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DOE OSTI2017

Co-produced natural ketolides methymycin and pikromycin inhibit bacterial growth by preventing synthesis of a limited number of proteins

Antibiotics methymycin (MTM) and pikromycin (PKM), co-produced by Streptomyces venezuelae, represent minimalist macrolide protein synthesis inhibitors. Unlike other macrolides, which carry several side chains, a single desosamine sugar is attached to the macrolactone ring of MTM and PKM. In addition, the macrolactone scaffold of MTM is smaller than in other macrolides. The unusual structure of MTM and PKM and their simultaneous secretion by S. venezuelae bring about the possibility that two compounds would bind to distinct ribosomal sites. However, by combining genetic, biochemical and crystallographic studies, we demonstrate that MTM and PKM inhibit translation by binding to overlapping sites in the ribosomal exit tunnel. Strikingly, while MTM and PKM readily arrest the growth of bacteria, ~40% of cellular proteins continue to be synthesized even at saturating concentrations of the drugs. Gel electrophoretic analysis shows that compared to other ribosomal antibiotics, MTM and PKM prevent synthesis of a smaller number of cellular polypeptides illustrating a unique mode of action of these antibiotics.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Seawater exposure causes hydraulic damage in dying Sitka-spruce trees

Sea-level rise is one of the most critical challenges facing coastal ecosystems under climate change. Observations of elevated tree mortality in global coastal forests are increasing, but the mechanism of salinity-stress induced tree mortality is currently unknown. We monitored progressive mortality and associated gas exchange and hydraulic shifts in Sitka-spruce (Picea sitchensis) trees located within a salinity gradient under an ecosystem-scale change of seawater exposure in Washington State, USA. Percentage of live foliated crown (PLFC) decreased and tree mortality increased with increasing soil salinity during the study period. A strong reduction in gas exchange and xylem hydraulic conductivity (Ks) occurred during tree death, with an increase of the percentage loss of xylem conductivity (PLC) and turgor loss point (ptlp). Hydraulic and osmotic shifts reflected that hydraulic function declined from seawater exposure and dying trees were unable to support osmotic adjustment. Constrained gas exchange was strongly related to hydraulic damage at both stem and leaf levels. Significant correlations between foliar sodium (Na+) concentration and gas exchange and key hydraulic parameters (Ks, PLC and ptlp ) suggest that cellular injury related to the toxic effects of ion accumulation impacted the physiology of these dying trees. This study provides evidence of toxic effects on cellular function that become manifest in all aspects of plant functioning, leading to unfavourable osmotic and hydraulic conditions.

Zhang, Hongxia↗

DOE OSTI2024

Cynipid wasps systematically reprogram host metabolism and restructure cell walls in developing galls

Many insects have evolved the ability to manipulate plant growth to generate extraordinary structures called galls, in which insect larva can develop while being sheltered and feeding on the plant. In particular, cynipid (Hymenoptera: Cynipidae) wasps have evolved to form morphologically complex galls and generate an astonishing array of gall shapes, colors, and sizes. However, the biochemical basis underlying these remarkable cellular and developmental transformations remains poorly understood. A key determinant in plant cellular development is cell wall deposition that dictates the physical form and physiological function of newly developing cells, tissues, and organs. However, it is unclear to what degree cell walls are restructured to initiate and support the formation of new gall tissue. Here, we characterize the molecular alterations underlying gall development using a combination of metabolomic, histological, and biochemical techniques to elucidate how valley oak (Quercus lobata) leaf cells are reprogrammed to form galls. Strikingly, gall development involves an exceptionally coordinated spatial deposition of lignin and xylan to form de novo gall vasculature. Our results highlight how cynipid wasps can radically change the metabolite profile and restructure the cell wall to enable the formation of galls, providing insights into the mechanism of gall induction and the extent to which plants can be entirely reprogrammed to form unique structures and organs.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

From sensing to acclimation: The role of membrane lipid remodeling in plant responses to low temperatures

