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Search Results: 1 - 10 of 208730 matches for " Dustin L. Higashi "
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N. elongata Produces Type IV Pili That Mediate Interspecies Gene Transfer with N. gonorrhoeae
Dustin L. Higashi, Nicolas Biais, Nathan J. Weyand, Al Agellon, Jennifer L. Sisko, Lewis M. Brown, Magdalene So
PLOS ONE , 2011, DOI: 10.1371/journal.pone.0021373
Abstract: The genus Neisseria contains at least eight commensal and two pathogenic species. According to the Neisseria phylogenetic tree, commensals are basal to the pathogens. N. elongata, which is at the opposite end of the tree from N. gonorrhoeae, has been observed to be fimbriated, and these fimbriae are correlated with genetic competence in this organism. We tested the hypothesis that the fimbriae of N. elongata are Type IV pili (Tfp), and that Tfp functions in genetic competence. We provide evidence that the N. elongata fimbriae are indeed Tfp. Tfp, as well as the DNA Uptake Sequence (DUS), greatly enhance N. elongata DNA transformation. Tfp allows N. elongata to make intimate contact with N. gonorrhoeae and to mediate the transfer of antibiotic resistance markers between these two species. We conclude that Tfp functional for genetic competence is a trait of a commensal member of the Neisseria genus. Our findings provide a mechanism for the horizontal gene transfer that has been observed among Neisseria species.
Genome Sequencing Reveals Widespread Virulence Gene Exchange among Human Neisseria Species
Pradeep Reddy Marri,Mary Paniscus,Nathan J. Weyand,María A. Rendón,Christine M. Calton,Diana R. Hernández,Dustin L. Higashi,Erica Sodergren,George M. Weinstock,Steven D. Rounsley,Magdalene So
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0011835
Abstract: Commensal bacteria comprise a large part of the microbial world, playing important roles in human development, health and disease. However, little is known about the genomic content of commensals or how related they are to their pathogenic counterparts. The genus Neisseria, containing both commensal and pathogenic species, provides an excellent opportunity to study these issues. We undertook a comprehensive sequencing and analysis of human commensal and pathogenic Neisseria genomes. Commensals have an extensive repertoire of virulence alleles, a large fraction of which has been exchanged among Neisseria species. Commensals also have the genetic capacity to donate DNA to, and take up DNA from, other Neisseria. Our findings strongly suggest that commensal Neisseria serve as reservoirs of virulence alleles, and that they engage extensively in genetic exchange.
The 5 Hallmarks of Biomaterials Success: An Emphasis on Orthopaedics  [PDF]
Dustin L. Williams, Brad M. Isaacson
Advances in Bioscience and Biotechnology (ABB) , 2014, DOI: 10.4236/abb.2014.54035
Abstract:

Over the past 200 years, there has been significant advancements in the fields of bioengineering and orthopaedics. Investigators, clinicians and manufactures are learning that the success of implant systems is not limited to a single factor, but a combination of variables that must work in unison to provide stability and high survivorship. Innovations continue to advance these fields and include: biomimetic alterations, three-dimensional, patient-specific reconstructions and novel coatings to mitigate aseptic loosening or other pathologies. However, implant systems continue to fail in clinical practice since they do not adhere to key fundamental principles. Therefore, this article is intended to highlight 5 hallmarks of biomaterials that should be considered during design, surgery, and post-operative rehabilitation.

