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Disease Research

Introduction to Disease Research

Current Research

Diabetes research

Article 1

Insufficient beta-cell mass and insulin production are characteristic of Type 1 and Type 2 diabetes. Islet mass can be expanded ex vivo or in vivo and used as a substantial treatments for either form of diabetes. Cholecystokinin (CCK) stimulates beta-cell proliferation in human islets, while retaining beta-cell function.

Therapy that causes the expansion of beta-cells in the absence of alpha-cell proliferation places diabetic patients at risk for hypoglycemia. However, this can potentially be avoided with concomitant alpha-cell expansion. As a result, the co-expansion of alpha-cells and beta-cells via CCK-dependent pathways may restore hypoglycemic responsiveness and insulin independence, for patients that receive islet transplants.

CCK overexpression triggers beta-cell proliferation through a unique pathway. This impact on beta-cell proliferation implies that CCK has a potentially therapeutic value for islet cell transplants or enhancing beta-cell mass. Researchers hope to better understand the signaling pathways that activate the factors of cell cycle machinery that respond to CCK, in order to identify other therapeutic agents.

Link to article:

http://mend.endojournals.org/cgi/rapidpdf/me.2008-0255v1

Article 2

Elevated circulating ferritin concentrations are associated with a higher risk of Type 2 diabetes and metabolic syndrome in middle-aged and elderly Chinese people that were independent of obesity, inflammation, adipokines and other risk factors. This highlights the significant role of iron overload for metabolic diseases especially in a country with a relatively high prevalence of iron deficiency.

There is a strong positive association between elevated iron stores, and the risks of Type 2 diabetes, impaired fasting glucose (IFG) and metabolic syndrome in middle-aged and elderly Chinese men and women. There is also a substantial positive correlation between ferritin and HbA1c in diabetic participants for both Chinese men and women. These facts highlight the crucial role of iron overload as a significant independent risk factor for metabolic diseases in subgroups of developing countries.

There are substantially different bodily ferritin concentrations among different geographic locations of China. These differences in iron levels may be attributed to multivariate dietary patterns.

There is also a strong positive association between elevated plasma ferritin concentrations and the risk of Type 2 diabetes, IFG and metabolic syndrome in both Chinese men and women.

Elevated ferritin concentrations frequently cluster with well-established risk factors of Type 2 diabetes that include obesity, metabolic syndrome and inflammation.

Inflammation is a suggested regulator of ferritin mRNA and protein levels, as well as ferritin secretion. It is believed that ferritin may increase the risk of type 2 diabetes, IFG or metabolic syndrome via a pathway that does not significantly overlap with C-reactive protein (CRP) and other inflammatory diseases.

Metabolic syndrome is a strong risk factor for Type 2 diabetes, but this syndrome has scarcely been researched in the ferritin-diabetes associated studies. Elevated ferritin concentrations may increase the risk of Type 2 diabetes via mechanisms that do not include the metabolic syndrome.

Link to article:

http://jcem.endojournals.org/cgi/rapidpdf/jc.2008-1159v1

Research on Cisplatin Uptake

Cellular resistance to cisplatin potentially comprises of mechanisms that limit cisplatin uptake, accumulation, altered cellular repair mechanisms, and changes that promote cell survival. Altered cisplating uptake has been proven to occur from a reduction in membrane-binding/transport proteins, and endocytosis. It has also been proven that up-regulation of SIRT1 is a contributor of cisplatin resistance that is in association with altered mitochondrial metabolism.

A cell will obtain the majority of its energy via oxidative phosphorylation, which is executed in the mitochondria. Any alteration in the glycolytic metabolic pathway of a cell could substantially affect biosynthetic reactions and cellular susceptibility, when under stress.

The mitochondria serve as ideal targets for cisplatin toxicity, because previous research has exhibited that cells with a reduced number of mitochondria are substantially more resistant to cisplatin than normal cells.

The disruption of glucose uptake is associated with abnormal mitochondrial function.

Over expression of SIRT1 results in reduced oxygen consumption as well as altered mitochondrial bioenergenesis. Oxygen consumption is linked to the generation of reactive oxygen species, and reactive oxygen species levels are correlated with cisplatin toxicity.

When glucose is limited, there is an over expression of SIRT1, which increases the cisplatin resistance of parent cells. However, reducing glucose metabolism has also proven to result in SIRT1 over expression with elevated cisplatin resistance.

Researchers have surmised a mechanism by which the over expression of SIRT1 contributes to cisplatin resistance. When incorporated into DNA or RNA, cisplatin perturbs nucleic acid structure and function, which results in the destruction of tumor cells. SIRT1 deacetylates the p53 protein that recognizes and binds to DNA that has been modified by cisplatin. The SIRT1-induced deacetylation results in reduced p53-dependent apoptosis, when DNA is damaged. The interaction of p53 and SIRT1 after platination may molecularly link DNA damage and p52-mediated DNA repair.

SIRT1 is involved in cellular metabolism, and requires NAD+ to function. Researchers have discovered that NAD+ concentration is higher in cells that overexpress SIRT1. Low NAD+ concentrations would allegedly diminish SIRT1 deacetylase activity, and increase the likelihood of cell senescence or apoptosis via p53 acetylation. The NAD+ dependence of the deacetylase activity of SIRT1 seems to imply that SIRT1 may regulate NAD+ metabolism. Specifically, SIRT1 might serve as a bridge that coordinates the metabolic status with regulation of key target genes that are involved in cancer resistance to cisplatin.

SIRT1 may be a central modulator of cellular processes that require energy such as fatty acid synthesis, protein synthesis and cell growth. If this is true, then over expression of SIRT1 could potentially result in a higher apoptotic threshold.

