Biomedical subjects

Kyongbum Lee

Publications and source records attributed to Kyongbum Lee.

11 recordsLinked to original sources

PubMed2026

DURABLE: A Workflow for Determining Corrosion-Driving and Protective Microbial Mechanisms.

Microbiologically influenced corrosion (MIC) threatens global infrastructure, causing billions of dollars in annual losses. Its persistence stems from unresolved mechanisms─particularly the metabolites produced by microorganisms that drive or inhibit corrosion─and the microbial community structures. Progress has been hindered by the absence of systematic workflows to rapidly and accurately identify MIC-relevant microorganisms and their functions. Here, we present DURABLE (Detection of Unique Corrosion Resistant or Accelerating Biologics in a Laboratory Environment), a pipeline that couples high-throughput microbial screening with genomic and metabolic workflows. We applied the DURABLE workflow to six diesel tank samples and revealed fuel-dependent microbial community structures, which showed greater diversity and evenness in bacterial communities than their fungal counterparts. The workflow used carbon steel beads to rapidly screen over 80 bacterial isolates for corrosive activity, reducing assay time to approximately 2 days compared with the conventional 30-day metal coupon test. More than 40 isolates were identified as corrosive. Further testing using mass spectrometry analysis revealed corrosion-associated metabolites, which were further validated using electrochemical assays. Thus, DURABLE achieved a ∼15-fold increase in screening speed and provided a scalable and mechanistic framework for dissecting MIC dynamics. We expect this advance will enable the development of precision mitigation strategies in hydrocarbon fuel infrastructure.

Bacteria↗

PubMed2007

Effects of forced uncoupling protein 1 expression in 3T3-L1 cells on mitochondrial function and lipid metabolism.

Obesity-related increase in body fat mass is a risk factor for many diseases, including type 2 diabetes. Controlling adiposity by targeted modulation of adipocyte enzymes could offer an attractive alternative to current dietary approaches. Brown adipose tissue, which is present in rodents but not in adult humans, expresses the mitochondrial uncoupling protein 1 (UCP1) that promotes cellular energy dissipation as heat. Here, we report on the direct metabolic effects of forced UCP1 expression in white adipocytes derived from a murine (3T3-L1) preadipocyte cell line. After stable integration, the ucp1 gene product was continuously expressed during differentiation and reduced the total lipid accumulation by approximately 30% without affecting other adipocyte markers, such as cytosolic glycerol-3-phosphate dehydrogenase activity and leptin production. The expression of UCP1 also decreased glycerol output and increased glucose uptake, lactate output, and the sensitivity of cellular ATP content to nutrient removal. However, oxygen consumption and beta-oxidation were minimally affected. Together, our results suggest that the reduction in intracellular lipid by constitutive expression of UCP1 reflects a downregulation of fat synthesis rather than an upregulation of fatty acid oxidation.

3T3-L1 Cells↗

PubMed2006

An algorithm for modularity analysis of directed and weighted biological networks based on edge-betweenness centrality.

MOTIVATION: Modularity analysis is a powerful tool for studying the design of biological networks, offering potential clues for relating the biochemical function(s) of a network with the 'wiring' of its components. Relatively little work has been done to examine whether the modularity of a network depends on the physiological perturbations that influence its biochemical state. Here, we present a novel modularity analysis algorithm based on edge-betweenness centrality, which facilitates the use of directional information and measurable biochemical data.

Algorithms↗

PubMed2006

Extracellular matrix remodeling--methods to quantify cell-matrix interactions.

Tissue turnover during wound healing, regeneration or integration of biomedical materials depends on the rate and extent of materials trafficking into and out of cells involved in extracellular matrix (ECM) remodeling. To exploit these processes, we report the first model for matrix trafficking in which these issues are quantitatively assessed for cells grown on both native collagen (normal tissue) and denatured collagen (wound state) substrates. Human fibroblasts more rapidly remodeled denatured versus normal collagen type I to form new ECM. Fluxes to and from the cells from the collagen substrates and the formation of new ECM were quantified using radioactively labeled substrates. The model can be employed for the systematic and quantitative study of the impact of a broad range of physiological factors and disease states on tissue remodeling, integrating extracellular matrix structures and cell biology.

Cell Communication↗

PubMed2005

Identification of distributed metabolic objectives in the hypermetabolic liver by flux and energy balance analysis.

A methodology for inferring distributed metabolic objectives from time series flux data is developed by combining metabolic flux analysis, pathway identification, free energy balances, and nested optimization. This methodology is used to investigate the metabolic response of the rat liver to burn injury-induced whole body inflammation. Gibbs free energy changes were computed for stoichiometrically balanced sequences of reactions, or pathways, rather than individual reactions, to account for energetic coupling between reactions. Systematic enumeration of pathways proceeded by elementary flux mode (EFM) analysis. Together with stoichiometric balances and external metabolite flux measurements, the DeltaG(PATH)(o) criterion provided sufficient constraints to solve a series of nested optimization problems on the metabolic goal functions and associated flux distributions of fasted livers during the first-week time course of burn injury. The optimization results suggest that there is a consistent metabolic goal function for the liver that is insensitive to the changing metabolic burdens experienced by the liver during the first-week time course. As defined by the goal function coefficients, the global metabolic objective was to distribute the metabolic resources between amino acid metabolism and ketone body synthesis. These findings point to a role for the time-invariant structure of the metabolic reaction network, expressed as stoichiometric and thermodynamic constraints, in shaping the cellular metabolic objective.

