Measuring perceived differences in surface texture due to changes in higher order statistics.

We investigate the ability of humans to perceive changes in the appearance of images of surface texture caused by the variation of their higher order statistics. We incrementally randomize their phase spectra while holding their first and second order statistics constant in order to ensure that the change in the appearance is due solely to changes in third and other higher order statistics. Stimuli comprise both natural and synthetically generated naturalistic images, with the latter being used to prevent observers from making pixel-wise comparisons. A difference scaling method is used to derive the perceptual scales for each observer, which show a sigmoidal relationship with the degree of randomization. Observers were maximally sensitive to changes within the 20%-60% randomization range. In order to account for this behavior we propose a biologically plausible model that computes the variance of local measurements of phase congruency.

Testing limits on matte surface color perception in three-dimensional scenes with complex light fields.

We investigated limits on the human visual system's ability to discount directional variation in complex lights field when estimating Lambertian surface color. Directional variation in the light field was represented in the frequency domain using spherical harmonics. The bidirectional reflectance distribution function of a Lambertian surface acts as a low-pass filter on directional variation in the light field. Consequently, the visual system needs to discount only the low-pass component of the incident light corresponding to the first nine terms of a spherical harmonics expansion [Basri, R., Jacobs, D. (2001). Lambertian reflectance and linear subspaces. In: International Conference on Computer Vision II, pp. 383-390; Ramamoorthi, R., Hanrahan, P., (2001). An efficient representation for irradiance environment maps. SIGGRAPH 01. New York: ACM Press, pp. 497-500] to accurately estimate surface color. We test experimentally whether the visual system discounts directional variation in the light field up to this physical limit. Our results are consistent with the claim that the visual system can compensate for all of the complexity in the light field that affects the appearance of Lambertian surfaces.

Past trials influence perception of ambiguous motion quartets through pattern completion.

There are many celebrated examples of ambiguous perceptual configurations such as the Necker cube that abruptly and repeatedly "switch" among possible perceptual states. When such ambiguous configurations are presented intermittently, observers tend to see the same perceptual state on successive trials. The outcome of each trial apparently serves to "prime" the outcome of the following. We sought to determine how long the influence of a past trial persists by using ambiguous motion quartets as stimuli. We found large, significant effects of all four most recent trials, but the results were not consistent with any priming model. The results could be explained instead as perceptual completion of two kinds of temporal patterns, repeating and alternating. We conclude that the visual system does not passively remember perceptual state: it analyzes recent perceptual history and attempts to predict what will come next. These predictions can alter what is seen.

The effect of perceived surface orientation on perceived surface albedo in binocularly viewed scenes.

We examined how observers discount perceived surface orientation in estimating perceived albedo (lightness). Observers viewed complex rendered scenes binocularly. The orientation of a test patch was defined by depth cues of binocular disparity and linear perspective. On each trial, observers first estimated the orientation of the test patch in the scene by means of a gradient probe and then matched its perceived albedo to a reference scale. We found that observers' perception of orientation was nearly veridical and that they substantially discounted perceived orientation in estimating perceived albedo.

Characterization of nuclease-resistant ribozymes directed against hepatitis B virus RNA.

Hepatitis B virus (HBV) is responsible for > 350 million cases of chronic hepatitis B worldwide and 1.2 million deaths each year. To explore the use of ribozymes as a novel therapy for HBV infection, nuclease-resistant ribozymes that target highly conserved regions of HBV RNA were screened in cell culture. These synthetic ribozymes have the potential to cleave all four major HBV RNA transcripts and to block the HBV lifecycle by cleavage of the pregenomic RNA. A number of the screened ribozymes demonstrate activity in cell culture systems, as measured by decreased levels of HBV surface antigen, HBV e antigen and HBV DNA. In addition, a lead anti-HBV ribozyme maintains activity against a lamivudine-resistant HBV variant in cell culture. Treatment of HBV transgenic mice with lead anti-HBV ribozymes significantly reduced viraemia compared with saline-treated animals and was as effective as treatment with lamivudine. In conclusion, the therapeutic use of a ribozyme alone or in combination with current therapies (lamivudine or interferons) may lead to improved HBV therapy.

