Results for “Gene expression” — Page 71

NASA NTRSDate not supplied

Batch Effect Correction Methods for NASA GeneLab Transcriptomic Datasets

RNA sequencing (RNA-seq) data from space biology experiments promise to yield invaluable insights into the effects of spaceflight on terrestrial biology. However, sample numbers from each study are low due to limited crew availability, hardware, and space. To increase statistical power, spaceflight RNA-seq datasets from different missions are often aggregated together. However, this can introduce technical variation or "batch effects", often due to differences in sample handling, sample processing, and sequencing platforms. Several computational methods have been developed to correct for technical batch effects, thereby reducing their impact on true biological signals. In this study, we combined 7 mouse liver RNA-seq datasets from NASA GeneLab (part of the NASA Open Science Data Repository) to evaluate several common batch effect correction methods (ComBat and ComBat-seq from the sva R package, and Median Polish, Empirical Bayes, and ANOVA from the MBatch R package). We quantitatively evaluated the ability of these methods to correct for technical batch variables in space biology RNA-seq data using the following criteria: BatchQC, principal component analysis, dispersion separability criterion, log fold change correlation, and differential gene expression analysis. Each batch variable / correction method combination was then assessed using a custom scoring approach to identify the optimal correction method for the combined dataset, by geometrically probing the space of all allowable scoring functions to yield an aggregate volume-based scoring measure. Finally, we describe the way in which the GeneLab multi-study analysis and visualization portal will allow users to examine the presence or absence of batch effects using multiple metrics. If the user chooses to perform batch effect correction, the scoring approach described here can be implemented to identify the optimal correction method to use for their specific combined dataset prior to analysis.

Lauren M. Sanders↗

NASA NTRSDate not supplied

Comparison of Radiation-Induced Damage Between Livers From Control and Chimeric Mice

Assessment of human health risks associated with space radiation exposure is based largely on the knowledge gained from studies in which animals, mostly rodents, are exposed to high-LET radiation on the ground. It has been recognized that translation of animal results to meaningful implications for human disease can be challenging, particularly for certain risk categories such as the high-LET radiation effects in the central nervous system (CNS). Considering limitations in utilizing non-human primates and clinical studies in humans, chimeric animals can potentially bridge the knowledge gap between rodents and humans. In a chimeric animal, a specific organ or a cell type is replaced with respective human cells that are functional. In this pilot study, we used PXB mice whose livers contain >90% human cells. These mice were exposed to gamma rays to investigate DNA damage and transcriptomics changes in the chimeric livers. Results obtained from PXB mice were compared to non-engrafted control animals from the same background strain that were exposed to identical conditions. Staining of the liver tissues with H&E indicated that the human liver tissue in chimeric mice responded differently than the mouse liver tissue to gamma radiation on the cellular level, as evidenced by differences in inflammation and cellular damage seen on histopathology. The gene expression data collected from the liver samples will also be presented, which potentially offers an explanation for the differential responses.

Honglu Wu↗

NASA NTRSDate not supplied

Comparison of Radiation-Induced Damage Between Livers From Control and Chimeric Mice

Assessment of human health risks associated with space radiation exposure is based largely on the knowledge gained from studies in which animals, mostly rodents, are exposed to high-LET radiation on the ground. It has been recognized that translation of animal results to meaningful implications for human disease can be challenging, particularly for certain risk categories such as the high-LET radiation effects in the central nervous system (CNS). Considering limitations in utilizing non-human primates and clinical studies in humans, chimeric animals can potentially bridge the knowledge gap between rodents and humans. In a chimeric animal, a specific organ or a cell type is replaced with respective human cells that are functional. In this pilot study, we used PXB mice whose livers contain >90% human cells. These mice were exposed to gamma rays to investigate DNA damage and transcriptomics changes in the chimeric livers. Results obtained from PXB mice were compared to non-engrafted control animals from the same background strain that were exposed to identical conditions. Staining of the liver tissues with H&E indicated that the human liver tissue in chimeric mice responded differently than the mouse liver tissue to gamma radiation on the cellular level, as evidenced by differences in inflammation and cellular damage seen on histopathology. The gene expression data collected from the liver samples will also be presented, which potentially offers an explanation for the differential responses.

