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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 145 records · Page 8
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Optimization of Application-Driven Development of In Vitro Neuromuscular Junction Models

Neuromuscular junctions (NMJs) are specialized synapses responsible for signal transduction between motor neurons (MNs) and skeletal muscle tissue. Malfunction at this site can result from developmental disorders, toxic environmental exposures, and neurodegenerative diseases leading to severe neurological dysfunction. Exploring these conditions in human or animal subjects is restricted by ethical concerns and confounding environmental factors. Therefore, in vitro NMJ models provide exciting opportunities for advancements in tissue engineering. In the last two decades, multiple NMJ prototypes and platforms have been reported, and each model system design is strongly tied to a specific application: exploring developmental physiology, disease modeling, or high-throughput screening. Directing the differentiation of stem cells into mature MNs and/or skeletal muscle for NMJ modeling has provided critical cues to recapitulate early-stage development. Patient-derived inducible pluripotent stem cells provide a personalized approach to investigating NMJ disease, especially when disease etiology cannot be resolved down to a specific gene mutation. Having reproducible NMJ culture replicates is useful for high-throughput screening to evaluate drug toxicity and determine the impact of environmental threat exposures. Cutting-edge bioengineering techniques have propelled this field forward with innovative microfabrication and design approaches allowing both two-dimensional and three-dimensional NMJ culture models. Many of these NMJ systems require further validation for broader application by regulatory agencies, pharmaceutical companies, and the general research community. In this summary, we present a comprehensive review on the current state-of-art research in NMJ models and discuss their ability to provide valuable insight into cell and tissue interactions.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS2003

In Vitro Studies on Space Radiation-Induced Delayed Genetic Responses: Shielding Effects

Understanding the radiation risks involved in spaceflight is of considerable importance, especially with the long-term occupation of ISS and the planned crewed exploration missions. Several independent causes may contribute to the overall risk to astronauts exposed to the complex space environment, such as exposure to GCR as well as SPES. Protons and high-Z energetic particles comprise the GCR spectrum and may exert considerable biological effects even at low fluence. There are also considerable uncertainties associated with secondary particle effects (e.g. HZE fragments, neutrons etc.). The interaction of protons and high-LET particles with biological materials at all levels of biological organization needs to be investigated fully in order to establish a scientific basis for risk assessment. The results of these types of investigation will foster the development of appropriately directed countermeasures. In this study, we compared the biological responses to proton irradiation presented to the target cells as a monoenergetic beam of particles of complex composition delivered to cells outside or inside a tissue phantom head placed in the United States EVA space suit helmet. Measurements of chromosome aberrations, apoptosis, and the induction of key proteins were made in bone marrow from CBA/CaJ and C57BL/6 mice at early and late times post exposure to radiation at 0, 0.5, 1 and 2 Gy while inside or outside of the helmet. The data showed that proton irradiation induced transmissible chromosomal/genomic instability in haematopoietic stem cells in both strains of mice under both irradiation conditions and especially at low doses. Although differences were noted between the mouse strains in the degree and kinetics of transforming growth factor-beta 1 and tumor necrosis factor-alpha secretion, there were no significant differences observed in the level of the induced instability under either radiation condition, or for both strains of mice. Consequently, when normalized to physical dose, the monoenergetic proton field present inside the helmet-protected phantom produced equivalent biological responses, when compared to unshielded cells, as measured by the induction of delayed genetic effects in murine haematopoietic stem cells.

Kadhim, Munira A.↗
NASA NTRS2019

Deciphering the Effects of Microgravity Cell by Cell

Forces generated by gravity have a profound impact on the behavior of cells in tissues affecting the course of the cell cycle and differentiation fate of progenitors in mammalian tissues. These cells are contributing to normal tissue regenerative health and defence against disease. In Human space exploration context, it is extremely important to determine spaceflight provoked changes in tissue's regenerational capabilities. Microgravity experienced during spaceflight causes unloading and mechanical disuse on all orthostatic support tissues, therefore impacting stem cell fate and lineage commitment decisions. Investigating how ESCs respond to mechanical stimulation is a platform for fundamental developmental and regeneration research applicable for spaceflight. However, the gene expression programs associatiated with early committment stem cell pathways in response to physical stimulation are not readily known. Single-cell RNA-seq technologies have recently revolutionized the world of molecular biology by providing the capability to assess gene expression pattern within a single cell. Our method isolates and separately barcodes mRNAs from thousands of single cells and sequences their expressomes. Understanding regenerative processes on a molecular level would not only help reduce long-term spaceflight impact on health, but also may enable the development of novel tissue regenerative approaches to tissue degeneration on Earth.