Illuminant cues in surface color perception: tests of three candidate cues.

Many recent computational models of surface color perception presuppose information about illumination in scenes. The models differ primarily in the physical process each makes use of as a cue to the illuminant. We evaluated whether the human visual system makes use of any of three of the following candidate illuminant cues: (1) specular highlight, (2) full surface specularity [Lee, H. C. (1986). Method for computing the scene-illuminant chromaticity from specular highlights. Journal of the Optical Society of America A, 3(10), 1694-1699; D'Zmura, M., & Lennie, P. (1986). Mechanisms of color constancy. Journal of the Optical Society of America A, 3(10), 1662-1672], and (3) uniform background. Observers viewed simulated scenes binocularly in a computer-controlled Wheatstone stereoscope. All simulated scenes contained a uniform background plane perpendicular to the observer's line of sight and a small number of specular, colored spheres resting on the uniform background. Scenes were rendered under either standard illuminant D65 or standard illuminant A. Observers adjusted the color of a small, simulated test patch to appear achromatic. In a series of experiments we perturbed the illuminant color signaled by each candidate cue and looked for an influence of the changed cue on achromatic settings. We found that the specular highlight cue had a significant influence, but that the influence was asymmetric: greater when the base illuminant, CIE standard Illuminant A, was perturbed in the direction of Illuminant D65 than vice versa. Neither the full surface specularity cue nor the background cue had any observable influence. The lack of influence of the background cue is likely due to the placement of the test patch in front of the background rather than, as is typical, embedded in the background.

Quantitative determination of a chemically modified hammerhead ribozyme in blood plasma using 96-well solid-phase extraction coupled with high-performance liquid chromatography or capillary gel electrophoresis.

Versatile bioanalytical assays to detect chemically stabilized hammerhead ribozyme and putative ribozyme metabolites from plasma are described. The extraction protocols presented are based on serial solid-phase extractions performed on a 96-well plate format and are compatible with either IEX-HPLC or CGE back-end analysis. A validation of both assays confirmed that both the HPLC and the CGE methods possess the required linearity, accuracy, and precision to accurately measure concentrations of hammerhead ribozyme extracted from plasma. These methods should be of general use to detect and quantitate ribozymes from other biological fluids such as serum and urine.

Daylight, biochrome surfaces, and human chromatic response in the Fourier domain.

We first report Fourier analyses of a collection of 348 daylight spectral power distributions and 1,695 biochrome surface reflectance functions. The power spectra of the daylights are low pass with more than 99% of spectral power below 1 cycle/300 nm and 99.9% below 3 cycles/300 nm. The power spectra of reflectance functions are also low pass with more than 99% of spectral power below 4 cycles/300 nm and 99.9% below 11 cycles/300 nm. Consequently, the resulting color signals are typically low pass with, for our samples, an estimated frequency cutoff of 5 cycles/300 nm. Theoretical and experimental data concerning human chromatic response in the frequency domain show that this limit corresponds to the highest frequency that the color system can resolve. The implications for normal and abnormal human color vision are discussed.

Bioactivity of anti-angiogenic ribozymes targeting Flt-1 and KDR mRNA.

Vascular endothelial growth factor (VEGF) and its receptors Flt-1 and KDR play important roles in physiological and pathological angiogenesis. Ribozymes that target the VEGF receptor mRNAs were developed and their biological activities in cell culture and an animal model were assessed. Ribozymes targeting Flt-1 or KDR mRNA sites reduced VEGF-induced proliferation of cultured human vascular endothelial cells and specifically lowered the level of Flt-1 or KDR mRNA present in the cells. Anti- Flt-1 and KDR ribozymes also exhibited anti-angiogenic activity in a rat corneal pocket assay of VEGF-induced angiogenesis. This report illustrates the anti-angiogenic potential of these ribozymes as well as their value in studying VEGF receptor function in normal and pathophysiologic states.

