Biomedical subjects

Christina Chan

Publications and source records attributed to Christina Chan.

At least 19 recordsLinked to original sources

PubMed2005

Palmitic and stearic fatty acids induce Alzheimer-like hyperphosphorylation of tau in primary rat cortical neurons.

Epidemiological studies suggest that high fat diets significantly increase the risk of Alzheimer's disease (AD). In addition, the AD brain is characterized by high fatty acid content compared to that of healthy subjects. Nevertheless, the basic mechanism relating elevated fatty acids and the pathogenesis of AD remains unclear. The present study examines the role of fatty acids in causing hyperphosphorylation of the tau protein, one of the characteristic signatures of AD pathology. Hyperphosphorylation of tau disrupts the cell cytoskeleton and leads to neuronal degeneration. Here, primary rat cortical neurons and astrocytes were treated with saturated free fatty acids (FFAs), palmitic and stearic acids. There was no change in the levels of phosphorylated tau in rat cortical neurons treated directly with these FFAs. The conditioned media from FFA-treated astrocytes, however, caused hyperphosphorylation of tau in the cortical neurons at AD-specific phospho-epitopes. Co-treatment of neurons with N-acetyl cysteine, an antioxidant, reduced FFA-induced hyperphosphorylation of tau. The present results establish a central role of FFAs in causing hyperphosphorylation of tau through astroglia-mediated oxidative stress.

Alzheimer Disease↗

PubMed2004

Controlling primary hepatocyte adhesion and spreading on protein-free polyelectrolyte multilayer films.

The development of new methods for fabricating thin films that provide precise control of the three-dimensional topography and cell adhesion could lead to significant advances in the fields of tissue engineering and biosensors. This Communication describes the successful attachment and spreading of primary hepatocytes on polyelectrolyte multilayer (PEM) films without the use of adhesive proteins such as collagen or fibronectin. We demonstrate that the attachment and spreading of primary hepatocytes can be controlled using this layer-by-layer deposition of ionic polymers. In our study, we used synthetic polymers, namely poly(diallyldimethylammonium chloride) (PDAC) and sulfonated poly(styrene) (SPS) as the polycation and polyanion, respectively, to build the multilayers. Primary hepatocytes attached and spread preferentially on SPS surfaces over PDAC surfaces. SPS patterns were formed on PEM surfaces, either by microcontact printing of SPS onto PDAC surfaces or vice versa, to obtain patterns of primary hepatocytes. PEM is a useful technique for fabricating controlled co-cultures with specified cell-cell and cell-surface interactions on a protein-free environment, thus providing flexibility in designing cell-specific surfaces for tissue engineering applications.

Animals↗

PubMed2004

Integrating gene expression and metabolic profiles.

Recent advances in high throughput technologies have generated an abundance of biological information, such as gene expression, protein-protein interaction, and metabolic data. These various types of data capture different aspects of the cellular response to environmental factors. Integrating data from different measurements enhances the ability of modeling frameworks to predict cellular function more accurately and can lead to a more coherent reconstruction of the underlying regulatory network structure. Different techniques, newly developed and borrowed, have been applied for the purpose of extracting this information from experimental data. In this study, we developed a framework to integrate metabolic and gene expression profiles for a hepatocellular system. Specifically, we applied genetic algorithm and partial least square analysis to identify important genes relevant to a specific cellular function. We identified genes 1) whose expression levels quantitatively predict a metabolic function and 2) that play a part in regulating a hepatocellular function and reconstructed their role in the metabolic network. The framework 1) preprocesses the gene expression data using statistical techniques, 2) selects genes using a genetic algorithm and couples them to a partial least squares analysis to predict cellular function, and 3) reconstructs, with the assistance of a literature search, the pathways that regulate cellular function, namely intracellular triglyceride and urea synthesis. This provides a framework for identifying cellular pathways that are active as a function of the environment and in turn helps to uncover the interplay between gene and metabolic networks.

Algorithms↗

PubMed2004

Selective depositions on polyelectrolyte multilayers: self-assembled monolayers of m-dPEG acid as molecular template.

This paper describes the fabrication of self-assembled monolayer (SAM) patterns of m-d-poly(ethylene glycol) (m-dPEG) acid molecules onto polyelectrolyte multilayers (PEMs). The patterned SAMs on PEMs were created by ionic interactions using microcontact printing (microCP) technique. The created m-dPEG acid monolayer patterns on PEMs act as resistive templates, and thus further depositions of consecutive poly(anion)/poly(cation) pairs of charged particles result in the formation of three-dimensional (3-D) patterned PEM films or selective particle depositions atop the original multilayer thin films. In this study, we illustrate nonlithographic methods of patterning and controlling 3-D PEM architectures and selective particle depositions. We investigated the effect of variables--the choice of solvent, concentration, pH, substrate pretreatment, and stamp contact times--on microcontact printing of m-dPEG acid molecules onto PEM films to determine the optimal conditions for these parameters to achieve efficient transfer of m-dPEG acid patterns onto PEMs. Among the variables, the pH of the m-dPEG acid ink solution played the most important role in the transfer efficiency of the patterns onto the multilayer films. The patterned films were characterized by optical microscopy and atomic force microscopy (AFM).