Animals↗

PubMed2005

Identification of neutrophil gelatinase-associated lipocalin (NGAL) as a discriminatory marker of the hepatocyte-secreted protein response to IL-1beta: a proteomic analysis.

The liver is the major source of proteins used throughout the body for various functions. Upon injury or infection, an acute phase response (APR) is initiated in the liver that is primarily mediated by inflammatory cytokines such as interleukin-1beta (IL-1beta) and interleukin-6. Among others, the APR is characterized by an altered protein synthetic profile. We used two-dimensional gel electrophoresis to study the dynamics of changes in protein synthesis in hepatocytes exposed to these inflammatory cytokines. Protein profiles were quantified using image analysis and further analyzed using multivariate statistical methods. Our results indicate that IL-1beta and IL-6 each induces secreted protein responses with distinct dynamics and dose-dependence. Parallel stimulation by IL-1beta and IL-6 results in a protein pattern indistinguishable from the IL-1beta pattern, indicating a dominant effect of IL-1beta over IL-6 at the doses tested. Multidimensional scaling (MDS) of correlation distances between protein secretion levels revealed two protein pairs that are robustly co-secreted across the various cytokine stimulation conditions, suggesting shared regulatory pathways. Finally, we also used multivariate alternating conditional expectation (MACE) to identify transformation functions that discriminated the cytokine-stimulated and untreated hepatocyte-secreted protein profiles. Our analysis indicates that the expression of neutrophil gelatinase-associated lipocalin (NGAL) was sufficient to discriminate between IL-1beta and IL-6 stimulation. The combination of proteomics and multivariate analysis is expected to provide new information on the cellular regulatory networks involved in generating specific cellular responses.

Acute-Phase Proteins↗

PubMed2004

Identification of optimal classification functions for biological sample and state discrimination from metabolic profiling data.

MOTIVATIONS: Classification of biological samples for diagnostic purposes is a difficult task because of the many decisions involved on the number, type and functional manipulations of the input variables. This study presents a generally applicable strategy for systematic formulation of optimal diagnostic indexes. To this end, we develop a novel set of computational tools by integrating regression optimization, stepwise variable selection and cross-validation algorithms. RESULTS: The proposed discrimination methodology was applied to plasma and tissue (liver) metabolic profiling data describing the time progression of liver dysfunction in a rat model of acute hepatic failure generated by d-galactosamine (GalN) injection. From the plasma data, our methodology identified seven (out of a total of 23) metabolites, and the corresponding transform functions, as the best inputs to the optimal diagnostic index. This index showed better time resolution and increased noise robustness compared with an existing metabolic index, Fischer's BCAA/AAA molar ratio, as well as indexes generated using other commonly used discriminant analysis tools. Comparison of plasma and liver indexes found two consensus metabolites, lactate and glucose, which implicate glycolysis and/or gluconeogenesis in mediating the metabolic effects of GalN.

Algorithms↗

PubMed2003

Profiling of dynamic changes in hypermetabolic livers.

The liver plays an important role in the overall negative nitrogen balance leading to muscle wasting commonly observed in patients following many conditions, including severe injury, cancer, and diabetes. In order to study changes in liver metabolism during the establishment of such catabolic states, we used a rat skin burn injury model that induces hypermetabolism and muscle wasting. At various times during the first week following the injury, livers were isolated and perfused in a recirculating system under well-defined conditions. We applied a steady-state metabolic flux analysis model of liver metabolism and then used k-means clustering to objectively group together reaction flux time profiles. We identified six distinct groups of reactions that were differentially responsive: (1) pentose phosphate pathway (PPP); (2) amino acid oxidation reactions leading to the formation of tricarboxylic acid (TCA) cycle intermediates; (3) gluconeogenesis; (4) TCA-cycle and mitochondrial oxidation; (5) lipolysis, beta-oxidation, and ketone body formation; and (6) urea-cycle. Burn injury sequentially upregulated the urea-cycle, the PPP, and the TCA-cycle, in order, while beta-oxidation and gluconeogenesis remained unchanged. The upregulation of the PPP was transient, whereas the rise in urea- and TCA-cycle fluxes was sustained. An ATP balance predicted an increased production of ATP and energy expenditure starting on day 3 post-burn, which correlated with the induction of the oxidative phosphorylation uncoupler uncoupling protein-2. We conclude that metabolic profiling using flux analysis and clustering analysis is a useful methodology to characterize the differential activation of metabolic pathways in perfused organs and to identify specific key pathways that are sensitive to a stimulus or insult without making a priori assumptions.

Adaptation, Physiological↗

PubMed2003

Metabolic flux analysis of cultured hepatocytes exposed to plasma.