In vivo imaging approaches in animal models of rheumatoid arthritis
Michael L Dustin
Arthritis Research & Therapy , 2003, DOI: 10.1186/ar768
Abstract: The early phase of exploration of the lymphoid system generated a wealth of information about anatomy and in vivo responses. Our ability to define molecular structures in the context of the anatomy of the in vivo immune response, first with antibodies and more recently with tools of molecular genetics, has increased the ability to incisively test hypotheses through in vivo experimentation. This is leading to a renaissance in a variety of in vivo studies, mostly focused around genetic manipulations. The molecular genetics tools are also complemented by new technologies to image the movements and interactions of cells in vivo.The present review will focus on emerging technologies that allow in vivo imaging of specific cells or molecules using noninvasive methods or direct microscopic imaging of single cells in the in vivo environment using minimally invasive methods. Microscopic imaging has the advantage of being able to study single cells in action. Invasiveness in this case refers specifically to the need for surgical procedures to expose tissues for high-resolution imaging of cells or molecules of interest. The advantages and limitations of each approach are discussed with a specific emphasis on imaging in joints and on work directly relevant to rheumatoid arthritis. This information is summarized in Table 1.Imaging of events in intact live animals is a powerful approach primarily because it allows studies over time with minimal perturbation of the experiment. These methods also couple in powerful ways with molecule genetics technologies that allow in situ labeling of cell populations expressing specific genes. The present review will also discuss recent studies in this area with direct relevance to animal models of rheumatoid arthritis.The expression of luciferase has for many years been a powerful tool in gene expression studies. This is because the substrates in the luciferase reaction generate no signal (light) in the absence of luciferase. Instruments that det
The immunological synapse
Michael L Dustin
Arthritis Research & Therapy , 2002, DOI: 10.1186/ar559
Abstract: The immunological synapse (IS) is a specialized cell–cell junction between a thymus-derived lymphocyte (T cell) and an antigen-presenting cell (APC) [1,2]. Initiation of an antigen-specific immune response is based on the interaction between T-cell receptors (TCRs) and major histocompatibility complex proteins that have bound antigenic peptides (MHCps) [3,4]. Because the TCRs and MHCps are attached to the surface of the T cell and the APC, respectively, the initiation of an immune response requires a molecular grasp between the T cell and the APC – a synapse. A current focus of research on the IS is to determine how this supramolecular structure contributes to T-cell sensitivity and to the fidelity of the T-cell response. Four areas in which the concept of the IS is contributing to our understanding of T-cell activation are the coordination of antigen recognition and T-cell migration; the role of the cytoskeleton in T-cell activation; the mechanism of sensitive antigen recognition by T cells; and the integration of the adaptive and innate immune responses.The formation of the IS has been followed over time in live T cells interacting with planar bilayers [2] and studied at specific time points in fixed cell–cell conjugates [5]. The T cell forms an adhesion zone with the antigen-presenting bilayer; this zone is then surrounded by areas of close contact where TCR can reach the MHCp. If the TCR engagement exceeds a threshold rate and level, the T cell stops migrating and forms a ring of engaged TCRs at the periphery of the nascent IS (Fig. 1a). This pattern takes ~30 seconds to form and corresponds to the peak of TCR-associated tyrosine phosphorylation and Ca2+ mobilization. Within a few more seconds, the sites of TCR engagement move from the periphery of the contact area to the center of the contact area to form the mature IS (Fig. 1b). During this time, the disk-like region of LFA-1–ICAM-1 (intercellular adhesion molecule-1) interaction appears to give way to the cen
Development of Simultaneous HPLC-Fluorescence Assay of Phenol and Chlorophenols in Tap Water after Pre-Column Derivatization with 3-Chlorocarbonyl-6,7-dimethoxy-1- methyl-2(1H)-quinoxalinone  [PDF]
Yasuhiko Higashi
Detection (Detection) , 2016, DOI: 10.4236/detection.2016.41003
Abstract: Chlorophenols (2-chlorophenol, 4-chlorophenol, 2,4-dichlorophenol, 2,6-dichlorophenol and 2,4, 6-trichlorophenol) may be presented in natural waters or drinking water as a result of disinfection processes involving chlorination, or as contaminants derived from domestic products, industrial operations and agricultural chemicals. A previous HPLC-UV method for determination of phenol and five chlorophenols in tap water using 4-fluoro-7-nitro-2,1,3-benzoxadiaole as a UV labeling reagent shows limited sensitivity. Here, we present an improved HPLC-fluorescence detection method for simultaneous determination of phenol and the above chlorophenols in tap water after pre-column derivatization with 3-chlorocarbonyl-6,7-dimethoxy-1-methyl-2(1H)-quino- xalinone (DMEQ-COCl), using a short, narrow column (50 × 2.1 mm i.d., packed with 5 μm particles of C18 material) to improve the sensitivity. Standard samples containing the compounds are derivatized with DMEQ-COCl in borate buffer (pH 9.0) at room temperature for 3 mins. The response is linear in the concentration range of 0.01 - 0.05 to 0.5 mg/L with r2 values ≥0.9967 for all compounds. The lower limits of detection are 0.001 to 0.008 mg/L, and the coefficients of variation are less than 8.8%. The recovery values from tap water spiked with standard samples are satisfactory. The present method is suitable for examining whether or not tap water samples are contaminated with phenol and chlorophenols in excess of regulatory values.
