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DOE OSTI2022

Expanding the Scope of Bacterial CRISPR Activation with PAM-Flexible dCas9 Variants

CRISPR-Cas transcriptional tools have been widely applied for programmable regulation of complex biological networks. In comparison to eukaryotic systems, bacterial CRISPR activation (CRISPRa) has stringent target site requirements for effective gene activation. While genes may not always have an NGG protospacer adjacent motif (PAM) at the appropriate position, PAM-flexible dCas9 variants can expand the range of targetable sites. Here we systematically evaluate a panel of PAM-flexible dCas9 variants for their ability to activate bacterial genes. We observe that dxCas9-NG provides a high dynamic range of gene activation for sites with NGN PAMs while dSpRY permits modest activity across almost any PAM. Similar trends were observed for heterologous and endogenous promoters. For all variants tested, improved PAM-flexibility comes with the trade-off that CRISPRi-mediated gene repression becomes less effective. Weaker CRISPR interference (CRISPRi) gene repression can be partially rescued by expressing multiple sgRNAs to target many sites in the gene of interest. Furthermore, our work provides a framework to choose the most effective dCas9 variant for a given set of gene targets, which will further expand the utility of CRISPRa/i gene regulation in bacterial systems.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRSDate not supplied

Evaluation of Aerobic Standards for Lunar Surface Extravehicular Activities

Introduction: As NASA prepares to return to the Moon, astronauts will need to be physically primed to successfully execute Extravehicular Activities (EVA) on the Lunar surface. Compared to past Apollo missions, Artemis missions will include EVAs of increased physical demand, frequency, intensity, and duration, thus requiring adequate fitness to successfully and safely complete mission objectives. The physical demand associated with partial gravity (g) EVAs on the Moon is expected to be greater compared to microgravity EVAs based on initial workload estimation. Currently, aerobic fitness standards for partial g EVAs are not well supported by high-fidelity data and require further research for establishing standards to protect crew health and performance during Lunar surface missions. Therefore, the aim of this investigation is to characterize metabolic data from Lunar analog simulations and in-flight crew population aerobic capacity data to validate the current NASA 3001 standard for celestial partial g aerobic fitness (aerobic capacity (VO_2pk) ≥36.5ml/kg/min). Methods: In order to evaluate aerobic fitness requirements for Lunar EVAs, the following were performed: 1) preliminary analysis of long-duration (6 hr) EVA analog simulations in the Neutral Buoyancy Laboratory (NBL) and the Active Response Gravity Offload System (ARGOS) to evaluate expected metabolic rates for 1/6 g EVAs (NBL: n=1 female; ARGOS: n=1 male) and 2) assessment of the current NASA 3001 celestial surface EVA aerobic standard (aerobic capacity (VO_2pk) ≥36.5ml/kg/min) with data from an ISS astronaut population (n=30 male + 13 female) captured before and during space flight (flight day 15). Preliminary Results: Average fractional aerobic capacity during simulated EVAs were 33%±7% VO_2pk and 23.3±7% VO_2pk in the NBL and ARGOS, respectively. This was within a previously predicted 30–40% sustainable work rate. Average metabolic rates for some tasks performed in the NBL, such as traverse (40.4% VO_2pk) and ingress (47.1% VO_2pk) were higher than the predicted sustainable work range. In ARGOS, the tasks with the greatest metabolic rates were object relocation (34.2% VO_2pk) and incapacitated crew rescue (27.0% VO_2pk). Characterization of ISS crewmember aerobic capacity determined that the average preflight VO_2pk was 42.1±5.4 ml/kg/min for females and 37.5±5.4 ml/kg/min for males. At preflight, 21.5% of crewmembers were below the 36.5 ml/kg/min in-mission aerobic standard for celestial surface EVA as outlined in NASA-STD-3001. In-flight, both female and male crewmembers experienced reductions in VO_2pk (11.7% and 10.9%, respectively), such that, during the mission, 62% of crewmembers were below the standard aerobic capacity level for celestial surface EVAs. Conclusions: Our preliminary data suggest that while average metabolic rates for simulated Lunar EVA fall within the 30–40% sustainable work range, task specific metabolic rates exceed this range and may indicate that greater fitness is necessary for more strenuous tasks expected to be performed on the Lunar surface. Additionally, deconditioning due to space flight results in most crewmembers falling below the current celestial partial g EVA standard, which may increase risk to crew health and performance and completing mission objectives for surface missions. Further research is necessary in Artemis-specific analog environments to validate the current NASA-3001 aerobic standard for celestial EVAs. Additionally, work is ongoing to validate the current NASA-3001 strength standard for celestial EVAs.

