Engineering Commons^™

Design Of Variable Stiffness Energy Storage Walking Assist Hip Exoskeleton And Simulation Of Assistance Effect, Bingshan Hu, Ke Cheng, Sheng Lu, Hongliu Yu

Journal of System Simulation

Abstract: Passive energy storage walking assist exoskeleton makes full use of the human’s own energy, reducing energy consumption when walking. Aiming at the present passive energy storage walking assist exoskeleton adopts fixed stiffness joint, a passive variable stiffness energy storage walking assist hip exoskeleton is designed, on the base of joint energy flow characteristics in the process of people walking and the change of stiffness characteristics. The human-exoskeletons coupling model is established, and the optimal stiffness that minimizes the power consumption of the human body walking on a flat surface, as well as the total metabolism and the main thigh …

Reciprocal Regulation Of Transketolase-Like 1 And Hypoxia-Inducible Factor 1 Alpha In Metabolic Reprogramming And Growth Of Diffuse Midline Glioma, H3 K27m-Mutant, Christopher Andrew Waker

Browse all Theses and Dissertations

Diffuse midline glioma, K27M-mutant (DMG) are intractable brain tumors, primarily occurring in the pediatric and adolescent population. Patients have a median survival of less than one year after diagnosis. A lack of therapeutic targets has been a barrier to improvement in patient survival. Irradiation therapy improves symptoms while chemotherapy and surgical intervention, for the most part, have not yet demonstrated utility for treatment. DMG are characterized by a histone H3 mutation that results in the genome-wide loss of epigenetic-repressive marks on K27 and is associated with the misexpression of genes, like Cancer/Testis antigens. A member of this group, transketolase-like 1 …

Monitoring Fgf1-Treated Skin Wounds With Label-Free Multiphoton Microscopy, Gianna Busch

Biomedical Engineering Undergraduate Honors Theses

Chronic skin wounds pose a significant threat to public health, affecting as many as 5.5 million people in the United States every year and costing the healthcare system $10 billion annually^1,2. These wounds are associated with prolonged inflammation, poor vascularization, increased infection risk, and high mortality rates^3,4. There is significant interest in developing and testing a variety of biologics to promote wound healing. However, quantitative evaluations of healing are typically limited to measurements of wound size, and there is a critical need to develop quantitative biomarkers sensitive to different aspects of the healing process. Label-free multiphoton …

In Vivo Optical Metabolic Imaging Of Long-Chain Fatty Acid Uptake In Orthotopic Models Of Triple-Negative Breast Cancer, Megan C. Madonna, Joy E. Duer, Joyce V. Lee, Jeremy Williams, Baris Avsaroglu, Caigang Zhu, Riley Deutsch, Roujia Wang, Brian T. Crouch, Matthew D. Hirschey, Andrei Goga, Nirmala Ramanujam

Biomedical Engineering Faculty Publications

Targeting a tumor’s metabolic dependencies is a clinically actionable therapeutic approach; however, identifying subtypes of tumors likely to respond remains difficult. The use of lipids as a nutrient source is of particular importance, especially in breast cancer. Imaging techniques offer the opportunity to quantify nutrient use in preclinical tumor models to guide development of new drugs that restrict uptake or utilization of these nutrients. We describe a fast and dynamic approach to image fatty acid uptake in vivo and demonstrate its relevance to study both tumor metabolic reprogramming directly, as well as the effectiveness of drugs targeting lipid metabolism. Specifically, …

Nmr Metabolomics For Optimizing Cell-Free Protein Synthesis, Angela M. Campo

Browse all Theses and Dissertations

Cell-Free Protein Synthesis (CFPS) has been utilized by biochemists to produce a variety of chemicals and therapeutics. While CFPS has spawned research in the biochemistry and medical communities, there are still unknown issues with interlaboratory variability with the technique. This work explored the black box nature of CFPS reactions by analyzing the CFPS reactions in situ with Nuclear Magnetic Resonance (NMR) spectroscopy. Aim 1 developed the protocol for conducting NMR experiments on E. coli cell-free reactions as well as a data analysis pipeline. This was accomplished with 1H NMR, capturing metabolite changes over time. The 1D NOESY experiment proved to …