Simple HPLC-Fluorescence Determination of Raspberry Ketone in Fragrance Mist after Pre-Column Derivatization with 4-Hydrazino-7-nitro-2,1,3-benzoxadiazole  [PDF]
Yasuhiko Higashi
Journal of Analytical Sciences, Methods and Instrumentation (JASMI) , 2016, DOI: 10.4236/jasmi.2016.62006
Abstract: Raspberry ketone {RK, 4-(4-hydroxyphenyl)butan-2-one} is a natural compound contained in raspberry, and is added to cosmetics for skin whitening. It is very important to measure the RK level in cosmetics for quality assessment, since RK structurally resembles 4-(4-hydroxyphenyl)-2-butanol, which causes leukoderma on consumers’ skin. Here, we present a simple HPLC-fluorescence method for determination of RK in a fragrance mist by pre-column derivatization with 4-hydrazino-7-nitro-2,1,3-benzoxadiazole hydrazine (NBD-H), which reacts with the carbonyl group of RK. The NBD-RK derivative was eluted from a reversed-phase ODS column, and detected with excitation at 470 nm and emission at 550 nm. The retention time of NBD-RK derivative obtained by reaction with NBD-H at 80°C for 20 min was 10.3 min. The standard curve was linear in the range of 0.2 to 10 μg/mL, with a correlation coefficient (r2) value of 0.9980. The lower limit of detection was 0.018 μg/mL (absolute amount of 1.8 pmol). The coefficients of variation were less than 8.1%. The content of RK in fragrance mist (1.00 mL) was 1.18 ± 0.07 mg (range: 1.12 to 1.28 mg, n = 5). Recovery tests were satisfactory (83.9% ± 3.9%; range: 79.6 to 88.8%, n = 5).
Improved Method for Determination of Raspberry Ketone in Fragrance Mist by HPLC-Fluorescence Analysis after Pre-Column Derivatization with 4-(N,N-Dimethylaminosulfonyl)-7-(N-chloroformylmethyl-N-methylamino)-2,1,3-benzoxadiazole  [PDF]
Yasuhiko Higashi
Journal of Analytical Sciences, Methods and Instrumentation (JASMI) , 2018, DOI: 10.4236/jasmi.2018.82002
Abstract:

Raspberry ketone {RK, 4-(4-hydroxyphenyl)butan-2-one} is structurally resembles 4-(4-hydroxyphenyl)-2-butanol, which causes leukoderma on consumers’ skin. Therefore, it is important to measure in cosmetics for quality assessment. Very recently, an HPLC-fluorescence method for determination of RK in a fragrance mist by pre-column derivatization with 4-hydrazino-7-nitro-2,1,3-benzoxadiazole hydrazine was established. However, the derivatization conditions (80°C, 20 min) were severe. In this study, an improved pre-column derivatization with 4-(N,N-dimethylaminosulfonyl)-7-(N-chloro-formylmethyl-N-methylamino)-2,1,3-benzoxadiazole (DBD-COCl) is presented by HPLC-fluorescence method for determination of RK. The DBD-CO-RK derivative was eluted from a reversed-phase ODS column, and detected with excitation at 440 nm and emission at 543 nm. Derivatization was performed at room temperature for 3 min. The retention time of DBD-CO-RK derivative was 16.8 min. The standard curve was linear in the range of 0.05 to 2.5 μg/mL, with a correlation coefficient (r2) value of 0.9988. The lower limit of detection was 0.01 μg/mL (absolute amount of 0.3 pmol). The coefficients of variation were less than 10.0%. The content of RK in fragrance mist (1.00 mL) was 1.20 ± 0.08 mg (range, 1.10 to 1.31 mg,

Cooperative Retraction of Bundled Type IV Pili Enables Nanonewton Force Generation
Nicolas Biais,Beno?t Ladoux,Dustin Higashi,Magdalene So,Michael Sheetz
PLOS Biology , 2012, DOI: 10.1371/journal.pbio.0060087
