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NASA spinoffs to bioengineering and medicine

The societal and economic benefits derived from the application of aerospace technology to improved health care are examined, and examples of the space-technology spinoffs are presented. Special attention is given to the applications of aerospace technology from digital image processing, space medicine and biology, microelectronics, optics and electrooptics, and ultrasonic imaging. The role of the NASA Technology Application Team in helping the potential technology users to identify and evaluate the technology transfer opportunities and to apply space technology in the field of medicine is discussed.

Rouse, Doris J.↗

NASA spinoffs to bioengineering and medicine

Through the active transfer of technology, the National Aeronautics and Space Administration (NASA) Technology Utilization (TU) Program assists private companies, associations, and government agencies to make effective use of NASA's technological resources to improve U.S. economic competitiveness and to provide societal benefit. Aerospace technology from areas such as digital image processing, space medicine and biology, microelectronics, optics and electrooptics, and ultrasonic imaging have found many secondary applications in medicine. Examples of technology spinoffs are briefly discussed to illustrate the benefits realized through adaptation of aerospace technology to solve health care problems. Successful implementation of new technologies increasingly requires the collaboration of industry, universities, and government, and the TU Program serves as the liaison to establish such collaborations with NASA. NASA technology is an important resource to support the development of new medical products and techniques that will further advance the quality of health care available in the U.S. and worldwide.

Biomedical Engineering/instrumentation/trends↗

Nanoscale Bioengineering Solutions for Space Exploration the Nanopore Sequencer

Characterization of biological systems at the molecular level and extraction of essential information for nano-engineering design to guide the nano-fabrication of solid-state sensors and molecular identification devices is a computational challenge. The alpha hemolysin protein ion channel is used as a model system for structural analysis of nucleic acids like DNA. Applied voltage draws a DNA strand and surrounding ionic solution through the biological nanopore. The subunits in the DNA strand block ion flow by differing amounts. Atomistic scale simulations are employed using NASA supercomputers to study DNA translocation. with the aim to enhance single DNA subunit identification. Compared to protein channels, solid-state nanopores offer a better temporal control of the translocation of DNA and the possibility to easily tune its chemistry to increase the signal resolution. Potential applications for NASA missions, besides real-time genome sequencing include astronaut health, life detection and decoding of various genomes. http://phenomrph.arc.nasa.gov/index.php

Ioana, Cozmuta↗

Biomaterials Out of Thin Air: In Situ, On-Demand Printing of Advanced Biocomposites: A New Materials Design and Production Technique Using 3D-Printed Arrays of Bioengineered Cells

