Peptide-Drug Conjugates: An Emerging Direction for the Next Generation of Peptide Therapeutics.

Building on recent advances in peptide science, medicinal chemists have developed a hybrid class of bioconjugates, called peptide-drug conjugates, that demonstrate improved efficacy compared to peptides and small molecules independently. In this Perspective, we discuss how the conjugation of synergistic peptides and small molecules can be used to overcome complex disease states and resistance mechanisms that have eluded contemporary therapies because of their multi-component activity. We highlight how peptide-drug conjugates display a multi-factor therapeutic mechanism similar to that of antibody-drug conjugates but also demonstrate improved therapeutic properties such as less-severe off-target effects and conjugation strategies with greater site-specificity. The many considerations that go into peptide-drug conjugate design and optimization, such as peptide/small-molecule pairing and chemo-selective chemistries, are discussed. We also examine several peptide-drug conjugate series that demonstrate notable activity toward complex disease states such as neurodegenerative disorders and inflammation, as well as viral and bacterial targets with established resistance mechanisms.

Eye movements and event segmentation: Eye movements reveal age-related differences in event model updating.

People spontaneously segment continuous ongoing actions into sequences of events. Prior research found that gaze similarity and pupil dilation increase at event boundaries and that older adults segment more idiosyncratically than do young adults. We used eye tracking to explore age-related differences in gaze similarity (i.e., the extent to which individuals look at the same places at the same time as others) and pupil dilation at event boundaries. Older and young adults watched naturalistic videos of actors performing everyday activities while we tracked their eye movements. Afterward, they segmented the videos into subevents. Replicating prior work, we found that pupil size and gaze similarity increased at event boundaries. Thus, there were fewer individual differences in eye position at boundaries. We also found that young adults had higher gaze similarity than older adults throughout an entire video and at event boundaries. This study is the first to show that age-related differences in how people parse continuous everyday activities into events may be partially explained by individual differences in gaze patterns. Those who segment less normatively may do so because they fixate less normative regions. Results have implications for future interventions designed to improve encoding in older adults. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Identification of potential CD8+ epitopes in pp62 polyprotein of African swine fever virus using computational immunology.

The highly infectious African swine fever virus (ASFV) is currently the only known DNA arbovirus within the Asfarviridae family which primarily infects domestic pigs and wild boars. African swine fever (ASF) leads to a mortality rate of up to 100% which has caused massive socio-economic losses worldwide. Previous research indicates that ASFV's virulence can be attributed to polyprotein pp62, which plays a crucial role in viral assembly and core maturation. This particular study utilized in silico analysis to identify highly conserved cytotoxic T-cell epitopes in pp62 that can potentially serve as key components for future ASFV vaccines. To achieve this, the researchers retrieved, clustered, and aligned the peptide sequences of pp62. Subsequently, the aligned sequences were analyzed to identify epitopes that bind promiscuously to the swine major histocompatibility complex I (MHC I) alleles and exhibiting MHC IC50 values < 500 nM. Additionally, peptide sequences with positive proteasome and TAP scores were considered. Potential cross-reactivity was assessed by comparing the peptide sequences against available proteome sequences of Sus scrofa domesticus in various databases. Furthermore, molecular docking was conducted to evaluate the binding of candidate epitopes with swine leukocyte antigen-1*0401 (SLA-1*0401). The dissociation constants, binding energies, root mean square deviation, and root mean square fluctuation values for the SLA-epitope complexes were compared with a positive reference. In the course of the study, 21 highly conserved CD8+ epitopes were identified, out of which four were further assessed for their potential immunogenicity. The results demonstrated that the highly conserved CD8+ epitopes discovered in this study are promising for integration into future ASFV vaccine formulations. As preliminary data, it is anticipated that these findings will subsequently undergo in vitro and in vivo studies in the future.

The role of individual differences in resistance to persuasion on memory for political advertisements.

When people see political advertisements on a polarized issue they take a stance on, what factors influence how they respond to and remember the adverts contents? Across three studies, we tested competing hypotheses about how individual differences in social vigilantism (i.e., attitude superiority) and need for cognition relate to intentions to resist attitude change and memory for political advertisements concerning abortion. In Experiments 1 and 2, we examined participants' intentions to use resistance strategies to preserve their pre-existing attitudes about abortion, by either engaging against opposing opinions or disengaging from them. In Experiment 3, we examined participants' memory for information about both sides of the controversy presented in political advertisements. Our results suggest higher levels of social vigilantism are related to greater intentions to counterargue and better memory for attitude-incongruent information. These findings extend our understanding of individual differences in how people process and respond to controversial social and political discourse.

