ATP-generating Pathways Research Articles

AMPK inhibitor compound C suppresses cell proliferation by induction of apoptosis and autophagy in human colorectal cancer cells

AMP-activated protein kinase (AMPK) is a main regulator of energy metabolism through the inhibition of biosynthetic pathways and enhancement of ATP-generating pathways. However, targeting AMPK as anti-tumor therapy remains controversial. In this study, we examined the effect of compound C, a small molecule inhibitor of AMPK, on the proliferation of several human colorectal cancer cell lines with diverse characteristics. Four human colorectal cancer cell lines (HCT116, DLD-1, SW480, and KM12C) were treated with compound C. Cell viability was determined by MTS assay. Cell cycle prolife was analyzed by flow cytometry. Acidic vesicular organelles were detected by acridine orange staining. Protein levels were measured by western blotting. Compound C inhibited the growth of four cell lines in a dose-dependent manner and caused G(2) /M arrest. Compound C increased sub-G(1) cell population and induced chromatin condensation and cleavage of PARP in HCT116 and KM12C cells, while it induced acidic vesicular formation and conversion of LC3-I to autophagosome-associated LC3-II in DLD-1 and SW480 cells. Survivin, an anti-apoptotic protein, was down-regulated in all cell lines treated with compound C. Compound C induces apoptotic or autophagic death in colorectal cancer cells and the preferred death mode is cell type-dependent.

Inhibition of Glycolytic Enzymes Mediated by Pharmacologically Activated p53

Unique sensitivity of tumor cells to the inhibition of glycolysis is a good target for anticancer therapy. Here, we demonstrate that the pharmacologically activated tumor suppressor p53 mediates the inhibition of glycolytic enzymes in cancer cells in vitro and in vivo. We showed that p53 binds to the promoters of metabolic genes and represses their expression, including glucose transporters SLC2A12 (GLUT12) and SLC2A1 (GLUT1). Furthermore, p53-mediated repression of transcription factors c-Myc and HIF1α, key drivers of ATP-generating pathways in tumors, contributed to ATP production block. Inhibition of c-Myc by p53 mediated the ablation of several glycolytic genes in normoxia, whereas in hypoxia down-regulation of HIF1α contributed to this effect. We identified Sp1 as a transcription cofactor cooperating with p53 in the ablation of metabolic genes. Using different approaches, we demonstrated that glycolysis block contributes to the robust induction of apoptosis by p53 in cancer cells. Taken together, our data suggest that tumor-specific reinstatement of p53 function targets the "Achilles heel" of cancer cells (i.e. their dependence on glycolysis), which could contribute to the tumor-selective killing of cancer cells by pharmacologically activated p53.

Adrenergic Regulation of AMP-activated Protein Kinase in Brown Adipose Tissue in Vivo

AMP-activated protein kinase (AMPK), an evolutionarily conserved serine-threonine kinase that senses cellular energy status, is activated by stress and neurohumoral stimuli. We investigated the mechanisms by which adrenergic signaling alters AMPK activation in vivo. Brown adipose tissue (BAT) is highly enriched in sympathetic innervation, which is critical for regulation of energy homeostasis. We performed unilateral denervation of BAT in wild type (WT) mice to abolish neural input. Six days post-denervation, UCP-1 protein levels and AMPK α2 protein and activity were reduced by 45%. In β(1,2,3)-adrenergic receptor knock-out mice, unilateral denervation led to a 25-45% decrease in AMPK activity, protein expression, and Thr(172) phosphorylation. In contrast, acute α- or β-adrenergic blockade in WT mice resulted in increased AMPK α Thr(172) phosphorylation and AMPK α1 and α2 activity in BAT. But short term blockade of α-adrenergic signaling in β(1,2,3)-adrenergic receptor knock-out mice resulted in decreased AMPK activity in BAT, which strongly correlated with enhanced phosphorylation of AMPK on Ser(485/491), a site associated with inhibition of AMPK activity. Both PKA and AKT inhibitors attenuated AMPK Ser(485/491) phosphorylation resulting from α-adrenergic blockade and prevented decreases in AMPK activity. In vitro mechanistic studies in BAT explants showed that the effects of α-adrenergic blockade appeared to be secondary to inhibition of oxygen consumption. In conclusion, adrenergic pathways regulate AMPK activity in vivo acutely via alterations in Thr(172) phosphorylation and chronically through changes in the α catalytic subunit protein levels. Furthermore, AMPK α Ser(485/491) phosphorylation may be a novel mechanism to inhibit AMPK activity in vivo and alter its biological effects.