N.C. Strock↗

NASA NTRS1984

The role of the Shuttle Engineering Simulator in the Solar Max rescue mission

Background information concerning the makeup of the SES, the role it traditionally plays in support of the Shuttle Program, its capabilities and the specific work performed on the simulator during the sixteen months proceedings the Solar Max repair mission is presented. The SES plays a significant role in the preparations for the STS 41-C Solar Max repair mission and during the mission in the development of the plan and procedures to retrieve the satellite after the MMU docking attempts failed. The development of the on orbit simulation capabilities required to support this mission is to a very large extent simply an expansion of the capabilities designed into the SES to support previous engineering simulation requirements for the Space Shuttle program.

St.john, R. H.↗

NASA NTRS1994

Biomedical Use of Aerospace Personal Cooling Garments

Personal thermoregulatory systems are required during extravehicular activity (EVA) to remove the metabolic heat generated by the suited astronaut. The Extravehicular and Protective Systems (STE) Branch of NASA Ames Research Center has developed advanced concepts or liquid cooling garments for both industrial and biomedical applications for the past 25 years. Examples of this work include: (1) liquid cooled helmets for helicopter pilots and race car drivers; (2) vests for fire and mine rescue personnel; (3) bras to increase the definition of tumors during thermography; (4) lower body garments for young women with erythomelaigia; and (5) whole body garments used by patients with multiple sclerosis (MS). The benefits of the biomedical application of artificial thermoregulation received national attention through two recent events: (1) the liquid-cooled garment technology was inducted into the United States Space Foundation's Space Technology Hall of Fame (1993); and (2) NASA has signed a joint Memorandum of Understanding with the Multiple Sclerosis Association (1994) to share this technology for use with MS patient treatment. The STE Branch is currently pursuing a program to refine thermoregulatory design in light of recent technology developments that might be applicable for use by several medical patient populations. Projects have been initiated to apply thermoregulatory technology for the treatment and/or rehabilitation of patients with spinal cord injuries, multiple sclerosis, migraine headaches, and to help prevent the loss of hair during chemotherapy.

Webbon, Bruce W.↗

NASA NTRS1984

SARSAT - Using space for the search and rescue of lives in distress

A major cooperative effort between west and east bloc nations in developing a new space system for detecting ships and aircraft in distress is discussed. The compatible working of the SARSAT system developed by Canada, France, and the US and the COSPAS system developed by the Soviet bloc is summarized, and the SARSAT system is described. The method of location determination is presented, and the ELT and EPIRB, space segment, local user terminal, and mission control center subsystems of the SARSAT system are surveyed, showing block diagrams and data on spacecraft orbital parameters and ELT/EPIRB BECON characteristics. The current problem of ground-based interference is discussed. An outline of future development activities leading to the implementation of a fully operational system is included.

Flatow, F.↗

NASA NTRS1975

Portable Oxygen Subsystem (POS)

Concept selection, design, fabrication, and testing of a Portable Subsystem (POS) for use in space shuttle operations are described. Tradeoff analyses were conducted to determine the POS concept for fabrication and testing. The fabricated POS was subjected to unmanned and manned tests to verify compliance with statement of work requirements. The POS used in the development program described herein met requirements for the three operational modes -- prebreathing, contaminated cabin, and personnel rescue system operations.

Source record↗

NASA NTRS2001

Restoration, Enhancement, and Distribution of the ATLAS-1 Imaging Spectrometric Observatory (ISO) Space Science Data Set

The primary goal of the funded task was to restore and distribute the ISO ATLAS-1 space science data set with enhanced software and database utilities. The first year was primarily dedicated to physically transferring the data from its original format to its initial CD archival format. The remainder of the first year was devoted to the verification of the restored data set and database. The second year was devoted to the enhancement of the data set, especially the development of IDL utilities and redesign of the database and search interface as needed. This period was also devoted to distribution of the rescued data set, principally the creation and maintenance of a web interface to the data set. The final six months was dedicated to working with NSSDC to create a permanent, off site, hive of the data set and supporting utilities. This time was also used to resolve last minute quality and design issues.

