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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 127 records · Page 7
All sourcesNASA NTRSDOE OSTI
DOE OSTI2025

Developing a media formulation to sustain ex vivo chloroplast function

Chloroplasts are critical organelles in plants and algae responsible for accumulating biomass through photosynthetic carbon fixation and cellular maintenance through metabolism in the cell. Chloroplasts are increasingly appreciated for their role in biomanufacturing, as they can produce many useful molecules, and a deeper understanding of chloroplast regulation and function would provide more insight for the biotechnological applications of these organelles. However, traditional genetic approaches to manipulate chloroplasts are slow, and generation of transgenic organisms to study their function can take weeks to months, significantly delaying the pace of research. To develop chloroplasts themselves as a quicker and more defined platform, we isolated chloroplasts from the green algae, Chlamydomonas reinhardtii, and examined their photosynthetic function after extraction. Combined with a metabolic modeling approach using flux-balance analysis, we identified key metabolic reactions essential to chloroplast function and leveraged this information into reagents that can be used in a “chloroplast media” capable of maintaining chloroplast photosynthetic function over time ex vivo compared to buffer alone. We envision this could serve as a model platform to enable more rapid design-build-test-learn cycles to study and improve chloroplast function in combination with genetic modifications and potentially as a starting point for the bottom-up design of a synthetic organelle-containing cell.

Chlamydomonas reinhardtii↗
DOE OSTI2024

Genomic surveillance indicates clonal replacement of hypervirulent Klebsiella pneumoniae ST881 and ST29 lineage strains in vivo

The emergence of hypervirulent Klebsiella pneumoniae (hvKp) poses a significant public health threat, particularly regarding its carriage in the healthy population. However, the genomic epidemiological characteristics and population dynamics of hvKp within a single patient across distinct infection episodes remain largely unknown. This study aimed to investigate the clonal replacement of hvKp K2-ST881 and K54-ST29 lineage strains in a single patient experiencing multiple-site infections during two independent episodes. Two strains, designated EDhvKp-1 and EDhvKp-2, were obtained from blood and cerebrospinal fluid during the first admission, and the strain isolated from blood on the second admission was named EDhvKp-3. Whole-genome sequencing, utilizing both short-read Illumina and long-read Oxford Nanopore platforms, was conducted. In silico multilocus sequence typing (MLST), identification of antimicrobial resistance and virulence genes, and the phylogenetic relationship between our strains and other K. pneumoniae ST881 and ST29 genomes retrieved from the public database were performed. Virulence potentials were assessed through a mouse lethality assay. Our study indicated that the strains were highly susceptible to multiple antimicrobial agents. Plasmid sequence analysis confirmed that both virulence plasmids, pEDhvKp-1 (166,008 bp) and pEDhvKp-3 (210,948 bp), belonged to IncFIB type. Multiple virulence genes, including rmpA , rmpA2 , rmpC , rmpD , iroBCDN , iucABCD , and iutA , were identified. EDhvKp-1 and EDhvKp-2 showed the closest relationship to strain 502 (differing by 51 SNPs), while EDhvKp-3 exhibited 69 SNPs differences compared to strain TAKPN-1, which all recovered from Chinese patients in 2020. In the mouse infection experiment, both ST881 EDhvKp-1 and ST29 EDhvKp-3 displayed similar virulence traits, causing 90 and 100% of the mice to die within 72 h after intraperitoneal infection, respectively. Our study expands the spectrum of hvKp lineages and highlights genomic alterations associated with clonal switching between two distinct lineages of hvKP that successively replaced each other in vivo . The development of novel strategies for the surveillance, diagnosis, and treatment of high-risk hvKp is urgently needed.

Liu, Ning↗
DOE OSTI2024

Activity of Cannabidiol on Ex Vivo Amino Acid Fermentation by Bovine Rumen Microbiota

Amino-acid-fermenting bacteria are wasteful organisms within the rumens of beef cattle that remove dietary amino nitrogen by producing ammonia, which is then excreted renally. There are currently no on-label uses for the control of this microbial guild, but off-label use of broad-spectrum antimicrobials has shown efficacy, which contributes to antimicrobial resistance. Plant-derived antimicrobials supplemented into the diets of cattle may offer worthwhile alternatives. This study sought to investigate the role of cannabidiol (CBD) as a terpenophenolic antimicrobial. Ex vivo cell suspensions were harvested from the rumen fluid of Angus × Holstein steers in non-selective media with amino acid substrates. The suspensions were treated with five concentrations of CBD (860 μg mL−1–0.086 μg mL−1) and incubated (24 h), after which ammonia production and viable number of cells per substrate and treatment were measured. The data demonstrated a ~10–15 mM reduction in ammonia produced at the highest concentration of CBD and negligible changes in the viable number of amino-acid-fermenting bacteria. CBD does not appear to be a biologically or economically viable terpenophenolic candidate for the control of amino acid fermentation in beef cattle.