Structure-Activity Relationships of the Antimicrobial Peptide Natural Product Apidaecin.

With the growing crisis of antimicrobial resistance, it is critical to continue to seek out new sources of novel antibiotics. This need has led to renewed interest in natural product antimicrobials, specifically antimicrobial peptides. Nonlytic antimicrobial peptides are highly promising due to their unique mechanisms of action. One such peptide is apidaecin (Api), which inhibits translation termination through stabilization of the quaternary complex of the ribosome-apidaecin-tRNA-release factor. Synthetic derivatives of apidaecin have been developed, but structure-guided modifications have yet to be considered. In this work, we have focused on modifying key residues in the Api sequence that are responsible for the interactions that stabilize the quaternary complex. We present one of the first examples of a highly modified Api peptide that maintains its antimicrobial activity and interaction with the translation complex. These findings establish a starting point for further structure-guided optimization of Api peptides.

Adsorption kinetics of polyacrylamide-based polyelectrolyte onto a single silica particle studied using microfluidics and optical tweezers.

HYPOTHESIS: Polyelectrolyte adsorption is considered important in tuning the surface property and the fate of particles; however, often studied on macroscopic surfaces. To gain insights into how polyelectrolytes are adsorbed onto a single particle, it is imperative to utilize techniques capable of elucidating adsorption kinetics on a single-particle level in a controlled flow field. EXPERIMENTS: The polyelectrolyte adsorption kinetics was investigated by electrophoretic mobility measurements combined with the kinetics study onto a single-particle viewpoint using microfluidics and optical tweezers. We directly evaluated the thickness, δH, of adsorbed polyelectrolyte onto a negatively-charged silica particle to deduce the adsorbed polyelectrolyte's conformation. The effect of charge density and salt concentrations were studied. FINDINGS: All polyelectrolytes exhibited dependence of δH on salt concentration. The attractive interactions control the cationic polyelectrolytes adsorption process. The δH depends on charge density indicating more loops and tail confirmation for the weakly-charged polyelectrolytes. The anionic polyelectrolytes showed a dependence of the initial rate and saturation value of δH on salt concentrations, attributed to the repulsion between charged segments and the silica surfaces. Here, we present new insights into the polyelectrolyte adsorption kinetics, particularly the influence of electrostatic interaction from the single-particle perspective, inaccessible to conventional bulk measurements.

A reverse Monte Carlo algorithm to simulate two-dimensional small-angle scattering intensities.

Small-angle scattering (SAS) experiments are a powerful method for studying self-assembly phenomena in nanoscopic materials because of the sensitivity of the technique to structures formed by interactions on the nanoscale. Numerous out-of-the-box options exist for analysing structures measured by SAS but many of these are underpinned by assumptions about the underlying interactions that are not always relevant for a given system. Here, a numerical algorithm based on reverse Monte Carlo simulations is described to model the intensity observed on a SAS detector as a function of the scattering vector. The model simulates a two-dimensional detector image, accounting for magnetic scattering, instrument resolution, particle polydispersity and particle collisions, while making no further assumptions about the underlying particle interactions. By simulating a two-dimensional image that can be potentially anisotropic, the algorithm is particularly useful for studying systems driven by anisotropic interactions. The final output of the algorithm is a relative particle distribution, allowing visualization of particle structures that form over long-range length scales (i.e. several hundred nanometres), along with an orientational distribution of magnetic moments. The effectiveness of the algorithm is shown by modelling a SAS experimental data set studying finite-length chains consisting of magnetic nanoparticles, which assembled in the presence of a strong magnetic field due to dipole interactions.

Narrative Comprehension Guides Eye Movements in the Absence of Motion.