Germany, G. A.↗

NASA NTRS1989

Lifting-body option for a Space Station rescue vehicle

With the proposed development of the Space Station Freedom, the need for an assured crew return capability (ACRC) has been identified. During the past 2 years within NASA, there has been an ongoing activity to establish the design requirements for such a vehicle and to identify candidate vehicle concepts that can satisfy these requirements. From this activity, three primary design reference missions and four candidate vehicle concepts have been identified. In support of this activity, the emphasis at Langley has been to assess the potential application of a lifting-body concept derived from work on the HL-10 and X-24 programs in the late sixties. A vehicle systems analysis has been completed including wind-tunnel tests, aerodynamic heating predictions, six-degree-of-freedom entry simulations, and a global landing opportunities assessment. This report summarizes the results of work at Langley over the past 2 years to define the candidate lifting-body ACRC vehicle and provides insight into potential benefits of the higher cross range of this concept in accomplishing the Space Station rescue mission.

Hook, W. Ray↗

DOE OSTI2023

An Ensemble of Neural Networks for Moist Physics Processes, Its Generalizability and Stable Integration

Abstract With the recent advances in data science, machine learning has been increasingly applied to convection and cloud parameterizations in global climate models (GCMs). This study extends the work of Han et al. (2020, https://doi.org/10.1029/2020MS002076 ) and uses an ensemble of 32‐layer deep convolutional residual neural networks, referred to as ResCu‐en, to emulate convection and cloud processes simulated by a superparameterized GCM, SPCAM. ResCu‐en predicts GCM grid‐scale temperature and moisture tendencies, and cloud liquid and ice water contents from moist physics processes. The surface rainfall is derived from the column‐integrated moisture tendency. The prediction uncertainty inherent in deep learning algorithms in emulating the moist physics is reduced by ensemble averaging. Results in 1‐year independent offline validation show that ResCu‐en has high prediction accuracy for all output variables, both in the current climate and in a warmer climate with +4K sea surface temperature. The analysis of different neural net configurations shows that the success to generalize in a warmer climate is attributed to convective memory and the 1‐dimensional convolution layers incorporated into ResCu‐en. We further implement a member of ResCu‐en into CAM5 with real world geography and run the neural‐network‐enabled CAM5 (NCAM) for 5 years without encountering any numerical integration instability. The simulation generally captures the global distribution of the mean precipitation, with a better simulation of precipitation intensity and diurnal cycle. However, there are large biases in temperature and moisture in high latitudes. These results highlight the importance of convective memory and demonstrate the potential for machine learning to enhance climate modeling.

Meteorology & Atmospheric Sciences↗

NASA NTRS2016

Preserving, Enhancing, and Continuing the Scientific Legacy of the Apollo Sample Suite

From 1969 to 1972, Apollo astronauts collected 382 kg of rocks, soils, and core samples from six geologically diverse locations on the Moon. In the nearly 50 years since the samples were collected, over 3000 different studies have been conducted using the nearly 2200 different Apollo samples. Despite the maturity of the sample collection, many new studies of lunar samples are undertaken each year, with an average of more than 55 requests and more than 600 distinct subsamples allocated annually over the past five years. The Apollo samples are a finite resource, however. Although new studies are encouraged, it is important that new studies do not duplicate previous studies, and where possible, leverage previous results to inform and enhance the current studies. This helps to preserve the samples and scientific funding, both of which are precious resources. We have initiated several new efforts to rescue some of the early analyses from these samples, including unpublished analytical data. We are actively scanning NASA documentation in paper form that is related to the Apollo missions and sample processing, and we are collaborating with IEDA to establish a geochemical data base called MoonDB. To populate this database, we are actively working with about a dozen prominent lunar PIs to organize and transcribe years of both published and unpublished data, making it available to all researchers. This effort will also take advantage of new online analytical tools like PetDB. There have already been tangible results from the MoonDB data rescue effort. A pilot project involving the rescue of geochemical data of John Delano on Apollo pyroclastic glasses has already been referenced in multiple Apollo sample requests, and in fact, the compiled data was used as part of one of the new studies. Similarly, scanned sample handling reports have been utilized to find previously analyzed samples that were appropriate to fulfill new sample requests. We have also begun to image the Apollo samples using (1) micro‐CT scanning to document the interior structure of samples, and (2) comprehensive high resolution photography of samples, enabling high resolution 3D reconstructions of the samples. Both efforts will provide comprehensive access to these samples and allow for more targeted requests, and thus better curation of the samples for decades to come.