SIRT1 might also act as a metabolic sensor, as a result of its NAD+ dependence. If it does, then this might present a linkage between energy consumption to a transcriptional program that modulates cellular responses to stress. SIRT1 deacetylation of transcriptional complexes typically results in diminished transcriptional activation after deacetylating specific transcription factors.

SIRT1 could serve as a scaffold that tethers various transcriptional complexes. SIRT1 also confers resistance to etoposide via DNA repair augmentation. As a result, the over expression of SIRT1 may shift the transcription-dependent response of cisplatin resistance away from cell and towards cell survival.

When regulated by glucose homeostasis, SIRT1 may contribute to tumor cell resistance to cisplatin. This provides insight that could facilitate the development of methods to subjugate drug resistance of cancer patients.

Link to article:

http://www.ncbi.nlm.nih.gov/pubmed/18723829?ordinalpos=69&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Research on Single Nucleotide Polymorphisms

Two single nucleotide polymorphisms (SNPs) (rs490683 and rs9819506) are associated with changes in body weight and body mass index (BMI), within the promoter region of the human growth hormone secretagogue receptor (GHSR) gene. However, no evidence has been uncovered for the involvement of GHSR SNPs in body weight regulation. The SNPs rs490683 and rs9819506 are not unequivocally associated with measures of obesity or Type 2 diabetes. Rs490683 and rs509035 are in strong linkage disequilibrium (LD) with each other. Researchers speculate that this explains why none of the GHSR polymorphisms are statistically significantly associated with risk of Type 2 diabetes. It is speculated that individuals with the rs490683-GG genotype, where the NF-1 protein half site is intact, are endowed with enhanced GHSR expression, which potentially leads to increased receptor signaling that ultimately causes an increase in appetite. Researchers have discovered that individuals with the rs490683-GG genotype lose less weight during lifestyle interventions than do individuals with the rs490683-CC genotype. GHSR signals at approximately fifty percent activity, even when in the absence of an agonist, which implies that the control of the expression level of the receptor is directly correlated to signaling activity. GHSR expression in the hypothalamus increases, during prolonged fasting. This increased expression is suspected to contribute to the amplification of ghrelin action, which is expected to result in a ghrelin-independent increase in receptor signaling to produce an increase in appetite. The GHSR polymorphism rs490683 is proven to be associated with changes in body weight. For individuals that belong to the rs4906863-GG genotype, where the NF-1 site is intact, there is a supposed increase in ghrelin receptor expression. The alleles of rs519384 did not show any differential protein-binding. Therefore, this SNP is not a crucial region for GHSR gene expression. Polymorphisms in the GHSR gene are proven to be longitudinally associated with obesity and glucose metabolism. Genetic variation of the promoter potentially has an effect on appetite regulation via modulation of GHSR gene expression and ghrelin receptor signaling.

Link to article:

http://www.plosone.org/article/info:doi/10.1371/journal.pone.0002941

Research on Oxysterols

Examining the relationship between the concentrations of serum enzymatically and non-enzymatically generated oxysterols and measures of obesity, metabolic components that include insulin resistance, levels of blood pressure and serum lipids:

    Researchers have observed that oxysterol levels increase with increased measures of obesity that include body mass index (BMI) and WC, which is in further support of the already established associations between obesity and oxidative stress. Obesity and the metabolic syndrome result in an increase in CRP and atherosclerosis. These results are implicit of enhanced inflammatory stress and are associated with greater oxidative stress, and increased formation of oxysterols. Researchers hope that the relationship between adolescent obesity with greater oxidative stress in the form of oxysterols will be a relevant marker of adult cardiovascular risk, while monitoring obesity from childhood to adult life.

    Oxysterols act as modulators of insulin. Enzymatically produced oxysterols are established agonists of liver X receptors (LXRs), which are involved in enhanced insulin secretion, and the regulation of cholesterol metabolism. 7αOH is an enzymatically generated oxysterol. 7αOH has been observed to have a positive relationship with insulin, and is believed to have an impact on insulin secretion.

Oxysterols are present as early markers of oxidative stress-mediated deregulations that involve cholesterol metabolism. Oxysterol formation might serve as an important link between hypercholesterolemia and metabolic derangements that are present in the progression to vascular dysfunctions. As a result, oxysterols are capable of being a metabolic linkage to vascular risk factors that agglomerate during adolescence.

Link to article:

http://www.ncbi.nlm.nih.gov/pubmed/18713821?ordinalpos=104&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Research on Hepatocytes

Fatty livers are associated with insulin resistance, atherosclerosis and the metabolic syndrome. Fatty livers also predict future cardiovascular events. There is support that insulin resistance is a strong underlying mechanism for dyslipidemia that increases LDL particle density and lowers HDL-cholesterol. Researchers have also suggested that fat accumulation in the liver has an independent effect on this form of dyslipidemia. The liver is also involved in immune responses. Hepatocytes comprise of approximately two thirds of total liver cells, with biliary epithelial cells, sinusoidal cells, Kupffer cells, stellate cells, dendritic cells, and lymphocytes represent the other cell types. Kupffer cells and lymphocytes are the main cell types that are involved in hepatic immune responses. Kupffer cells represent the largest group of fixed macrophages in the body. They account for approximately twenty percent of nonparenchymal cells in the liver. Kupffer cells that are in the liver clear endotoxins (lipopolysaccharides) from the passing blood, phagocyte debris in microorganisms. Kupffer cells also produce cytokines, which have a significant role in cell differentiation and cell proliferation. The gastrointestinal tract is thought to be crucial for hepatic inflammation and the pathophysiology of non-alcoholic steatohepatitis (NASH).

