Quantitative Analysis Of Interaction Research Articles

Abstract C022: T cell movement through complex obstacles presented by the microenvironment of pancreatic tumor spheroids

Abstract Immunotherapy has emerged as a promising approach for treating pancreatic cancer, however its efficacy against solid tumors has been largely limited. In order to be effective, cell-based immunotherapies are required to be in close proximity with the target cancerous cells. However, owing to its dense desmoplastic stroma, many regions of a pancreatic tumor, have areas within the tumor volumes that exclude anti-tumor immune cells. Therefore, we need to define mechanisms used by therapeutic T cells to navigate effectively in some tumor regions and the barriers that limit their infiltration and distribution in other regions of the tumor microenvironment (TME). In this work, we have advanced our knowledge of T cell migration and infiltration through complex TMEs by analyzing T cell dynamics in 3D engineered in vitro environments that can replicate key TME features of pancreatic ductal adenocarcinoma (PDA). Using our state-of the-art imaging techniques with multiphoton excitation (MPE) and second harmonic generation (SHG), we have monitored the migration of engineered human mesothelin reactive CAR-T cells through fibrous microenvironments and their interactions with pancreatic tumor spheroids. Quantitative analysis of interactions between CAR-T cells and carcinoma cells, in terms of velocity, directionality and mean square displacement (MSD) were performed, with the aim of utilizing these phenotypes as “design criteria” for the development of more effective engineered T cell therapies. The therapeutic efficacy of engineered T cells was also evaluated in standard 2D killing assays and then new 3D assays of increased complexity, both of which revealed mesothelin specific cancer cell death. Overall, we are starting to define the physical and molecular mechanisms that result in resistance to immunotherapy against pancreatic cancer, and we anticipate that our work will be applicable to other solid tumors with similar challenges and address key unanswered question in cancer biology. Citation Format: Shambojit Roy, Zhongming Chen, Guhan Qian, Paolo P. Provenzano. T cell movement through complex obstacles presented by the microenvironment of pancreatic tumor spheroids [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C022.

Interactional patterns in the online language classroom: a quantitative analysis across proficiency levels and lesson types

Online second language instruction has boomed in recent years, aided by technological affordances and the forced changes in instructional modality resulting from the COVID-19 pandemic. This transformation has underscored the critical role of interaction in online pedagogy. Research suggests that increasing opportunities for interaction between students and instructors is essential for fostering second language acquisition (SLA). However, little research exists quantifying the production of diverse types of interaction in online language instruction, particularly among experienced instructors. The present study utilizes an interactionist framework to perform a quantitative analysis of interaction in online Spanish language coursework, categorized according to interaction initiation type: Instructor-Prompted Participation (IPP), Unprompted Oral Participation (UOP), Unprompted Text Participation (UTP), i.e., chat usage, and interaction length (as exhibited by both the number of student turns in an interaction and the average length of those turns). Data includes 18 h of video recordings of synchronous L2 Spanish language instruction across proficiency levels and lesson types at a distance-learning university in the UK. Lesson types included grammar workshops and exam preparation. Results show that the interactional patterns in online language courses are influenced by proficiency level and lesson type. Lower proficiency students engaged in interaction routines more frequently, while the ability to engage in extended discourse was contingent upon the specific activity/lesson type. The study helps address the dearth of research on interaction and language teaching in languages other than English (LOTE).

Theoretical analysis of stress intensity factors and interactions of collinear flaws with unequal lengths in brittle solids under uniaxial tensile conditions

