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51.
Byounghoon Hwang Funita P. Phan Kevin McCool Eun Young Choi Jinsam You Adam Johnson Anjon Audhya Shigeki Miyamoto 《PloS one》2015,10(3)
NF-κB essential modulator, NEMO, plays a key role in canonical NF-κB signaling induced by a variety of stimuli, including cytokines and genotoxic agents. To dissect the different biochemical and functional roles of NEMO in NF-κB signaling, various mutant forms of NEMO have been previously analyzed. However, transient or stable overexpression of wild-type NEMO can significantly inhibit NF-κB activation, thereby confounding the analysis of NEMO mutant phenotypes. What levels of NEMO overexpression lead to such an artifact and what levels are tolerated with no significant impact on NEMO function in NF-κB activation are currently unknown. Here we purified full-length recombinant human NEMO protein and used it as a standard to quantify the average number of NEMO molecules per cell in a 1.3E2 NEMO-deficient murine pre-B cell clone stably reconstituted with full-length human NEMO (C5). We determined that the C5 cell clone has an average of 4 x 105 molecules of NEMO per cell. Stable reconstitution of 1.3E2 cells with different numbers of NEMO molecules per cell has demonstrated that a 10-fold range of NEMO expression (0.6–6x105 molecules per cell) yields statistically equivalent NF-κB activation in response to the DNA damaging agent etoposide. Using the C5 cell line, we also quantified the number of NEMO molecules per cell in several commonly employed human cell lines. These results establish baseline numbers of endogenous NEMO per cell and highlight surprisingly normal functionality of NEMO in the DNA damage pathway over a wide range of expression levels that can provide a guideline for future NEMO reconstitution studies. 相似文献
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Phan Tu Quy Nguyen Khoa Hien Nguyen Chi Bao Doan Thanh Nhan Dang Van Khanh Nguyen Thi Ai Nhung Truong Quy Tung Nguyen Dinh Luyen Duong Tuan Quang 《Luminescence》2015,30(3):325-329
A new rhodamine–ethylenediamine–nitrothiourea conjugate (RT) was synthesized and its sensing property as a fluorescent chemodosimeter toward metal ions was explored in water media. Analytical results from absorption and fluorescence spectra revealed that the addition of Hg2+ ions to the aqueous solution of the chemodosimeter RT caused a distinct fluorescence OFF–ON response with a remarkable visual color change from colorless to pink; however, no clear spectral and color changes were observed from other metal ions including: Zn2+, Cu2+, Cd2+, Pb2+, Ag+, Fe2+, Cr3+, Co3+, Ni2+, Ca2+, Mg2+, K+ and Na+. The sensing results and the molecular structure suggested that a Hg2+‐induced a desulfurization reaction and cyclic guanylation of the thiourea moiety followed by ring‐opening of the rhodamine spirolactam in RT are responsible for a distinct fluorescence turn‐on signal, indicating that RT is a remarkably sensitive and selective chemodosimeter for Hg2+ ions in aqueous solution. Hg2+ within a concentration range from 0.1 to 25 μM can be determined using RT as a chemodosimeter and a detection limit of 0.04 μM is achieved. Copyright © 2014 John Wiley & Sons, Ltd. 相似文献
54.
Kathleen M. Yoder Mimi L. Phan Kai Lu David S. Vicario 《Developmental neurobiology》2015,75(3):302-314
Songbirds learn individually unique songs through vocal imitation and use them in courtship and territorial displays. Previous work has identified a forebrain auditory area, the caudomedial nidopallium (NCM), that appears specialized for discriminating and remembering conspecific vocalizations. In zebra finches (ZFs), only males produce learned vocalizations, but both sexes process these and other signals. This study assessed sex differences in auditory processing by recording extracellular multiunit activity at multiple sites within NCM. Juvenile female ZFs (n = 46) were reared in individual isolation and artificially tutored with song. In adulthood, songs were played back to assess auditory responses, stimulus‐specific adaptation, neural bias for conspecific song, and memory for the tutor's song, as well as recently heard songs. In a subset of females (n = 36), estradiol (E2) levels were manipulated to test the contribution of E2, known to be synthesized in the brain, to auditory responses. Untreated females (n = 10) showed significant differences in response magnitude and stimulus‐specific adaptation compared to males reared in the same paradigm (n = 9). In hormone‐manipulated females, E2 augmentation facilitated the memory for recently heard songs in adulthood, but neither E2 augmentation (n = 15) nor E2 synthesis blockade (n = 9) affected tutor song memory or the neural bias for conspecific song. The results demonstrate subtle sex differences in processing communication signals, and show that E2 levels in female songbirds can affect the memory for songs of potential suitors, thus contributing to the process of mate selection. The results also have potential relevance to clinical interventions that manipulate E2 in human patients. © 2014 Wiley Periodicals, Inc. Develop Neurobiol 75: 302–314, 2015 相似文献
55.