Abstract Low temperatures pose a dramatic challenge to plant viability. Chilling and freezing disrupt cellular processes, forcing metabolic adaptations reflected in alterations to membrane compositions. Understanding the mechanisms of plant cold tolerance is increasingly important due to anticipated increases in the frequency, severity, and duration of cold events. This review synthesizes current knowledge on the adaptive changes of membrane glycerolipids, sphingolipids, and phytosterols in response to cold stress. We delve into key mechanisms of low-temperature membrane remodeling, including acyl editing and headgroup exchange, lipase activity, and phytosterol abundance changes, focusing on their impact at the subcellular level. Furthermore, we tabulate and analyze current gycerolipidomic data from cold treatments of Arabidopsis, maize, and sorghum. This analysis highlights congruencies of lipid abundance changes in response to varying degrees of cold stress. Ultimately, this review aids in rationalizing observed lipid fluctuations and pinpoints key gaps in our current capacity to fully understand how plants orchestrate these membrane responses to cold stress.

Plant Sciences↗

DOE OSTI2020

Modular cell-free expression plasmids to accelerate biological design in cells

Industrial biotechnology aims to produce high-value products from renewable resources. This can be challenging because model microorganisms—organisms that are easy to use like Escherichia coli—often lack the machinery required to utilize desired feedstocks like lignocellulosic biomass or syngas. Non-model organisms, such as Clostridium, are industrially proven and have desirable metabolic features but have several hurdles to mainstream use. Namely, these species grow more slowly than conventional laboratory microbes, and genetic tools for engineering them are far less prevalent. To address these hurdles for accelerating cellular design, cell-free synthetic biology has matured as an approach for characterizing non-model organisms and rapidly testing metabolic pathways in vitro. Unfortunately, cell-free systems can require specialized DNA architectures with minimal regulation that are not compatible with cellular expression. In this work, we develop a modular vector system that allows for T7 expression of desired enzymes for cell-free expression and direct Golden Gate assembly into Clostridium expression vectors. Utilizing the Joint Genome Institute’s DNA Synthesis Community Science Program, we designed and synthesized these plasmids and genes required for our projects allowing us to shuttle DNA easily between our in vitro and in vivo experiments. We next validated that these vectors were sufficient for cell-free expression of functional enzymes, performing on par with the previous state-of-the-art. Lastly, we demonstrated automated six-part DNA assemblies for Clostridium autoethanogenum expression with efficiencies ranging from 68% to 90%. We anticipate this system of plasmids will enable a framework for facile testing of biosynthetic pathways in vitro and in vivo by shortening development cycles.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Repeated exposure to eucalyptus wood smoke alters pulmonary gene and metabolic profiles in male Long-Evans rats

Abstract Exposure to wildfire smoke is associated with both acute and chronic cardiopulmonary illnesses, which are of special concern for wildland firefighters who experience repeated exposure to wood smoke. It is necessary to better understand the underlying pathophysiology by which wood smoke exposure increases pulmonary disease burdens in this population. We hypothesize that wood smoke exposure produces pulmonary dysfunction, lung inflammation, and gene expression profiles associated with future pulmonary complications. Male Long-Evans rats were intermittently exposed to smoldering eucalyptus wood smoke at 2 concentrations, low (11.0 ± 1.89 mg/m3) and high (23.7 ± 0.077 mg/m3), over a 2-week period. Whole-body plethysmography was measured intermittently throughout. Lung tissue and lavage fluid were collected 24 h after the final exposure for transcriptomics and metabolomics. Increasing smoke exposure upregulated neutrophils and select cytokines in the bronchoalveolar lavage fluid. In total, 3446 genes were differentially expressed in the lungs of rats in the high smoke exposure and only 1 gene in the low smoke exposure (Cd151). Genes altered in the high smoke group reflected changes to the Eukaryotic Initiation Factor 2 stress and oxidative stress responses, which mirrored metabolomics analyses. xMWAS-integrated analysis revealed that smoke exposure significantly altered pathways associated with oxidative stress, lung morphogenesis, and tumor proliferation pathways. These results indicate that intermittent, 2-week exposure to eucalyptus wood smoke leads to transcriptomic and metabolic changes in the lung that may predict future lung disease development. Collectively, these findings provide insight into cellular signaling pathways that may contribute to the chronic pulmonary conditions observed in wildland firefighters.