Single cell sequencing↗
DOE OSTI2022

Deep Learning‐Assisted Automated Single Cell Electroporation Platform for Effective Genetic Manipulation of Hard‐to‐Transfect Cells

Abstract Genome engineering of cells using CRISPR/Cas systems has opened new avenues for pharmacological screening and investigating the molecular mechanisms of disease. A critical step in many such studies is the intracellular delivery of the gene editing machinery and the subsequent manipulation of cells. However, these workflows often involve processes such as bulk electroporation for intracellular delivery and fluorescence activated cell sorting for cell isolation that can be harsh to sensitive cell types such as human‐induced pluripotent stem cells (hiPSCs). This often leads to poor viability and low overall efficacy, requiring the use of large starting samples. In this work, a fully automated version of the nanofountain probe electroporation (NFP‐E) system, a nanopipette‐based single‐cell electroporation method is presented that provides superior cell viability and efficiency compared to traditional methods. The automated system utilizes a deep convolutional network to identify cell locations and a cell‐nanopipette contact algorithm to position the nanopipette over each cell for the application of electroporation pulses. The automated NFP‐E is combined with microconfinement arrays for cell isolation to demonstrate a workflow that can be used for CRISPR/Cas9 gene editing and cell tracking with potential applications in screening studies and isogenic cell line generation.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Human myoblasts differentiate in various mesenchymal lineages and inhibit allogeneic T cell proliferation through an indolamine 2,3 dioxygenase dependent pathway

Muscle stem cells (MuSC) are considered as a reliable source of therapeutic cells to restore diseased muscles. However in most cases, injected MuSC-derived myoblasts are rapidly destroyed by the host immune response, which impairs the beneficial effect. By contrast, human mesenchymal stromal cells (MSC), have been reported to exhibit potent immune regulatory functions. Thus, we investigated, in vitro, the multipotent differentiation- and immunosuppressive capacities of human myoblasts and compared these features with those of human MSC. Myoblasts shared numerous cell surface markers with MSC, including CD73, CD90, CD105 and CD146. Both cell type were negative for HLA-DR and CD45, CD34 and CD31. CD56, a myogenic marker, was expressed by myoblasts exclusively. Myoblasts displayed multipotent potential capabilities with differentiation in chondrocytes, adipocytes and osteoblasts in vitro. Myoblasts also inhibited allogenic T cell proliferation in vitro in a dose dependent manner, very similarly to MSC. This effect was partly mediated via the activation of indolamine 2,3 dioxygenase enzyme (IDO) after IFNγ exposure. Altogether, these data demonstrate that human myoblasts can differentiate in various mesenchymal linages and exhibit powerful immunosuppressive properties in vitro. Such features may open new therapeutic strategies for MuSC-derived myoblasts.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

A conserved molecular logic for neurogenesis to gliogenesis switch in the cerebral cortex

During development, neural stem cells in the cerebral cortex, also known as radial glial cells (RGCs), generate excitatory neurons, followed by production of cortical macroglia and inhibitory neurons that migrate to the olfactory bulb (OB). Understanding the mechanisms for this lineage switch is fundamental for unraveling how proper numbers of diverse neuronal and glial cell types are controlled. We and others recently showed that Sonic Hedgehog (Shh) signaling promotes the cortical RGC lineage switch to generate cortical oligodendrocytes and OB interneurons. During this process, cortical RGCs generate intermediate progenitor cells that express critical gliogenesis genes Ascl1, Egfr, and Olig2. The increased Ascl1 expression and appearance of Egfr + and Olig2 + cortical progenitors are concurrent with the switch from excitatory neurogenesis to gliogenesis and OB interneuron neurogenesis in the cortex. While Shh signaling promotes Olig2 expression in the developing spinal cord, the exact mechanism for this transcriptional regulation is not known. Furthermore, the transcriptional regulation of Olig2 and Egfr has not been explored. Here, we show that in cortical progenitor cells, multiple regulatory programs, including Pax6 and Gli3, prevent precocious expression of Olig2, a gene essential for production of cortical oligodendrocytes and astrocytes. We identify multiple enhancers that control Olig2 expression in cortical progenitors and show that the mechanisms for regulating Olig2 expression are conserved between the mouse and human. Our study reveals evolutionarily conserved regulatory logic controlling the lineage switch of cortical neural stem cells.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