We have completed the proof of concept described in our Phase I proposal, a two-material array of nonstructural proteins. We created an implementation of each step in our technology concept and demonstrated its critical functionality. The biological chassis and printing hardware we created as part of this work can be re-used for future work by inserting a material coding region upstream of the fluorescent tag. Overall, we showed that our technology concept is sound. The mission benefit analyses, as described in our Phase I proposal, are complete and contained in this report. These calculations show that our technology can save hundreds of kilograms of upmass for a potential planetary human habit construction mission: the mass per habitat module can be reduced by approximately one third if the biomaterials are manufactured on Earth and included in the mission upmass, and the full 240 kg per module can be saved if the materials are derived entirely from in situ resources. Mass savings between these two extremes is expected for an actual mission, depending on the level of in situ resource extraction technology. We have shown that continued advancement of this technology concept for use in a space mission environment is justified. Our survey of future development pathways proved extremely informative in light of the lessons learned from our proof of concept work and mission scenario analyses. For example, we were able for the first time to distinguish between the levels of functionality provided by production of structural proteins, other polymers such as polysaccharides, and true organic-inorganic composites such as bone and mineralized shell. This new information represents a significant advance in formulating specific applications, and key enabling technologies, for our proposed concept. We surveyed potential collaborations with other projects and synergies with enabling technologies that are developing. We have received requests for collaboration from other institutions, including labs at Stanford University and Drexel University. We have also received visits from industry, including Organovo, a tissue engineering company, and Autodesk, a major 3D and materials design software company. Finally, we have been in touch with the team behind the 2013 NIAC Phase ll 'Super Ball Bot-Structures for Planetary Landing and Exploration' and are planning to develop our biomaterial printing technology with the goal of enabling tensegrity-based rovers such as theirs to use lighter, more robust materials. A smooth transition from TRL 2 to TRL 3 assumes that the implementations of the technology concept which demonstrate critical functionality are also pathways for future development; while this is the case for most hardware or software projects, the multidisciplinary nature of our project, particularly the biological aspect of it, means that this is not always true. For example, as part of this work we showed that although there are large number of known genetic parts that correspond to non-structural materials, this is not true for sequences for structural organic proteins, let alone biominerals. These realizations allowed us to further subdivide our concept into more detailed development areas, some of which are clearly established at TRL 3, others of which were newly identified sub-technologies moved from TRL 1 to TRL 2. Similarly, although a single feasibility /benefit analysis is sufficient for advancement from TRL 2 to TRL 3, not all potential benefits to a technology concept as broad in scope as ours are apparent at TRL 2. Both our future pathways survey and our proof of concept work highlighted that the true mass savings potential of our technology concept cannot be quantified without modification of existing materials modelling tools to take into account the possibility of positional materials properties customization. Therefore, we have simultaneously both advanced one potential set of applications of our technology concept from TRL 2 to TRL 3 and also identified a previously unknown set of applications and advanced it from TRL 1 to TRL 2. Overall, we have moved the original formulation of our concept forward from TRL 2 to TRL 3, and the expanded formulation of it presented in this document has been advanced from a combination of TRL 1 and early 1RL 2 to an overall late TRL 2. We have also identified the key areas necessary for both short-term and long-term advancement, and made recommendations for specific future work in the most promising directions. With future work on a 1-2 year timeframe to continue advancement to overall TRL 3, we will be well positioned to begin work on a specific space mission technology insertion path.

Biology↗

System and method for all optical electrode interface for bioengineering application

The present disclosure relates to an electro-optical modulator system having a source laser which generates an input optical signal. The input optical signal is received by an electro-optical module. The electro-optical module is implantable into an anatomy and includes a plurality of pixels. Each pixel has associated therewith an electrode and an optical modulator subsystem. The electrode receives electrical signals from the anatomy. The optical modulator subsystem receives the input optical signal and modulates the input optical signal to generate modulated optical output signals in relation to the received electrical signals. A detector subsystem may be used to receive and collect the modulated optical output signals.

Patra, Susant K.↗

AlgaeOrtho, a bioinformatics tool for processing ortholog inference results in algae

Introduction: Microalgae constitute a prominent feedstock for producing biofuels and biochemicals by virtue of their prolific reproduction, high bioproduct accumulation, and the ability to grow in brackish and saline water. However, naturally occurring wild type algal strains are rarely optimal for industrial use; therefore, bioengineering of algae is necessary to generate superior performing strains that can address production challenges in industrial settings, particularly the bioenergy and bioproduct sectors. One of the crucial steps in this process is deciding on a bioengineering target: namely, which gene/protein to differentially express. These targets are often orthologs which are defined as genes/proteins originating from a common ancestor in divergent species. Although bioinformatics tools for the identification of protein orthologs already exist, processing the output from such tools is nontrivial, especially for a researcher with little or no bioinformatics experience. Methods: The present study introduces AlgaeOrtho, a user-friendly tool that builds upon the SonicParanoid orthology inference tool (based on an algorithm that identifies potential protein orthologs based on amino acid sequences) and the PhycoCosm database from JGI (Joint Genome Institute) to help researchers identify orthologs of their proteins of interest in multiple diverse algal species. Results: The output of this application includes a table of the putative orthologs of their protein of interest, a heatmap showing sequence similarity (%), and an unrooted tree of the putative protein orthologs. Notably, the tool would be instrumental in identifying novel bioengineering targets in different algal strains, including targets in not-fully annotated algal species, since it does not depend on existing protein annotations. We tested AlgaeOrtho using three case studies, for which orthologs of proteins relevant to bioengineering targets, were identified from diverse algal species, demonstrating its ease of use and utility for bioengineering researchers. Discussion: This tool is unique in the protein ortholog identification space as it can visualize putative orthologs, as desired by the user, across several algal species.