Toxicology↗

DOE OSTI2017

Predicting Transport of 3,5,6-Trichloro-2-Pyridinol Into Saliva Using a Combination Experimental and Computational Approach

A combination experimental and computational approach was developed to predict chemical transport into saliva. A serous-acinar chemical transport assay was established to measure chemical transport with non-physiological (standard cell culture medium) and physiological (using surrogate plasma and saliva medium) conditions using 3,5,6-trichloro-2-pyridinol (TCPy) a metabolite of the pesticide chlorpyrifos. High levels of TCPy protein binding was observed in cell culture medium and rat plasma resulting in different TCPy transport behaviors in the two experimental conditions. In the non-physiological transport experiment, TCPy reached equilibrium at equivalent concentrations in apical and basolateral chambers. At higher TCPy doses, increased unbound TCPy was observed, and TCPy concentrations in apical and basolateral chambers reached equilibrium faster than lower doses, suggesting only unbound TCPy is able to cross the cellular monolayer. In the physiological experiment, TCPy transport was slower than non-physiological conditions, and equilibrium was achieved at different concentrations in apical and basolateral chambers at a comparable ratio (0.034) to what was previously measured in rats dosed with TCPy (saliva:blood ratio: 0.049). A cellular transport computational model was developed based on TCPy protein binding kinetics and accurately simulated all transport experiments using different permeability coefficients for the two experimental conditions (1.4 vs 0.4 cm/hr for non-physiological and physiological experiments, respectively). The computational model was integrated into a physiologically based pharmacokinetic (PBPK) model and accurately predicted TCPy concentrations in saliva of rats dosed with TCPy. Overall, this study demonstrates an approach to predict chemical transport in saliva potentially increasing the utility of salivary biomonitoring in the future.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

DOE OSTI2025

Variation in Flooding Tolerance in Populus deltoides ‘D-124’ and P. trichocarpa x P. deltoides Hybrid ‘52–225’

Flooding poses a substantial challenge to plant survival and productivity, particularly in riparian genus like Populus. This study examines the physiological, morphological, metabolic, and molecular responses of Populus deltoides ‘D-124’ and P. trichocarpa x P. deltoides hybrid clone ‘52–225’ under control and inundated conditions to identify differences in flooding tolerance. Under flooding conditions, physiological and cellular stress was more pronounced in P. deltoides ‘D-124’ than in the hybrid clone ‘52–225,’ as evidenced by lower transpiration (E), photosynthesis (A), and chlorophyll content. In contrast, ‘52–225’ showed reduced ROS accumulation suggesting better cellular function under stress. Morphologically, ‘52–225’ produced more shoot-born roots, which likely enhance oxygen transport and metabolic activity during flooding. Metabolite profiling revealed both overlapping and distinct patterns of sugar and amino acid accumulation between genotypes. Gene expression analysis revealed that flooding-responsive genes, including ALCOHOL DEHYDROGENASE 1 and HYPOXIA RESPONSIVE ERF 2, were activated in both genotypes, with a more pronounced response noted in ‘52–225.’ These findings extend our understanding of flooding tolerance mechanisms in Populus by connecting physiological traits, stress responses, and genetic regulation. This research contributes to the development of more flooding-resilient poplar varieties, with potential applications in breeding and restoration programs for flooding-prone environments.

Flooding stress↗

DOE OSTI2018

Comprehensive High Throughput One‐Pot Analysis of Glycoproteins through Tool for Rapid Analysis of Glycopeptide by Permethylation (TRAP) Method.