Abstract: The causative agent of gonorrhea, Neisseria gonorrhoeae, bears retractable filamentous appendages called type IV pili (Tfp). Tfp are used by many pathogenic and nonpathogenic bacteria to carry out a number of vital functions, including DNA uptake, twitching motility (crawling over surfaces), and attachment to host cells. In N. gonorrhoeae, Tfp binding to epithelial cells and the mechanical forces associated with this binding stimulate signaling cascades and gene expression that enhance infection. Retraction of a single Tfp filament generates forces of 50–100 piconewtons, but nothing is known, thus far, on the retraction force ability of multiple Tfp filaments, even though each bacterium expresses multiple Tfp and multiple bacteria interact during infection. We designed a micropillar assay system to measure Tfp retraction forces. This system consists of an array of force sensors made of elastic pillars that allow quantification of retraction forces from adherent N. gonorrhoeae bacteria. Electron microscopy and fluorescence microscopy were used in combination with this novel assay to assess the structures of Tfp. We show that Tfp can form bundles, which contain up to 8–10 Tfp filaments, that act as coordinated retractable units with forces up to 10 times greater than single filament retraction forces. Furthermore, single filament retraction forces are transient, whereas bundled filaments produce retraction forces that can be sustained. Alterations of noncovalent protein–protein interactions between Tfp can inhibit both bundle formation and high-amplitude retraction forces. Retraction forces build over time through the recruitment and bundling of multiple Tfp that pull cooperatively to generate forces in the nanonewton range. We propose that Tfp retraction can be synchronized through bundling, that Tfp bundle retraction can generate forces in the nanonewton range in vivo, and that such high forces could affect infection.
Cooperative Retraction of Bundled Type IV Pili Enables Nanonewton Force Generation
Nicolas Biais,Beno?t Ladoux,Dustin Higashi,Magdalene So,Michael Sheetz
PLOS Biology , 2008, DOI: 10.1371/journal.pbio.0060087
Abstract: The causative agent of gonorrhea, Neisseria gonorrhoeae, bears retractable filamentous appendages called type IV pili (Tfp). Tfp are used by many pathogenic and nonpathogenic bacteria to carry out a number of vital functions, including DNA uptake, twitching motility (crawling over surfaces), and attachment to host cells. In N. gonorrhoeae, Tfp binding to epithelial cells and the mechanical forces associated with this binding stimulate signaling cascades and gene expression that enhance infection. Retraction of a single Tfp filament generates forces of 50–100 piconewtons, but nothing is known, thus far, on the retraction force ability of multiple Tfp filaments, even though each bacterium expresses multiple Tfp and multiple bacteria interact during infection. We designed a micropillar assay system to measure Tfp retraction forces. This system consists of an array of force sensors made of elastic pillars that allow quantification of retraction forces from adherent N. gonorrhoeae bacteria. Electron microscopy and fluorescence microscopy were used in combination with this novel assay to assess the structures of Tfp. We show that Tfp can form bundles, which contain up to 8–10 Tfp filaments, that act as coordinated retractable units with forces up to 10 times greater than single filament retraction forces. Furthermore, single filament retraction forces are transient, whereas bundled filaments produce retraction forces that can be sustained. Alterations of noncovalent protein–protein interactions between Tfp can inhibit both bundle formation and high-amplitude retraction forces. Retraction forces build over time through the recruitment and bundling of multiple Tfp that pull cooperatively to generate forces in the nanonewton range. We propose that Tfp retraction can be synchronized through bundling, that Tfp bundle retraction can generate forces in the nanonewton range in vivo, and that such high forces could affect infection.
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