09 BIOMASS FUELS↗

Optimization of Energy Flow Through Synthetic Metabolic Modules and Regulatory Networks in a Model Photosynthetic Eukaryotic Microbe

Diatoms are important oceanic photosynthetic microorganisms, responsible for approximately 20% of all carbon fixation on Earth. Additionally, diatoms are attractive metabolic engineering candidates. This investigation leveraged Systems Biology techniques to advance our understanding of and potential to engineer diatoms for bioproducts of societal interest, such as biofuels. Systems Biology combines computational and experimental techniques to understand an organism at the whole-cell level as well as characterize the interface between the organism and its environment. Our research resulted in major contributions to the understanding of systems-level light-driven metabolism. Diatoms have a unique evolutionary history. As a result, existing understanding of photosynthetic metabolism may not be applicable to diatom physiology. We generated, validated, and published a high-quality metabolic model of diatom metabolism facilitating novel insights into these unique organisms. Additionally, we overcame a consistent challenge in the modeling of photosynthetic organisms by integrating light into metabolic models. This result allows realistic assessments of how the light environment affects cell physiology and metabolism, a necessary advancement for bioengineering. Next, we generated several methodologies for integrating large-scale datasets with models of metabolism. These “big data” approaches begin to address a constant challenge in biology, which is translating data into information and understanding. Finally, we built web-based databases for housing, distributing, and visualizing systems-level biology data. These projects created an online ecosystem for sharing, analyzing, and characterizing metabolic information and is currently being used by public and private institutions. Photosynthetic bioengineering has the promise to create high-energy products with minimal inputs. Systems Biology has a long history of facilitating bioengineering of cellular metabolism. However, photosynthetic organisms are only sparsely present in this history. This project and its outputs have accelerated the application of proven Systems Biology techniques to light-driven metabolism. The resulting methodologies will accelerate public and private endeavors leveraging photosynthetic systems in the bioengineering of bioproducts such as biofuels or carbon sequestration.

09 BIOMASS FUELS↗

Direct SERDS sensing of molecular biomarkers in plants under field conditions

Molecular biomarkers such as microRNAs (miRNAs) play important roles in regulating various developmental processes in plants. Understanding these pathways will help bioengineer designing organisms for efficient biomass accumulation. Current methods for RNA analysis require sample extraction and multi-step sample analysis, hindering work in field studies. Recent work in the incorporation of nanomaterials for plant bioengineering research is leading the way of an agri-tech revolution. As an example, surface-enhanced Raman scattering (SERS)–based sensors can be used to monitor RNA in vivo. However, the use of SERS in the field has been limited due to issues with observing Raman signal over complex background. To this end, shifted-excitation Raman difference spectroscopy (SERDS) offers an effective solution to extract the SERS signal from high background based on a physical approach. In this manuscript, we report the first application of SERDS on SERS sensors. We investigated this technique on SERS sensor developed for the detection of a microRNA biomarker, miR858. We tested the technique on in vitro samples and validated the technique by detecting the presence of exogenous miR858 in plants directly under ambient light in a growth chamber. The possibility of moving the detection of nucleic acid targets outside the constraints of laboratory setting enables numerous important bioengineering applications. Finally, such applications can revolutionize biofuel production and agri-tech through the use of nanotechnology-based monitoring of plant growth, plant health, and exposure to pollution and pathogens.