O-GlcNAc transferase Ogt regulates embryonic neuronal development through modulating Wnt/β-catenin signaling

Abstract Ogt-mediated O-GlcNAcylation is enriched in the nervous system and involves in neuronal development, brain function and neurological diseases. However, the roles of Ogt and O-GlcNAcylation in embryonic neurogenesis have remained largely unknown. Here, we show that Ogt is highly expressed in embryonic brain, and Ogt depletion reduces the proliferation of embryonic neural stem cells and migration of new born neurons. Ogt depletion in cultured hippocampal neurons impairs neuronal maturation, including reduced dendritic numbers and length, and immature development of spines. Mechanistically, Ogt depletion decreases the activity of Wnt/β-catenin signaling. Ectopic β-catenin rescues neuronal developmental deficits caused by Ogt depletion. Ogt also regulates human cortical neurogenesis in forebrain organoids derived from induced pluripotent stem cells. Our findings reveal the essential roles and mechanisms of Ogt-mediated O-GlcNAc modification in regulating mammalian neuronal development.

Shen, Hui↗
DOE OSTI2023

Intra- and inter-molecular regulation by intrinsically-disordered regions governs PUF protein RNA binding

PUF proteins are characterized by globular RNA-binding domains. They also interact with partner proteins that modulate their RNA-binding activities. Caenorhabditis elegans PUF protein fem-3 binding factor-2 (FBF-2) partners with intrinsically disordered Lateral Signaling Target-1 (LST-1) to regulate target mRNAs in germline stem cells. Here, we report that an intrinsically disordered region (IDR) at the C-terminus of FBF-2 autoinhibits its RNA-binding affinity by increasing the off rate for RNA binding. Moreover, the FBF-2 C-terminal region interacts with its globular RNA-binding domain at the same site where LST-1 binds. This intramolecular interaction restrains an electronegative cluster of amino acid residues near the 5' end of the bound RNA to inhibit RNA binding. LST-1 binding in place of the FBF-2 C-terminus therefore releases autoinhibition and increases RNA-binding affinity. This regulatory mechanism, driven by IDRs, provides a biochemical and biophysical explanation for the interdependence of FBF-2 and LST-1 in germline stem cell self-renewal.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Structural maturation of myofilaments in engineered 3D cardiac microtissues characterized using small angle x-ray scattering

Abstract Understanding the structural and functional development of human-induced pluripotent stem-cell-derived cardiomyocytes (hiPSC-CMs) is essential to engineering cardiac tissue that enables pharmaceutical testing, modeling diseases, and designing therapies. Here we use a method not commonly applied to biological materials, small angle x-ray scattering, to characterize the structural development of hiPSC-CMs within three-dimensional engineered tissues during their preliminary stages of maturation. An x-ray scattering experimental method enables the reliable characterization of the cardiomyocyte myofilament spacing with maturation time. The myofilament lattice spacing monotonically decreases as the tissue matures from its initial post-seeding state over the span of 10 days. Visualization of the spacing at a grid of positions in the tissue provides an approach to characterizing the maturation and organization of cardiomyocyte myofilaments and has the potential to help elucidate mechanisms of pathophysiology, and disease progression, thereby stimulating new biological hypotheses in stem cell engineering.

36 MATERIALS SCIENCE↗
NASA NTRS1983

Late skin damage in rabbits and monkeys after exposure to particulate radiations

Preliminary results are reported of experiments on the late effects of exposure to particulate radiations on stem cell populations. Skin biopsies were taken from the ears of rabbits irradiated 2-5 years previously with 530 MeV/amu Ar ions (LET 90 keV/micron), or 365 MeV/amu Ne ions (LET 35 keV micron), and from the chests and inner thighs of rhesus monkeys irradiated 16-18 years previously with 32-MeV protons (LET about 1.2 keV/micron). Skin fibroblast cultures obtained from the biopsy samples in rabbits were observed to undergo dose-dependent decreases in in vitro life span, with estimated survival curves showing the effects of Ar-ion irradiation to be more severe than those of Ne-ion irradiation. In addition, the healing of the biopsy wound was observed to become slower as radiation dose increased. In the monkey, radiation reduced the average number of fibroblasts at the time of cessation of growth in culture. Results thus demonstrate the capacity of skin sampling to reveal stem cell destruction, and have important implications for astronauts and other persons at risk of particle exposure with regard to healing responses to trauma or surgery.