Interactions between multiple flaws with different sizes in brittle solids are a key factor that induce failure. Previous studies have focused on the initiation, propagation and coalescence behaviors of multiple flaws in brittle solids. Nevertheless, there have been few theoretical or quantitative analyses of interactions between multiple flaws. In the present study, based on the Kachanov method, we first proposed theoretical analytical solutions of the stress intensity factors (SIFs) for two and three collinear flaws with unequal lengths under uniaxial tensile conditions. Then, a theoretical analysis of the effects of various geometric factors, such as flaw interval, flaw length and flaw number, was performed. The results supported the following findings: (1) For two collinear flaws, interactions between the primary and secondary flaws gradually weakened with increasing flaw interval, and the flaw length ratio gradually became the main factor that determined the interaction. (2) For three collinear flaws, when the interval between the primary and secondary flaws was not less than 1.5 times the length of the secondary flaw, the interaction had no effect on the propagation of the primary flaw. (3) The increase in the number of collinear flaws significantly enhanced the effect on propagation reinforcement between the flaws. Finally, numerical simulations were conducted to verify the results of the theoretical analysis, and good consistency was found. These research results are of great theoretical significance and engineering value for quantitatively understanding the failure mechanism of brittle solids containing multiple flaws and proposing effective measures to prevent cracking and ensure the safety of brittle solid structures.

Qualitative and quantitative analysis of interaction between cavitation patterns and vortices of a pitching hydrofoil from Lagrangian viewpoint

The evolution of transient flow structures and mass transport in cavitating flow around a pitching hydrofoil is investigated qualitatively and quantitatively, and the interaction between cavitation patterns and vortices is elucidated from Lagrangian viewpoint. First, turbulence effects are estimated by the density-corrected k–ω model to account for the local compressibility of the multiphase flow at Reynolds number Re=6.4×105. Then, the formation and evolution of vorticity structures during the whole pitching cycle are analyzed using Lagrangian averaged vorticity deviation method. By comparing the flow structures and hydrodynamic properties at varying angles of attack, the cavitating flow is divided into two distinct stages, namely multi-scale cloud cavitation phase from α+=10° to α−=8°, and traveling sheet cavitation phase from α−=8° to α+=10°. Specifically in cloud cavitation, the formation of the cavitation pattern is closely related to the development of the main vortex. Furthermore, the quantitative analysis method based on Lagrangian flow network is developed to deeply analyze the transport and mixing processes. Importantly, the coherence ratio and the mixing parameter are proposed as transport indicators to precisely quantify the spatial connectivity behavior. Finally, the correlations between vapor fraction, codelength, global coherence ratio and global mixing parameter are evaluated. As the conclusion, it is shown that Lagrangian methods are powerful tool for both qualitative and quantitative analysis, and the results obtained could provide a key and important understanding of the flow structure and changing mechanism between cavitation and vortices in marine hydro and propulsion systems.

The Effects of 3D Printing on Social Interactions in Inclusive Classrooms

ABSTRACT This research attempts to examine the role of 3D printer as a learning tool in inclusive education. It examines the quality of verbal and nonverbal interactions between students with disabilities (SWD) and students without disabilities (SWOD) in an inclusive classroom at primary schools, where 3D printers were used as a learning tool. The results presented in this paper are based on qualitative and quantitative analysis of interactions between 7 SWD and 31 SWOD. Even though both occurred, there was no statistical difference in generation of positive and negative verbal and nonverbal interactions between SWD and SWOD. Both, SWD and SWOD are of the opinion that cooperative learning with implementation of 3D printers contributes to their communication with peers, achieving better and durable knowledge in the subject where this learning approach was applied and they would like to learn in this way in the future.

Quantitative analyses of interactions between SpoVG and RNA/DNA

The Borrelia burgdorferi SpoVG protein has previously been found to be a DNA- and RNA-binding protein. To aid in the elucidation of ligand motifs, affinities for numerous RNAs, ssDNAs, and dsDNAs were measured and compared. The loci used in the study were spoVG, glpFKD, erpAB, bb0242, flaB, and ospAB, with particular focus on the untranslated 5′ portion of the mRNAs. Performing binding and competition assays yielded that the 5′ end of spoVG mRNA had the highest affinity while the lowest observed affinity was to the 5’ end of flaB mRNA. Mutagenesis studies of spoVG RNA and ssDNA sequences suggested that the formation of SpoVG-nucleic acid complexes are not entirely dependent on either sequence or structure. Additionally, exchanging uracil for thymine in ssDNAs did not affect protein-nucleic acid complex formation.

