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Infrared-visible sum frequency generation (SFG) vibrational spectroscopy, in combination with fluorescence microscopy, was employed to investigate the surface structure of lysozyme, fibrinogen, and bovine serum albumin (BSA) adsorbed on hydrophilic silica and hydrophobic polystyrene as a function of protein concentration. Fluorescence microscopy shows that the relative amounts of protein adsorbed on hydrophilic and hydrophobic surfaces increase in proportion with the concentration of protein solutions. For a given bulk protein concentration, a larger amount of protein is adsorbed on hydrophobic polystyrene surfaces compared to hydrophilic silica surfaces. While lysozyme molecules adsorbed on silica surfaces yield relatively similar SFG spectra, regardless of the surface concentration, SFG spectra of fibrinogen and BSA adsorbed on silica surfaces exhibit concentration-dependent signal intensities and peak shapes. Quantitative SFG data analysis reveals that methyl groups in lysozyme adsorbed on hydrophilic surfaces show a concentration-independent orientation. However, methyl groups in BSA and fibrinogen become less tilted with respect to the surface normal with increasing protein concentration at the surface. On hydrophobic polystyrene surfaces, all proteins yield similar SFG spectra, which are different from those on hydrophilic surfaces. Although more protein molecules are present on hydrophobic surfaces, lower SFG signal intensity is observed, indicating that methyl groups in adsorbed proteins are more randomly oriented as compared to those on hydrophilic surfaces. SFG data also shows that the orientation and ordering of phenyl rings in the polystyrene surface is affected by protein adsorption, depending on the amount and type of proteins.
An entry from the Cambridge Structural Database, the world’s repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
We investigated the antigenic requirements for restimulation of H-2- restricted cytolytic T lymphocytes (CTL) in vitro to determine whether H-2 I region-restricted helper T cells are required in these responses. In one set of experiments, we studied the in vitro response of (responder x nonresponder)F(1) female T cells to the male antigen H-Y. We chose to examine this response because it has been suggested that the defect in nonresponder strains is a failure of helper T cells to recognize H-Y in association with nonresponder I region determinants. However, we find that nonresponder male stimulator cells are as effective as F(1) male stimulator cells at inducing H-Y-specific CTL responses. This finding calls into question reports that secondary CTL responses to H-Y are dependent upon the activation of H-Y- specific helper T cells restricted to responder type I region determinants. In a second set of experiments, we examined the requirements for restimulation of H-2-restricted T cells specific for minor-histocompatibility antigens from long-term mixed lymphocyte cultures. These cultures were established by repeatedly restimulating cultures of specific T cells with H- 2-matched stimulator cells expressing foreign minor histocompatibility antigens. We found that H-2D-restricted T ceils, including CTL, could be restimulated with cells that were matched with the responding cells at only the D region genes. This response did not appear to result from positive allogeneic effects or from antigen processing and "representation" by responder type APC that might contaminate the cultures. Thus, we find no evidence for a requirement for I region-restricted helper T cells in these CTL responses. However, helper T cells are required because we find that CTL lines derived by limit-dilution cloning from these long-term MLC are absolutely dependent upon exogenous helper factors for growth. The most simple interpretation of these results is that the helper cells are restricted to H-2 antigens other than I region antigens or to antigens that code outside of the H-2 complex. Finally, we show that factor-dependent CTL lines must recognize their specific antigen to proliferate, even in the presence of exogenous factors. The requirement of activated CTL for antigen to proliferate provides an explanation for how specific CTL can be selectively enriched in MLC by specific antigen stimulation. Furthermore, it is at variance with reports that memory CTL or activated CTL require only interleukin 2 for restimulation.
Isotopic techniques are required for answering many fundamental pathophysiologic questions in diabetes research. Several isolopic methods and results are critically evaluated here, along with a description of recent methodologic advances. Included is the role of hepatic glucose production (HGP) in fasting hyperglycemia, the metabolic source of HGP (gluconeogenesis vs glycogenolysis), whether HGP is substrate-driven, and the tissue location and metabolic fate of glucose disposal. Progress in the measurement of HGP, gluconeogenesis, hepatic substrate fluxes, and hepatic uridine diphosphate-glucose metabolism is described. The uses of isotope-kinetic methods in identifying sites of drug action and genes contributing to non-insulin-dependent diabetes mellitus are also noted.