Bergtold, D. S.↗
NASA NTRS2016

The Effects of Spaceflight on Cellular Aging in Saccharomyces cerevisiae

The conditions encountered during spaceflight place unique stresses on physiological processes that oftentimes lead to deleterious effects. Identifying these effects and better understanding their molecular mechanisms will be essential in enabling long-duration space travel by humans. Studies in Saccharomyces cerevisiae suggest an aging model that involves the accumulation of toxic components, such as excess extrachromosomal rDNA and damaged mitochondria. This build-up then limits the replicative lifespan (the number of times a mother cell can form a new daughter cell). Remarkably, each new daughter cell emerges completely renewed from the senescing mother cell through an asymmetric distribution of aging determinants via mechanisms that are intricately linked to the budding process. When exposed to simulated microgravity, S. cerevisiae undergoes an altered budding process characterized by a breakdown in bud scar polarity. Because the budding process is critical to replicative aging, we hypothesize that the replicative lifespan may be affected by microgravity as well. To measure relative replicative aging rates, we will construct a strain of yeast in which daughter cells are inviable. In this strain, the Cre recombinase will be expressed under the control of the daughter cell specific promoter, pSCW11, and LoxP sites will be inserted at both flanks of two essential genes involved in the cell cycle, UBC9 and CDC20, using a CRISPRCas9 system. Thus, UBC9 and CDC20 will be excised from daughter cells, leading to cell-cycle arrest and eventual death. To mimic the low shear conditions encountered in microgravity, this strain will be grown in rotating wall vessels. The number of viable mother cells will be monitored over time, and this rate will be compared to cells growing in standard conditions. Because asymmetric division also occurs in mammalian cells (e.g. in neural stem cells), this study will provide insight into how cellular aging rates may change in mammals and will help empower humans to thrive in space for extended and even indefinite periods of time.

aging↗
DOE OSTI2021

Tunicamycin-induced endoplasmic reticulum stress inhibits chemoresistance of FaDu hypopharyngeal carcinoma cells in 3D collagen I cultures and in vivo

Highlights: • 3D collagen I culture induces chemoresistance of FaDu hypopharyngeal carcinoma cells. • Tunicamycin-induced ER stress inhibits chemoresistance in collagen I cultures. • Tunicamycin-induced ER stress potentiates anticancer efficacy in vivo. • Tunicamycin-induced ER stress inhibits TGF-β1 and modifies integrin β1 glycosylation. The prognosis in patients with advanced head and neck squamous cell carcinoma (HNSCC) is widely affected by the resistance to chemotherapy. As a culture scaffold, collagen I was showed to promote CSC (cancer stem cell) properties of cancer cells which could be used as in vitro models to study the chemoresistance in HNSCC. Endoplasmic reticulum (ER) stress is a cellular stress condition which could affect tumor progression and promote the anti-tumor effects of certain drugs. However, the impact of ER stress on collagen I induced CSC properties and chemoresistance of HNSCC cells has not been addressed. In this study we investigated the effects of tunicamycin (TM) induced ER stress on the stemness and sensitivity to chemotherapeutic drugs of FaDu hypopharyngeal carcinoma cells in 3D (three-dimensional) collagen I cultures and mouse xenograft models. Our study revealed that Collagen I scaffold promoted CSC properties and increased G1 population of FaDu cells in 3D cultures, accompanied by maturation of integrin β1 and enhanced activated TGF-β1 concentration. Compared to 2D (two-dimensional) cultured cells, cells in 3D Collagen I scaffold exhibited significantly increased resistance to chemotherapeutic drugs of cisplatin and paclitaxel. Further analysis revealed that TM induced ER stress preferentially attenuated chemoresistance of FaDu cells in 3D collagen I, downregulated their CSC properties and TGF-β1 concentration and resulted in deglycosylation of integrin β1. TM was further evaluated in the mouse xenograft models and showed significant tumor growth inhibition in combination with paclitaxel than either TM or paclitaxel alone. Taken together, Our findings suggest that TM-induced ER stress potentiates anticancer efficacy of FaDu cells in 3D cultures and in vivo, and highlight implications for targeting chemotherapy-resistant cancer stem cells under ER stress conditions.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Drosophila intestinal homeostasis requires CTP synthase

CTP synthase (CTPS) senses all four nucleotides and forms filamentous structures termed cytoophidia in all three domains of life. How CTPS and cytoophidia function in a developmental context, however, remains underexplored. We report that CTPS forms cytoophidia in a subset of cells in the Drosophila midgut. We found that cytoophidia exist in intestinal stem cells (ISC) and enteroblasts in similar proportions. Both refeeding after starvation and feeding with dextran sulfate sodium (DSS) induce ISC proliferation and elongate cytoophidia. Knockdown of CTPS inhibits ISC proliferation. Remarkably, disruption of CTPS cytoophidia inhibits DSS-induced ISC proliferation. Taken together, these data suggest that both the expression level and the filament-form property of CTPS are crucial for intestinal homeostasis in Drosophila.