Honglu Wu↗

NASA NTRS2024

Impact of Simulated Microgravity on Mesenchymal Stem Cell Proliferation

Background Long term exposure to microgravity has adverse effects on human body such as bone loss. Due to the vital role of mesenchymal stem cell (MSC) in bone regeneration, studying MSC under microgravity conditions has been of interest during the past decade. However, high cost and limited access to space flight can dampen enthusiasm of researchers. Microgravity simulators provide a way to investigate the effects of microgravity in human physiology. Methods In this study, we compared MSC cultures under simulated microgravity using random positioning machine (RPM) and Gravite 3D simulators. Two culture durations, 1 week and 2 weeks, were selected for MSC cultures without media change. To evaluate impact of simulated microgravity on cell proliferation, cell counting was performed using trypan blue exclusion assay. Additionally, RNA samples and conditioned media were collected for further analysis. Results After 1 week and 2 weeks, no cell morphology change was observed from RPM and Gravite cultures compared to control. Notably, distinct lower cell confluence at the edge of the flasks on the RPM was observed, which could be caused by fluid sheer forces. MSCs cultured on both microgravity simulators showed decreased cell proliferation after 1-week culture. After 2-week culture, cells grown on the RPM showed no significant difference in cell proliferation compared with control while cells cultured on the Gravite bioreactor showed significantly enhanced MSC proliferation. Conclusion Type of microgravity simulator and duration of culture significantly affect MSC proliferation. Further investigation into how simulated microgravity influences MSC gene expression and cytokine secretion related to bone loss is currently on going in our lab.

Cuiping Zhang↗

NASA NTRSDate not supplied

Optimizing a Small RNAseq Analysis Pipeline for NASA GeneLab Using Open-Source Tools and Libraries

Small RNA sequencing (small RNAseq) is a powerful tool for studying the regulation of gene expression in various organisms. Small RNAseq has been leveraged in space biology research to study how expression of small RNAs, e.g. micro RNAs (miRNAs), small interfering RNAs (siRNAs), and piwi-interacting RNAs (piRNAs), change upon exposure to the space environment. NASA GeneLab currently hosts small RNAseq raw data derived from space-relevant experiments on the Open Science Data Repository (OSDR). To maximize the accessibility of these data to the scientific community, in addition to hosting raw data, which is only interpretable by bioinformaticians, GeneLab plans to process all small RNAseq datasets and make those processed data available to the scientific community via the OSDR. In this study, we present the development of the GeneLab standardized pipeline for processing small RNAseq datasets. Using human, plant, and synthetic small RNAseq datasets, we interrogate various open-source software and publicly available databases to evaluate their accuracy and reproducibility in each step of the pipeline. For quality control and adapter detection and trimming, we evaluated TrimGalore!, FASTX, SeqKit, and DNApi methods to optimize alignment to reference genomes. We compared BWA, Bowtie, and Bowtie2 to determine the optimal alignment tool. For each alignment tool we also assessed various reference databases, including Ensembl reference genomes and different types of small RNA reference databases, including genome, hairpin, and miRNA references from the miRbase and MirGeneDB databases. To quantify the aligned data, we compared SAMtools, HTSeq, and RSEM for counting alignment events from each alignment tool used. Finally, we evaluated various tools, including DESeq2 and EdgeR, for data normalization and subsequent differential expression analysis. We will present the results from our comparative analyses for each pipeline step and propose a consensus pipeline for processing small RNAseq data derived from various organisms exposed to the space environment.