A core folding model for catalysis by the hammerhead ribozyme accounts for its extraordinary sensitivity to abasic mutations.

Introducing abasic nucleotides at each of 13 positions in the conserved core of the hammerhead ribozyme causes a large decrease in the extent of catalysis [Peracchi, A., et al. (1996) Proc. Natl. Acad. Sci. U.S.A. 93, 11522]. This extreme sensitivity to structural defects is in contrast to the behavior of protein enzymes and larger ribozymes. Several additional differences in the behavior of the hammerhead relative to that of protein enzymes and larger ribozymes are described herein. The deleterious effects of the abasic mutations are not relieved by lowering the temperature, by increasing the concentration of monovalent or divalent metal ions, or by adding polyamines, in contrast to effects observed with protein enzymes and large RNA enzymes. In addition, the abasic mutations do not significantly weaken substrate binding. These results and previous observations are all accounted for by a "core folding" model in which the stable ground state structure of the hammerhead ribozyme complexed with the substrate is a partially folded state that must undergo an additional folding event to achieve its catalytic conformation. We propose that the peculiar behavior of the hammerhead arises because the limited structural interconnections in a small RNA enzyme do not allow the ground state to stably adopt the catalytic conformation; within the globally folded catalytic conformation, limited structural interconnections may further impair catalysis by hampering the precise alignment of active site functional groups. This behavior represents a basic manifestation of the well-recognized interconnection between folding and catalysis.

Structural variation induced by different nucleotides at the cleavage site of the hammerhead ribozyme.

The hammerhead ribozyme is capable of cleaving RNA substrates at 5' UX 3' sequences (where the cleavage site, X, can be A, C, or U). Hammerhead complexes containing dC, dA, dI, or rG nucleotides at the cleavage site have been studied by NMR. The rG at the cleavage site forms a Watson-Crick base pair with C3 in the conserved core of the hammerhead, indicating that rG substrates inhibit the cleavage reaction by stabilizing an inactive conformation of the molecule. Isotope-edited NMR experiments on the hammerhead complexes show that there are different short proton-proton distances between neighboring residues depending upon whether there is a dC or dA at the cleavage site. These NMR data demonstrate that there are significant differences in the structure and/or dynamics of the active-site residues in these hammerhead complexes. Molecular dynamics calculations were used to model the conformations of the cleavage-site variants consistent with the NMR data. The solution conformations of the hammerhead ribozyme-substrate complexes are compared with the X-ray structure of the hammerhead ribozyme and are used to help understand the thermodynamic and kinetic differences among the cleavage-site variants.

The structural basis of hammerhead ribozyme self-cleavage.

We have captured an 8.7 A conformational change that takes place in the cleavage site of the hammerhead ribozyme during self-cleavage, using X-ray crystallography combined with physical and chemical trapping techniques. This rearrangement brings the hammerhead ribozyme from the ground state into a conformation that is poised to form the transition state geometry required for hammerhead RNA self-cleavage. Use of a 5'-C-methylated ribose adjacent to the cleavage site permits this ordinarily transient conformational change to be kinetically trapped and observed crystallographically after initiating the hammerhead ribozyme reaction in the crystal. Cleavage of the corresponding unmodified hammerhead ribozyme in the crystal under otherwise identical conditions is faster than in solution, indicating that we have indeed trapped a catalytically relevant intermediate form of this RNA enzyme.

Identification of a subpopulation of human renal microvascular endothelial cells with capacity to form capillary-like cord and tube structures.