Journal Article↗

PubMed2004

Inferring pathways and networks with a Bayesian framework.

Numerous mathematical methods have been adapted and developed to quantitatively reverse engineer biological networks, for example, signal transduction pathways, from experimental micro-array data. Compared with stochastic methods, such as Boolean networks, and deterministic methods, such as thermodynamic or differential equation-based models, Bayesian network analysis has the ability to assess, with scoring metrics, causal relations based on conditional probabilities and thus permit hypothesis testing. The goal of this paper is to illustrate the integration of several Bayesian based techniques into a unified Bayesian framework that can infer hepatocellular networks from metabolic data. Reverse engineering of pathways and networks provides a framework for predictive modeling and hypotheses testing to gain deeper insight into living organisms, disease mechanisms, and targeted therapeutics. Evaluating this methodology initially against the known biochemical network provides confidence in the networks that are uncovered from the experimental data using this framework. From the metabolic data we inferred the known sub-networks, such as the tricarboxylic acid (TCA) and urea cycles. In addition, we combined the relationships learned from the data and our current knowledge of the biological system to postulate several alternative metabolic sub-network models that can predict a particular cellular function, such as intracellular triglyceride accumulation.

Algorithms↗

PubMed2004

Inverse modeling using multi-block PLS to determine the environmental conditions that provide optimal cellular function.

MOTIVATIONS: Tissue engineering constitutes an important field with its potential of addressing the current shortage in organ availability. To successfully develop tissue-engineered organs, it is crucial to understand how to maintain the cells under conditions that maximize their ability to perform their physiological roles, regardless of the environment, whether the cells are part of an extracorporeal system, such as the bioartificial liver assist device, or an implantable tissue-engineered device. Our goals are to (1) provide insight into how cells will behave when confronted with changes in its environment and (2) determine the optimal environmental factors to achieve a desired level of cellular function. RESULTS: Diverse sets of environmental factors were used to systematically perturb the metabolic behavior associated with pre-conditioning and plasma supplementation. To probe metabolic state of hepatocytes, metabolic flux analysis was used to obtain the metabolic profile. We applied a multi-block partial least square (MPLS) model to relate environmental factors and fluxes to levels of intracellular lipids and urea synthesis. The MPLS model identified: (1) the most influential environmental factors and (2) how the metabolic pathways are altered by these factors. Finally, we inverted the MPLS model to determine the concentrations and types of environmental factors required to obtain the most economical solution for achieving optimal levels of cellular function for practical situations.

Amino Acids↗

PubMed2004

Hepatic tissue engineering for adjunct and temporary liver support: critical technologies.

The severe donor liver shortage, high cost, and complexity of orthotopic liver transplantation have prompted the search for alternative treatment strategies for end-stage liver disease, which would require less donor material, be cheaper, and less invasive. Hepatic tissue engineering encompasses several approaches to develop adjunct internal liver support methods, such as hepatocyte transplantation and implantable hepatocyte-based devices, as well as temporary extracorporeal liver support techniques, such as bioartificial liver assist devices. Many tissue engineered liver support systems have passed the "proof of principle" test in preclinical and clinical studies; however, they have not yet been found sufficiently reliably effective for routine clinical use. In this review we describe, from an engineering perspective, the progress and remaining challenges that must be resolved in order to develop the next generation of implantable and extracorporeal devices for adjunct or temporary liver assist.

Biomedical Technology↗

PubMed2004

Extracting novel information from gene expression data.

Data from high throughput technologies, such as DNA microarrays, necessitated the development of new computational methodologies for analyzing the high dimensional information contained within the gene expression data. Liao's group suggested the use of network component analysis to predict transcription factor activities by integrating gene expression data from Escherichia coli with known connectivity information between their genes and transcription factors. This introduces an approach for obtaining novel information from gene expression data.

Computational Biology↗

PubMed2003

Direct extraction of Sabin poliovirus genomes from human fecal samples using a guanidine thiocyanate extraction method.