SmallRNAseq, NASA GeneLab, quality control, adapte↗

NASA NTRS2024

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

There are numerous applications for microalgae in spaceflight missions and on Earth, such as oxygen production, carbon dioxide removal, nutrition, wastewater processing, and biofuel production. Space Algae-2 aims to test the genetic stability of Arthrospira platensis, commonly known as spirulina, during six-months of continuous culture on the International Space Station. Long-duration exposure to ionizing radiation and microgravity may impact growth, nutrient composition, and genetic stability. The high protein, vitamin, antioxidant content, and radiation resistance make spirulina a promising candidate for bioregenerative life support systems. A concept of operations was developed to grow and harvest algal biomass in space. Preflight testing experiments optimized conditions for an extended growth period in a gas permeable bioreactor bag. Preflight and post-harvest storage methods were developed in addition to a novel cryopreservation method. After sample return, multi-omics analysis will be conducted to determine the mutation rate, gene expression, and the protein and metabolite profile. The concept of operations for Space Algae-2 was tested during a lunar mission simulation within a semi-controlled environment. During a six-day lunar analog mission at the Hawai’i Space Exploration Analog and Simulation (HI-SEAS) A. platensis was successfully grown using flight-like hardware. The cyanobacteria were harvested and used to supplement bread as an example of spirulina biomass utilization. Overall, the data collected from Space Algae-2 will inform potential bioengineering of spirulina for space and terrestrial applications.

Algae↗

NASA NTRSDate not supplied

Mechanical Unloading Effects Hematopoiesis and Subsequent Red Blood Cell Life Cycle Evaluated at Singe Cell Resolution

Red blood cell formation or erythropoiesis in the bone marrow and exit into circulation are known to be affected by mechanical loading. Specifically, space anemia is a condition observed during prolonged microgravity unloading in which the red blood cell (RBC) mass in circulation is reduced. However, the mechanisms causing space anemia are yet to be clearly established, but may include increased hemolysis of RBC in circulation, decreased RBC production or a combination of both factors. The presentation of space anemia is comparable to chronic anemia conditions associated with aging, disuse, and poor health. Molecular dysregulation in the RBC's life cycle is poorly defined and is a significant concern in the context of disuse anemia as mechanotransduction significantly alters of gene expression, protein synthesis and cell signaling pathways. Our study analyzes mouse single cell RNA sequencing libraries from the bone marrow, circulating blood, and spleen and integrates the RBC lineage populations from the tissues into a RBC life cycle atlas. This study also probes how age at time of spaceflight contributes to the molecular signature of space anemia. The overarching objective of the study is to develop a mechanistic understanding of space anemia and translate the findings to a better understanding of aging, disuse, and poor health-related anemia to defining the role of mechanical loading in health and devise potential therapeutic interventions. Important findings of our study are: 1) that mechanical unloading in microgravity alters the proportion of developmental RBC lineage cells, skewing toward progenitor, indicating that fewer RBCs may be present in blood; 2) that late reticulocyte number is elevated in the bone marrow of older animals, suggesting a failure to enter circulation; and 3) that the molecular signature of the circulating RBCs is not significantly altered in microgravity. However, expression markers for eryptosis and erythrophagocytosis are elevated in circulating and spleen RBCs, suggesting spaceflight may shorten the lifetime of RBCs after exit from the marrow. In conclusion our results suggest early stages of erythropoiesis proliferation and differentiation in the marrow are key factors in space anemia and mechanical loading may be a necessary step of RBC entry in circulation.

blood↗

NASA NTRSDate not supplied

Plausible Biological Mechanisms Underlying Sex Differences in Radiation-Induced Lung Cancer Risk

Some epidemiological studies suggest that women are at greater risk for radiation-induced lung cancer than men, but this observation is not consistent across all studies. A scientific committee formed by the National Council on Radiation Protection and Measurement evaluated evidence for a sex difference in lung cancer risk from radiation exposuresby reviewing animal studies and assessing the biological plausibility that such a difference might exist. The committee identified four mechanisms that could potentially result in greater radiogenic lung cancer risks for women. The first of these is that radiation exposure increases the relative risk for a molecular subtype of spontaneous lung cancer that occurs predominantly in women. The second is that is that sex chromosome or gene expression differences between men and women places women at greater risk. The third is that hormonal differences between men and women, particularly for estrogen levels, puts women at greater risk. The fourth mechanism is that sex differences in immune system function underlie a sex difference in radiation associated lung cancer risk.None of these mechanisms hasyet been proven to play a role in radiogenic lung cancer risk.