Functional Importance Of Lipin Phosphorylation, Stephanie Elizabeth Hood

Graduate Theses and Dissertations

Highly conserved throughout evolution, lipins are dual functioning proteins found from yeast to humans. Functioning in the cytoplasm as phosphatidate phosphatase enzymes (PAP), lipins produce diacylglycerol that serves as a precursor for neutral fats and membrane phospholipids. Alternatively, nuclear lipins are responsible for the regulation of metabolic genes. Interestingly, both the mammalian lipin 1 paralog and the single Drosophila Lipin ortholog are highly phosphorylated proteins. Target of rapamycin (TOR) has previously been identified as one of the kinases that controls the subcellular localization of both lipin 1 and Drosophila Lipin. However, other serine and threonine kinases are predicted to be …

In Vivo Metabolic And Vascular Response To Hypoxia In Twist Knockdown Murine Breast Cancer, Brandon Sturgill

Graduate Theses and Dissertations

Twist transcription factor is often overexpressed in aggressive tumors. Although needed in early embryonic development for organogenesis, Twist is known to induce an epithelial to mesenchymal transition in cells. In cancer, epithelial to mesenchymal transitions can lead to increased motility and invasiveness. It has also been linked to metabolic reprogramming and increased metastatic risk. Furthermore, metabolic preferences can increase proliferation, enhance metastatic potential, and influence the site of metastasis. We hypothesize that Twist directly affects the metabolism of cancer cells. We expect to see in vivo what we have seen in vitro; Twist overexpression should promote a shift away from …

Metabolomics Of Sorghum Roots During Nitrogen Stress Reveals Compromised Metabolic Capacity For Salicylic Acid Biosynthesis, Amy M. Sheflin, Dawn Chiniquy, Chaohui Yuan, Emily Goren, Indrajit Kumar, Max Braud, Thomas Brutnell, Andrea L. Eveland, Susannah Tringe, Peng Liu, Stephen Kresovich, Ellen Marsh, Daniel P. Schachtman, Jessica E. Prenni

Nebraska Center for Biotechnology: Faculty and Staff Publications

Sorghum (Sorghum bicolor [L.] Moench) is the fifth most productive cereal crop worldwide with some hybrids having high biomass yield traits making it promising for sustainable, economical biofuel production. To maximize biofuel feedstock yields, a more complete understanding of metabolic responses to low nitrogen (N) will be useful for incorporation in crop improvement efforts. In this study, 10 diverse sorghum entries (including inbreds and hybrids) were field-grown under low and full N conditions and roots were sampled at two time points for metabolomics and 16S amplicon sequencing. Roots of plants grown under low N showed altered metabolic profiles at …

Modeling Cell Line-Specific Recruitment Of Signaling Proteins To The Insulin-Like Growth Factor 1 Receptor, Keesha E. Erickson, Dipak Barua, For Full List Of Authors, See Publisher's Website.

Chemical and Biochemical Engineering Faculty Research & Creative Works

Receptor tyrosine kinases (RTKs) typically contain multiple autophosphorylation sites in their cytoplasmic domains. Once activated, these autophosphorylation sites can recruit downstream signaling proteins containing Src homology 2 (SH2) and phosphotyrosine-binding (PTB) domains, which recognize phosphotyrosine-containing short linear motifs (SLiMs). These domains and SLiMs have polyspecific or promiscuous binding activities. Thus, multiple signaling proteins may compete for binding to a common SLiM and vice versa. To investigate the effects of competition on RTK signaling, we used a rule-based modeling approach to develop and analyze models for ligand-induced recruitment of SH2/PTB domain-containing proteins to autophosphorylation sites in the insulin-like growth factor 1 …

Curation Of A Cho Dg44 Genome Scale Model And Application To Support Cell Culture Development Process, Cyrielle Calmels, Laetitia Malphettes, Mikael Rørdam Andersen