Biotechnology & Applied Microbiology↗
DOE OSTI2020

PARP Theranostic Auger Emitters Are Cytotoxic in BRCA Mutant Ovarian Cancer and Viable Tumors from Ovarian Cancer Patients Enable Ex-Vivo Screening of Tumor Response

Theranostics are emerging as a pillar of cancer therapy that enable the use of single molecule constructs for diagnostic and therapeutic application. As poly adenosine diphosphate (ADP)-ribose polymerase 1 (PARP-1) is overexpressed in various cancer types, and is localized to the nucleus, PARP-1 can be safely targeted with Auger emitters to induce DNA damage in tumors. Here, we investigated a radioiodinated PARP inhibitor, [125I]KX1, and show drug target specific DNA damage and subsequent killing of BRCA1 and non-BRCA mutant ovarian cancer cells at sub-pharmacological concentrations several orders of magnitude lower than traditional PARP inhibitors. Furthermore, we demonstrated that viable tumor tissue from ovarian cancer patients can be used to screen tumor radiosensitivity ex-vivo, enabling the direct assessment of therapeutic efficacy. Finally, we showed tumors can be imaged by single-photon computed tomography (SPECT) with PARP theranostic, [123I]KX1, in a human ovarian cancer xenograft mouse model. These data support the utility of PARP-1 targeted radiopharmaceutical therapy as a theranostic option for PARP-1 overexpressing ovarian cancers.

BRCA1↗
DOE OSTI2021

Coadministration of a Clinically Relevant Dexamethasone Dosage With Ablative Radiation Therapy Reduces Peripheral Lymphocytes But Does Not Alter In Vivo Intratumoral Lymphocyte Phenotype or Inhibit Efficacy of Radiation Therapy in a Murine Colorectal Tumor Model

Dexamethasone is commonly given during radiation therapy (RT) to manage toxicities. Our study examines if dexamethasone coadministration with RT inhibits the RT-induced antitumor T cell response in mouse.

62 RADIOLOGY AND NUCLEAR MEDICINE↗
DOE OSTI2021

In vivo visualization of butterfly scale cell morphogenesis in Vanessa cardui

Many organisms exhibit functional micro- and nanoscale materials with structural definition and performance that challenge synthetic fabrication techniques, yet we know little about the processes that enable their formation. Using butterfly scales as a model system for functional biomaterials, we establish a timeline of scale formation and quantify relevant structural parameters for developing painted lady butterflies. We overcome challenges of previous efforts by imaging structure formation directly in living organisms, which allows us to continuously observe the evolving wing tissue and the fine details of individual scale cells. Visualization of scale structure formation in live butterflies forms the basis for modeling the underlying biomechanical processes and opens avenues for their translation into advanced fabrication strategies.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Enhanced X-ray diffraction of in vivo-grown μNS crystals by viscous jets at XFELs

μNS is a 70 kDa major nonstructural protein of avian reoviruses, which cause significant economic losses in the poultry industry. They replicate inside viral factories in host cells, and the μNS protein has been suggested to be the minimal viral factor required for factory formation. Thus, determining the structure of μNS is of great importance for understanding its role in viral infection. In the study presented here, a fragment consisting of residues 448–605 of μNS was expressed as an EGFP fusion protein in Sf9 insect cells. EGFP-μNS (448–605) crystallization in Sf9 cells was monitored and verified by several imaging techniques. Cells infected with the EGFP-μNS (448–605) baculovirus formed rod-shaped microcrystals (5–15 µm in length) which were reconstituted in high-viscosity media (LCP and agarose) and investigated by serial femtosecond X-ray diffraction using viscous jets at an X-ray free-electron laser (XFEL). The crystals diffracted to 4.5 Å resolution. A total of 4227 diffraction snapshots were successfully indexed into a hexagonal lattice with unit-cell parameters a = 109.29, b = 110.29, c = 324.97 Å. The final data set was merged and refined to 7.0 Å resolution. Preliminary electron-density maps were obtained. While more diffraction data are required to solve the structure of μNS (448–605) , the current experimental strategy, which couples high-viscosity crystal delivery at an XFEL with in cellulo crystallization, paves the way towards structure determination of the μNS protein.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Shortwave‐Infrared‐Emitting Nanoprobes for CD8 Targeting and In Vivo Imaging of Cytotoxic T Cells in Breast Cancer