Zeigler, Ryan↗

NASA NTRS2016

Human and Robotic Exploration Missions to Phobos Prior to Crewed Mars Surface Missions

Phobos is a scientifically significant destination that would facilitate the development and operation of the human Mars transportation infrastructure, unmanned cargo delivery systems and other Mars surface systems. In addition to developing systems relevant to Mars surface missions, Phobos offers engineering, operational, and public engagement opportunities that could enhance subsequent Mars surface operations. These opportunities include the use of low latency teleoperations to control Mars surface assets associated with exploration science, human landing‐site selection and infrastructure development which may include in situ resource utilization (ISRU) to provide liquid oxygen for the Mars Ascent Vehicle (MAV). A human mission to Mars' moons would be preceded by a cargo predeploy of a surface habitat and a pressurized excursion vehicle (PEV) to Mars orbit. Once in Mars orbit, the habitat and PEV would spiral to Phobos using solar electric propulsion based systems, with the habitat descending to the surface and the PEV remaining in orbit. When a crewed mission is launched to Phobos, it would include the remaining systems to support the crew during the Earth‐Mars transit and to reach Phobos after insertion in to Mars orbit. The crew would taxi from Mars orbit to Phobos to join with the predeployed systems in a spacecraft that is based on a MAV, dock with and transfer to the PEV in Phobos orbit, and descend in the PEV to the surface habitat. A static Phobos surface habitat was chosen as a baseline architecture, in combination with the PEV that was used to descend from orbit as the main exploration vehicle. The habitat would, however, have limited capability to relocate on the surface to shorten excursion distances required by the PEV during exploration and to provide rescue capability should the PEV become disabled. To supplement exploration capabilities of the PEV, the surface habitat would utilize deployable EVA support structures that allow astronauts to work from portable foot restraints or body restrain tethers in the vicinity of the habitat. Prototype structures were tested as part of NEEMO 20. PEVs would contain closed loop guidance and provide life support and consumables for two crew for 2 weeks plus reserves. The PEV has a cabin that uses the exploration atmosphere of 8.2 psi with 34% oxygen, enabling use of suit ports for rapid EVA with minimal oxygen prebreathe as well as dust control by keeping the suits outside the pressurized volume. When equipped with outriggers and control moment gyros, the PEV enables EVA tasks of up to 8 pounds of force application without the need to anchor. Tasks with higher force requirements can be performed with PEV propulsion providing the necessary thrust to react forces. Exploration of Phobos builds heavily from the developments of the cis‐lunar proving ground, and significantly reduces Mars surface risk by facilitating the development and testing of habitats, MAVs, and pressurized rover cabins that are all Mars surface forward. A robotic precursor mission to Phobos and Deimos is also under consideration and would need to launch in 2022 to support a 2031 human Phobos mission.

Gernhardt, Michael L.↗

NASA NTRS2016

Human and Robotic Exploration Missions to Phobos Prior to Crewed Mars Surface Missions

Gernhardt, Michael L.↗

NASA NTRS2017

Emergency Locator Transmitter Survivability and Reliability Study

A comprehensive study of Emergency Locator Transmitter (ELT) performance was conducted over a three year period concluding in 2016 in support of the Search and Rescue (SAR) Mission Office at National Aeronautics and Space Administration (NASA) Goddard Space Flight Center (GSFC). The study began with a review of reported performance cited in a collection of works published as early as 1980 as well as analysis of a focused set of contemporary aviation crash reports. Based on initial research findings, a series of subscale and fullscale system tests were performed at NASA Langley Research Center (LaRC) with the goals of investigating ELT system failure modes and developing recommended improvements to the Radio Technical Commission for Aeronautics (RTCA) Minimum Operational Performance Specification (MOPS) that will result in improved system performance. Enhanced performance of ELT systems in aviation accidents will reduce unnecessary loss of human life and make SAR operations safer and less costly by reducing the amount of time required to locate accident sites.