Quantitatively probing interactions between membrane with adaptable wettability and oil phase in oil/water separation

The membrane method based on adaptive wettability shows great advantages in oil-water separation. At present, researches focus on the excellent application performance of the membrane material, while the quantitative analysis of interactions in oil-water separation is rarely recognized. Herein, we constructed an adaptable wettability membrane with multiple polymer networks by polydopamine (PDA) and mussel-inspired amphiphilic polymer. Based on the Owens three-probe liquid method, the surface energy of the modified membrane was verified to meet the adaptive wettability conditions, with surface energies (γS) of 147.6 mJ m−2 (superhydrophilic/underwater superoleophobic) and 49.87 mJ m−2 (superhydrophobic/superoleophobic), respectively. The adhesion or repulsion of the membrane to the oil phase under different conditions during the separation process was quantified by the chemical probe AFM technique. In addition, the oil-water selective separation mechanism was further analyzed in a simplified membrane microchannel model. The results show that the different wetting produces capillary additional pressure in opposite directions, resulting in different energies to be overcome when the oil or water passes through the microchannels, thus achieving selective separation.

Quantitative Analysis of the Interactive Influence for Harmonic Emission in Wind Farms

In the process of integrating large-scale wind farms into the power system, the harmonic interaction among wind farms causes potential safe and stable operation threats to the power grid. To effectively control the harmonics of the wind power grid-connected system, quantitative analysis of the harmonic interaction is very necessary and meaningful. Therefore, this paper firstly explains the harmonic interaction among wind farms from the perspective of the primary and secondary emission. We use the concept of harmonic impedance to analyze the mechanism of harmonic interaction and proposed a calculation method for quantitative analysis of the harmonic interaction. Using the calculation method, the influence caused by the wind farm self-impedance, the contact impedance, and Static Var Generator to the harmonic interaction are quantitatively analyzed. Finally, taking multiple wind farms in actual operation as a case, the effectiveness of the method is verified by time domain simulation.

Studies on Protein-RNA:DNA Hybrid Interactions by Microscale Thermophoresis (MST).

Microscale thermophoresis (MST) is a technology that allows for quantitative analysis of interactions between biomolecules with low sample consumption. MST uses localized temperature fields to measure the diffusion rates of the free and bound states of a fluorescently labeled protein, and to determine the dissociation constant KD by fitting of the binding isotherm with a 1:1 binding model. Here, we describe the use of MST for quantitative analysis of the interaction of the N-terminal his-tagged 6-methyladenine (m6A) reader protein YTHDF2 with m6A modified and unmodified RNA, in single-strand configuration or with RNA:DNA hybrid substrates. The described protocol is also suitable for studies of interactions with proteins binding to double-stranded RNA or DNA substrates.

Startup influencing factor investigation and quantitative interaction analysis on a loop thermosyphon with multiple evaporators

Loop thermosyphon with multiple evaporators has unique startup features and interaction characteristics between evaporators which are worth studying. In this paper, the startup influencing factors of a loop thermosyphon with multiple evaporators are studied experimentally and visually. Meanwhile, the interaction between evaporators is evaluated quantitatively using a newly defined parameter called interaction ratio, which represents percentage change in temperature divided by percentage change in heating power. The results show that the temperature of evaporators often has a local peak value during the startup, at which moment the boiling becomes fully developed. Also, the effect of filling ratio, heating power value and distribution on the local peak temperature is analyzed. For each evaporator, interaction ratio on itself is always the largest. Among the evaporators, interaction ratios of the evaporator in the middle are the largest because the competition and crowding of its flow with the other evaporators are more intense. With the increase of filling ratio, interaction ratios on itself decrease while interaction ratios on the others increase.