High molecular weight (MW) polymers have shown promise in terms of improving the properties and the efficacy of low MW therapeutics. However, new systems that are highly biocompatible, are biodegradable, have well-defined MW, and have multiple functional groups for drug attachment are still needed. The biological evaluation of a library of eight polyester dendrimer-poly(ethylene oxide) (PEO) bow-tie hybrids is described here. The group of evaluated polymers was designed to include a range of MWs (from 20000 to 160000) and architectures with the number of PEO arms ranging from two to eight. In vitro experiments revealed that the polymers were nontoxic to cells and were degraded to lower MW species at pH 7.4 and pH 5.0. Biodistribution studies with (125)I-radiolabeled polymers showed that the high MW carriers (>40000) exhibited long circulation half-lives. Comparison of the renal clearances for the four-arm versus eight-arm polymers indicated that the more branched polymers were excreted more slowly into the urine, a result attributed to their decreased flexibility. Due to their essentially linear architecture that does not provide for good isolation of the iodinated phenolic moieties, the polymers with "two arms" were rapidly taken up by the liver. The biodistributions of two long-circulating high MW polymers in mice bearing subcutaneous B16F10 tumors were evaluated, and high levels of tumor accumulation were observed. These new carriers are therefore promising for applications in drug delivery and are also useful for improving our understanding of the effect of polymer architecture on pharmacokinetic properties.
Characterization of the functional organization of the photochemical apparatus in the light sensitive chlorophyll b-deficient oil yellow-yellow green (OY-YG) mutant of maize (Zea mays) is presented. Spectrophotometric and kinetic analysis revealed substantially lower amounts of the light harvesting complex of photosystem II (LHCII-peripheral) in high light-grown OY-YG thylakoids. However, accumulation of a tightly bound LHCII appears unaffected by the lesion. Changes in photosystem (PS) stoichiometry include lower amounts of PSII with characteristic fast kinetics (PSII(alpha)) and a substantial accumulation of PSII centers with characteristic slow kinetics (PSII(beta)) in the thylakoid membrane of the OY-YG mutant. Thus, PSII(beta) is the dominant photosystem in the mutant chloroplasts. In contrast to wild type, roughly 80% of the mutant PSII(beta) centers are functionally coupled to the plastoquinone pool and are probably localized in the appressed regions of the thylakoid membrane. These centers, designated PSII(beta)-Q(B)-reducing (Q(B) being the secondary electron quinone acceptor of PSII), are clearly distinct from the typical PSII(beta)-Q(B)-nonreducing centers found in the stroma lamellae of wild-type chloroplasts. It is concluded that the observed changes in the stoichiometry of electron-transport complexes reflect the existence of a regulatory mechanism for the adjustment of photosystem stoichiometry in chloroplasts designed to correct any imbalance in light absorption by the two photosystems.
Abstract Assembly of TCRα and TCRδ genes from the TCRα/δ locus is tightly controlled for the proper generation of αβ and γδ T cells. Of >100 shared variable gene segments in the TCRα/δ locus, only a few are predominantly used for the TCRδ gene assembly, while most are for TCRα. However, the importance and mechanisms of the selective variable gene rearrangement for T cell development are not fully understood. We report herein that the development of a tissue-specific γδ T cell population is critically affected by recombination signal sequence-associated restriction on the variable gene usage for TCRδ assembly. We found that the development of substitute skin γδ T cells in mice deficient of the TCRγ3 gene, which is used in wild-type skin γδ T cells, was drastically affected by the strain background. A Vγ2+ skin γδ T cell population developed in mice of the B6 but not the 129 strain backgrounds, due to a difference in the rearrangement of endogenous Vδ7+ TCRδ genes, which paired with the Vγ2+ TCRγ gene to generate the Vγ2/Vδ7+ skin γδ T cell precursors in fetal thymi of the B6 background mice. The defective TCRδ rearrangement of the 129-“Vδ7” gene was associated with specific variations in its recombination signal sequence, which renders it poorly compatible for rearrangement to Dδ genes. These findings provide the first direct evidence that recombination signal sequence-associated restriction on the variable gene usage for TCRα/δ gene assembly plays an important role in T cell development.
Article Free Access Share on PRoP: personal roving presence Authors: Eric Paulos Department of Electrical Engineering and Computer Sciences, University of California, Berkeley, Berkeley, CA Department of Electrical Engineering and Computer Sciences, University of California, Berkeley, Berkeley, CAView Profile , John Canny Department of Electrical Engineering and Computer Sciences, University of California, Berkeley, Berkeley, CA Department of Electrical Engineering and Computer Sciences, University of California, Berkeley, Berkeley, CAView Profile Authors Info & Claims CHI '98: Proceedings of the SIGCHI Conference on Human Factors in Computing SystemsJanuary 1998 Pages 296–303https://doi.org/10.1145/274644.274686Published:01 January 1998Publication History 94citation765DownloadsMetricsTotal Citations94Total Downloads765Last 12 Months92Last 6 weeks25 Get Citation AlertsNew Citation Alert added!This alert has been successfully added and will be sent to:You will be notified whenever a record that you have chosen has been cited.To manage your alert preferences, click on the button below.Manage my AlertsNew Citation Alert!Please log in to your account Save to BinderSave to BinderCreate a New BinderNameCancelCreateExport CitationPublisher SiteeReaderPDF