AMP-activated Protein Kinase Antagonizes Pro-apoptotic Extracellular Signal-regulated Kinase Activation by Inducing Dual-specificity Protein Phosphatases in Response to Glucose Deprivation in HCT116 Carcinoma

Mitogen-activated protein kinase (MAPK) pathways are involved in the regulation of cellular responses, including cell proliferation, differentiation, cell growth, and apoptosis. Because these responses are tightly related to cellular energy level, AMP-activated protein kinase (AMPK), which plays an essential role in energy homeostasis, has emerged as another key regulator. In the present study, we demonstrate a novel signal network between AMPK and MAPK in HCT116 human colon carcinoma. Glucose deprivation activated AMPK and three MAPK subfamilies, extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase (JNK), and p38 MAPK. Under these conditions, inhibition of endogenous AMPK by expressing a dominant-negative form significantly potentiated ERK activation, indicating that glucose deprivation-induced AMPK is specifically antagonizing ERK activity in HCT116 cells. Moreover, we provide novel evidence that AMPK activity is critical for p53-dependent expression of dual-specificity phosphatase (DUSP) 1 & 2, which are negative regulators of ERK. Notably, ERK exhibits pro-apoptotic effects in HCT116 cells under glucose deprivation. Collectively, our data suggest that AMPK protects HCT116 cancer cells from glucose deprivation, in part, via inducing DUSPs, which suppresses pro-apoptotic ERK, further implying that a signal network between AMPK and ERK is a critical regulatory point in coupling the energy status of the cell to the regulation of cell survival.

Phylogenetic Distributions and Histories of Proteins Involved in Anaerobic Pyruvate Metabolism in Eukaryotes

Protists that live in low oxygen conditions often oxidize pyruvate to acetate via anaerobic ATP-generating pathways. Key enzymes that commonly occur in these pathways are pyruvate:ferredoxin oxidoreductase (PFO) and [FeFe]-hydrogenase (H(2)ase) as well as the associated [FeFe]-H(2)ase maturase proteins HydE, HydF, and HydG. Determining the origins of these proteins in eukaryotes is of key importance to understanding the origins of anaerobic energy metabolism in microbial eukaryotes. We conducted a comprehensive search for genes encoding these proteins in available whole genomes and expressed sequence tag data from diverse eukaryotes. Our analyses of the presence/absence of eukaryotic PFO, [FeFe]-H(2)ase, and H(2)ase maturase sequences across eukaryotic diversity reveal orthologs of these proteins encoded in the genomes of a variety of protists previously not known to contain them. Our phylogenetic analyses revealed: 1) extensive lateral gene transfers of both PFO and [FeFe]-H(2)ase in eubacteria, 2) decreased support for the monophyly of eukaryote PFO domains, and 3) that eukaryotic [FeFe]-H(2)ases are not monophyletic. Although there are few eukaryote [FeFe]-H(2)ase maturase orthologs characterized, phylogenies of these proteins do recover eukaryote monophyly, although a consistent eubacterial sister group for eukaryotic homologs could not be determined. An exhaustive search for these five genes in diverse genomes from two representative eubacterial groups, the Clostridiales and the alpha-proteobacteria, shows that although these enzymes are nearly universally present within the former group, they are very rare in the latter. No alpha-proteobacterial genome sequenced to date encodes all five proteins. Molecular phylogenies and the extremely restricted distribution of PFO, [FeFe]-H(2)ases, and their associated maturases within the alpha-proteobacteria do not support a mitochondrial origin for these enzymes in eukaryotes. However, the unexpected prevalence of PFO, pyruvate:NADP oxidoreductase, [FeFe]-H(2)ase, and the maturase proteins encoded in genomes of diverse eukaryotes indicates that these enzymes have an important role in the evolution of microbial eukaryote energy metabolism.

Does diabetes therapy influence the risk of cancer?