Linking local water consumption in inland arid regions with imported virtual water: Approaches, application and actuators

The consumption of local blue-green physical water (blue-green PW) is linked to imported virtual water (imported VW) within economic production. The quantitative analysis of interactions between blue-green PW and imported VW is important for integrated management of water resources in inland arid areas. This study used the North Tianshan Mountains (NTM) as a case study to demonstrate the construction of a framework to account for local blue-green PW and imported VW in regional economic production using an input-output model. The results showed that the consumption and transfer of blue-green PW in the agricultural sector constitute the most important part of the water cycle in the NTM. Production consumed relatively less imported VW, 51.3% of which was by the secondary and tertiary industries. VW consumed by the primary industry in the NTM was closely correlated with that by the secondary and tertiary industries, whereas low consumption by the primary industry in the NTM and high demand for primary industry products from outside regions resulted in the direct outflow of large amount of blue-green VW. The current structure of trade in VW also contributed to increasing water shortages in the NTM. This study proposed that the enhancement of the cyclic strength of the primary industry in the local economic system is an effective measure to reduce water loss and improve water utilization efficiency. In addition, by considering the results of industrial linkage analysis of economic sectors, this study proposed that the government should promote the development of industries with large water storage potential, such as construction, manufacturing and forestry, and that adjustment of the structure of local industries should be accelerated so as to reduce the loss of VW during local economic production and trade flows of the primary industry.

Ligand Binding in Allosteric Flavoproteins: Part 1. Quantitative Analysis of the Interaction with NAD+ of the Apoptosis Inducing Factor (AIF) Harboring FAD in the Reduced State.

To perform their action, flavoproteins usually interact with a variety of low molecular weight partners, including electron transporters, yielding transient complexes whose tightness is often controlled by the redox state of the bound flavin cofactor. As a case study, here we describe the quantitative analysis of the redox-dependent interaction of the mammalian apoptosis inducing factor (AIF) with its NAD+ ligand. In particular, we report a protocol for the spectrophotometric titration of AIF in its reduced state under anaerobic conditions with NAD+, in order to determine the dissociation constant of the resulting complex.

Quantitative Analysis of Spatial Distribution Characteristics of Traditional Villages in Hunan Province

This study takes 658 traditional villages in Hunan province as the research object. Relying on ArcGIS geographic information system, the traditional villages in Hunan province were abstracted into point sets with attribute information. By means of average nearest neighbor distance analysis, kernel density analysis and intercity distribution comparison, the basic geographical characteristics of traditional villages in Hunan province were summarized, that is, significant agglomeration distribution, three agglomeration areas and uneven intercity distribution. Multiple types of natural geographical factors were superimposed, such as elevation, slope aspect, slope, river, water systems, annual precipitation and annual average temperature, so as to analyze the impact of elevation topography, river system and climate conditions on village site selection. With the help of geographic detector tool, the correlation between elevation, slope, water system, annual average temperature, annual precipitation and village distribution was quantitatively analyzed. Through the analysis of correlation degree, interaction and single factor characteristics, it is concluded that natural geographical factors are related to village distribution and multiple factors interact with each other. Based on the quantitative analysis of interactions between traditional villages and the natural environment, the distribution characteristics of traditional villages have been summarized, providing reference for the subsequent systematic research and protection of traditional villages.

Association with proteasome determines pathogenic threshold of polyglutamine expansion diseases

Expansion of glutamine residue track (polyQ) within soluble protein is responsible for eight autosomal-dominant genetic neurodegenerative disorders. These disorders affect cerebellum, striatum, basal ganglia and other brain regions. Each disease develops when polyQ expansion exceeds a pathogenic threshold (Qth). A pathogenic threshold is unique for each disease but the reasons for variability in Qth within this family of proteins are poorly understood. In the previous publication we proposed that polarity of the regions flanking polyQ track in each protein plays a key role in defining Qth value [1]. To explain the correlation between the polarity of the flanking sequences and Qth we performed quantitative analysis of interactions between polyQ-expanded proteins and proteasome. Based on structural and theoretical modeling, we predict that Qth value is determined by the energy of polar interaction of the flanking regions with the polyQ and proteasome. More polar flanking regions facilitate unfolding of α-helical polyQ conformation adopted inside the proteasome and as a result, increase Qth. Predictions of our model are consistent with Qth values observed in clinic for each of the eight polyQ-expansion disorders. Our results suggest that the agents that can destabilize polyQ α-helical structure may have a beneficial therapeutic effect for treatment of polyQ-expansion disorders.