09 BIOMASS FUELS↗

Multiomics Data Collection, Visualization, and Utilization for Guiding Metabolic Engineering

Biology has changed radically in the past two decades, growing from a purely descriptive science into also a design science. The availability of tools that enable the precise modification of cells, as well as the ability to collect large amounts of multimodal data, open the possibility of sophisticated bioengineering to produce fuels, specialty and commodity chemicals, materials, and other renewable bioproducts. However, despite new tools and exponentially increasing data volumes, synthetic biology cannot yet fulfill its true potential due to our inability to predict the behavior of biological systems. Here, we showcase a set of computational tools that, combined, provide the ability to store, visualize, and leverage multiomics data to predict the outcome of bioengineering efforts. We show how to upload, visualize, and output multiomics data, as well as strain information, into online repositories for several isoprenol-producing strain designs. We then use these data to train machine learning algorithms that recommend new strain designs that are correctly predicted to improve isoprenol production by 23%. This demonstration is done by using synthetic data, as provided by a novel library, that can produce credible multiomics data for testing algorithms and computational tools. In short, this paper provides a step-by-step tutorial to leverage these computational tools to improve production in bioengineered strains.

09 BIOMASS FUELS↗

Tissue Engineering Using Transfected Growth-Factor Genes

A method of growing bioengineered tissues includes, as a major component, the use of mammalian cells that have been transfected with genes for secretion of regulator and growth-factor substances. In a typical application, one either seeds the cells onto an artificial matrix made of a synthetic or natural biocompatible material, or else one cultures the cells until they secrete a desired amount of an extracellular matrix. If such a bioengineered tissue construct is to be used for surgical replacement of injured tissue, then the cells should preferably be the patient s own cells or, if not, at least cells matched to the patient s cells according to a human-leucocyteantigen (HLA) test. The bioengineered tissue construct is typically implanted in the patient's injured natural tissue, wherein the growth-factor genes enhance metabolic functions that promote the in vitro development of functional tissue constructs and their integration with native tissues. If the matrix is biodegradable, then one of the results of metabolism could be absorption of the matrix and replacement of the matrix with tissue formed at least partly by the transfected cells. The method was developed for articular chondrocytes but can (at least in principle) be extended to a variety of cell types and biocompatible matrix materials, including ones that have been exploited in prior tissue-engineering methods. Examples of cell types include chondrocytes, hepatocytes, islet cells, nerve cells, muscle cells, other organ cells, bone- and cartilage-forming cells, epithelial and endothelial cells, connective- tissue stem cells, mesodermal stem cells, and cells of the liver and the pancreas. Cells can be obtained from cell-line cultures, biopsies, and tissue banks. Genes, molecules, or nucleic acids that secrete factors that influence the growth of cells, the production of extracellular matrix material, and other cell functions can be inserted in cells by any of a variety of standard transfection techniques.