Cell Culture Engineering XVI

Genome scale models (GSM) have become a useful tool to connect different omics dataset into a single computational framework, thus giving a good overview of the flux distribution and metabolites interconnections in a specific environmental condition. A community genome-scale metabolic network reconstruction of Cricetulus griseus and cell line specific models have been recently developed 1. The main objectives with the use of the published CHO DG44 model were to enhance industrial bioprocess performance by suggesting genetic or metabolic targets, as well as strategies for medium optimization, and by bringing more fundamental knowledge about CHO cell metabolism. In a first step, …

Poly-Pathway Model Approach: Simulation Of Multiple Metabolic States, Erika Hagrot, Hildur Oddsdottir, Anders Forsgren, Veronique Chotteau

Cell Culture Engineering XV

Animal cell lines have a complex and flexible metabolism and can display varied metabolic behavior depending on the culture conditions. A model that simulate and predict these variations would be a precious tool in the development of media, feeds or processes. However, in order to function as a predictive tool such a model has to describe the multiple metabolic states that can occur for a variety of conditions. This leads to the challenge of identifying a flexible model structure of relevant metabolic pathways and kinetics. To address this challenge, we have introduced the poly-pathway model approach aiming at capturing …

Systems Analysis Of Cho Cell Metabolism For Enhanced Fed-Batch Process Performance: Identification Of Novel Growth Inhibitors And Their Control, Bhanu Chandra, Jaitahree Kale, Anamaria Ovalle, Gregory Hiller

Cell Culture Engineering XV

Over the past decade, process development efforts have led to attaining titers close to 3-4g/L on a routine basis in CHO fed-batch cultures. However, with the advent of biosimilars and the ever expanding therapeutic portfolio of innovator drugs, there is a growing need for enhancing the throughput of fed-batch cultures in order to meet the product requirements with the limited manufacturing capacity. The classical hurdle for higher productivities in fed-batch cultures is the sub-optimal metabolism of mammalian cells that causes accumulation of byproducts, including lactate and ammonia, resulting in growth inhibition. With a robust CHO expression system, our group has …

Investigating The Impact Of Process Optimization On Productivity, Product Quality, Cell Metabolism, And Intracellular Environment, Shailendra Singh

Cell Culture Engineering XV

One of the key goals in process development for monoclonal antibodies is to improve productivity and product quality as needed. The early stage cell culture process developed for an antibody had titers averaging 4 g/L and variable aggregates levels within cell culture. Through process optimization work, involving changes in media formulations, feeding strategy, and process parameters, the final optimized process achieved industry leading titers (greater than 10 g/L) with consistently lower aggregate levels. To understand the impact of process optimization on the CHO cell metabolism and intracellular environment we evaluated 4 conditions: Early stage cell culture process, 2 intermediate processes, …

Cho-Specific Recombinant Protein Glycosylation Reaction Network, Benjamin Kremkow, Kelvin Lee

Cell Culture Engineering XV

Protein glycosylation is one of the most important product quality attributes and impacts efficacy, half-life, and immunogenicity. Previous glycosylation models effectively simulated the key parts of the N-glycosylation pathway. Building upon these prior efforts as well as recent CHO-K1 and Chinese hamster (CH) genome sequencing efforts, we share a new model for CHO- and CH- glycosylation. The model contains all N-glycosylation-related genes and all of the metabolic genes associated with central carbon metabolism, nucleotide sugar precursor synthesis, and nucleotide sugar transport annotated from the CHO-K1 and CH genomes. The model predicts both intracellular and secreted glycans for both mAb and …

Poly-Pathway Model Approach: Simulation Of Multiple Metabolic States, Erika Hagrot

Cell Culture Engineering XV

Animal cell lines have a complex and flexible metabolism and can display varied metabolic behavior depending on the culture conditions. A model that simulate and predict these variations would be a precious tool in the development of media, feeds or processes. However, in order to function as a predictive tool such a model has to describe the multiple metabolic states that can occur for a variety of conditions. This leads to the challenge of identifying a flexible model structure of relevant metabolic pathways and kinetics. To address this challenge, we have introduced the poly-pathway model approach aiming at capturing …