A suspension cell‐based interaction platform for interrogation of membrane proteins

AbstractThe majority of clinically approved therapeutics target membrane proteins (MPs), highlighting the need for tools to study this important category of proteins. To overcome limitations with recombinant MP expression, whole cell screening techniques have been developed that present MPs in their native conformations. Whereas many such platforms utilize adherent cells, here we introduce a novel suspension cell‐based platform termed “biofloating” that enables quantitative analysis of interactions between proteins displayed on yeast and MPs expressed on mammalian cells, without need for genetic fusions. We characterize and optimize biofloating and illustrate its sensitivity advantage compared to an adherent cell‐based platform (biopanning). We further demonstrate the utility of suspension cell‐based approaches by iterating rounds of magnetic‐activated cell sorting selections against MP‐expressing mammalian cells to enrich for a specific binder within a yeast‐displayed antibody library. Overall, biofloating represents a promising new technology that can be readily integrated into protein discovery and development workflows.

Virtual ‘Experiential Expert’ Communities of Practice in Sharing Evidence-Based Prevention of Novel Psychoactive Substance (NPS) Use: the Portuguese Experience

We present findings from a unique virtual community of practice piloted to support a programme of prevention evidence and knowledge sharing among professional prevention practitioners as ‘experiential experts’ around tackling novel psychoactive substances (NPS) use in Portugal. A mixed-methods approach that combined quantitative analysis of interactions and qualitative content analysis of debates about NPS, NPS users, patterns of use and best practices in prevention of this type of drug use was conducted. Results show low and irregular interactions between members of this virtual community, but very rich discussions around sharing of experiences and problematizing practices. We discuss the layers of interaction between members, and the shared learning around policy and practice implications. Such virtual and collaborative work practices are not yet integrated within the drug prevention field where instead individualistic approaches tend to prevail and preclude the sharing of alternative solutions that shape different experiences. Our virtual community of NPS prevention experts provides a flagship for ongoing collaboration between research, generation of evidence informing policy and practice, professional training, support and shared learning. It underscores the need for an innovative and multi-disciplinary approach to sharing perspectives in tackling emerging and harmful drug trends.

Protein ligand interaction analysis against new CaMKK2 inhibitors by use of X-ray crystallography and the fragment molecular orbital (FMO) method

CaMKK2 (calcium/calmodulin dependent protein kinase kinase 2) is a serine/threonine protein kinase that regulates phosphorylation of CaM kinases (CaMKs) such as CaMKI, CaMKIV, and AMP-activated protein kinase (AMPK). From a pathological perspective, CaMKK2 plays a role in obesity, diabetes, and prostate cancer. Therefore, CaMKK2 is an attractive target protein for drug design. Here, we tried to find new CaMKK2 inhibitors by using ligand-based and structure-based drug design approaches. From the in silico hit compounds, we identified new inhibitors by using a CaMKK2 kinase assay. We solved X-ray crystallography structures of the CaMKK2–inhibitor complexes and performed Fragment Molecular Orbital (FMO) calculations to analyze the protein–ligand interactions, identify the key residues in inhibitor binding, and quantitatively measure their contribution. We experimentally determined five CaMKK2–inhibitor structures and calculated the binding energies of the inhibitors by the FMO method plus MM-PBSA (Molecular Mechanics Poisson-Boltzmann Surface Area) approach. The results showed a high correlation (R = −0.89) between experimentally measured inhibitory activity (pIC50) and the predicted ligand binding energy. We then quantitatively evaluated the contribution of each binding site residue in CaMKK2 by the IFIE (Inter-fragment Interaction Energy)/PIEDA (Pair Interaction Energy Decomposition Analysis) method. The IFIE values indicated that Lys194 and Glu236, which formed hydrogen bonds with the carboxylate groups of the inhibitors, were key residues for ligand binding. PIEDA revealed that the dispersion interaction of inhibitors with hydrophobic residues, such as Ile171, Phe267, and Leu319, contributed highly to ligand binding; we considered that this was due to CH-π interactions with methoxy groups and/or aromatic rings contained in our CaMKK2 inhibitor. These results from the quantitative interaction analysis by the FMO method are useful not only for future CaMMK2 inhibitor development but for application of the FMO method to in silico drug design.