Viewers' attentional selection while looking at scenes is affected by both top-down and bottom-up factors. However, when watching film, viewers typically attend to the movie similarly irrespective of top-down factors-a phenomenon we call the tyranny of film. A key difference between still pictures and film is that film contains motion, which is a strong attractor of attention and highly predictive of gaze during film viewing. The goal of the present study was to test if the tyranny of film is driven by motion. To do this, we created a slideshow presentation of the opening scene of Touch of Evil. Context condition participants watched the full slideshow. No-context condition participants did not see the opening portion of the scene, which showed someone placing a time bomb into the trunk of a car. In prior research, we showed that despite producing very different understandings of the clip, this manipulation did not affect viewers' attention (i.e., the tyranny of film), as both context and no-context participants were equally likely to fixate on the car with the bomb when the scene was presented as a film. The current study found that when the scene was shown as a slideshow, the context manipulation produced differences in attentional selection (i.e., it attenuated attentional synchrony). We discuss these results in the context of the Scene Perception and Event Comprehension Theory, which specifies the relationship between event comprehension and attentional selection in the context of visual narratives.

Order vs. Disorder: Cholesterol and Omega-3 Phospholipids Determine Biomembrane Organization.

Lipid structural diversity strongly affects biomembrane chemico-physical and structural properties in addition to membrane-associated events. At high concentrations, cholesterol increases membrane order and rigidity, while polyunsaturated lipids are reported to increase disorder and flexibility. How these different tendencies balance in composite bilayers is still controversial. In this study, electron paramagnetic resonance spectroscopy, small angle neutron scattering, and neutron reflectivity were used to investigate the structural properties of cholesterol-containing lipid bilayers in the fluid state with increasing amounts of polyunsaturated omega-3 lipids. Either the hybrid 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine or the symmetric 1,2-docosahexaenoyl-sn-glycero-3-phosphocholine were added to the mixture of the naturally abundant 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine and cholesterol. Our results indicate that the hybrid and the symmetric omega-3 phospholipids affect the microscopic organization of lipid bilayers differently. Cholesterol does not segregate from polyunsaturated phospholipids and, through interactions with them, is able to suppress the formation of non-lamellar structures induced by the symmetric polyunsaturated lipid. However, this order/disorder balance leads to a bilayer whose structural organization cannot be ascribed to either a liquid ordered or to a canonical liquid disordered phase, in that it displays a very loose packing of the intermediate segments of lipid chains.

Complete atrioventricular septal defect with absent or diminutive primum component: Incidence, anatomic characteristics, and outcomes.

BACKGROUND: Repair of complete atrioventricular septal defect with absent or diminutive primum defect is challenging because of atrial septal malposition and abnormal anatomy of the left atrioventricular valve. We sought to define the incidence, anatomy, and surgical outcomes of this entity. METHODS: We identified all patients in our institutional database presenting for complete atrioventricular septal defect repair from 2006 to 2018. Operative reports and echocardiograms were reviewed to determine the presence and size of the primum defect, atrioventricular valve anatomy, degree of atrioventricular valve regurgitation, repair method, and complications, including reoperation for atrioventricular valve regurgitation. Functionally univentricular patients and those receiving repair at an outside institution were excluded. RESULTS: Of 183 patients with complete atrioventricular septal defect, absent/diminutive primum defect occurred in 16 patients (8.7%; 10 absent, 6 diminutive). Six patients (38%) had leftward malposition of the atrium septum on the common atrioventricular valve. The rate of reoperation for left atrioventricular valve regurgitation was 31% (3 early, 2 late), for which preoperative predictors included leftward malposition of the atrial septum onto the common atrioventricular valve (4/6 patients with malposition required reoperation, P = .036, Fisher exact test). One patient exhibiting this risk factor died. The overall rate of moderate or greater left atrioventricular valve regurgitation on the most recent postoperative echocardiogram was 13% (2/16 patients; median follow-up, 141 days; range, 3-2236 days). CONCLUSIONS: Complete atrioventricular septal defect with absent or diminutive primum defect is a unique variant of complete atrioventricular septal defect for which the risk of reoperation for left atrioventricular valve regurgitation after complete repair is high and risk factors include leftward malposition of the atrial septum on the common atrioventricular valve.

Hollow Gold Nanosphere Templated Synthesis of PEGylated Hollow Gold Nanostars and Use for SERS Detection of Amyloid Beta in Solution.