To permit rapid and efficient detection of Sabin poliovirus type 3 from human fecal samples, we developed a guanidine thiocyanate (GuSCN) extraction and reverse transcriptase polymerase chain reaction (RT-PCR) method. Using 10-fold serial dilutions from stock Sabin-Leon 12 a1b poliovirus type 3 at 10(7) TCID(50) per 0.1 ml, genome was detected to a dilution of 10(3) TCID(50) per 0.1 ml. A total of 40 archived fecal samples were examined using this GuSCN extraction method followed by RT-PCR. Fourteen of 20 poliovirus type 3 tissue culture-positive specimens (70%) and two of 20 tissue culture-negative specimens (10%) were detected by GuSCN extraction and RT-PCR. All positive and negative extraction and RT-PCR controls were identified accurately. This GuSCN extraction and RT-PCR technique is rapid, inexpensive, and can be readily adapted to identify genome sequences of other enterovirus types in large numbers of fecal samples. Moreover, the GuSCN technique extracts viral RNA directly from fecal samples, allowing observation of in vivo alterations of genome sequences. Further studies are underway to examine the development of revertant point mutations in the Sabin poliovirus type 3 genome following oral administration of trivalent Sabin Oral Poliovirus Vaccine to humans.

Animals↗

PubMed2003

Splinter removal.

Splinter injuries are common, but larger and deeper splinters are often difficult and painful to remove at home. These splinters often present as a foreign body embedded in the superficial or subcutaneous soft tissues. Whenever possible, reactive objects like wood, thorns, spines, and vegetative material should be removed immediately, before inflammation or infection occurs. Superficial horizontal splinters are generally visible on inspection or easily palpated. A horizontal splinter is exposed completely by incising the skin over the length of the long axis of the splinter, and removed by lifting it out with forceps. A subungual splinter may be removed by cutting out a V-shaped piece of the nail. The point of the V is at the proximal tip of the splinter, which is grasped and removed, taking particular care not to push the splinter further into the nail bed. Removal of an elusive splinter can be challenging and may require the use of imaging modalities for better localization. Deeper splinters, especially those close to important structures such as nerves, tendons, blood vessels, or vital organs, should be referred for removal.

Foreign Bodies↗

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

Measurement of cell-mediated immunity with a Varicella-Zoster Virus-specific interferon-gamma ELISPOT assay: responses in an elderly population receiving a booster immunization.

An interferon-gamma ELISPOT assay has been developed for assessment of cellular immune responses to Varicella-Zoster Virus (VZV) in large, multi-center clinical vaccine trials. We show that the assay performed best when testing peripheral blood mononuclear cells (PBMCs) that had been isolated and then frozen on the same day as blood was drawn, and that freezing PBMCs from blood that was stored overnight before processing resulted in dramatically reduced responses. This assay was used to monitor cell-mediated immunity (CMI) in response to a booster immunization with an investigational live, attenuated VZV vaccine in an elderly population that had been vaccinated 8-10 years previously. The booster vaccine elicited a 1.6- to 1.7-fold rise in the VZV-specific cellular immune response as measured by the ELISPOT assay. The increase from pre to post booster vaccination response was more pronounced (approximately 2.2-fold rise) in a subset of subjects who had received two prior immunizations with a live, attenuated vaccine.

Aged↗

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

Varicella vaccination in children with nephrotic syndrome: a report of the Southwest Pediatric Nephrology Study Group.

OBJECTIVE: To evaluate the safety and immunogenicity of varicella vaccine in children with nephrotic syndrome, including those taking low-dose, alternate-day prednisone. STUDY DESIGN: Prospective, open-label, multicenter clinical trial of varicella vaccine in a 2-dose regimen in US and Canadian children (12 months to <18 years) with nephrotic syndrome. Varicella Zoster Virus (VZV) antibody levels were measured after the first and second vaccine dose and yearly for 2 years. Patients were monitored for adverse reactions to vaccine, exposure to varicella, dermatomal zoster, and chickenpox. RESULTS: Twenty-nine children, mean age 4.9 (SD 1.9) years, 45% receiving every-other-day steroids, received 2 vaccine doses. All patients seroconverted and had VZV antibody levels considered protective against breakthrough varicella (>or=5 gpELISA units) after 2 doses. At 2-year follow-up, all patients retained detectable antibody, and 91% (21 of 23) had levels >or=5 gpELISA units. There were no adverse events associated with vaccination. CONCLUSIONS: Varicella vaccine was generally well tolerated and highly immunogenic in children with nephrotic syndrome, including those on low-dose, alternate-day prednisone.

Anti-Inflammatory Agents↗

PubMed2002

Use of statistical models for evaluating antibody response as a correlate of protection against varicella.