Endothelial specialization is a prominent feature within distinct capillary beds of organs such as mammalian kidney, yet immunological markers for functionally distinct subpopulations of cultured endothelial cells from tissue sources such as kidney have not been available. We developed a simple and reproducible isolation and culture procedure to recover human renal microvascular endothelial cells (HRMEC) from the cortex of unused donor kidneys. This procedure yields highly purified preparations of cells that display endothelial markers that include Factor VIII antigen, acetyl-LDL receptors, and determinants that bind Ulex europaeus lectin. HRMEC assemble into capillary-like cord and tube structures when plated on the surface of basement membrane-like matrix (BMM) in media containing phorbol myristate acetate. To further define subpopulations of HRMEC, we generated a panel of monoclonal antibodies and screened for those recognizing cell surface determinants. One monoclonal antibody recovery from this screen recognized a cell surface protein expressed on a subpopulation of HRMEC that we have designated PEC-1 (pioneer endothelial cell antigen-1). Cells expressing PEC-1 extended long, interconnecting filopodial processes in response to phorbol myristate acetate and assembled into capillary-like structures when plated on BMM. Anti-PEC-1 immunoprecipitated proteins of 25 and 27 kDa. Magnetic bead separation of PEC-1 (+) cells selected cells that assemble into capillary-like cord and tube structures. The remaining PEC-1 (-) HRMEC population formed matrix adherent patches. In the kidney, the PEC-1 determinant is expressed on a small subpopulation of microvascular glomerular cells and is prominently expressed on the apical membrane of proximal tubule cells. The PEC-1 determinant discriminates among subpopulations of HRMEC, identifying a subpopulation that contributes to assembly of capillary-like structures.

Biophysical characterization of zinc ejection from HIV nucleocapsid protein by anti-HIV 2,2'-dithiobis[benzamides] and benzisothiazolones.

HIV nucleocapsid protein (NCp7) has been suggested as a possible target for 2,2'-dithiobis-[benzamide] and benzisothiazolone agents that inhibit viral replication in infected cells (Rice et al. Science 1995, 270, 1194-1197). The solution behavior of these compounds and the mechanistic events leading to removal of Zn from HIV nucleocapsid protein in vitro has been studied by electrospray ionization mass spectrometry, 500 MHz one- and two-dimensional nuclear magnetic resonance spectroscopy, and circular dichroism spectroscopy. We demonstrate that (1) Zn ejection is accompanied by formation of covalent complexes formed between the 2,2'-dithiobis[benzamide] monomers and Cys residues of Zn-depleted NCp7, (2) the rate of Zn ejection is faster for the C-terminal Zn finger and slower for the N-terminal finger, (3) Zn ejection results in a loss of structural integrity of the NCp7 protein, and (4) there is no appreciable interaction between a nonreactive isostere of the lead 2,2'-dithiobis[benzamide] and NCp7 in buffered aqueous solution. These findings are discussed in terms of the mechanism of action of Zn ejection by aromatic 2,2'-dithiobis[benzamides].

A video-input device that displays high-luminance, high-resolution color images.

We describe the design and performance of a bright, full-color, video-input display monitor that can produce image luminances as high as 50,000 cd/m2 for images 15 x 10 deg in size. We have constructed this device by rearranging the components of a commercially available projection television. Our display device allows an experimenter to evaluate human visual responses to complex spectro-spatio-temporal patterns presented at naturally occurring light levels.

The in vitro ejection of zinc from human immunodeficiency virus (HIV) type 1 nucleocapsid protein by disulfide benzamides with cellular anti-HIV activity.