Link to article:

http://www.ncbi.nlm.nih.gov/pubmed/18723451?ordinalpos=71&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Research on Cardiac metabolism

Histone deacetylase (HDAC)-3 is a prominent regulator of cardiac energy metabolism. Despite the high degree of homology among HDAC1, HDAC2, and HDAC3, HDAC3 clearly has a significant role in the maintenance of cardiac function.

    Under basal conditions, HDAC3, together with the nuclear receptor corepressor/silencing mediator for retinoic acid and thyroid hormone receptors (NCoR/SMRT) or the retinoblastoma (Rb) complex, is recruited by PPARs and other nuclear receptors to the promoters of target genes to facilitate the transcriptional repression by nuclear receptors. Mitochondrial derangements from the loss of HDAC3 in cardiomyocytes resemble those seen in diabetic cardiomyopathies. The increased oxidation of fatty acids that occurs in diabetic hearts causes enhanced reducing equivalents to the electron transport chain, which results in free radical production, mitochondrial uncoupling, and decreased cardiac efficiency.

Link to article:

http://www.jci.org/articles/view/35847#SEC3

Research on Ketone bodies

A low carbohydrate diet has been proven to be an effective treatment for severe pharmacoresistant therapy. In order to reveal any therapeutic applications of a ketogenic diet, researchers have tested for effects of physiological ketone bodies on neuronal firing rates and excitability. Spontaneous neuronal firing rates of neurons in slices of rat or mouse substantia nigra pars reticulata were reduced by physiological content of ketone bodies (ß-hydroxybutyrate or acetoacetate). This region is believed to control seizure generalization, by acting as a “seizure gate.” This ketone effect was amplified in larger cells that fired more rapidly. Ketone body effects were able to be eliminated via pharmacological extirpation or gene knock-out of KATP channels. Researchers suggest that epilepsy may be treatable by enhancing the natural activity-limiting function of KATP neuron channels by ketone bodies or glycolytic restriction.

Ketone bodies were discovered to reduce SNr neuron-firing rates while requiring the presence of functional KATP channels and GABAB signaling. Researchers proposed that this requirement is a result from opening KATP channels, which inhibits GABAB of SNr neuron firing. KATP channels have also been reported to be activated by G-proteins and G-protein-coupled receptors. Specifically, KATP channels were discovered to be activated by GABAB¬ receptors in central neurons. This indicates that some degree of GABAB modulation is essential for determining the point at which these channels may be activated. It is speculated that this activation point may be triggered by direct G-protein modulation or phosphoinositide lipid level modulation.

During ketosis, ketone bodies are capable of partially or mostly replacing glucose functionality as a fuel source for the brain. Ketogenic diet studies in rodents revealed that the glycolytic pathway substrate levels were altered, when analyzing whole-brain extracts. Therefore, glycolysis modification was the identifiable difference between glucose and ketone body metabolism. Effective compartmentation of ATP molecules produced from glycolysis may potentially play a significant role in controlling and fuelling plasma membrane processes. Glycolytic enzymes are also believed to be closely associated with KATP channels that may support this ATP compartmentation.

Direct connections between levels of electrical activity and ATP consumption at the plasma membrane were observed. These connections allow for KATP channels to play an anticonvulsant role, and predict a correlation between the strength of ketone body effects and initial neuronal firing rates. It is proposed that a ketogenic diet may function as an epilepsy treatment by reducing glycolytic ATP production. This reduction is accomplished by lowering electrical activity levels needed to trigger this protective mechanism. Pertinence of the glycolytic inhibition mechanism through a specific diet is supported by dietary therapy conducted on humans, known as the Low Glycemic Index Treatment. When adhering to a ketogenic diet, this treatment may potentially moderate or lower the rate of glycolysis.

References:

http://www.jneurosci.org/cgi/content/full/27/14/3618

Research on Cardiolipin

Cardiolipin (CL) has been discovered to be substantially depleted in diabetic myocardium six weeks after streptozotocin (STZ) injections. Accompanying this depletion was an increase in triacylglycerol content and changes in polar lipid molecular species. Left in a diabetic state for more than six weeks, the myocardium experienced complementary lipid indicators of diabetic cardiomyopathy. This made researchers incapable of identifying time-dependent diabetic myocardium lipid alterations. Recent advancements in shotgun lipidomics have allowed for the identification of abundant losses of CL molecules in STZ-treated hearts, during the early stages of diabetes. These early stages also indicated the remodeling of any remaining CL molecules. Specifically, there was a nigh 16-fold increase in 18:2-22:6-22:6-22:6 CL content. These metabolic CL modifications precede diabetic myocardial triacylglycerol accumulation, and occur within a few days of inducing a diabetic state. Identifiably, ob/ob mice exhibited a significant redistribution from 18:2 fatty acids (FA)-containing CL molecules to 22:6 FA-containing CL molecules. Altogether, these results illustrate the early diabetic CL remodeling and hydrolysis alterations congruous with CL content fluctuation within precipitating mitochondrial dysfunction, during diabetic cardiomyopathy.

Any alteration in the content or structure of CL molecules has been proven to affect the biophysical environment of mitochondrial membranes in a way that modified the potentials and activities of mitochondrial transmembrane enzymes. These modifications evidently altered the overall dynamics and bioenergetics of mitochondria. Inferentially, quantitative and qualitative CL changes are expected to contribute to mitochondrial dysfunction in diabetic hearts. These occurrences, however, did not necessitate any physical functional mitochondrial alterations, during early stages of diabetes.