Hollow gold nanospheres (HGNs) have been used as the template for seed-mediated growth of multibranched hollow gold nanostars (HNS). The HGNs were synthesized via anerobic reduction of cobalt chloride to cobalt nanoparticles and then formation of a gold shell via galvanic replacement followed by the oxidation of the cobalt core. We obtained control of the inner core size of the HGNs by increasing the size of the sacrificial cobalt core and by varying the ratio of B(OH)3/BH4 using boric acid rather than 48 h aged borohydride. We synthesized the HNS by reducing Au3+ ions in the presence of Ag+ ions using ascorbic acid, creating a spiky morphology that varied with the Au3+/Ag+ ratio. A broadly tunable localized surface plasmon resonance was achieved through control of both the inner core and the spike length. Amyloid beta (Aß) was conjugated to the HNS by using a heterobifunctional PEG linker and identified by the vibrational modes associated with the conjugated ring phenylalanine side chain. A bicinchoninic acid assay was used to determine the concentration of Aß conjugated to HNS as 20 nM, which is below the level of Aß that negatively affects long-term potentiation. Both the core size and spike length were shown to affect the optical properties of the resulting nanostructures. This HGN templated method introduced a new parameter for enhancing the plasmonic properties of gold nanostars, namely, the addition of a hollow core. Hollow gold nanostars are highly desirable for a wide range of applications, including high sensitivity disease detection and monitoring.

K-Carrageenan Stimulates Pre-Osteoblast Proliferation and Osteogenic Differentiation: A Potential Factor for the Promotion of Bone Regeneration?

Current cell-based bone tissue regeneration strategies cannot cover large bone defects. K-carrageenan is a highly hydrophilic and biocompatible seaweed-derived sulfated polysaccharide, that has been proposed as a promising candidate for tissue engineering applications. Whether κ-carrageenan can be used to enhance bone regeneration is still unclear. In this study, we aimed to investigate whether κ-carrageenan has osteogenic potential by testing its effect on pre-osteoblast proliferation and osteogenic differentiation in vitro. Treatment with κ-carrageenan (0.5 and 2 mg/mL) increased both MC3T3-E1 pre-osteoblast adhesion and spreading at 1 h. K-carrageenan (0.125-2 mg/mL) dose-dependently increased pre-osteoblast proliferation and metabolic activity, with a maximum effect at 2 mg/mL at day three. K-carrageenan (0.5 and 2 mg/mL) increased osteogenic differentiation, as shown by enhanced alkaline phosphatase activity (1.8-fold increase at 2 mg/mL) at day four, and matrix mineralization (6.2-fold increase at 2 mg/mL) at day 21. K-carrageenan enhanced osteogenic gene expression (Opn, Dmp1, and Mepe) at day 14 and 21. In conclusion, κ-carrageenan promoted MC3T3-E1 pre-osteoblast adhesion and spreading, metabolic activity, proliferation, and osteogenic differentiation, suggesting that κ-carrageenan is a potential osteogenic inductive factor for clinical application to enhance bone regeneration.

Magnetic-field-controlled spin fluctuations and quantum critically in Sr3Ru2O7.

When the transition temperature of a continuous phase transition is tuned to absolute zero, new ordered phases and physical behaviour emerge in the vicinity of the resulting quantum critical point. Sr3Ru2O7 can be tuned through quantum criticality with magnetic field at low temperature. Near its critical field Bc it displays the hallmark T-linear resistivity and a [Formula: see text] electronic heat capacity behaviour of strange metals. However, these behaviours have not been related to any critical fluctuations. Here we use inelastic neutron scattering to reveal the presence of collective spin fluctuations whose relaxation time and strength show a nearly singular variation with magnetic field as Bc is approached. The large increase in the electronic heat capacity and entropy near Bc can be understood quantitatively in terms of the scattering of conduction electrons by these spin-fluctuations. On entering the spin-density-wave ordered phase present near Bc, the fluctuations become stronger suggesting that the order is stabilised through an "order-by-disorder" mechanism.

Knowledge guides attention to goal-relevant information in older adults.