Predictive Engineering Of Cho Cells Using Systems Biology Models, Nathan Lewis

Cell Culture Engineering XV

Decades of bioprocess optimization have resulted in substantial improvements in recombinant protein production. However, some proteins remain difficult to express, and there is an increasing awareness of the need for improved control of critical quality attributes of recombinant protein drugs. To enable cell engineering efforts to enhance protein production and control product quality, we have enumerated the CHO cell parts through genome sequencing efforts,^1,2 and are now providing context to these parts by reconstructing genome-scale networks of the secretory pathway, glycosylation, and metabolism CHO (Figure 1). Using these models, which account for the activities of more than 2000 genes, …

Deletion Of Nfnab In Thermoanaerobacterium Saccharolyticum And Its Effect On Metabolism, Jonathan Lo, Tianyong Zheng, Daniel G. Olson, Natalie Ruppertsberger, Shital Tripathi, Adam Guss, Lee Lynd

Dartmouth Scholarship

NfnAB catalyzes the reversible transfer of electrons from reduced ferredoxin and NADH to 2 NADP⁺. The NfnAB complex has been hypothesized to be the main enzyme for ferredoxin oxidization in strains of Thermoanaerobacterium saccharolyticum engineered for increased ethanol production. NfnAB complex activity was detectable in crude cell extracts of T. saccharolyticum. Activity was also detected using activity staining of native PAGE gels. The nfnAB gene was deleted in different strains of T. saccharolyticum to determine its effect on end product formation. In wild-type T. saccharolyticum, deletion of nfnAB resulted in a 46% increase in H₂ formation but …

Cofactor Specificity Of The Bifunctional Alcohol And Aldehyde Dehydrogenase (Adhe) In Wild-Type And Mutant Clostridium Thermocellum And Thermoanaerobacterium Saccharolyticum, Tianyong Zheng, Daniel G. Olson, Liang Tian, Yannick J. Bomble, Michael Himmel, Jonathan Lo, Shuen Hon, A. Joe Shaw, Johannes P. Van Dijken, Lee Lynd

Dartmouth Scholarship

Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are thermophilic bacteria that have been engineered to produce ethanol from the cellulose and hemicellulose fractions of biomass, respectively. Although engineered strains of T. saccharolyticum produce ethanol with a yield of 90% of the theoretical maximum, engineered strains of C. thermocellum produce ethanol at lower yields (∼50% of the theoretical maximum). In the course of engineering these strains, a number of mutations have been discovered in their adhE genes, which encode both alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) enzymes. To understand the effects of these mutations, the adhE genes from six strains of C. …

Going From Microbial Ecology To Genome Data And Back: Studies On A Haloalkaliphilic Bacterium Isolated From Soap Lake, Washington State, Melanie R. Mormile

Biological Sciences Faculty Research & Creative Works

Soap Lake is a meromictic, alkaline (∼pH 9.8) and saline (∼14-140 g liter^-1) lake located in the semiarid area of eastern Washington State. Of note is the length of time it has been meromictic (at least 2000 years) and the extremely high sulfide level (∼140 mM) in its monimolimnion. As expected, the microbial ecology of this lake is greatly influenced by these conditions. A bacterium, Halanaerobium hydrogeniformans, was isolated from the mixolimnion region of this lake. Halanaerobium hydrogeniformans is a haloalkaliphilic bacterium capable of forming hydrogen from 5- and 6-carbon sugars derived from hemicellulose and cellulose. Due …

Effects Of Local Ph On The Formation And Regulation Of Cristae Morphologies, Dong Hoon Song, Jonghyun Park, Martin A. Philbert, Ann Marie Sastry, Wei Lu

Mechanical and Aerospace Engineering Faculty Research & Creative Works

Cristae, folded subcompartments of the inner mitochondrial membrane (IMM), have complex and dynamic morphologies. Since cristae are the major site of adenosine triphosphate synthesis, morphological changes of cristae have been studied in relation to functional states of mitochondria. In this sense, investigating the functional and structural significance of cristae may be critical for understanding progressive mitochondrial dysfunction. However, the detailed mechanisms of the formation and regulation of these cristae structures have not been fully elucidated. Among the hypotheses concerning the regulation of cristae morphologies, we exclusively investigate the effects of the local pH gradient on the cristae morphologies by using …