A significant biomarker for diabetic cardiomyopathy was triacylglycerol (TG) accumulation. The identified quantitative and qualitative changes in CL content have been proven to inhibit mitochondrial function and, more specifically, to attenuate the mitochondria’s ability to remove any TG excess in diabetic hearts. This defect is believed to be onset by electron transport chain inefficiency as a result of nonphysiological CL content. This inefficiency is also thought to increase negative redox potential and reduce mitochondrial fatty acid -oxidation flux.

References:

http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=17487985

Research on analytical techniques

Advancements in the field of metabolomics have furthered the research and understanding of disease states pertinent to metabolic pathways. Specifically, metabolomics information improves the capability of identifying drug off-target effects. The progression of metabolomics is largely a result of advancements in high-precision high-throughput mass spectrometry. As the field of metabolomics continues to develop, there is an apparent need to apply bioinformatics tools and methods. Researchers have systematically analyzed and quantified more than 800 metabolites in blood plasma samples of diabetic mice under rosiglitazone treatment. Metabolic phenotypes of diabetes and medication were proven to be capable of statistically objective recovery. Methylglutarylcarnitine content in healthy and diabetic mice were both oppositely impacted when treated with rosiglitazone. Metabolite concentration ratio analysis revealed new potential biomarkers of diabetes, which include N-hydroxyacyloylsphingosyl-phosphocholines SM(OH)28:0 and SM(OH)26:0. A hierarchical clustering technique on partial eta-squared values was employed to identify functionally related groups of metabolites that indicated shifts from lysophosphatidylcholines to phosphatidylcholines. These shifts implied diabetes disease states.

Researchers conducted bioinformatics analyses comprising of a standard experimental setting that included independent “state” and “medication” factors. Targeted quantitative metabolomics was employed, and covered a wide range of analytes. Researchers first focused on reproducing already identified metabolic effects of rosiglitazone on diabetes. This “medication” factor expectedly impacted non-esterified fatty acid metabolism and decreased long-chain acylcarnitine concentrations. The next experimental objective involved identifying new diabetes related compounds and/or pathways. The observed effects on this “state” factor indicated the presence of previously unidentified metabolites involved in experimental rosiglitazone treatment of diabetes. These groups of metabolites prominently consisted of glycero-phosphocholines and sphingomyelins, and were not observed during any single metabolite analysis. Researchers also made an effort to identify metabolites that might enhance the effects of rosiglitazone or provide a better understanding of diabetes and its effects on metabolic pathways and mechanisms. The detection of new metabolites, including glycero-phosphocholines, raises the possibility of interpreting the functionality of entire metabolite classes in disease states, and medicated disease states.

References:

http://endo.endojournals.org/cgi/rapidpdf/en.2007-1747v1

Research on Metabolic Profiling of Ovarian Carcinoma

Metabolic Profiling of Ovarian Carcinoma

Denkert C, Budczies J, Kind T, Weichert W, Tablack P, Sehouli J, Niesporek S, Koensgen D, Dietel M, Fiehn O (2006) GC-TOF based metabolic profiling reveals different metabolite patterns in invasive ovarian carcinomas and ovarian borderline tumors. Cancer Res. 66, 10795-10804

Summary: In this study, the researchers determined that there are indeed several differences in metabolism between malignant ovarian carcinomas and borderline ovarian tumors. They conducted gas chromatography time-of-flight mass spectrometry to identify metabolites in homogenized tumor samples, and then conducted several statistical analyses to prove that approximately 50 metabolites can be used as biomarkers to differentiate between malignant carcinomas and borderline tumors. The researchers also discovered that the increased presence of nitrogen-donating amino acids and molecules important in protein synthesis in the malignant samples indicate a higher metabolic turnover rate, or increased cell replication.

Terms: • metabolite profiling: determining the presence and amount of each metabolite in a sample

• principal component analysis (PCA): "a technique used to reduce multidimensional data sets to lower dimensions for analysis" (Wikipedia)

• differential regulation: comparison of metabolic regulation in two different systems or one system under two different circumstances

• flux: "the rate of flow of energy or particles across a given surface" (WordNet)

• permutation (in statistical analysis): "rearrangement of the elements of a set" (American Heritage Dictionary)

Relevance: This study demonstrates that the presence of certain metabolites can determine the malignancy of aberrant tumor cells in ovarian tissue. One noted change was an increase in metabolites involved in the tricarboxylic acid cycle (TCA), which indicates an increased need for energy by malignant tumor cells. This is similar to what we discussed in class about the fed and starved states regulating the glycolysis and gluconeogenesis cycles, with respect to the concentration of chemical energy (ATP).

Research on Dynamics of the Cellular Metabolome during Human Cytomegalovirus Infection

Dynamics of the Cellular Metabolome during Human Cytomegalovirus Infection

http://www.pubmedcentral.nih.gov/picrender.fcgi?tool=pmcentrez&artid=1698944&blobtype=pdf

Summary: When a human cell is infected with cytomegalovirus (HCMV), what happens to the metabolic components in the glycolysis pathway, tricarboxylic acid cycle and pyrimidine biosynthesis? This was answered by the use of liquid chromatography-tandem mass spectrometry, hydrophilic interaction and selected reaction monitoring. 63 different metabolites were tested for their levels in a cultured (vitro) fibroblast that was infected with Human Cytomegalovirus. The results of the experiment were that the levels of the metabolites were higher in the infected cells than the uninfected cells, thus the infection changes metabolic homeostasis.

Terms: Metabolic flux- The rate that metabolites flow through a metabolic pathway.

Biopolymers- Produced by living organism examples starch, proteins, peptides, DNA and RNA

Glycolysis- The breaking of a glucose molecule, through a series of enzymes and products to produce two pyruvate molecules and four ATP.

Tricarboxylic acid cycle- A metabolic pathway in aerobic organisms that uses carbohydrates, fats and proteins and converts them to carbon dioxide, water and energy.