Checkpoint immunotherapy has made great strides in the treatment of solid tumors, but many patients do not respond to immune checkpoint inhibitors. Identification of tumor‐infiltrating cytotoxic T cells (CTLs) has the potential to stratify patients and monitor immunotherapy responses. In this study, the design of cluster of differentiation (CD8 + ) T cell‐targeted nanoprobes that emit shortwave infrared (SWIR) light in the second tissue‐transparent window for noninvasive, real‐time imaging of CTLs in murine models of breast cancer is presented. SWIR‐emitting rare‐earth nanoparticles encapsulated in human serum albumin are conjugated with anti‐CD8α to target CTLs with high specificity. CTL targeting is validated in vitro through binding of nanoprobes to primary mouse CTLs. The potential for the use of SWIR fluorescence intensity to determine CTL presence is validated in two syngeneic mammary fat pad tumor models, EMT6 and 4T1, which differ in immune infiltration. SWIR imaging using CD8‐targeted nanoprobes successfully identifies the presence of CTLs in the more immunogenic EMT6 model, while imaging confirms the lack of substantial immune infiltration in the nonimmunogenic 4T1 model. In this work, the opportunity for SWIR imaging using CD8‐targeted nanoprobes to assess CTL infiltration in tumors for the stratification and monitoring of responders to checkpoint immunotherapy is highlighted.

Shah, Jay V.↗
DOE OSTI2024

Enhancing 2‐Pyrone Synthase Efficiency by High‐Throughput Mass‐Spectrometric Quantification and In Vitro/In Vivo Catalytic Performance Correlation

Abstract Engineering efficient biocatalysts is essential for metabolic engineering to produce valuable bioproducts from renewable resources. However, due to the complexity of cellular metabolic networks, it is challenging to translate success in vitro into high performance in cells. To meet such a challenge, an accurate and efficient quantification method is necessary to screen a large set of mutants from complex cell culture and a careful correlation between the catalysis parameters in vitro and performance in cells is required. In this study, we employed a mass‐spectrometry based high‐throughput quantitative method to screen new mutants of 2‐pyrone synthase (2PS) for triacetic acid lactone (TAL) biosynthesis through directed evolution in E. coli . From the process, we discovered two mutants with the highest improvement (46 fold) in titer and the fastest k cat (44 fold) over the wild type 2PS, respectively, among those reported in the literature. A careful examination of the correlation between intracellular substrate concentration, Michaelis‐Menten parameters and TAL titer for these two mutants reveals that a fast reaction rate under limiting intracellular substrate concentrations is important for in‐cell biocatalysis. Such properties can be tuned by protein engineering and synthetic biology to adopt these engineered proteins for the maximum activities in different intracellular environments.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Construction of the Bioconjugate Py‐Macrodipa‐PSMA and Its In Vivo Investigations with Large 132/135 La 3+ and Small 47 Sc 3+ Radiometal Ions

To harness radiometals in clinical settings, a chelator forming a stable complex with the metal of interest and targets the desired pathological site is needed. Toward this goal, we previously reported a unique set of chelators that can stably bind to both large and small metal ions, via a conformational switch. Within this chelator class, py-macrodipa is particularly promising based on its ability to stably bind several medicinally valuable radiometals including large 132/135 La 3+ , 213 Bi 3+ , and small 44 Sc 3+ . Here, in this study, we report a 10-step organic synthesis of its bifunctional analogue py-macrodipa-NCS, which contains an amine-reactive −NCS group that is amenable for bioconjugation reactions to targeting vectors. The hydrolytic stability of py-macordipa-NCS was assessed, revealing a half-life of 6.0 d in pH 9.0 aqueous buffer. This bifunctional chelator was then conjugated to a prostate-specific membrane antigen (PSMA)-binding moiety, yielding the bioconjugate py-macrodipa-PSMA, which was subsequently radiolabeled with large 132/135 La 3+ and small 47 Sc 3+ , revealing efficient and quantitative complex formation. The resulting radiocomplexes were injected into mice bearing both PSMA-expressing and PSMA-non-expressing tumor xenografts to determine their biodistribution patterns, revealing delivery of both 132/135 La 3+ and 47 Sc 3+ to PSMA+ tumor sites. However, partial radiometal dissociation was observed, suggesting that py-macrodipa-PSMA needs further structural optimization.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