Madry, Henning↗

Influence of Load Profile on Biomechanics of the Squat and Deadlift

Long duration space travel will expose astronauts to extended periods of reduced gravity. Since gravity is not present to assist loading, astronauts will use resistive and aerobic exercise regimes for the duration of the space flight to counteract the effect reduced gravity has on the body. Astronauts will exercise on a flywheel based device on the second Orion Exploration Mission (EM2). The effect that the flywheel load profile has on biomechanics is unknown when compared to free weights or a simulated free weight profile. The purpose of this evaluation is to compare the differences in lower body kinematics and kinetics between the flywheel and free weight profile. Test subjects were instrumented with reflective markers for motion capture data collection while exercising on the Device for Aerobic and Resistive Training (DART) prototype developed by TDA Research, Inc. of Wheat Ridge, CO. Exercises performed were either the squat while wearing a harness or deadlift while grasping a T-bar, both of which interfaced with the DART through a rope cable. The DART, a motorized device controlled via software, is capable of producing different load profiles. Profiles include simulated free weights with adjustable eccentric overload capability and flywheel. Test variables included the applied device load and the load profile, both set through the DART software interface. Motion capture data was collected with a 12 camera system (Smart-DX, BTS Bioengineering, Brooklyn, NY). Bilateral ground reaction force data were collected with force plates (P6000, BTS Bioengineering). DART cable force was recorded through an internal load cell. Data was collected from a total of four test subjects. The three repetition maximum was determined for each test subject for both squat and deadlift. High, medium and low loads were determined based on this maximum. The test subject performed five repetitions of each exercise at each load and each test trial was repeated twice. Cadence was controlled during exercising. Biomechanical data were used to drive the models utilizing the OpenSim software platform. Subject specific models were scaled to match the anthropometrics of the test subjects and used to estimate the peak joint angle, joint range of motion, the peak joint moment and the joint moment impulse of the lower extremity joints and the lumbar joint.These are preliminary results because the data analysis is ongoing. There was a lot of inter-subject variability, however, in general, there was a greater peak lumbar flexion angle for the flywheel squat exercise. There was a trend towards a greater range of motion at the hip for the squat exercise with free weight. There was also a greater hip extension, lumbar extension and ankle plantar flexion impulse moment during the squat exercise with free weight. During the deadlift exercise, the peak knee flexion angle and peak knee extension moment were greater when using the free weight profile. For deadlift, the hip extension, lumbar extension and ankle plantar flexion impulse moment tended to be greater with the free weight profile. Overall, the kinematic and kinetic outcomes for the flywheel profile were either statistically the same as free weight profile in many cases, significantly reduced in a few cases, and rarely higher.

Bioengineering↗

Two-Scale 13 C Metabolic Flux Analysis for Metabolic Engineering

Accelerating the Design-Build-Test-Learn (DBTL) cycle in synthetic biology is critical to achieving rapid and facile bioengineering of organisms for the production of, e.g., biofuels and other chemicals. The Learn phase involves using data obtained from the Test phase to inform the next Design phase. As part of the Learn phase, mathematical models of metabolic fluxes give a mechanistic level of comprehension to cellular metabolism, isolating the principle drivers of metabolic behavior from the peripheral ones, and directing future experimental designs and engineering methodologies. Furthermore, the measurement of intracellular metabolic fluxes is specifically noteworthy as providing a rapid and easy-to-understand picture of how carbon and energy flow throughout the cell. Here, we present a detailed guide to performing metabolic flux analysis in the Learn phase of the DBTL cycle, where we show how one can take the isotope labeling data from a 13 C labeling experiment and immediately turn it into a determination of cellular fluxes that points in the direction of genetic engineering strategies that will advance the metabolic engineering process.For our modeling purposes we use the Joint BioEnergy Institute (JBEI) Quantitative Metabolic Modeling (jQMM) library, which provides an open-source, python-based framework for modeling internal metabolic fluxes and making actionable predictions on how to modify cellular metabolism for specific bioengineering goals. It presents a complete toolbox for performing different types of flux analysis such as Flux Balance Analysis, 13 C Metabolic Flux Analysis, and it introduces the capability to use 13 C labeling experimental data to constrain comprehensive genome-scale models through a technique called two-scale 13 C Metabolic Flux Analysis (2S- 13 C MFA) [1]. In addition to several other capabilities, the jQMM is also able to predict the effects of knockouts using the MoMA and ROOM methodologies. The use of the jQMM library is illustrated through a step-by-step demonstration, which is also contained in a digital Jupyter Notebook format that enhances reproducibility and provides the capability to be adopted to the user's specific needs. As an open-source software project, users can modify and extend the code base and make improvements at will, providing a base for future modeling efforts.