Pyrimidine pathway- The making of pyrimidine nucleotides examples thymine, uracil and cytosine in DNA and RNA.

Hydrophilic interaction chromatography- A tool used to separate compounds by hydrophobicity.

Selected Reaction Monitoring scan event- (SRM) A tool in which a specific product ion is detected, by its specific mass fragment.

Relevance: This paper tested the metabolites in the glycolysis pathway. They found a increase in hexose phosphate, FBP, DHAP, 3-P glycerate and PEP when a cell is infected with the Human Cytomegalovirus compared to a mock-infected cell. The paper also talked about the different ways a metabolite level can change or a change in flux.

Research on Effect of Phenylalanine Metabolites on the Activities of Enzymes of Ketone-Bodies

Effect of Phenylalanine Metabolites on the Activities of Enzymes of Ketone-Body Utilization in Brain of Suckling Rats http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1164225&blobtype=pdf

Summary: The main focus of this resource is to identify why people with defects in their phenylketonuria develop brain abnormalities. Phenylketonuria is caused by a deficiency of phenylalanine hydroxylase enzyme. Phenylalanine and its derivatives build up to toxic levels in the body and interfere with other pathways since they cannot be converted to tyrosine fast enough or at all. If the harmful metabolites are detected it may be possible to treat phenylketonuria in another way other than the current treatment of a low phenylalanine diet.

Terms: extrahepatic- outside of, or unrelated to the liver (http://cancerweb.ncl.ac.uk/cgi-bin/omd?extrahepatic)

thiolysis- is a reaction with a thiol (R-SH) that cleaves one compound into two. This reaction is similar to hydrolysis, which involves water instead of a thiol. This reaction is seen in β-oxidation of fatty acids. (http://en.wikipedia.org/wiki/Thiolysis)

acetoacetyl-CoA thiolase- (also called thiolase II) is specific for the thiolysis of acetoacetyl-CoA and involved in biosynthetic pathways such as poly -hydroxybutyrate synthesis or steroid biogenesis. (https://bioinformatics.ccr.buffalo.edu/cgi-bin/software/pfam/getdesc?name=Thiolase_N)

3-oxo acid CoA-transferase- An enzyme catalyzing the reversible conversion of acetoacetyl-CoA and succinate into succinyl-CoA and acetoacetate; malonyl-CoA can substitute for succinyl-CoA and a few other 3-oxo acids for the acetoacetate; an important step in order for the ketone bodies to serve as a fuel for extrahepatic tissues. (http://cancerweb.ncl.ac.uk/cgi-bin/omd?3-oxoacid-CoA+transferase)

3-hydroxybutyrate dehydrogenase- Catalysis of the reaction: (R)-3-hydroxybutanoate + NAD+ = acetoacetate + NADH + H+ (http://www.reactome.org/cgi-bin/eventbrowser?DB=gk_current&FOCUS_SPECIES=Gallus%20gallus&ID=11646&) Citrate synthase- The enzyme citrate synthase exists in nearly all living cells and stands as a pace-making enzyme in the first step of the Citric Acid Cyle (or Krebs Cycle). Citrate synthase catalyzes the condensation reaction of the two-carbon acetate residue from acetyl coenzyme A and a molecule of four-carbon oxaloacetate to form the six-carbon citrate (http://en.wikipedia.org/wiki/Citrate_synthase)

Relevance: The research done by this group used the Lineweaver-Burke Plot to determine the effects of phenylpyruvate, phenylacetate, and o-hydroxyphenylacetate (all phenylalanine metabolites) on 3-hydroxybutyrate dehydrogenase. The plots show that they work as inhibitors. Lineweaver-Burke plots were also used to detect how phenylalanine and its metabolites affect Acetoacteyl-CoA thiolase activity. They found from these plots that phenylpyruvate is the only strong inhibitor out of the phenylalanine metabolites. Phenylpyruvate has been shown to competitively inhibit citrate synthase activity which we learned means that it must somehow bind to the active site of the enzyme and block the intended substrate from getting to it. The Lineweaver-Burke plots demonstrate this by the increased slope (Km/Vmax) when phenylalanine is added. Apparently, hexokinase is also inhibited by phenylpyruvate which would mean that glucose would not be able to be converted to glucose-6-phosphate and there could be problems with completing the glycolysis pathway. Pyruvate carboxylase is also inhibited by phenylpyruvate which means that gluconeogenesis would have difficulty occurring since Pyruvate would not be able to be converted to oxaloacetate. Phosphogluconate dehydrogenase is another enzyme inhibited by phenylpyruvate which would mean that the pentose phosphate pathway would be inhibited also since 6-phosogluconate wouldn’t be as readily converted to d-ribulose 5-phosphate. Even though we have not talked about acetyl-CoA carboxylase yet in class, one of the components of it is biotin, which we learned is a cofactor in gluconeogenesis and is also a cofactor for this enzyme. The general idea is similar to Box10-2 which talks about diseases resulting from lipid accumulations except Phenylketonuria is a disease from accumulations of amino acids.

Research on Environmental Metabolomics

Environmental Metabolomics

http://www.actionbioscience.org/genomic/viant.html

Summary: This particular resource is an article dealing with Environmental Metabolomics as applied to the events of disease and toxicity in wildlife. It gives a general description of Metabolomics as a whole but goes more deeply into several investigations regarding specific organisms and their diseases. The overview covers the applications of Metabolomics to human concerns initially, discussing the detection of coronary heart disease using NMR metabolomics, the prediction of subarachnoid hemorrhages by analysis of cerebral spinal fluid, and sample classification methods for progressive neurological disease.