Hepatic metabolism can be investigated using metabolic flux analysis (MFA), which provides a comprehensive overview of the intracellular metabolic flux distribution. The characterization of intermediary metabolism in hepatocytes is important for all biotechnological applications involving liver cells, including the development of bioartificial liver (BAL) devices. During BAL operation, hepatocytes are exposed to plasma or blood from the patient, at which time they are prone to accumulate intracellular lipids and exhibit poor liver-specific functions. In a prior study, we found that preconditioning the primary rat hepatocytes in culture medium containing physiological levels of insulin, as opposed to the typical supraphysiological levels found in standard hepatocyte culture media, reduced lipid accumulation during subsequent plasma exposure. Furthermore, supplementing the plasma with amino acids restored hepatospecific functions. In the current study, we used MFA to quantify the changes in intracellular pathway fluxes of primary rat hepatocytes in response to low-insulin preconditioning and amino acid supplementation. We found that culturing hepatocytes in medium containing lower physiological levels of insulin decreased the clearance of glucose and glycerol with a concomitant decrease in glycolysis. These findings are consistent with the general notion that low insulin, especially in the presence of high glucagon levels, downregulates glycolysis in favor of gluconeogenesis in hepatocytes. The MFA model shows that, during subsequent plasma exposure, low-insulin preconditioning upregulated gluconeogenesis, with lactate as the primary precursor in unsupplemented plasma, with a greater contribution from deaminated amino acids in amino acid-supplemented plasma. Concomitantly, low-insulin preconditioning increased fatty acid oxidation, an effect that was further enhanced by amino acid supplementation to the plasma. The increase in fatty acid oxidation reduced intracellular triglyceride accumulation. Overall, these findings are consistent with the notion that the insulin level in medium culture presets the metabolic machinery of hepatocytes such that it directly impacts on their metabolic behavior during subsequent plasma culture.

Amino Acids↗

PubMed2003

Metabolic flux analysis of hepatocyte function in hormone- and amino acid-supplemented plasma.

Understanding the metabolic and regulatory pathways of hepatocytes is important for biotechnological applications involving liver cells. Previous attempts to culture hepatocytes in plasma yielded poor functional results. Recently we reported that hormone (insulin and hydrocortisone) and amino acid supplementation reduces intracellular lipid accumulation and restores liver-specific function in hepatocytes exposed to heparinized human plasma. In the current study, we performed metabolic flux analysis (MFA) using a simplified metabolic network model of cultured hepatocytes to quantitively estimate the changes in lipid metabolism and relevant intracellular pathways in response to hormone and amino acid supplementation. The model accounts for the majority of central carbon and nitrogen metabolism, and assumes pseudo-steady-state with no metabolic futile cycles. We found that beta-oxidation and tricarboxylic acid (TCA) cycle fluxes were upregulated by both hormone and amino acid supplementation, thus enhancing the rate of lipid oxidation. Concomitantly, hormone and amino acid supplementation increased gluconeogenic fluxes. This, together with an increased rate of glucose clearance, caused an increase in predicted glycogen synthesis. Urea synthesis was primarily derived from ammonia and aspartate generated through transamination reactions, while exogenous ammonia removal accounted for only 3-6% of the urea nitrogen. Amino acid supplementation increased the endogenous synthesis of oxaloacetate, and in turn that of aspartate, a necessary substrate for the urea cycle. These findings from MFA provide cues as to which genes/pathways relevant to fatty acid oxidation, urea production, and gluconeogenesis may be upregulated by plasma supplementation, and are consistent with current knowledge of hepatic amino acid metabolism, which provides further credence to this approach for evaluating the metabolic state of hepatocytes under various environmental conditions.

Albumins↗

PubMed2003

Induction of a hypermetabolic state in cultured hepatocytes by glucagon and H2O2.

Stress hormones and pro-inflammatory cytokines are putative signals triggering increased energy expenditure or "hypermetabolism" commonly observed in inflammatory states. Cytokines also cause the release of reactive oxidants by immune cells resident in tissues in vivo. Therefore, we hypothesized that oxidative stress plays a role in the induction of hypermetabolism. We examined the effect of glucagon (1.0 nM), a catabolic stress hormone, and the oxidant H(2)O(2) (1.0 mM) on the metabolism of stable hepatocyte cultures for 4 days. Combined H(2)O(2) and glucagon treatment, but not H(2)O(2) or glucagon used alone, increased the hepatocyte oxygen uptake rate 25% above control untreated cells after a lag-time of 72 h. The same treatment also increased the expression of mitochondrial uncoupling protein-2 (UCP2). These effects were significantly inhibited by the antioxidant N-acetylcysteine (5mM) and the pentose phosphate pathway (PPP) inhibitor dehydroepianderosterone (200 microM). Glucagon alone induced urea synthesis and H(2)O(2) alone induced the PPP. These findings show, for the first time, that oxidative stress, in combination with glucagon, increases metabolic energy expenditure in cultured cells, and that this effect may be mediated by UCP-2. Furthermore, the results implicate the PPP in the induction of the hypermetabolic response.

Animals↗