95% CI, 2.63-30.4). These results support the need for further cervical screening of HPV- older women, as many of the cases were HPV- at baseline.
Adult↗