radiation↗

NASA NTRSDate not supplied

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

There are numerous applications for microalgae in spaceflight missions and on Earth, such as, oxygen production, carbon dioxide removal, nutrition, wastewater processing, and biofuel production. Space Algae-2 aims to test the genetic stability of Arthrospira platensis, commonly known as spirulina, during six-months of continuous culture in spaceflight on the International Space Station. Long-duration exposure to ionizing radiation and microgravity may impact growth, nutrient composition, and genetic stability. The high protein, vitamin, antioxidant content, and radiation resistance make spirulina a promising candidate for bioregenerative life support systems during long-duration missions. A concept of operations was developed to grow and harvest algal biomass in space. Preflight testing experiments were conducted to optimize conditions for an extended growth period in a gas permeable bioreactor bag. Preflight and post-harvest storage methods were developed in addition to a novel cryopreservation method. After sample return, multi-omics analysis will be conducted to determine the mutation rate, gene expression, and protein and metabolite profile. The concept of operations for Space Algae-2 was tested at HI-SEAS (Hawai’i Space Exploration Analog and Simulation) during a six-day lunar analog mission (EMMIHS23, EuroMoonMars, International MoonBase Alliance, HI-SEAS, 2023). A. platensis was grown in the semi-controlled environment using flight-like hardware and solar powered LED lights. Then, the biomass was harvested and used to supplement bread as an example of A. platensis utilization. Overall, the data collected from Space Algae-2 will inform potential bioengineering of spirulina for space and terrestrial applications.

Algae↗

NASA NTRS2024

Designing Payload and Spaceflight Operations for Plants From Extreme 1 Terrestrial Environments

Terrestrial plants from the edges of the limits of life are likely to harbor genes that confer an advantage in deep space environments. These plants are seemingly capable of performing mission critical functions under prevailing deep space conditions while informing directed gene manipulation in target plant species. However, their adaptations to physiologically extreme habitats may hinder efficacy of routine laboratory techniques established for model plants. Here we present the development of Antarctic moss Ceratodon purpureus payload and flight operations for the ARTEMOSS experiment to the ISS astute of limited physical space and crew time. We demonstrate that the hydrophobic surface of Antarctic moss impedes chemical tissue fixation and precludes usage of RNAlater coupled with payload hardware deployed in standard plant spaceflight experiments. We show that deep-freezing the moss tissue on Petri plates provides adequate tissue fixation and allows for extraction of high-quality RNA suitable for gene expression profiling. We replaced hardware with stacks of Petri plates housing Antarctic moss and chemical fixation with deep-freeze in cryogenic GLACIER freezer. Our design can be translated to other plant species, expanding current techniques of experimentation with plants from extreme terrestrial environments aimed toward advancing human space exploration.

Agata K Zupanska↗

NASA NTRSDate not supplied

Space Algae-2 Ground and Lunar Analog Studies in Preparation for Long-Duration Propagation of Cyanobacteria in Spaceflight