Type 2 diabetes is associated with three of the five leading causes of cancer mortality in the USA—carcinoma of the colon, pancreas and breast (postmenopausal) [1]. The excess risk for each cancer is ~30% (colon), ~50% (pancreas) and ~20% (breast) [2–4]. Type 1 diabetes carries an excess cancer risk of ~20%, but involves a different range of tumours [5]. The major cancers linked with type 2 diabetes are also associated with obesity or insulin resistance, suggesting that factors other than glucose play an important role [6]. These observations, although wellattested, have attracted relatively little interest within the world of diabetes. This is partly due to the dominant role of cardiovascular disease, which largely accounts for the twofold increase in mortality associated with type 2 diabetes [7], and partly, perhaps, because cancer has seemed unavoidable. The latter assumption can no longer be considered correct, for several studies have shown metformin to be associated with a lower risk of cancer than insulin or sulfonylureas [8–10]. Bowker and colleagues examined the relationship between diabetes treatment and mortality in a health database from Saskatchewan, and found that cancer mortality was almost doubled among insulin users (HR 1.9, 95% CI 1.5–2.4, p<0.0001) relative to metformin users, and that sulfonylureas were also associated with increased mortality (HR 1.3, 95% CI 1.1–1.6, p=0.012) [9]. A study published in this issue of Diabetologia confirms these observations, while showing that cancer risk in metformintreated patients is similar to that in patients who have not as yet received medication for diabetes. Furthermore, the paper suggests that the effect of metformin may be tumour-specific, in that its use was associated with a reduced risk of cancer of the colon and pancreas, but not of cancer of the breast or prostate [10]. The antitumour effect of metformin seems to be mediated via its ability to increase the AMP-activated protein kinase (AMPK) signalling pathway [11]. AMPK, which is activated by a rise in the AMP:ATP ratio, plays a key role in cellular energy balance. Activation restores levels of ATP by switching on ATP-generating pathways and switching off ATP-consuming pathways, and this enzyme is thought to mediate many of the metabolic actions of metformin [12]. Increased AMPK activity also leads to an inhibition of the downstream mammalian target of rapamycin (mTOR) complex; mTOR kinase integrates U. Smith The Lundberg Laboratory for Diabetes Research, Department of Molecular and Clinical Medicine, Sahlgrenska University Hospital, Goteborg, Sweden

Energy metabolism among eukaryotic anaerobes in light of Proterozoic ocean chemistry.

Recent years have witnessed major upheavals in views about early eukaryotic evolution. One very significant finding was that mitochondria, including hydrogenosomes and the newly discovered mitosomes, are just as ubiquitous and defining among eukaryotes as the nucleus itself. A second important advance concerns the readjustment, still in progress, about phylogenetic relationships among eukaryotic groups and the roughly six new eukaryotic supergroups that are currently at the focus of much attention. From the standpoint of energy metabolism (the biochemical means through which eukaryotes gain their ATP, thereby enabling any and all evolution of other traits), understanding of mitochondria among eukaryotic anaerobes has improved. The mainstream formulations of endosymbiotic theory did not predict the ubiquity of mitochondria among anaerobic eukaryotes, while an alternative hypothesis that specifically addressed the evolutionary origin of energy metabolism among eukaryotic anaerobes did. Those developments in biology have been paralleled by a similar upheaval in the Earth sciences regarding views about the prevalence of oxygen in the oceans during the Proterozoic (the time from ca 2.5 to 0.6 Ga ago). The new model of Proterozoic ocean chemistry indicates that the oceans were anoxic and sulphidic during most of the Proterozoic. Its proponents suggest the underlying geochemical mechanism to entail the weathering of continental sulphides by atmospheric oxygen to sulphate, which was carried into the oceans as sulphate, fueling marine sulphate reducers (anaerobic, hydrogen sulphide-producing prokaryotes) on a global scale. Taken together, these two mutually compatible developments in biology and geology underscore the evolutionary significance of oxygen-independent ATP-generating pathways in mitochondria, including those of various metazoan groups, as a watermark of the environments within which eukaryotes arose and diversified into their major lineages.

The AMP-activated protein kinase: more than an energy sensor

The AMPK (AMP-activated protein kinase) is a highly conserved eukaryotic protein serine/threonine kinase. It mediates a nutrient signalling pathway that senses cellular energy status and was appropriately called the fuel gauge of the cell. At the cellular level, AMPK controls energy homoeostasis by switching on catabolic ATP-generating pathways, while switching off anabolic ATP-consuming processes. Its effect on energy balance extends to whole-body energy homoeostasis, because, in the hypothalamus, it integrates nutritional and hormonal signals that control food intake and body weight. The interest in AMPK also stems from the demonstration of its insulin-independent stimulation of glucose transport in skeletal muscle during exercise. Moreover, the potential importance of AMPK in metabolic diseases is supported by the notion that AMPK mediates the anti-diabetic action of biguanides and thiazolidinediones and that it might be involved in the metabolic syndrome. Finally, the more recent demonstration that AMPK activation could occur independently of changes in cellular energy status, suggests that AMPK action extends to the control of non-metabolic functions.