Several disulfide benzamides have been shown to possess wide-spectrum antiretroviral activity in cell culture at low micromolar to submicromolar concentrations, inhibiting human immunodeficiency virus (HIV) type 1 (HIV-1) clinical and drug-resistant strains along with HIV-2 and simian immunodeficiency virus [Rice, W. G., Supko, J. G., Malspeis, L., Buckheit, R. W., Jr., Clanton, D., Bu, M., Graham, L., Schaeffer, C. A., Turpin, J. A., Domagala, J., Gogliotti, R., Bader, J. P., Halliday, S. M., Coren, L., Sowder, R. C., II, Arthur, L. O. & Henderson, L. E. (1995) Science 270, 1194-1197]. Rice and coworkers have proposed that the compounds act by "attacking" the two zinc fingers of HIV nucleocapsid protein. Shown here is evidence that low micromolar concentrations of the anti-HIV disulfide benzamides eject zinc from HIV nucleocapsid protein (NCp7) in vitro, as monitored by the zinc-specific fluorescent probe N-(6-methoxy-8-quinoyl)-p-toluenesulfonamide (TSQ). Structurally similar disulfide benzamides that do not inhibit HIV-1 in culture do not eject zinc, nor do analogs of the antiviral compounds with the disulfide replaced with a methylene sulfide. The kinetics of NCp7 zinc ejection by disulfide benzamides were found to be nonsaturable and biexponential, with the rate of ejection from the C-terminal zinc finger 7-fold faster than that from the N-terminal. The antiviral compounds were found to inhibit the zinc-dependent binding of NCp7 to HIV psi RNA, as studied by gel-shift assays, and the data correlated well with the zinc ejection data. Anti-HIV disulfide benzamides specifically eject NCp7 zinc and abolish the protein's ability to bind psi RNA in vitro, providing evidence for a possible antiretroviral mechanism of action of these compounds. Congeners of this class are under advanced preclinical evaluation as a potential chemotherapy for acquired immunodeficiency syndrome.

Testing the indeterminacy of linear color mechanisms from color discrimination data.

It have previously been reported that, for some choices of the fixed spatial and temporal characteristics of test stimuli, it was possible to estimate the spectral sensitivities of chromatic mechanisms from chromatic discrimination data alone. If mechanism sensitivities could be reliably estimated for any choice of test stimuli characteristics, the influence of spatial and temporal factors on chromatic discrimination performance could be directly measured. Previous studies, using test stimuli with other spatio-temporal characteristics, have found equi-discrimination contours whose ellipsoidal shapes seem to preclude estimation of mechanisms. Since there is no commonly-accepted method for testing the adequacy of ellipsoidal fits of chromatic equi-discrimination contours, it is possible that alternative psychophysical procedures combined with more powerful statistical tests could detect the pattern of deviations from ellipticality reported previously. In this paper, we described psychophysical tests and statistical analyses that, taken together, provide a more powerful test of the indeterminacy of mechanisms than previous methods. We develop a method based on analysis of residuals for detecting the pattern of deviations from ellipticality. We apply these tests under fixed experimental conditions similar to those in which other researchers have found ellipsoidal equi-discrimination contours. For these conditions, for any of the tests performed, we do not reject the hypothesis that equi-discrimination surfaces are ellipsoidal.

Proximity judgments in color space: tests of a Euclidean color geometry.

We describe two tests of the hypothesis that human judgments of the proximity of colors are consistent with a Euclidean geometry on color matching space. The first test uses proximity judgments to measure the angle between any two intersecting lines in color space. Pairwise estimates of the angles between three lines in a plane were made in order to test the additivity of angles. Three different color proximity tasks were considered. Additivity failed for each of the three proximity tasks. Secondly, we tested a prediction concerning the growth of the variability of judgments of similarity with the distance between the test and reference stimuli. The Euclidean hypothesis was also rejected by this test. The results concerning the growth of variability are consistent with the assumption that observers use a city-block metric when judging the proximity of colored lights.

Measurement and modeling of depth cue combination: in defense of weak fusion.

Various visual cues provide information about depth and shape in a scene. When several of these cues are simultaneously available in a single location in the scene, the visual system attempts to combine them. In this paper, we discuss three key issues relevant to the experimental analysis of depth cue combination in human vision: cue promotion, dynamic weighting of cues, and robustness of cue combination. We review recent psychophysical studies of human depth cue combination in light of these issues. We organize the discussion and review as the development of a model of the depth cue combination process termed modified weak fusion (MWF). We relate the MWF framework to Bayesian theories of cue combination. We argue that the MWF model is consistent with previous experimental results and is a parsimonious summary of these results. While the MWF model is motivated by normative considerations, it is primarily intended to guide experimental analysis of depth cue combination in human vision. We describe experimental methods, analogous to perturbation analysis, that permit us to analyze depth cue combination in novel ways. In particular these methods allow us to investigate the key issues we have raised. We summarize recent experimental tests of the MWF framework that use these methods.