There are numerous applications for microalgae in spaceflight missions and on Earth, such as oxygen production, carbon dioxide removal, nutrition, wastewater processing, and biofuel production. Space Algae-2 aims to test the genetic stability of Arthrospira platensis, commonly known as spirulina, during six-months of continuous culture on the International Space Station. Long-duration exposure to ionizing radiation and microgravity may impact growth, nutrient composition, and genetic stability. The high protein, vitamin, antioxidant content, and radiation resistance make spirulina a promising candidate for bioregenerative life support systems. A concept of operations was developed to grow and harvest algal biomass in space. Preflight testing experiments optimized conditions for an extended growth period in a gas permeable bioreactor bag. Preflight and post-harvest storage methods were developed in addition to a novel cryopreservation method. After sample return, multi-omics analysis will be conducted to determine the mutation rate, gene expression, and the protein and metabolite profile. The concept of operations for Space Algae-2 was tested during a lunar mission simulation within a semi-controlled environment. During a six-day lunar analog mission at the Hawai’i Space Exploration Analog and Simulation (HI-SEAS) A. platensis was successfully grown using flight-like hardware. The cyanobacteria were harvested and used to supplement bread as an example of spirulina biomass utilization. Overall, the data collected from Space Algae-2 will inform potential bioengineering of spirulina for space and terrestrial applications.

Algae↗

NASA NTRSDate not supplied

Evaluating the Efficacy of Conditional Variational Autoencoders in Generating Synthetic Single Nuclei RNA-Seq Data for Space Biology Research

Astronauts are subject to unique stressors during spaceflight, leading to changes in their cellular function. However, neither astronauts nor model organisms respond the same to spaceflight, and research implicates a contribution of omics components in differential responses. Understanding how gene expression affects astronaut health is critical for the success of long-term space missions, prompting interest in developing personalized predictive models leveraging artificial intelligence (AI) and machine learning (ML) techniques. Developing such models requires extensive data, which is challenging to obtain and share. This study explores the use of conditional variational autoencoders (CVAEs) to synthetically generate single-nuclei RNA-seq (snRNA-seq) data. CVAEs build on standard variational autoencoders (VAEs) by conditioning data generation on covariates like sample identity and mission parameters, enhancing the relevance of generated data for specific contexts. For our work, we built two CVAEs with varying degrees of sparsity to optimize both interpretability and generative power. We train and validate models on existing snRNA-seq data collected from the brain tissue of mice subjected to spaceflight conditions and their ground control counterparts. We evaluate model performance using statistical tests and visualizations to compare synthetic data to real data. We aim to demonstrate that these prototype CVAE architectures could be used in future space biology work and that this is a method worth further exploring.

Sarah Golts↗

NASA NTRSDate not supplied

Nasa Genelab - Knowledge Graph Fabric Enables Deep Biomedical Analysis of Multi-Omics Datasets

The limited number of astronauts and human samples from long-duration space missions pose significant challenges for studying the health risks associated with spaceflight and developing new treatments. As a result, much of our understanding of the biological impact of space travel relies on samples from model organisms. NASA GeneLab, integrated into Open Science Data Repository (OSDR) is a centralized multi-omics resource containing almost 1000 datasets from over 500 space-related studies from human and model organism samples. Previous studies have demonstrated that human phenotypes and physiological changes caused by spaceflight can be identified by connecting gene expression data from model organisms flown in space to a biomedical knowledge graph (SPOKE). In this work, we present a data fabric connecting OSDR datasets to SPOKE that empowers biomedical analyses through the GeneLab visualization portal. This collaboration is funded by NSF’s Proto-OKN program.

data fabric↗

DOE OSTI2020

Rapid identification of microorganisms

Methods of labeling, identifying and differentiating microorganisms using functionalized Buckyballs are provided herein. The invention further provides methods for imaging or inhibiting gene expression using functionalized Buckyballs of the invention. The invention also provides a system for labeling, identifying and differentiating microorganisms.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Small RNAs at the Interface of the Plant-Fungal Interactions

Fungi and plants interact in a myriad of ways. These interactions range from mutualism to parasitism, and yet, many plant species appear to use similar tools to deal with both. One recent observation is the role that small RNAs play in mediating the conversation between plant and fungus. Increasingly, many studies demonstrate these RNAs are pivotal modulators, reprogramming gene expression and cellular processes essential to the biogenesis of these inter-kingdom relationships.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

BETO 2021 Peer Review - Enhancing Acetogen Formate Utilization to Value-Added Products 2.3.2.112