Stimson, Chad M.↗

NASA NTRS2010

NASA Contingency Shuttle Crew Support (CSCS) Medical Operations

The genesis of the space shuttle began in the 1930's when Eugene Sanger came up with the idea of a recyclable rocket plane that could carry a crew of people. The very first Shuttle to enter space was the Shuttle "Columbia" which launched on April 12 of 1981. Not only was "Columbia" the first Shuttle to be launched, but was also the first to utilize solid fuel rockets for U.S. manned flight. The primary objectives given to "Columbia" were to check out the overall Shuttle system, accomplish a safe ascent into orbit, and to return back to earth for a safe landing. Subsequent to its first flight Columbia flew 27 more missions but on February 1st, 2003 after a highly successful 16 day mission, the Columbia, STS-107 mission, ended in tragedy. With all Shuttle flight successes come failures such as the fatal in-flight accident of STS 107. As a result of the STS 107 accident, and other close-calls, the NASA Space Shuttle Program developed contingency procedures for a rescue mission by another Shuttle if an on-orbit repair was not possible. A rescue mission would be considered for a situation where a Shuttle and the crew were not in immediate danger, but, was unable to return to Earth or land safely. For Shuttle missions to the International Space Station (ISS), plans were developed so the Shuttle crew would remain on board ISS for an extended period of time until rescued by a "rescue" Shuttle. The damaged Shuttle would subsequently be de-orbited unmanned. During the period when the ISS Crew and Shuttle crew are on board simultaneously multiple issues would need to be worked including, but not limited to: crew diet, exercise, psychological support, workload, and ground contingency support

Adams, Adrien↗

DOE OSTI2021

Exosomes derived from thymic stromal lymphopoietin-treated dendritic cells regulate T helper 17/regulatory T cell differentiation via miR-21/Smad7 axis

Highlights: • Exosomes from TSLP-treated DCs promote Th17 differentiation. • Exosomes from TSLP-treated DCs inhibit Tregs differentiation. • MiR-21 is highly expressed in exosomes from TSLP-treated DCs. • Exosomal miR-21 regulates Th17/Treg differentiation by targeting Smad7. Thymic stromal lymphopoietin (TSLP) is associated with fungal keratitis. This work aims to investigate whether TSLP can regulate T helper (Th) 17 and regulatory T cell (Treg) differentiation. We separated dendritic cells (DCs) from peripheral blood of healthy volunteers. DCs were treated with TSLP to activate DCs, and exosomes were obtained. CD{sup +} T cells were incubated with exosomes from TSLP-treated DCs. We found that exosomes from TSLP-treated DCs notably promoted the proportions of Th17 cells and inhibited the proportions of Tregs in the CD4{sup +} T cells. Moreover, exosomes from TSLP-treated DCs enhanced the expression of retinoid-related orphan receptor γt (RORγt) and interleukin 17 (IL-17), and repressed the expression of forkhead box protein P3 (Foxp3) and interleukin 10 (IL-10) in the CD4{sup +} T cells. Furthermore, miR-21 was highly expressed in exosomes from TSLP-treated DCs. Exosomes from TSLP-treated miR-21-silenced DCs promoted Treg differentiation and suppressed Th17 differentiation. Smad7 up-regulation repressed Th17 differentiation and enhanced Treg differentiation, which was abolished by miR-21 overexpression. Smad7 overexpression rescued the effect of exosomes from TSLP-treated DCs on Th17/Treg differentiation. In conclusion, our article confirms that TSLP induces DCs to deliver miR-21 by secreting exosomes, and thus miR-21 regulates Th17/Treg differentiation by inhibiting Smad7. Thus, this work further reveals the biological role of miR-21 in fungal keratitis.