Self-assembly of robust gold nanoparticle monolayer architectures for quantitative protein interaction analysis by LSPR spectroscopy

Localized surface plasmon resonance (LSPR) detection offers highly sensitive label-free detection of biomolecular interactions. Simple and robust surface architectures compatible with real-time detection in a flow-through system are required for broad application in quantitative interaction analysis. Here, we established self-assembly of a functionalized gold nanoparticle (AuNP) monolayer on a glass substrate for stable, yet reversible immobilization of Histidine-tagged proteins. To this end, one-step coating of glass substrates with poly-L-lysine graft poly(ethylene glycol) functionalized with ortho-pyridyl disulfide (PLL-PEG-OPSS) was employed as a reactive, yet biocompatible monolayer to self-assemble AuNP into a LSPR active monolayer. Site-specific, reversible immobilization of His-tagged proteins was accomplished by coating the AuNP monolayer with tris-nitrilotriacetic acid (trisNTA) PEG disulfide. LSPR spectroscopy detection of protein binding on these biocompatible functionalized AuNP monolayers confirms high stability under various harsh analytical conditions. These features were successfully employed to demonstrate unbiased kinetic analysis of cytokine-receptor interactions.Graphical abstract

Crystal structure, quantum chemical and Hirshfeld surface analysis of N-ethyl-6-methyl-2-oxo-4-phenyl-1,2,3,4-tetrahydropyrimidine-5-carboxamide

Abstract The molecular structure of N-ethyl-6-methyl-2-oxo-4-phenyl-1,2,3,4-tetrahydropyrimidine-5- carboxamide (C14H16N3O2) has been investigated via single crystal X-ray diffraction. The compound crystallizes in the monoclinic, P21 space group with unit cell parameters a = 11.5273(8), b = 7.4347(5), c = 16.1243(11) A, and β = 104.941(7)° with two independent molecules in the asymmetric unit. The dihedral angles formed between mean plane of pyrimidine moiety with phenyl ring is 82.09° and 83.17° in respectively in two molecules indicates their conformations are slightly different. The interplay of intermolecular C H…O and N H…N, hydrogen bonds found to be effective in stabilizing the molecular structure. The PIXEL Coulomb London Pauli (CLP) module used to estimate energetics of interactions among molecules in crystal packing. Quantitative analysis of interactions also presented using Hirshfeld fingerprint plots. Further, reaction sites in the molecule are represented in molecular electrostatic potential maps (MEP) plot.

A concise method for quantitative analysis of interactions between lipids and membrane proteins

Although interactions between lipids and membrane proteins (MPs) have been considered crucially important for understanding the functions of lipids, lack of useful and convincing experimental methods has hampered the analysis of the interactions. Here, we developed a surface plasmon resonance (SPR)-based concise method for quantitative analysis of lipid-MP interactions, coating the sensor chip surface with self-assembled monolayer (SAM) with C6-chain. To develop this method, we used bacteriorhodopsin (bR) as an MP, and examined its interaction with various types of lipids. The merits of using C6-SAM-modified sensor chip are as follows: (1) alkyl-chains of SAM confer a better immobilization of MPs because of the efficient preconcentration due to hydrophobic contacts; (2) SAM provides immobilized MPs with a partial membranous environment, which is important for the stabilization of MPs; and (3) a thinner C6-SAM layer (1 nm) compared with MP size forces the MP to bulge outward from the SAM surface, allowing extraneously injected lipids to be accessible to the hydrophobic transmembrane regions. Actually, the amount of bR immobilized on C6-SAM is 10 times higher than that on a hydrophilic CM5 sensor chip, and AFM observations confirmed that bR molecules are exposed on the SAM surface. Of the lipids tested, S-TGA-1, a halobacterium-derived glycolipid, had the highest specificity to bR with a nanomolar dissociation constant. This is consistent with the reported co-crystal structure that indicates the formation of several intermolecular hydrogen bonds. Therefore, we not only reproduced the specific lipid-bR recognition, but also succeeded in its quantitative evaluation, demonstrating the validity and utility of this method.