The article discussed the basics of Metabolomics in terms of disease diagnosis and monitoring. It discussed how Metabolomics uses Bioinformatics to manage the sheer volumes of metabolic information gleaned in this system of study and how the tools of the trade tend more towards chemistry concerns. It especially went over the advantages of metabolomics for disease diagnosis because it works on phenotype and the effects of disease and toxicity on phenotype are readily apparent. The main focus for Environmental Metabolomics is disease and toxicity occurrence and formation in various forms of wildlife. The article used several examples to show the uses that Metabolomics has in this area. It discussed the identification of cancer in Marine Catfish, chemical risk assessment methods in sentinel species of fish, mammals and especially earthworms and to monitor withering disease in the Californian Red Abalone. It also made it clear that further developments and applications are inevitable as this field expands.

Terms:

Hemolymph - Another term for blood

Toxicity - The level of damage done to a biological system of a chemical or substance during exposure. The state of being capable of doing damage to a biological system by a chemical or substance during exposure.

Metabolome - The collection of all the metabolites within a cell and their interactions.

Angiography - a method of diagnosing and treating medical conditions by taking images of major blood vessels, oftentimes with a contrast material added.

Subarachnoid hemorrhage - a sudden hemorrhage of a blood vessel or vessels over the surface of the brain

Amyotrophic lateral sclerosis - A progressive neurological disease in which loss of motor nerve cells in the spine and brain produces muscle paralysis and eventually death. Often referred to as "Lou Gehrig's disease".

Aquaculture - The growing and harvesting of marine life for food or other purposes.

Withering syndrome - A disease that infects several species of abalone (a common shellfish) through infection in the epithelium of the intestinal tract by an intracellular prokaryote, with morphological characteristics of the class Proteobacteria, order Rickettsiales and family Rickettsiaceae. A new genus and species has been suggested for this organism (Candidatus Xenohaliotis californiensis). The disease causes lethargy, atrophy of the foot muscle, retraction of visceral muscles and eventual death.

Relevance: We discussed the effects of certain metabolites being denied the cell and also how different conditions effect the workings of glycolysis and gluconeogenesis in terms of regulation. This article ties into this since diseases and toxic compounds can impact the regulation of many metabolic pathways and sometimes involve the denial of certain metabolites for a pathway.

Research on regulation of central metabolic pathways in a fungal pathogen

Niche-specific regulation of central metabolic pathways in a fungal pathogen

(http://www.pubmedcentral.nih.gov/articlerender.fcgi?&pubmedid=16681837)

Summary: The focus of this article is on how Candida albicans adjusts its metabolic pathways in relation to carbon at different stages of a systemic infection to maintain paradox. The inspiration for this study was the paradox of how the glyoxylate cycle, known to be required for Candida albicans virulence, could be used during bloodstream infections when glyoxylate cycle genes are repressed by the glucose levels in blood. The answer was found to be the adaptability of metabolic pathways in Candida albicans; glyoxylate and gluconeogenesis are actually upregulated as the organism is phagocytosed by neutrophils and macrophages and downregulated in favor of glycolysis at other stages.

Terms: -The glyoxylate pathway occurs in only a few vertabrates and is much more common in plants and microorganisms, such as candida albicans. It allows the synthesis of carbohydrates from fats.

-Candida albicans is a common diploid fungus that is rarely harmful to humans except in cases of immunocompromised patients, where it can cause dangerous and even fatal systemic infections.

-Candidiasis is a clinical term for a fungal infection of Candida species. Candida albicans are a part of normal flora for humans that can sometimes result in superficial skin, oral, and vaginal infections.

-Candidemia is a more severe, systemic infection of Candida that is often found in HIV and transplant patients. It can spread through the bloodstream and infect the kidneys with fatal consequences if left untreated.

-PCK1 is a key control gene that regulates gluconeogenesis. In the study, its expression was monitored to determine at what points of infection candida albicans was relying on gluconeogenesis.

Relevance: The information in this article related to what we've studied in the course to date because it was a practical examination of how pathways are used in vivo. It also gives an indication of the depth of detail involved in metabolomics. Even a relatively simple organism like Candida albicans has to undergo a complex shift in metabolic pathways to survive and thrive within an animal host.

Research on Human Serum Metabolite in Health and Disease

Human Serum Metabolite in Health and Disease

http://www.husermet.org/

Summary: The main aim of this study is to establish 'baseline' ranges of metabolites in human serum that are caused by such 'disease-independent' factors such as age, gender, lifestyle diet etc. These baselines from numerous healthy individuals would then be compared to metabolite values from patients in an attempt to determine whether metabolite values can be used as a prediction tool for disease as well as possibly monitor patient response to treatment. The two main diseases being focused on are Alzheimer's Disease and Ovarian cancer.

Terms:

Metabolite - A substance necessary for or taking part in a particular metabolic process.

Biomarker – A specific physical trait used to measure or indicate the effects or progress of a disease or condition

Metabolome – The complete set of small-molecule metabolites to be found within a biological sample

Pathophysiology - the functional changes associated with a disease.

Transcriptome – The complete set of al mRNA molecules produced by a cell.

FTICR – Fourier transform ion cyclotron resonance mass spectrometry. This is a type of mass spectrometer for determining the mass-to-charge ratio (m/z) of ions based on the cyclotron frequency of the ions in a fixed magnetic field.

Relevance: This research relates to our brief class discussion on using a patient's metabolic profile as a tool in the diagnosis of disease, as well as prediction/monitoring of the effectiveness of drugs given to the patient. Analysis of a patient's metabolic profile can be utilised as an additional diagnostic tool, especially when other clinical symptoms are conflicting or not apparent. For example, the clinical distinction between Alzheimer's disease and normal age-related memory-loss is tenuous and subjective – identification of a metabolite unique to Alzheimer's disease would prove to be valuable in early diagnosis and treatment.