Glycoproteins plays key roles in various cellular functions such as protein folding, protein targeting, receptor function, cell–cell recognition, protein stability, signaling, and other cellular functions. Currently a comprehensive and universal tool for the automated analysis of glycopeptides through their tandem mass spectrometry and data processing is a tedious process. We recently developed a method, Tool for Rapid Analysis of glycopeptide by Permethylation (TRAP), that simplifies glycoprotein analysis by integrating glycan sequencing and glycopeptide analysis in a single experiment. We employed this approach for the detailed analysis of glycosidic linkages, composition, glycan sequencing, glycosylation site mapping and sequencing of peptide moiety of glycopeptides of glycoproteins from both prokaryotes and eukaryotes. Here we are presenting an easy to use, robust, suitable for large as well as small sample sets and also cost‐effective glycoprotein analysis tool through TRAP approach. A permethylation protocol which can be automated and suitable for reliable permethylation of glycopeptides from the protease digest of glycoproteins was developed. Initial studies on glycopeptides from a model glycoprotein bovine RNAse B indicates that improved sensitivity and throughput of permethylation reactions can be achieved in conducting the reaction in microscale. This approach would enable convenient, rapid, and reliable high‐throughput screening of protein glycosylation and can be employed as a cost‐effective kit for glycosylated biomarker discoveries. In order to improvise the comprehensive glycosylation analysis and throughput, we have also developed an LC‐MS/MS platform for the analysis of glycopeptides processed through TRAP. We have conducted the analysis of standard glycoproteins such as bovine RNase B, human transferrin, and bovine fetuin and our preliminary results indicate that TRAP hyphened with LC‐MS/MS enables characterization of isomers of each glycoform at each glycosylation sites of complex glycoprotein mixtures. Support or Funding Information This research was supported by the National Institutes of Health (NIH) funded research, Grant R21GM122633; resource for biomedical glycomics, Grant P41GM103490; instrument grant, Grant S10OD018530; and U.S. Department of Energy grant, Grant DE‐SC0015662, to the Complex Carbohydrate Research Center. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .

Shajahan, Asif↗

DOE OSTI2017

A bioarchitectonic approach to the modular engineering of metabolism

Dissociating the complexity of metabolic processes into modules is a shift in focus from the single gene/gene product to functional and evolutionary units spanning the scale of biological organization. When viewing the levels of biological organization through this conceptual lens, modules are found across the continuum: domains within proteins, co-regulated groups of functionally associated genes, operons, metabolic pathways and (sub)cellular compartments. Combining modules as components or subsystems of a larger system typically leads to increased complexity and the emergence of new functions. By virtue of their potential for 'plug and play' into new contexts, modules can be viewed as units of both evolution and engineering. Through consideration of lessons learned from recent efforts to install new metabolic modules into cells and the emerging understanding of the structure, function and assembly of protein-based organelles, bacterial microcompartments, a structural bioengineering approach is described: one that builds from an architectural vocabulary of protein domains. Further, this bioarchitectonic approach to engineering cellular metabolism can be applied to microbial cell factories, used in the programming of members of synthetic microbial communities or used to attain additional levels of metabolic organization in eukaryotic cells for increasing primary productivity and as the foundation of a green economy.This article is part of the themed issue 'Enhancing photosynthesis in crop plants: targets for improvement'.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

Paraburkholderia madseniana sp. nov., a phenolic acid-degrading bacterium isolated from acidic forest soil

RP11 T was isolated from forest soil following enrichment with 4-hydroxybenzoic acid. Cells of RP11 T are aerobic, non-sporulating, exhibit swimming motility, and are rods (0.8 µm by 1.4 µm) that often occur as diplobacillus or in short chains (3–4 cells). Optimal growth on minimal media containing 4-hydroxybenzoic acid (µ=0.216 hr -1 ) occurred at 30 °C, pH 6.5 or 7.0 and 0% salinity. Comparative chemotaxonomic, genomic and phylogenetic analyses revealed the isolate was distinct from its closest relative type strains identified as Paraburkholderia aspalathi LMG 27731 T , Paraburkholderia fungorum LMG 16225 T and Paraburkholderia caffeinilytica CF1 T . Strain RP11 T is genetically distinct from P. aspalathi , its closest relative, in terms of 16S rRNA gene sequence similarity (98.7%), genomic average nucleotide identity (94%) and in silico DNA–DNA hybridization (56.7%±2.8). The composition of fatty acids and substrate utilization pattern differentiated strain RP11 T from its closest relatives, including growth on phthalic acid. Strain RP11 T encoded the greatest number of aromatic degradation genes of all eleven closely related type strains and uniquely encoded a phthalic acid dioxygenase and paralog of the 3-hydroxybenzoate 4-monooxygenase. The only ubiquinone detected in strain RP11 T was Q-8, and the major cellular fatty acids were C 16:0 , 3OH-C 16:0 , C 17:0 cyclo, C 19:0 cyclo ω8c, and summed feature 8 (C 18:1 ω7c/ω6c). On the basis of this polyphasic approach, it was determined that strain RP11 T represents a novel species from the genus Paraburkholderia for which the name Paraburkholderia madseniana sp. nov. is proposed. The type strain is RP11 T (=DSM 110123 T =LMG 31517 T ).