In vaccine clinical trials, humoral antibody responses are often used to measure the effect of a vaccine because they correlate with a vaccine's protective efficacy against the target disease. While the concept of a correlate of protection usually refers to establishing a protective level of antibody titre, identifying a clear-cut value is often impossible because vaccine efficacy is not related solely to the antibody titre. We propose examining the relationship between disease protection and the whole distribution of antibody responses rather than a single cut-off level. In particular, we use failure-time models to estimate the relationship between long-term disease breakthroughs and primary antibody responses after vaccination. We apply these models to show that the varicella antibody response measured by glycoprotein enzyme-linked immunosorbent assay 6 weeks after vaccination strongly correlate with protection against varicella (chickenpox); we used 7-year follow-up data from children who received one dose of a live attenuated varicella (Oka/Merck) vaccine. In addition, we explore the potential use of these models to predict long-term disease breakthrough rates and to estimate the predicted vaccine efficacy of a similar varicella vaccine made with a modified manufacturing process.

Antibodies, Viral↗

PubMed2002

Varicella vaccination in children with chronic renal failure. A report of the Southwest Pediatric Nephrology Study Group.

Children with kidney disease are at risk for serious varicella-related complications. To evaluate the safety and immunogenicity of a two-dose regimen of varicella vaccine in children (aged 1-19 years) with chronic renal insufficiency and on dialysis, the Southwest Pediatric Nephrology Study Group (SPNSG) undertook an open-label, multi-center, prospective 3-year clinical trial. Ninety-six patients without history of varicella were enrolled. Fifty (mean age 4.2 years) had no detectable varicella zoster virus (VZV) antibody; 98% seroconverted after the two-dose regimen. At 1, 2, and 3 years' follow-up, all patients studied maintained VZV antibody, including 16 who received a transplant. No significant vaccine-associated adverse events were seen. One subject developed mild varicella (10-50 maculopapular lesions) 16 months post transplant. In multivariate regression analysis, patients vaccinated after age 6 years had VZV antibody levels 73% (95% confidence interval 33%-89%) lower than patients vaccinated before age 6 years after controlling for gender, estimated glomerular filtration rate, and dialysis treatment. Adjusted analysis also showed that VZV antibody levels were lower after kidney transplantation, but this appeared to be a transient phenomenon. In this study, varicella vaccination with a two-dose regimen of varicella vaccine was generally well tolerated and highly immunogenic in children with chronic kidney disease.

Adolescent↗

PubMed2002

Metabolic pre-conditioning of cultured cells in physiological levels of insulin: generating resistance to the lipid-accumulating effects of plasma in hepatocytes.

Understanding the regulation of hepatocyte lipid metabolism is important for several biotechnological applications involving liver cells. During exposure of hepatocytes to plasma, as is the case in extracorporeal bioartificial liver assist devices, it has been reported that hepatic-specific functions, e.g., albumin and urea synthesis and diazepam removal, are dramatically compromised and hepatocytes progressively accumulate cytoplasmic lipid droplets. We hypothesized that the composition of hepatocyte culture medium significantly affects lipid metabolism during subsequent plasma exposure. Rat hepatocytes were cultured in medium containing either physiological (50 microU/mL) or supra-physiological (500 mU/mL) insulin levels for 1 week and then exposed to human plasma supplemented with or without amino acids. We found that insulin's anabolic effects, such as stimulation of triglyceride storage, were carried over from the pre-conditioning to the plasma exposure period. While hepatocytes cultured in high insulin medium accumulated large quantities of triglycerides during subsequent plasma exposure, culture in low insulin medium largely prevented lipid accumulation. Urea and albumin secretion, as well as the ammonia removal rate, were largely unaffected by insulin but increased with amino acid supplementation. Thus, hepatocyte metabolism during plasma exposure can be modulated by medium pre-conditioning and supplements added to plasma.

Albumins↗

PubMedDate not supplied

Application of multivariate analysis to optimize function of cultured hepatocytes.

Understanding the metabolic and regulatory pathways of hepatocytes is important for biotechnological applications involving liver cells, including the development of bioartificial liver (BAL) devices. To characterize intermediary metabolism in the hepatocytes, metabolic flux analysis (MFA) was applied to elucidate the changes in intracellular pathway fluxes of primary rat hepatocytes exposed to human plasma and to provide a comprehensive snapshot of the hepatic metabolic profile. In the current study, the combination of preconditioning and plasma supplementation produced distinct metabolic states. Combining the metabolic flux distribution obtained by MFA with methodologies such as Fisher discriminant analysis (FDA) and partial least squares or projection to latent structures (PLS) provided insights into the underlying structure and causal relationship within the data. With the aid of these analyses, patterns in the cellular response of the hepatocytes that contributed to the separation of the different hepatic states were identified. Of particular interest was the recognition of distal pathways that strongly correlated with a particular hepatic function. The hepatic functions investigated were intracellular triglyceride accumulation and urea production. This study illustrates a framework for optimizing hepatic function and a possibility of identifying potential targets for improving hepatic functions.

Algorithms↗