Research on Mitochondrial Disorders Using Intracellular ATP Reporters

New Insights into the Bioenergetics of Mitochondrial Disorders Using Intracellular ATP Reporters http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=196555

Summary: In this paper, the scientists are trying to see if ATP is compartmentalized within cells and different pools of ATP sustain different cell functions. They also wanted to see, more specifically which cell compartments are more depleted of ATP and if certain ATP dependent functions are affected differently in mitochondrial diseases. To show this, the experiment used wild-type firefly luciferase and made chimeric constructs with the required mutations which were then inserted into human cell lines using a mammalian expression vector pCDNA3.0. The results showed that the ATP levels decreased markedly in the mutant forms under oxidative conditions suggesting that ATP is transported from the mitochondria to other energy extensive compartments even under mitochondrial OXPHOS.

Terms:

Luciferase – It is the generic name for enzymes used in nature for bioluminescence.

Phosphocreatine – A phosphorylated creatine molecule that is an important energy store in the skeletal muscle.

Lactic acidosis – It is a condition caused by the buildup of lactic acid in the body and leads to acidification of the blood.

Osteosarcoma – It is the most common type if malignant bone cancer.

Retinitis pigmentosa – Consists of a group of inherited disorders in which abnormalities of the photoreceptors or the retinal pigment epithelium of the retina lead to progressive vision loss.

Relevance: The article relates to what we have been doing in class because it deals with the production of energy, ATP. Here we see how mutations in the mitochondria affect the production of ATP in the different cellular compartments. Also it shows that there is indeed a different concentration of ATP in the different cellular organelles.

Research on Metabolite profiling of human colon carcinoma

Metabolite profiling of human colon carcinoma--deregulation of TCA cycle and amino acid turnover. http://www.ncbi.nlm.nih.gov/pubmed/18799019?ordinalpos=16&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Metabolomics identifies perturbations in human disorders of propionate metabolism

http://www.ncbi.nlm.nih.gov/pubmed/17951291?ordinalpos=38&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Articles and Web Pages for Review and Inclusion

Peer-Reviewed Article #1:

Metabolomics: moving towards personalized medicine

Italian Journal of Pediatrics 2009, 35:30

Reviewed by Sara B

Main Focus

Identify the main focus of the resource. Possible answers include specific organisms, database design, integration of information, but there are other possibilities as well.

New Terms

New Term 1: Definition. (source: http://)
New Term 2: Definition. (source: http://)
New Term 3: Definition. (source: http://)
New Term 4: Definition. (source: http://)
New Term 5: Definition. (source: http://)
New Term 6: Definition. (source: http://)
New Term 7: Definition. (source: http://)
New Term 8: Definition. (source: http://)
New Term 9: Definition. (source: http://)
New Term 10: Definition. (source: http://)

Summary

Enter your article summary here. Please note that the punctuation is critical at the start (and sometimes at the end) of each entry. It should be 300-500 words. What are the main points of the article? What questions were they trying to answer? Did they find a clear answer? If so, what was it? If not, what did they find or what ideas are in tension in their findings?

Relevance to a Traditional Metabolism Course

Enter a 100-150 word description of how the material in this article connects to a traditional metabolism course. Does the article relate to particular pathways (e.g., glycolysis, the citric acid cycle, steroid synthesis, etc.) or to regulatory mechanisms, energetics, location, integration of pathways? Does it talk about new analytical approaches or ideas? Does the article show connections to the human genome project (or other genome projects)?

Peer-Reviewed Article #2:

Metabolomics Reveals Metabolic Biomarkers of Crohn's Disease

PLoS One. 2009; 4(7): e6386. Reviewed by Divya Kumar

Main Focus

This article was focused on the comparison of metabolites from numerous metabolic pathways in the feces of patients with Crohn's Disease and in healthy people. The purpose of this study was to determine whether the metabolite composition of the feces could be used to diagnose Crohn's Disease, as there are currently no adequate non-intrusive methods of diagnosing and monitoring the disease.

New Terms

Dysbiosis: An imbalance in the intestinal bacteria that precipitates changes in the normal activities of the gastrointestinal tract or vagina, possibly resulting in health problems. (source: http://medical-dictionary.thefreedictionary.com/Dysbiosis)
Microbiome: The entourage of associated microflora in a host.(source: http://www.nature.com/nrmicro/journal/v3/n5/glossary/nrmicro1152_glossary.html)
Serological: Of or relating to serology, the branch of medical science that deals with serums; especially with blood serums and disease (source: http://wordnetweb.princeton.edu/perl/webwn?s=serology)
Crohn's Disease: Crohn's disease is an ongoing disorder that causes inflammation of the digestive tract, also referred to as the gastrointestinal (GI) tract. Crohn's disease can affect any area of the GI tract, from the mouth to the anus, but it most commonly affects the lower part of the small intestine, called the ileum. The swelling extends deep into the lining of the affected organ. The swelling can cause pain and can make the intestines empty frequently, resulting in diarrhea.(source:http://digestive.niddk.nih.gov/ddiseases/pubs/crohns/)
Cohort study: A type of study involving a group of people sharing common characteristics, usually done to test an association between cause an effect. (source: http://en.wikipedia.org/wiki/Cohort_study)
Prostaglandin: A potent substance that acts like a hormone and is found in many bodily tissues (and especially in semen); produced in response to trauma and may affect blood pressure, metabolism and smooth muscle activity (source: http://wordnetweb.princeton.edu/perl/webwn?s=prostaglandin)
Discordant: A twin pair (or set of individuals) in which one member exhibits a certain trait and the other does not. (source:http://www.kumc.edu/gec/gloss.html)
Concordant: Presence of the same trait in both members of a pair of twins (or set of individuals).(source: http://www.kumc.edu/gec/gloss.html)
Mononuclear cells: Lymphocytes, dendritic cells and monocytes/macrophages. These are usually found in the blood, but also in tissues.(source: http://www.nature.com/nrm/journal/v4/n1/glossary/nrm1005_glossary.html)