Microbiology↗

DOE OSTI2020

Paraburkholderia solitsugae sp. nov. and Paraburkholderia elongata sp. nov., phenolic acid-degrading bacteria isolated from forest soil and emended description of Paraburkholderia madseniana

Two bacterial strains, 1N T and 5N T , were isolated from hemlock forest soil using a soluble organic matter enrichment. Cells of 1N T (0.65×1.85 µm) and 5N T (0.6×1.85 µm) are Gram-stain-negative, aerobic, motile, non-sporulating and exist as single rods, diplobacilli or in chains of varying length. During growth in dilute media (≤0.1× tryptic soy broth; TSB), cells are primarily motile with flagella. At higher concentrations (≥0.3× TSB), cells of both strains increasingly form non-motile chains, and cells of 5N T elongate (0.57×~7 µm) and form especially long filaments. Optimum growth of 1N T and 5N T occurred at 25–30 °C, pH 6.5–7.0 and <0.5% salinity. Results of comparative chemotaxonomic, genomic and phylogenetic analyses revealed that 1N T and 5N T were distinct from one another and their closest related type strains: Paraburkholderia madseniana RP11 T , Paraburkholderia aspalathi LMG 27731 T and Paraburkholderia caffeinilytica CF1 T . The genomes of 1N T and 5N T had an average nucleotide identity (91.6 and 91.3%) and in silico DNA–DNA hybridization values (45.8%±2.6 and 45.5%±2.5) and differed in functional gene content from their closest related type strains. The composition of fatty acids and patterns of substrate use, including the catabolism of phenolic acids, also differentiated strains 1N T and 5N T from each other and their closest relatives. The only ubiquinone present in strains 1N T and 5N T was Q-8. The major cellular fatty acids were C 16 : 0 , 3OH-C 16 : 0 , C 17 : 0 cyclo, C 19 : 0 cyclo ω8 c and summed features 2 (3OH-C 14 : 0 / C 16 : 1 iso I), 3 (C 16 : 1 ω6 c /ω7 c ) and 8 (C 18 : 1 ω7 c /ω6 c ). A third bacterium, strain RL16-012-BIC-B, was isolated from soil associated with shallow roots and was determined to be a strain of P. madseniana (ANI, 98.8%; 16S rRNA gene similarity, 100%). Characterizations of strain RL16-012-BIC-B (DSM 110723=LMG 31706) led to proposed emendments to the species description of P. madseniana . Our polyphasic approach demonstrated that strains 1N T and 5N T represent novel species from the genus Paraburkholderia for which the names Paraburkholderia solitsugae sp. nov. (type strain 1N T =DSM 110721 T =LMG 31704 T ) and Paraburkholderia elongata sp. nov. (type strain 5N T =DSM 110722 T =LMG 31705 T ) are proposed.