Proteome Changes in Bovine Longissimus Thoracis Muscle During the Early Postmortem Storage Period

Postmortem changes in protein composition up to 24 h in bovine longissimus thoracis muscle were investigated by two-dimensional gel electrophoresis and MALDI-TOF MS/MS. A total of 47 spots were significantly changed the first 24 h postmortem. The 39 identified proteins can be divided into five groups: metabolic enzymes, defense and stress proteins, structural proteins, proteolytic enzymes, and unclassified proteins. The identified metabolic enzymes are all associated with ATP-generating pathways, either the glycolytic pathway or energy metabolism. In addition, several defense and stress proteins were changed in abundance in this study. These findings contribute to a better understanding of the biochemical processes during postmortem storage of meat.

Understanding the molecular basis of the interaction between NDPK-A and AMPK alpha 1.

Nucleoside diphosphate kinase (NDPK) (nm23/awd) belongs to a multifunctional family of highly conserved proteins (approximately 16 to 20 kDa) including two well-characterized isoforms (NDPK-A and -B). NDPK catalyzes the conversion of nucleoside diphosphates to nucleoside triphosphates, regulates a diverse array of cellular events, and can act as a protein histidine kinase. AMP-activated protein kinase (AMPK) is a heterotrimeric protein complex that responds to the cellular energy status by switching off ATP-consuming pathways and switching on ATP-generating pathways when ATP is limiting. AMPK was first discovered as an activity that inhibited preparations of acetyl coenzyme A carboxylase 1 (ACC1), a regulator of cellular fatty acid synthesis. We recently reported that NDPK-A (but not NDPK-B) selectively regulates the alpha1 isoform of AMPK independently of the AMP concentration such that the manipulation of NDPK-A nucleotide trans-phosphorylation activity to generate ATP enhanced the activity of AMPK. This regulation occurred irrespective of the surrounding ATP concentration, suggesting that "substrate channeling" was occurring with the shielding of NDPK-generated ATP from the surrounding medium. We speculated that AMPK alpha1 phosphorylated NDPK-A during their interaction, and here, we identify two residues on NDPK-A targeted by AMPK alpha1 in vivo. We find that NDPK-A S122 and S144 are phosphorylated by AMPK alpha1 and that the phosphorylation status of S122, but not S144, determines whether substrate channeling can occur. We report the cellular effects of the S122 mutation on ACC1 phosphorylation and demonstrate that the presence of E124 (absent in NDPK-B) is necessary and sufficient to permit both AMPK alpha1 binding and substrate channeling.

Nucleoside diphosphate kinase A as a controller of AMP-kinase in airway epithelia

This review integrates recent understanding of a novel role for NDPK-A in two related directions: Firstly, its role in an airway epithelial cell when bound to the luminal (apical) membrane and secondly in the cytosol of many different cells (epithelial and non-epithelial) where an isoform-specific interaction occurs with a regulatory partner, AMPKα1. Thus NDPK-A is present in both a membrane and cytosolic environment but in the apical membrane, its roles are not understood in detail; preliminary data suggest that it co-localises with the cystic fibrosis protein (CFTR). In cytosol, we find that NDPK-A is coupled to the catalytic alpha1 isoform of the AMP-activated protein kinase (AMPKα subunit), which is part of a heterotrimeric protein complex that responds to cellular energy status by switching off ATP-consuming pathways and switching on ATP-generating pathways when ATP is limiting. We find that ATP is located within this complex and ‘fed’ from NDPK to AMPK without ever ‘seeing’ bulk solution. Importantly, the reverse can also happen such that AMPK activity can be made to decline when NDPK-A ‘steals’ ATP from AMPK. Thus we propose a novel paradigm in NDPK-A function by suggesting that AMP-kinase can be regulated by NDPK-A, independently of AMP.