Zhuo Li Jon D. Gabbard Scott Johnson Daniel Dlugolenski Shannon Phan S. Mark Tompkins Biao He 《PloS one》2015,10(3)
H7N9 has caused fatal infections in humans. A safe and effective vaccine is the best way to prevent large-scale outbreaks in the human population. Parainfluenza virus 5 (PIV5), an avirulent paramyxovirus, is a promising vaccine vector. In this work, we generated a recombinant PIV5 expressing the HA gene of H7N9 (PIV5-H7) and tested its efficacy against infection with influenza virus A/Anhui/1/2013 (H7N9) in mice and guinea pigs. PIV5-H7 protected the mice against lethal H7N9 challenge. Interestingly, the protection did not require antibody since PIV5-H7 protected JhD mice that do not produce antibody against lethal H7N9 challenge. Furthermore, transfer of anti-H7 serum did not protect mice against H7N9 challenge. PIV5-H7 generated high HAI titers in guinea pigs, however it did not protect against H7N9 infection or transmission. Intriguingly, immunization of guinea pigs with PIV5-H7 and PIV5 expressing NP of influenza A virus H5N1 (PIV5-NP) conferred protection against H7N9 infection and transmission. Thus, we have obtained a H7N9 vaccine that protected both mice and guinea pigs against lethal H7N9 challenge and infection respectively. 相似文献
56.
Knutsen AK Chang YV Grimm CM Phan L Taber LA Bayly PV 《Journal of biomechanical engineering》2010,132(10):101004
Folding of the cerebral cortex is a critical phase of brain development in higher mammals but the biomechanics of folding remain incompletely understood. During folding, the growth of the cortical surface is heterogeneous and anisotropic. We developed and applied a new technique to measure spatial and directional variations in surface growth from longitudinal magnetic resonance imaging (MRI) studies of a single animal or human subject. MRI provides high resolution 3D image volumes of the brain at different stages of development. Surface representations of the cerebral cortex are obtained by segmentation of these volumes. Estimation of local surface growth between two times requires establishment of a point-to-point correspondence ("registration") between surfaces measured at those times. Here we present a novel approach for the registration of two surfaces in which an energy function is minimized by solving a partial differential equation on a spherical surface. The energy function includes a strain-energy term due to distortion and an "error energy" term due to mismatch between surface features. This algorithm, implemented with the finite element method, brings surface features into approximate alignment while minimizing deformation in regions without explicit matching criteria. The method was validated by application to three simulated test cases and applied to characterize growth of the ferret cortex during folding. Cortical surfaces were created from MRI data acquired in vivo at 14 days, 21 days, and 28 days of life. Deformation gradient and Lagrangian strain tensors describe the kinematics of growth over this interval. These quantitative results illuminate the spatial, temporal, and directional patterns of growth during cortical folding. 相似文献
57.
Forthal DN Gilbert PB Landucci G Phan T 《Journal of immunology (Baltimore, Md. : 1950)》2007,178(10):6596-6603
Nonneutralizing Abs may play a role in protecting animals and humans from lentiviral infections. We explored the Ab-dependent, cell-mediated virus inhibition (ADCVI) Ab response to recombinant gp120 (rgp120) vaccination in sera from 530 participants in the Vax 004 trial. Serum ADCVI activity was measured against a clinical R5 strain of HIV-1 using peripheral blood mononuclear effector cells from healthy donors. The level of vaccine-induced ADCVI activity correlated inversely with the rate of acquiring HIV infection following vaccination, such that for every 10% increase in ADCVI activity, there was a 6.3% decrease in the hazard rate of infection (p=0.019). Some vaccinated individuals also mounted an ADCVI response against two other clinical R5 strains of HIV-1. However, ADCVI activity correlated poorly with neutralizing or CD4-gp120-blocking Ab activity measured against laboratory strains. Finally, the degree to which the ADCVI Ab response predicted the rate of infection was influenced by polymorphisms at the FcgammaRIIa and FcgammaRIIIa gene loci. These data indicate that rgp120 vaccination can elicit Abs with antiviral activity against clinical strains of HIV-1. However, such activity requires the presence of FcR-bearing effector cells. Our results provide further evidence that ADCVI may play a role in preventing HIV infection. 相似文献
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59.
We describe the construction of six novel plasmid-based IPTG-inducible expression vectors for Bacillus subtilis and related species. While one vector allows intracellular production of recombinant proteins, the second provides a strong
secretion signal. The third vector allows addition of the c-Myc epitope tag, and the remaining three vectors provide the purification tags His and Strep. The versatility of all six
vectors was demonstrated by the insertion of several reporter genes and by their regulated overexpression. Recombinant proteins
with a His- or Strep-tag could be purified to near homogeneity in a single step. 相似文献
60.
Primers that contain portions noncomplementary to the target region are usually used to add to the PCR product a utility sequence such as a restriction site or a universal probe binding site. We have demonstrated that primers with short 5'AT-rich overhangs increase real-time PCR fluorescent signal. The improvement is particularly significant for difficult to amplify templates, such as highly variable viral sequences or bisulfite-treated DNA. 相似文献