Microbiology↗

DOE OSTI2023

Sulfurospirillum diekertiae sp. nov., a tetrachloroethene-respiring bacterium isolated from contaminated soil

Two anaerobic, tetrachloroethene- (PCE-) respiring bacterial isolates, designated strain ACS DCE T and strain ACS TCE , were characterized using a polyphasic approach. Cells were Gram-stain-negative, motile, non-spore-forming and shared a vibrioid- to spirillum-shaped morphology. Optimum growth occurred at 30°C and 0.1–0.4% salinity. The pH range for growth was pH 5.5–7.5, with an optimum at pH 7.2. Hydrogen, formate, pyruvate and lactate as electron donors supported respiratory reductive dechlorination of PCE to cis-1,2-dichloroethene (cDCE) in strain ACS DCE T and of PCE to trichloroethene (TCE) in strain ACS TCE . Both strains were able to grow with pyruvate under microaerobic conditions. Nitrate, elemental sulphur, and thiosulphate were alternative electron acceptors. Autotrophic growth was not observed and acetate served as carbon source for both strains. The major cellular fatty acids were C 16:1 ω7c, C 16:0 , C 14:0 and C 18:1 ω7c. Both genomes feature a circular plasmid. Strains ACS DCE T and ACS TCE were previously assigned to the candidate species 'Sulfurospirillum acididehalogenans'. Here, based on key genomic features and pairwise comparisons of whole-genome sequences, including average nucleotide identity, digital DNA–DNA hybridization and average amino acid identity, strains ACS DCE T and ACS TCE , 'Ca. Sulfurospirillum diekertiae' strains SL2-1 and SL2-2, and the unclassified Sulfurospirillum sp. strain SPD-1 are grouped into one distinct species separate from previously described Sulfurospirillum species. Compared to Sulfurospirillum multivorans and Sulfurospirillum halorespirans, which dechlorinate PCE to cDCE without substantial TCE accumulation, these five strains produce TCE or cDCE as the end product. In addition, some cellular fatty acids (e.g., C 16:0 3OH, C 17:0 iso 3OH, C 17:0 2OH) were detected in strains ACS DCE T and ACS TCE but not in other Sulfurospirillum species. On the basis of phylogenetic, physiological and phenotypic characteristics, 'Ca. Sulfurospirillum acididehalogenans' and 'Ca. Sulfurospirillum diekertiae' are proposed to be merged into one novel species within the genus Sulfurospirillum, for which the name Sulfurospirillum diekertiae sp. nov. is proposed. Finally, the type strain is ACS DCE T (=JCM 33349 T = KCTC 15819 T =CGMCC 1.5292 T ).

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2019

Nonequilibrium physics in biology

Life is characterized by a myriad of complex dynamic processes allowing organisms to grow, reproduce, and evolve. Physical approaches for describing systems out of thermodynamic equilibrium have been increasingly applied to living systems, which often exhibit phenomena not found in those traditionally studied in physics. Spectacular advances in experimentation during the last decade or two, for example, in microscopy, single-cell dynamics, in the reconstruction of subcellular and multicellular systems outside of living organisms, and in high throughput data acquisition, have yielded an unprecedented wealth of data on cell dynamics, genetic regulation, and organismal development. These data have motivated the development and refinement of concepts and tools to dissect the physical mechanisms underlying biological processes. Notably, landscape and flux theory as well as active hydrodynamic gel theory have proven useful in this endeavor. Together with concepts and tools developed in other areas of nonequilibrium physics, significant progress has been made in unraveling the principles underlying efficient energy transport in photosynthesis, cellular regulatory networks, cellular movements and organization, embryonic development and cancer, neural network dynamics, population dynamics and ecology, as well as aging, immune responses, and evolution. In this paper, recent advances in nonequilibrium physics are reviewd and their application to biological systems is surveyed. Many of these results are expected to be important cornerstones as the field continues to build our understanding of life.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2019

Interaction and Regulation Between Lipid Mediator Phosphatidic Acid and Circadian Clock Regulators