13C Metabolic flux analysis↗

Cytoprotection, proliferation and epidermal differentiation of adipose tissue-derived stem cells on emu oil based electrospun nanofibrous mat

Electrospun nanofibrous scaffolds containing natural substances with wound healing properties such as Emu oil (EO) may have a great potential for increasing the efficiency of stem cell-based skin bioengineering. For this purpose, EO blended PCL/PEG electrospun nanofibrous mats were successfully fabricated and characterized using FE-SEM, FTIR and Universal Testing Machine. The efficiency of the scaffolds in supporting the adherence, cytoprotection, proliferation and differentiation of adipose tissue-derived stem cells (ADSCs) to keratinocyte was evaluated. GC/MS and HPLC were used to determine the composition of pure EO, which revealed to be mainly fatty acids and carotenoids. FE-SEM and cell proliferation assays showed that adhesion and proliferation of ADSCs on EO-PCL/PEG nanofibers was significantly higher than on PCL/PEG nanofibers. Additionally, EO-PCL/PEG nanofibers with free radical scavenging properties conferred a cytoprotective effect against cell-damaging free radicals, while the ability to support cell adhesion and growth was maintained or even improved. Immunostaining of ADSCs on EO-PCL/PEG nanofibers confirmed the change in morphology of ADSCs from spindle to polygonal shape suggesting their differentiation toward an epidermal linage. Moreover, the expression levels of the keratin 10, filaggrin, and involucrin that are involved in epidermal differentiation were upregulated in a stage-specific manner. This preliminary study shows that EO-PCL/PEG nanofibers could be a good candidate for the fabrication of wound dressings and skin bioengineered substitutes with ADSCs. - Highlights: • EO-PCL/PEG nanofibers showed excellent biocompatibility. • EO-PCL/PEG nanofibers conferred a cytoprotective effect against free radicals. • EO with nanofibrous scaffolds promoted epidermal differentiation of ADSCs. • EO-PCL/PEG with ADSCs provide an efficient cell–scaffold construct.

60 APPLIED LIFE SCIENCES↗

A machine learning approach to predict metabolic pathway dynamics from time-series multiomics data

New synthetic biology capabilities hold the promise of dramatically improving our ability to engineer biological systems. However, a fundamental hurdle in realizing this potential is our inability to accurately predict biological behavior after modifying the corresponding genotype. Kinetic models have traditionally been used to predict pathway dynamics in bioengineered systems, but they take significant time to develop, and rely heavily on domain expertise. Here, we show that the combination of machine learning and abundant multiomics data (proteomics and metabolomics) can be used to effectively predict pathway dynamics in an automated fashion. The new method outperforms a classical kinetic model, and produces qualitative and quantitative predictions that can be used to productively guide bioengineering efforts. This method systematically leverages arbitrary amounts of new data to improve predictions, and does not assume any particular interactions, but rather implicitly chooses the most predictive ones.

59 BASIC BIOLOGICAL SCIENCES↗

Enhancing lipid production in plant cells through automated high-throughput genome engineering and phenotyping

Plant bioengineering is a time-consuming and labor-intensive process with no guarantee of achieving desired traits. Here, we present a fast, automated, scalable, high-throughput pipeline for plant bioengineering (FAST-PB) in maize (Zea mays) and Nicotiana benthamiana. FAST-PB enables genome editing and product characterization by integrating automated biofoundry engineering of callus and protoplast cells with single-cell matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). We first demonstrated that FAST-PB could streamline Golden Gate cloning, with the capacity to construct 96 vectors in parallel. Using FAST-PB in protoplasts, we found that PEG2050 increased transfection efficiency by over 45%. For proof-of-concept, we established a reporter-gene-free method for CRISPR editing and phenotyping via mutation of high chlorophyll fluorescence 136. We show that diverse lipids were enhanced up to 6-fold using CRISPR activation of lipid controlling genes. In callus cells, an automated transformation platform was employed to regenerate plants with enhanced lipid traits through introducing multigene cassettes. Lastly, FAST-PB enabled high-throughput single-cell lipid profiling by integrating MALDI-MS with the biofoundry, protoplast, and callus cells, differentiating engineered and unengineered cells using single-cell lipidomics. Furthermore, these innovations massively increase the throughput of synthetic biology, genome editing, and metabolic engineering and change what is possible using single-cell metabolomics in plants.

59 BASIC BIOLOGICAL SCIENCES↗