Interactions Of Peptide Triazole Thiols With Env Gp120 Induce Irreversible Breakdown And Inactivation Of Hiv-1 Virions, Arangassery Bastian, Mark Contarino, Lauren D. Bailey, Rachna Aneja, Diogo Rodrigo Magalhaes Moreira, Kevin Freedman, Karyn Mcfadden, Caitlin Duffy, Ali Emileh

Dartmouth Scholarship

Background: We examined the underlying mechanism of action of the peptide triazole thiol, KR13 that has been shown previously to specifically bind gp120, block cell receptor site interactions and potently inhibit HIV-1 infectivity.

Results: KR13, the sulfhydryl blocked KR13b and its parent non-sulfhydryl peptide triazole, HNG156, induced gp120 shedding but only KR13 induced p24 capsid protein release. The resulting virion post virolysis had an altered morphology, contained no gp120, but retained gp41 that bound to neutralizing gp41 antibodies. Remarkably, HIV-1 p24 release by KR13 was inhibited by enfuvirtide, which blocks formation of the gp41 6-helix bundle during membrane fusion, while …

Atypical Glycolysis In Clostridium Thermocellum, Jilai Zhou, Daniel G. Olson, D. Aaron Argyros, Yu Deng, Walter M. Van Gulik, Johannes P. Van Dijken, Lee R. Lynd

Dartmouth Scholarship

Cofactor specificities of glycolytic enzymes in Clostridium thermocellum were studied with cellobiose-grown cells from batch cultures. Intracellular glucose was phosphorylated by glucokinase using GTP rather than ATP. Although phosphofructokinase typically uses ATP as a phosphoryl donor, we found only pyrophosphate (PPi)-linked activity. Phosphoglycerate kinase used both GDP and ADP as phosphoryl acceptors. In agreement with the absence of a pyruvate kinase sequence in the C. thermocellum genome, no activity of this enzyme could be detected. Also, the annotated pyruvate phosphate dikinase (ppdk) is not crucial for the generation of pyruvate from phosphoenolpyruvate (PEP), as deletion of the ppdk gene did …

An In Silico Liver: Model Of Gluconeogenesis, Elie R. Chalhoub

ETD Archive

An in silico liver was developed in attempt to represent the in vivo state of the fasted liver. It featured two conceptual models. The first one represented carbohydrate metabolism of the human liver, which included the heterogeneous nature of the liver by incorporating spatial variation of key enzyme activities. This model was able to predict the overall fluxes in tissue and the effect of high intensity exercise on the various hepatic fluxes. A second model of hepatic metabolism was developed to represent the complex interplay between gluconeogenesis, lipid metabolism, and alcohol metabolism in the fasted rat liver. Biochemical pathways are …

Slides: Appropriate Sustainable Energy Technologies: A Light To The World, Lakshman D. Guruswamy, Jason B. Aamodt, Blake Feamster

2012 Energy Justice Conference and Technology Exposition (September 17-18)

Presenter: Jason Aamodt, Attorney; Adjunct Professor, University of Tulsa

15 slides

Improved Tumor Contrast Achieved By Single Time Point Dual-Reporter Fluorescence Imaging, Kenneth M. Tichauer, Kimberley S. Samkoe, Kristian J. Sexton, Jason R. Gunn, Tayyaba Hasan, Brian W. Pogue

Dartmouth Scholarship

In this study, we demonstrate a method to quantify biomarker expression that uses an exogenous dual-reporter imaging approach to improve tumor signal detection. The uptake of two fluorophores, one nonspecific and one targeted to the epidermal growth factor receptor (EGFR), were imaged at 1 h in three types of xenograft tumors spanning a range of EGFR expression levels (n  =  6 in each group). Using this dual-reporter imaging methodology, tumor contrast-to-noise ratio was amplified by >6 times at 1 h postinjection and >2 times at 24 h. Furthermore, by as early as 20 min postinjection, the dual-reporter imaging signal …