Circadian clocks play important roles in regulating cellular metabolism, but the reciprocal effect that metabolism has on the clock is largely unknown in plants. Here we show that the central glycerolipid metabolite and lipid mediator phosphatidic acid (PA) interacts with and modulates the function of the core clock regulators LATE ELONGATED HYPOCOTYL (LHY) and CIRCADIAN CLOCK ASSOCIATED1 (CCA1) in Arabidopsis. PA reduced the ability of LHY and CCA1 to bind the promoter of their target gene TIMING OF CAB EXPRESSION1. Increased PA accumulation and inhibition of PA-producing enzymes had opposite effects on circadian clock outputs. Diurnal change in levels of several membrane phospholipid species, including PA, observed in wild-type was lost in the LHY and CCA1 double knockout mutant. Storage lipid accumulation was also affected in the clock mutants. These results indicate that the interaction of PA with the clock regulator may function as a cellular conduit to integrate the circadian clock with lipid metabolism.

bio-inspired↗

DOE OSTI2020

Phase-contrast 3D tomography of HeLa cells grown in PLLA polymer electrospun scaffolds using synchrotron X-rays

Advanced imaging is useful for understanding the three-dimensional (3D) growth of cells. X-ray tomography serves as a powerful noninvasive, nondestructive technique that can fulfill these purposes by providing information about cell growth within 3D platforms. There are a limited number of studies taking advantage of synchrotron X-rays, which provides a large field of view and suitable resolution to image cells within specific biomaterials. In this study, X-ray synchrotron radiation microtomography at Diamond Light Source and advanced image processing were used to investigate cellular infiltration of HeLa cells within poly L-lactide (PLLA) scaffolds. This study demonstrates that synchrotron X-rays using phase contrast is a useful method to understand the 3D growth of cells in PLLA electrospun scaffolds. Two different fiber diameter (2 and 4 µm) scaffolds with different pore sizes, grown over 2, 5 and 8 days in vitro , were examined for infiltration and cell connectivity. After performing visualization by segmentation of the cells from the fibers, the results clearly show deeper cell growth and higher cellular interconnectivity in the 4 µm fiber diameter scaffold. Finally, this indicates the potential for using such 3D technology to study cell–scaffold interactions for future medical use.

3D culture↗

DOE OSTI2022

High Performance Adaptive Physics Refinement to Enable Large-Scale Tracking of Cancer Cell Trajectory

The ability to track simulated cancer cells through the circulatory system, important for developing a mechanistic understanding of metastatic spread, pushes the limits of today's supercomputers by requiring the simulation of large fluid volumes at cellular-scale resolution. To overcome this challenge, we introduce a new adaptive physics refinement (APR) method that captures cellular-scale interaction across large domains and leverages a hybrid CPU-GPU approach to maximize performance. Through algorithmic advances that integrate multi-physics and multi-resolution models, we establish a finely resolved window with explicitly modeled cells coupled to a coarsely resolved bulk fluid domain. In this work we present multiple validations of the APR framework by comparing against fully resolved fluid-structure interaction methods and employ techniques, such as latency hiding and maximizing memory bandwidth, to effectively utilize heterogeneous node architectures. Collectively, these computational developments and performance optimizations provide a robust and scalable framework to enable system-level simulations of cancer cell transport.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Precoder Design for Physical-Layer Security and Authentication in Massive MIMO UAV Communications

Supporting reliable and seamless wireless connectivity for unmanned aerial vehicles (UAVs) has recently become a critical requirement to enable various different use cases of UAVs. Due to their widespread deployment footprint, cellular networks can support beyond visual line of sight (BVLOS) communications for UAVs. In this paper, we consider cellular connected UAVs (C-UAVs) that are served by massive multiple input-multiple-output (MIMO) links to extend coverage range, while also improving physical layer security and authentication. Here, we consider Rician channel and propose a novel linear precoder design for transmitting data and artificial noise (AN). We derive the closed-form expression of the ergodic secrecy rate of CUAVs for both conventional and proposed precoder designs. In addition, we obtain the optimal power splitting factor that divides the power between data and AN by asymptotic analysis. Then, we apply the proposed precoder design in the fingerprint embedding authentication framework, where the goal is to minimize the probability of detection of the authentication tag at an eavesdropper. In simulation results, we show the superiority of the proposed precoder in both secrecy rate and the authentication probability considering moderate and large number of antenna massive MIMO scenarios.

99 GENERAL AND MISCELLANEOUS↗