T cells get energized

Many cell types use AMPK (AMP-activated protein kinase) to signal that energy reserves are low so more energy generation is needed. But, say Tamas et al. on page 1665, T cells also use AMPK to signal that energy will be needed in the near future, for attacking target cells, proliferating or churning out cytokines. Figure 1 AMPK gets phosphorylated (and thus activated) after TCR ligation (α-CD3) in primary T cells. When cellular energy is in short supply—due, for example, to nutrient deficiency or metabolic stress—rising levels of AMP and falling levels of ATP trigger the activation of AMPK. AMPK then activates ATP-generating pathways, such as fatty acid oxidation. Tamas and colleagues now show that this pathway is also activated in T cells when their energy reserves dwindle. This is the first demonstration of physiological AMPK regulation in T cells. But T cells turned on AMPK not only in response to falling energy levels but also in anticipation of upcoming energy needs. “If a T cell fluxes calcium,” says senior author Doreen Cantrell, “it is indicating that it is about to activate and will need ATP.” And this Ca2+ flux, they found, triggered the activation of AMPK. Ca2+-induced AMPK activation required upstream Ca2+/calmodulin-dependent protein kinase kinases, which were not needed for AMPK activation in response to rising AMP/ATP ratios. PI3 kinase (PI3K)—an enzyme required for cytokine- and costimulation-induced energy generation in T cells—was also dispensable for AMPK activation. Determining the importance of the AMPK pathway, relative to these other energy-boosting pathways, awaits the development of conditional AMPK-deficient mice.

Differential utilization of two ATP-generating pathways is regulated by p53

A fundamental property of cancer cells is the preferential utilization of glycolysis over aerobic respiration to produce ATP. Renewed interest in understanding the mechanism underlying this metabolic shift in energy production is broadening our understanding of the relationship between cancer and cellular metabolism. In a recent article, Matoba et al. report that the p53 tumor suppressor regulates the expression of SCO2, a protein that is required for the assembly of cytochrome c oxidase (COX), a multimeric protein complex required for oxidative phosphorylation. The implication of these findings is that aerobic respiration is compromised in cells that lack functional p53.

Cancer's sweet tooth

Even in the presence of an adequate oxygen supply, many tumors metabolize the majority of the glucose they take up through glycolysis. It has been a long-held belief that this glycolytic phenotype is due to cancer-specific defects in mitochondrial oxidative phosphorylation. In this issue of Cancer Cell, Fantin et al. now report that most tumor cells have a substantial reserve capacity to produce ATP by oxidative phosphorylation when glycolysis is suppressed. These new data add to mounting evidence that the high rate of glycolysis exhibited by most tumors is required to support cell growth rather than to compensate for defect(s) in mitochondrial function.

A novel physical and functional association between nucleoside diphosphate kinase A and AMP-activated protein kinase alpha1 in liver and lung.

Nucleoside diphosphate kinase (NDPK, NM23/awd) belongs to a multifunctional family of highly conserved proteins (approximately 16-20 kDa) containing two well-characterized isoforms (NM23-H1 and -H2; also known as NDPK A and B). NDPK catalyses the conversion of nucleoside diphosphates into nucleoside triphosphates, regulates a diverse array of cellular events and can act as a protein histidine kinase. AMPK (AMP-activated protein kinase) is a heterotrimeric protein complex that responds to cellular energy status by switching off ATP-consuming pathways and switching on ATP-generating pathways when ATP is limiting. AMPK was first discovered as an activity that inhibited preparations of ACC1 (acetyl-CoA carboxylase), a regulator of cellular fatty acid synthesis. We report that NM23-H1/NDPK A and AMPK alpha1 are associated in cytosol from two different tissue sources: rat liver and a human lung cell line (Calu-3). Co-immunoprecipitation and binding assay data from both cell types show that the H1/A (but not H2/B) isoform of NDPK is associated with AMPK complexes containing the alpha1 (but not alpha2) catalytic subunit. Manipulation of NM23-H1/NDPK A nucleotide transphosphorylation activity to generate ATP (but not GTP) enhances the activity of AMPK towards its specific peptide substrate in vitro and also regulates the phosphorylation of ACC1, an in vivo target for AMPK. Thus novel NM23-H1/NDPK A-dependent regulation of AMPK alpha1-mediated phosphorylation is present in mammalian cells.

Crystallization of the glycogen-binding domain of the AMP-activated protein kinase beta subunit and preliminary X-ray analysis.

AMP-activated protein kinase (AMPK) is an intracellular energy sensor that regulates metabolism in response to energy demand and supply by adjusting the ATP-generating and ATP-consuming pathways. AMPK potentially plays a critical role in diabetes and obesity as it is known to be activated by metforin and rosiglitazone, drugs used for the treatment of type II diabetes. AMPK is a heterotrimer composed of a catalytic alpha subunit and two regulatory subunits, beta and gamma. Mutations in the gamma subunit are known to cause glycogen accumulation, leading to cardiac arrhythmias. Recently, a functional glycogen-binding domain (GBD) has been identified in the beta subunit. Here, the crystallization of GBD in the presence of beta-cyclodextrin is reported together with preliminary X-ray data analysis allowing the determination of the structure by single isomorphous replacement and threefold averaging using in-house X-ray data collected from a selenomethionine-substituted protein.