Electricity from a diversity of sources is increasingly utilized due to its low cost. However, since much of this energy is intermittently generated, there is a mismatch between energy demand and supply. Cheap intermittent energy offers opportunities to utilize this energy and generate value-added chemicals. Low-cost electricity can be used to chemically reduce CO2 to formate and methanol, which then can be upgraded to useful chemicals. To utilize formate for production of valuable chemicals, we decided to focus on Clostridium ljungdahlii as our host organism due to several advantages: it can already utilize formate under some conditions, it has the most developed genetic system for acetogens, and is a model organism for the Wood-Ljungdahl Pathway, the most efficient anaerobic carbon fixation pathway. Formate is a feedstock for a variety of bacteria and has several advantages to gaseous electrochemical products, including ease of storage and miscibility in liquid. Acetogens naturally take C1 compounds and convert them to higher chain products through acetyl-CoA, which is a precursor to many valuable products including carboxylic acids and alcohols. For this project, as a proof of concept, we are focusing on converting formate to butanol. This task primarily relies on two parts: improving formate utilization and installing a butanol pathway. First, we characterized the conditions of native formate utilization in C. ljungdahlii. We also have begun working on genetic tools to delete and express genes.

29 ENERGY PLANNING, POLICY, AND ECONOMY↗

DOE OSTI2021

A Robust Approach to Sequential Information Theoretic Planning

In many sequential planning applications a natural approach to generating high quality plans is to maximize an information reward such as mutual information (MI). Unfortunately, MI lacks a closed form in all but trivial models, and so must be estimated. In applications where the cost of plan execution is expensive, one desires planning estimates which admit theoretical guarantees. Through the use of robust M-estimators we obtain bounds on absolute deviation of estimated MI. Moreover, we propose a sequential algorithm which integrates inference and planning by maximally reusing particles in each stage. We validate the utility of using robust estimators in the sequential approach on a Gaussian Markov Random Field wherein information measures have a closed form. Lastly, we demonstrate the benefits of our integrated approach in the context of sequential experiment design for inferring causal regulatory networks from gene expression levels. Here, our method shows improvements over a recent method which selects intervention experiments based on the same MI objective.

Zheng, Sue↗

DOE OSTI2021

Effect of Cultivation Conditions on the Bioconversion of 4-Hydroxybenzoic Acid in Two White-Rot Fungi

White-rot fungi (WRF) are the most efficient organisms for lignin degradation in nature. A recent study has also shown the ability of WRF in funneling lignin-derived aromatics, including 4-hydroxybenzoic acid (4HBA) and vanillic acid, to central metabolism. However, it is unknown how the cultivation conditions affect the conversion of lignin-derived aromatic compounds. To address this, we performed multi-omic analyses and tracked the conversion of 4HBA in different cultivation conditions in two white-rot fungi: Trametes versicolor and Ceriporiopsis subvermispora. Specifically, we evaluated the effect of static and agitation cultivation conditions in the absence and the presence of antioxidants on fungal performance. We found that the comparison between static and agitation has the higher number of differentially expressed genes or protein abundances at the transcriptomic and proteomic levels, respectively, compared to the presence or the absence of antioxidants. Gene mapping on the enzymatic steps of the proposed 4HBA conversion pathway revealed that agitation causes upregulation of several oxidase genes such as aldehyde dehydrogenases, aldehyde oxidases, and hydroxylases in both fungi. Time-course experiments including 4HBA and cellobiose as carbon sources showed that T. versicolor exhibited higher 4HBA conversion levels in agitation (~110% more conversion, compared to static cultivation conditions) and in the presence of antioxidants (~10% more conversion, compared to the treatments without antioxidants). Interestingly, these trends were not found in C. subvermispora, suggesting different carbon metabolism in both species. Our findings are fundamental to better understand carbon sequestration from lignin in WRF, which will be key for future applications in the lignin valorization field.

29 ENERGY PLANNING, POLICY, AND ECONOMY↗