High Ethanol Titers From Cellulose By Using Metabolically Engineered Thermophilic, Anaerobic Microbes, D. Aaron Argyros, Shital A. Tripathi, Trisha F. Barrett, Stephen R. Rogers, Lawrence F. Feinberg, Daniel G. Olson, Justin M. Foden, Bethany B. Miller, Lee R. Lynd, David A. Hogsett, Nicky C. Caiazza

Dartmouth Scholarship

This work describes novel genetic tools for use in Clostridium thermocellum that allow creation of unmarked mutations while using a replicating plasmid. The strategy employed counter-selections developed from the native C. thermocellum hpt gene and the Thermoanaerobacterium saccharolyticum tdk gene and was used to delete the genes for both lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta). The Δldh Δpta mutant was evolved for 2,000 h, resulting in a stable strain with 40:1 ethanol selectivity and a 4.2-fold increase in ethanol yield over the wild-type strain. Ethanol production from cellulose was investigated with an engineered coculture of organic acid-deficient engineered strains of …

Electroporation-Induced Electrosensitization, Olga N. Pakhomova, Betsy W. Gregory, Vera A. Khorokhorina, Anglela M. Bowman, Shu Xiao, Andrei G. Pakhomov

Bioelectrics Publications

BACKGROUND: Electroporation is a method of disrupting the integrity of cell membrane by electric pulses (EPs). Electrical modeling is widely employed to explain and study electroporation, but even most advanced models show limited predictive power. No studies have accounted for the biological consequences of electroporation as a factor that alters the cell's susceptibility to forthcoming EPs.

METHODOLOGY/PRINCIPAL FINDINGS: We focused first on the role of EP rate for membrane permeabilization and lethal effects in mammalian cells. The rate was varied from 0.001 to 2,000 Hz while keeping other parameters constant (2 to 3,750 pulses of 60-ns to 9-micros duration, 1.8 …

Rapid Characterization And Engineering Of Natural Product Biosynthetic Pathways Via Dna Assembler, Zengyi Shao, Yunzi Luo, Huimin Zhao

Zengyi Shao

We report a synthetic biology strategy for rapid genetic manipulation of natural product biosynthetic pathways. Based on DNA assembler, this method synthesizes the entire expression vector containing the target biosynthetic pathway and the genetic elements required for DNA maintenance and replication in various hosts in a single-step manner through yeast homologous recombination, offering unprecedented flexibility and versatility in pathway manipulations.

A Molecular Dynamics Study On The Transport Of A Charged Biomolecule In A Polymeric Adsorbent Medium And Its Adsorption Onto A Charged Ligand, Enrico Riccardi, Jee-Ching Wang, Athanasios I. Liapis

Chemical and Biochemical Engineering Faculty Research & Creative Works

The transport of a charged adsorbate biomolecule in a porous polymeric adsorbent medium and its adsorption onto the covalently immobilized ligands have been modeled and investigated using molecular dynamics modeling and simulations as the third part of a novel fundamental methodology developed for studying ion-exchange chromatography based bioseparations. To overcome computational challenges, a novel simulation approach is devised where appropriate atomistic and coarse grain models are employed simultaneously and the transport of the adsorbate is characterized through a number of locations representative of the progress of the transport process. The adsorbate biomolecule for the system studied in this work changes …

Cloning And Heterologous Expression Of The Spectinabilin Biosynthetic Gene Cluster From Streptomyces Spectabilis, Zengyi Shao

Zengyi Shao

Spectinabilin is a rare nitrophenyl-substituted polyketide metabolite. Here we report the cloning and heterologous expression of the spectinabilin gene cluster from Streptomyces spectabilis. Unexpectedly, this gene cluster is evolutionarily closer to the aureothin gene cluster than to the spectinabilin gene cluster from Streptomyces orinoci. Moreover, the two nearly identical spectinabilin gene clusters use a distinctly different regulation mechanism.