How does viewers' knowledge guide their attention while they watch everyday events, how does it affect their memory, and does it change with age? Older adults have diminished episodic memory for everyday events, but intact semantic knowledge. Indeed, research suggests that older adults may rely on their semantic memory to offset impairments in episodic memory, and when relevant knowledge is lacking, older adults' memory can suffer. Yet, the mechanism by which prior knowledge guides attentional selection when watching dynamic activity is unclear. To address this, we studied the influence of knowledge on attention and memory for everyday events in young and older adults by tracking their eyes while they watched videos. The videos depicted activities that older adults perform more frequently than young adults (balancing a checkbook, planting flowers) or activities that young adults perform more frequently than older adults (installing a printer, setting up a video game). Participants completed free recall, recognition, and order memory tests after each video. We found age-related memory deficits when older adults had little knowledge of the activities, but memory did not differ between age groups when older adults had relevant knowledge and experience with the activities. Critically, results showed that knowledge influenced where viewers fixated when watching the videos. Older adults fixated less goal-relevant information compared to young adults when watching young adult activities, but they fixated goal-relevant information similarly to young adults, when watching more older adult activities. Finally, results showed that fixating goal-relevant information predicted free recall of the everyday activities for both age groups. Thus, older adults may use relevant knowledge to more effectively infer the goals of actors, which guides their attention to goal-relevant actions, thus improving their episodic memory for everyday activities.

Stabilizing the Solid-Electrolyte Interphase with Polyacrylamide for High-Voltage Aqueous Lithium-Ion Batteries.

The introduction of "water-in-salt" electrolyte (WiSE) concept opens a new horizon to aqueous electrochemistry that is benefited from the formation of a solid-electrolyte interphase (SEI). However, such SEI still faces multiple challenges, including dissolution, mechanical damaging, and incessant reforming, which result in poor cycling stability. Here, we report a polymeric additive, polyacrylamide (PAM) that effectively stabilizes the interphase in WiSE. With the addition of 5 molar % PAM to 21 mol kg-1 LiTFSI electrolyte, a LiMn2 O4 ∥L-TiO2 full cell exhibits enhanced cycling stability with 86 % capacity retention after 100 cycles at 1 C. The formation mechanism and evolution of PAM-assisted SEI was investigated using operando small angle neutron scattering and density functional theory (DFT) calculations, which reveal that PAM minimizes the presence of free water molecules at the anode/electrolyte interface, accelerates the TFSI- anion decomposition, and densifies the SEI.

Effect of shear flow on the hydrodynamic drag force of a spherical particle near a wall evaluated using optical tweezers and microfluidics.

The hydrodynamic drag force on a spherical particle in shear flow near-wall is investigated using optical tweezers and microfluidics. Simple shear flow is applied using a microfluidic channel at different volumetric flow rates. The hydrodynamic drag force exerted on the particle is detected from the displacement of the trapped particle. The effect of the wall is obtained from the force balance of the trapping and hydrodynamic drag force employing the exact solution of the theoretical model using the lubrication theory for a sphere near the wall. Here, we report the experimentally obtained hydrodynamic drag force coefficient under the influence of shear flow. The drag correction factor increases with decreasing distance from the wall due to the effect of the wall surface. We found that the calculated hydrodynamic drag force coefficient is in quantitative comparison with the theoretical prediction for a shear flow past a sphere near-wall. This study provides a straightforward investigation of the effect of the shear flow on the hydrodynamic drag force coefficient on a particle near the wall. Furthermore, these pieces of information can be used in various applications, particularly in optimizing microfluidic designs for mixing and separations of particles or exploiting the formation of the concentration gradient of particles perpendicular to flow directions caused by the non-linear hydrodynamic interactions.

Estimating different order polynomial logarithmic environmental Kuznets curves.