Summary

Crohn's Disease is an inflammatory bowel disease caused by a dysbiosis of intestinal microbes. Its etiology is largely unknown, but its main symptom is chronic inflammation of the gastrointestinal tract. Examining fecal extracts of test subjects has revealed that several metabolites, when present in the feces, are indicative of Crohn's Disease. A cohort study using 15 twins was done in this experiment. The sample cohort contained 7 healthy pairs of twins, 4 pairs that were discordant for predominantly colinic Crohn's Disease (CCD), 2 pairs discordant for predominantly ileal Crohn's Disease (ICD), and 2 pairs that were concordant for ICD Fecal samples from the subjects were collected, diluted and examined for metabolites using FT-ICR-MS analysis. The partial least squares (PLS) analysis used to examine the data from this experiment showed a clear difference among the metabolites in the feces of people with ICD, people with CCD, and healthy people. The data from this PLS analysis was the first to show a distinction between metabolites of healthy people and those who have Crohn's Disease, proving that metabolomics can be used as a method of accurately diagnosing Crohn's Disease. Metabolites from a wide range of pathways differentiated healthy from diseased individuals. A complete list of these metabolites is shown in Table 1 of the article. The presence of metabolites relating to tyrosine metabolism was of particular interest to the scientists because this confirms previous reports of increased transcripts of genes involved in tyrosine metabolism in peripheral blood mononuclear cells in patients with Crohn's Disease. The amino acids tryptophan and phenylalanine in the feces indicated that the individual had ICD. Because one subject with ICD interestingly had low tryptophan and phenylalanine levels in his/her feces and also had low E. coli levels in the gut, an interesting follow up study would be to look for links between E. coli abundance and amino acid levels in the gut. Also, it was found that several masses corresponding to metabolites within the bile acid biosynthesis pathway could be used to distinguish between the disease phenotypes. For example, masses corresponding to both saturated and unsaturated fatty acids were more prevalent in patients with ICD compared to the other two groups. The fact that arachidonic acid levels are high in colinic mucosa of patients is interesting because arachidonic acid is an inflammatory mediator and can attract leukocytes. This provides an explanation for the colinic inflammation that CD patients experience, and also offers a possible means of alleviating the negative symptoms of CD in patients. In general, bacteria that were more abundant in people with ICD were those that were associated with bile acids, and bacteria that were more abundant in individuals with healthy or CCD phenotypes corresponded more with phospholipids and flavin mononucleotides. This study introduces some exciting results that could likely lead to effective, non-invasive methods of detecting and monitoring Crohn's Disease, thus improving the quality of care received by those who suffer from the disease.

Relevance to a Traditional Metabolism Course

Crohn's Disease is an inflammatory bowel disease caused by a dysbiosis of intestinal microbes. Its etiology is largely unknown, but its main symptom is chronic inflammation of the gastrointestinal tract. Examining fecal extracts of test subjects has revealed that several metabolites, when present in the feces, are indicative of Crohn's Disease. A cohort study using 15 twins was done in this experiment. The sample cohort contained 7 healthy twin pairs, 4 pairs that were discordant for predominantly colinic Crohn's Disease (CCD), 2 pairs discordant for predominantly ileal Crohn's Disease (ICD), and 2 pairs that were concordant for ICD. Fecal samples from the subjects were collected, diluted and examined for metabolites using FT-ICR-MS analysis. The partial least squares (PLS) analysis used to examine the data from this experiment showed a clear difference among the metabolites in the feces of people with ICD, people with CCD, and healthy people. The data from this PLS analysis was the first to show a distinction between metabolites of healthy people and those who have Crohn's Disease, proving that metabolomics can be used as a method of accurately diagnosing Crohn's Disease. Metabolites from a wide range of pathways differentiated healthy from diseased individuals. A complete list of these metabolites is shown in Table 1 of the article. The presence of metabolites relating to tyrosine metabolism was of particular interest to the scientists because this confirms previous reports of increased transcripts of genes involved in tyrosine metabolism in peripheral blood mononuclear cells in patients with Crohn's Disease. The amino acids tryptophan and phenylalanine in the feces indicated that the individual had ICD. Because one subject with ICD interestingly had low tryptophan and phenylalanine levels in his/her feces and also had low E. coli levels in the gut, an interesting follow up study would be to look for links between E. coli abundance and amino acid levels in the gut. Also, it was found that several masses corresponding to metabolites within the bile acid biosynthesis pathway could be used to distinguish between the disease phenotypes. For example, masses corresponding to both saturated and unsaturated fatty acids were more prevalent in patients with ICD compared to the other two groups. The fact that arachidonic acid levels are high in colinic mucosa of patients is interesting because arachidonic acid is an inflammatory mediator and can attract leukocytes. This provides an explanation for the colinic inflammation that CD patients experience, and also offers a possible means of alleviating the negative symptoms of CD in patients. In general, bacteria that were more abundant in people with ICD were those that were associated with bile acids, and bacteria that were more abundant in individuals with healthy or CCD phenotypes corresponded more with phospholipids and flavin mononucleotides. This study introduces some exciting results that could likely lead to effective, non-invasive methods of detecting and monitoring Crohn's Disease, thus improving the quality of care received by those who suffer from the disease.