60 APPLIED LIFE SCIENCES↗
NASA NTRS2014

High-Density Spot Seeding for Tissue Model Formation

A method for making a tissue includes seeding cells at a selected concentration on a support to form a cell spot, incubating the cells to allow the cells to partially attach, rinsing the cells to remove any unattached cells, adding culture medium to enable the cells to proliferate at a periphery of the cell spot and to differentiate toward a center of the cell spot, and further incubating the cells to form the tissue. The cells may be C2C12 cells or other subclones of the C2 cell line, H9c2(2-1) cells, L6 cells, L8 cells, QM7 cells, Sol8 cells, G-7 cells, G-8 cells, other myoblast cells, cells from other tissues, or stem cells. The selected concentration is in a range from about 1 x 10(exp 5) cells/ml to about 1 x 10(exp 6) cells/ml. The tissue formed may be a skeletal muscle tissue, a cardiac muscle tissue, nerve tissue, or a bone tissue.

Marquette, Michele L.↗
NASA NTRS2013

Lab-On-Chip Clinorotation System for Live-Cell Microscopy Under Simulated Microgravity

Cells in microgravity are subject to mechanical unloading and changes to the surrounding chemical environment. How these factors jointly influence cellular function is not well understood. We can investigate their role using ground-based analogues to spaceflight, where mechanical unloading is simulated through the time-averaged nullification of gravity. The prevailing method for cellular microgravity simulation is to use fluid-filled containers called clinostats. However, conventional clinostats are not designed for temporally tracking cell response, nor are they able to establish dynamic fluid environments. To address these needs, we developed a Clinorotation Time-lapse Microscopy (CTM) system that accommodates lab-on- chip cell culture devices for visualizing time-dependent alterations to cellular behavior. For the purpose of demonstrating CTM, we present preliminary results showing time-dependent differences in cell area between human mesenchymal stem cells (hMSCs) under modeled microgravity and normal gravity.

Yew, Alvin G.↗
NASA NTRS2016

High-Density Spot Seeding for Tissue Model Formation

A model of tissue is produced by steps comprising seeding cells at a selected concentration on a support to form a cell spot, incubating the cells to allow the cells to partially attach, rinsing the cells to remove any cells that have not partially attached, adding culture medium to enable the cells to proliferate at a periphery of the cell spot and to differentiate toward a center of the cell spot, and further incubating the cells to form the tissue. The cells may be C2C12 cells or other subclones of the C2 cell line, H9c2(2-1) cells, L6 cells, L8 cells, QM7 cells, Sol8 cells, G-7 cells, G-8 cells, other myoblast cells, cells from other tissues, or stem cells. The selected concentration is in a range from about 1 x 10(exp 5) cells/ml to about 1 x 10(exp 6) cells/ml. The tissue formed may be a muscle tissue or other tissue depending on the cells seeded.

Marquette, Michele L.↗
NASA NTRS1980

Lab-On-Chip Clinorotation System for Live-Cell Microscopy Under Simulated Microgravity

Cells in microgravity are subject to mechanical unloading and changes to the surrounding chemical environment. How these factors jointly influence cellular function is not well understood. We can investigate their role using ground-based analogues to spaceflight, where mechanical unloading is simulated through the time-averaged nullification of gravity. The prevailing method for cellular microgravity simulation is to use fluid-filled containers called clinostats. However, conventional clinostats are not designed for temporally tracking cell response, nor are they able to establish dynamic fluid environments. To address these needs, we developed a Clinorotation Time-lapse Microscopy (CTM) system that accommodates lab-on- chip cell culture devices for visualizing time-dependent alterations to cellular behavior. For the purpose of demonstrating CTM, we present preliminary results showing time-dependent differences in cell area between human mesenchymal stem cells (hMSCs) under modeled microgravity and normal gravity.

Yew, Alvin G.↗