This paper contributes to the environmental literature by (i) demonstrating that the estimated coefficients and the statistical significance of the non-leading terms in quadratic, cubic, and quartic logarithmic environmental Kuznets curve (EKC) specifications are arbitrary and should therefore not be used to choose the preferred specification and (ii) detailing a proposed general-to-specific type methodology for choosing the appropriate specifications when attempting to estimate higher-order polynomials such as cubic and quartic logarithmic EKC relationships. Testing for the existence and shape of the well-known EKC phenomenon is a hot topic in the environmental economics literature. The conventional approach widely employs quadratic and cubic specifications and more recently also the quartic specification, where the variables are in logarithmic form. However, it is important that researchers understand whether the estimated EKC coefficients, turning points, and elasticities are statistically acceptable, economically interpretable, and comparable. In addition, it is vital that researchers have a clear structured non-arbitrary methodology for determining the preferred specification and hence shape of the estimated EKC. We therefore show mathematically and empirically the arbitrary nature of estimated non-leading coefficients in quadratic, cubic, and quartic logarithmic EKC specifications, being dependent upon the units of measurement chosen for the independent variables (e.g. dependent upon a rescaling of the variables such as moving from $m to $bn). Consequently, the practice followed in many previously papers, whereby the estimates of the non-leading terms are used in the decision to choose the preferred specification of an estimated EKC relationship, is incorrect and should not be followed since it potentially could lead to misleading conclusions. Instead, it should be based upon the sign and statistical significance of the estimated coefficients of the leading terms, the location of turning point(s), and the sign and statistical significance of the estimated elasticities. Furthermore, we suggest that researchers should follow a proposed general-to-specific type methodology for choosing the appropriate order of polynomials when attempting to estimate higher-order polynomial logarithmic EKCs.

Neonatal venoarterial extracorporeal membrane oxygenation cannulation technique.

Peripheral cannulation for initiation of neonatal venoarterial extracorporeal membrane oxygenation is an essential technical skill in the armamentarium of every congenital cardiac surgeon and is necessary for the care of newborns with congenital heart disease and other conditions presenting with cardiopulmonary failure. With certain exceptions, it is usually performed via a right neck cut-down to expose the right internal jugular vein and common carotid artery.   A strong understanding of the fundamental techniques, as well as strategies for avoiding common pitfalls, is critical to the success of cardiopulmonary rescue afforded by cannulation, especially considering the emergent conditions under which cannulation must usually be performed. We review here the operative approach and technique for neck cannulation for peripheral neonatal venoarterial extracorporeal membrane oxygenation.

Toxicological assessment and food allergy of silk fibroin derived from Bombyx mori cocoons.

Recent studies have demonstrated silk fibroin protein's (SF) ability to extend the shelf life of foods by mitigating the hallmarks of spoilage, namely oxidation and dehydration. Due to the potential for this protein to become more widespread, its safety was evaluated comprehensively. First, a bacterial reverse mutation test (Ames test) was conducted in five bacterial strains. Second, an in vivo erythrocyte test was conducted with Sprague Dawley rats at doses up to 1,000mg/kg-bw/day. Third, a range-finder study was conducted with Sprague Dawley rats at the highest consumption amount given solubility and oral gavage volume constrains (500mg/kg-bw/day). Fourth, a 28-day sub-chronic study in Sprague Dawley rats was conducted with the high dose set at 500mg/kg-bw/day, as limited by solubility of the protein in a single-gavage per-day study. Fifth, an in vitro pepsin digestion assay was performed to assess the potential for protein allergenicity. Sixth, allergenic potential was further assessed using liquid chromatography-mass spectroscopy for detection of allergenic insect proteins. Seventh, the SF protein sequences were subjected to bioinformatic analyses. Together, these studies raise no mutagenic, genotoxic, toxicological, or allergenic concerns with the oral consumption of silk fibroin.

Investigating Visual Crowding of Objects in Complex Real-World Scenes.

Visual crowding, the impairment of object recognition in peripheral vision due to flanking objects, has generally been studied using simple stimuli on blank backgrounds. While crowding is widely assumed to occur in natural scenes, it has not been shown rigorously yet. Given that scene contexts can facilitate object recognition, crowding effects may be dampened in real-world scenes. Therefore, this study investigated crowding using objects in computer-generated real-world scenes. In two experiments, target objects were presented with four flanker objects placed uniformly around the target. Previous research indicates that crowding occurs when the distance between the target and flanker is approximately less than half the retinal eccentricity of the target. In each image, the spacing between the target and flanker objects was varied considerably above or below the standard (0.5) threshold to either suppress or facilitate the crowding effect. Experiment 1 cued the target location and then briefly flashed the scene image before participants could move their eyes. Participants then selected the target object's category from a 15-alternative forced choice response set (including all objects shown in the scene). Experiment 2 used eye tracking to ensure participants were centrally fixating at the beginning of each trial and showed the image for the duration of the participant's fixation. Both experiments found object recognition accuracy decreased with smaller spacing between targets and flanker objects. Thus, this study rigorously shows crowding of objects in semantically consistent real-world scenes.