60 APPLIED LIFE SCIENCES↗

NASA NTRS1999

Fire and Ice - Safety, Comfort, and Getting the Firefighters' Job Done

Daily life for firefighters consists of working with life-threatening hazards in hostile environments. A major hazard is excessive ambient heat. New hazards have arisen from protective gear that was intended to increase survival time of firefighters while finding and rescuing victims. The insulation is so good now that a firefighter's metabolic heat buildup cannot escape. This forces body core temperatures to life threatening levels in about 20 minutes of moderate activity. Using NASA space suit technology, Oceaneering Space Systems developed a liquid cooling garment prototype that will remove up to 250 watts of metabolic heat. After testing and certification as an approved accessory for firefighter use, this garment will be available for use by any individual encapsulated in protective clothing. This demonstration will present a high surface area circulated liquid cooling garment displayed on a mannequin and available for attendees to try on to experience the effects of active cooling.

Foley, Tico↗

NASA NTRSDate not supplied

microRNA Based Countermeasure Mitigate Health Risks Associated with Space Radiation and Microgravity

From our earlier work, we demonstrated a circulating microRNA (miRNA) signature that is present and involved with the general increased health risks during spaceflight. From this work we demonstrated that this miRNA signature impacted the overall biology and health with both the microgravity and space radiation components of the space environment. We showed that this miRNA signature can be an optimal biomarker for health risk and also has potential to be utilized as a countermeasure to mitigate the damage caused by the space environment by utilizing a human 3D microvascular tissue model. By applying a novel self-delivery system to target 3 miRNAs (i.e. antagomirs) from our spaceflight miRNA signature impacting cardiovascular health risks, we were able to completely mitigate damage caused by exposure to simulated Galactic Cosmic Ray (GCR) irradiation. Here we further expand on the countermeasure experiments to uncover the specific novel biology involved with this countermeasure and in vivo experiments that demonstrates that these antagomirs rescue damage caused to certain organs due to both microgravity and space radiation. Specifically, the miRNAs rescued damage to the heart and immune suppression that occurred in addition to other key biology. In addition, we have also observed with the 3D microvascular tissue model improved DNA double strand break repair machinery which can also contribute to improved recovery and protection against damage caused by space radiation. This work expands on our previous work and further uncovers how a potential minimally invasive countermeasure can be used to mitigate space environment effects.

Afshin Beheshti↗

DOE OSTI2021

Persistent meanders and eddies lead to quasi-steady Lagrangian transport patterns in a weak western boundary current

The Brazil Current (BC) is a weak western boundary current flowing along the Southwestern Atlantic Ocean. It is frequently described as a flow with intense mesoscale activity and relatively low volume transport between 5.0 and 10.0 Sv. We use a 13-year eddy-resolving primitive-equation simulation to show that the presence of persistent meanders and eddies leads to characteristic quasi-steady Lagrangian transport patterns, aptly extracted through climatological Lagrangian Coherent Structures (cLCS). The cLCS position the surface expression of the BC core along the 2000-m isobath, in excellent visual agreement with high resolution satellite sea-surface temperature and the model Eulerian mean velocity. The cLCS deformation pattern also responds to zonally persistent cross-shelf SSH transition from positive (high) values near coastline to low (negative) values between 200- and 2000-m and back to positive (high) offshore from the 2000-m isobath. Zonally-paired cyclonic and anticyclonic structures are embedded in this transition, also causing the cLCS to deform into chevrons. An efficient transport barrier is identified close to the 200-m isobath confirmed by limited inshore movement of drogued drifters and accurately indicated by an along slope maxima of climatological strength of attraction. We also show that the persistent cyclonic and anticyclonic structures may induce localized cross-shelf transport. Regions of low climatological strength of attraction coincide with large shelves and with stagnant synthetic trajectories. We also show that cLCS accurately depict trajectories initiated at the location of Chevron’s spill (November 2011) as compared to synthetic and satellite-tracked trajectories, and the outline of the oil from that accident. There is also an agreement between the large-scale oil slicks reaching the Brazilian beaches (from August 2019 to February 2020) and the strength of climatological attraction at the coast. Our work also clarifies the influence of persistent mesoscale structures on the regional circulation. The identification and quantitative description of climatological Lagrangian coherent structures is expected to improve the effectiveness of future emergency response to oil spills, contingency planning, rescue operations, larval and fish connectivity assessment, drifter launch strategies, waste pollutant and marine debris dispersion and destination.

54 ENVIRONMENTAL SCIENCES↗