Activation of AMP-dependent protein kinase by hypoxia and hypothermia in the liver of frog Rana perezi

We have investigated different signaling molecules that could be activated by temperature acclimation and hypoxia, using an experimental approach consisting in submerging frogs in a water-filled box maintained at 2–4 °C at ambient oxygen levels or supplied with 98% N 2:2% CO 2 for normoxia or hypoxia conditions, respectively. The results obtained showed no significant changes in the expression of heat shock protein 70. The phosphorylation state of AMP-dependent activated protein kinase, the down-stream component of a protein kinase cascade that acts as an intracellular energy sensor, was significantly increased in both experimental conditions, showing higher values in the absence of oxygen. Similarly, the phosphorylation state of one of its known substrates, elongation factor 2, was also increased, consistent with the arrest of protein synthesis. These results point out an important role of this kinase, adjusting the rates of ATP-consuming and ATP-generating pathways, in the survival strategies to hypoxia and hypothermia.

Metabolic cold adaptation in Antarctic fishes: evidence from enzymatic activities of brain

To evaluate the concept of metabolic cold adaptation (MCA) in fishes, we compared – in brain, red muscle, and white muscle of Antarctic notothenioid fishes and tropical/subtropical fishes – the activities of two enzymes of ATP-generating pathways, citrate synthase (CS), an indicator of citric acid cycle activity (aerobic metabolism), and lactate dehydrogenase (LDH), an indicator of potential for ATP production through anaerobic glycolysis. Brain was chosen because, unlike locomotory muscle, its metabolic activity is not likely to be influenced by a species' level of activity or nutritional status, so MCA should be readily observed if present. CS and LDH activities in brain exhibited a high level of MCA, but compensation to temperature was not complete (48% for CS; 46% for LDH). CS and LDH activities in red and white muscle varied widely among species, according to the general level of locomotory activity. The 'mode of life'-related enzymatic activities in locomotory muscle show that study of MCA at the level of whole organism metabolism is fraught with difficulties and experimental ambiguities. In contrast, the low variation among species within each group in enzymatic activities in brain, and the large differences between groups in CS and LDH activity, show that brain is an excellent study system for evaluating metabolic compensation to temperature.

Age-associated changes in gene expression patterns in the duodenum and colon of rats

In humans, decreased intestinal motility, compromised nutritional status and increased risk of colon cancer are commonly associated with aging. Here, we used the cDNA microarray analysis to detect age-associated changes in duodenal and colonic gene expression in male Fischer 344 rats. The primary finding of this study is that the magnitude and direction of age-associated changes in gene expression differs in the colon and duodenum. In the colon, 56 genes showed altered expression, whereas expression of only 25 genes was altered in the duodenum. The magnitude of change was greater in the colon than in the duodenum. The direction of change also differed; in the aged colon, expression of 51 genes increased and only five genes decreased. In contrast, in the aged duodenum, only seven genes increased, whereas 18 genes decreased in expression. In the duodenum of aged rats, expression of genes involved in ATP-generating pathways is decreased. In contrast, in the colon of aged rats, expression of genes involved in energy generating pathways and in lipid oxidation is increased. In addition, in the aging colon, an increased expression of genes that show an aberrant regulation in colon cancer, including CD44, ras, and maspin is observed. Collectively, these findings provide clues to molecular events that may be related to compromised intestinal function and the high incidence of colon cancer in the aged population.

The action of thymol on oral bacteria.

Several effects of thymol, a plant-derived antimicrobial agent, on Porphyromonas gingivalis, Selenomonas artemidis and Streptococcus sobrinus were examined. The extremely rapid efflux of intracellular constituents evoked by thymol is consistent with its postulated membranotropic effects. Correlations between leakage-inducing concentrations of thymol and minimal inhibitory concentrations and minimal bactericidal concentrations suggest that membrane perforation is a principal mode of action of this substance. The thymol-induced decline in intracellular ATP in S. sobrinus appears to be entirely attributable to leakage, whereas in P. gingivalis thymol may also inhibit ATP-generating pathways. Relative changes in the transmembrane potential of resting cells of S. sobrinus pulsed with glucose are as sensitive to thymol as is leakage from this organism. The effects of thymol on transmembrane potential are probably secondary to those arising from leakage of intracellular substances.