Developmental changes in the balance of glycolytic ATP production and oxidative phosphorylation in ventricular cells: A simulation study.

The developmental program of the heart requires accurate regulation to ensure continuous circulation and simultaneous cardiac morphogenesis, because any functional abnormalities may progress to congenital heart malformation. Notably, energy metabolism in fetal ventricular cells is regulated in a manner that differs from adult ventricular cells: fetal cardiomyocytes generally have immature mitochondria and fetal ventricular cells show greater dependence on glycolytic ATP production. However, although various characteristics of energy metabolism in fetal ventricular cells have been reported, to our knowledge, a quantitative description of the contributions of these factors to fetal ventricular cell functions has not yet been established. Here, we constructed a mathematical model to integrate various characteristics of fetal ventricular cells and predicted the contribution of each characteristic to the maintenance of intracellular ATP concentration and sarcomere contraction under anoxic conditions. Our simulation results demonstrated that higher glycogen content, higher hexokinase activity, and lower creatine concentration helped prolong the time for which ventricular cell contraction was maintained under anoxic conditions. The integrated model also enabled us to quantitatively assess the contributions of factors related to energy metabolism in ventricular cells. Because fetal cardiomyocytes exhibit similar energy metabolic profiles to stem cell-derived cardiomyocytes and those in the failing heart, an improved understanding of these fetal ventricular cells will contribute to a better comprehension of the processes in stem cell-derived cardiomyocytes or under pathological conditions.

Prediction of the Standard Gibbs Energy of Transfer of Organic Ions Across the Interface between Two Immiscible Liquids.

The non-Bornian solvation model was applied for evaluation of the standard Gibbs energy (ΔGtr°,W→O) of transfer of organic ions from water (W) to organic solvent (O = nitrobenzene). The solvation energy of an ion in either W or O is basically formulated as the energy required for the formation of a nanosized ion­solvent interface around the ion; however, many organic ions with strongly charged groups (e.g., -SO3-, -CO2-, -NH3+) are preferentially hydrated in O. Here we divided the surface of an ion into "hydrated" and "non-hydrated" surfaces and then carried out regression analyses with experimental values of ΔGtr°,W→O. In the analyses, the local electric field on the surface of an organic ion was evaluated through density functional theory calculation. Good regression results were then obtained with the mean absolute error of 1.9 and 2.4 kJ mol-1 for 34 anions and 63 cations, respectively. These errors correspond to the error of ∼20 mV in the standard ion-transfer potential (ΔOWϕ°), being only two times larger than the typical experimental error (∼10 mV) in the voltammetric measurement. This non-Bornian model is promising for theoretical prediction of ΔGtr°,W→O (or ΔOWϕ°) for organic ions and possibly of the biomembrane permeability for ionic drugs.

Coextraction of water into nitrobenzene with organic ions.

Various organic anions (sulfonates (RSO3(-)), carboxylates (RCO2(-)), and phenolates (RO(-))) and ammonium cations (RNH3(+), R2NH2(+), and R3NH(+)) were distributed in the nitrobenzene (NB)-water system by using Crystal Violet and dipicrylaminate, respectively. The number of water molecules (n) being coextracted into NB with an ion was then determined by the Karl Fischer method. The n values determined and those reported previously showed the variation from 0.51 to 3.4, depending on not only the charged groups but also the noncharged R-groups. In this study, we focused our attention to the strong electric field on the charged group and its facilitation effect for binding water molecules in NB. The local electric field (Ei) on the surface of an organic ion was evaluated by using Gaussian09 program with a subprogram developed in our recent study. It was found that the n values showed a clear dependence on the average value of Ei on oxygen or hydrogen atoms, respectively, of an anionic or cationic group.

Gut-targeted immunonutrition boosting natural killer cell activity using Saccharomyces boulardii lysates in immuno-compromised healthy elderly subjects.

The aim of this study was to assess the immunomodulatory effect of KC-1317 (a symbiotic mixture containing Saccharomyces boulardii lysate in a cranberry, colostrum-derived lactoferrin, fragaria, and lactose mixture) supplementation in immune-compromised but otherwise healthy elderly subjects. A liquid formulation of KC-1317 was administered in a randomized controlled trial (RCT) fashion to healthy volunteers (65-79 years) previously selected for low natural killer (NK) cell activity, and this parameter was checked at the completion of the study. A significant improvement in NK cell activity of KC-1317 consumers was observed as compared to placebo at the end of 2 months. Although preliminary, these beneficial immune-modulatory effects of KC-1317 in aged individuals might indicate its employment within a wider age-management strategy.

Contribution of quantitative changes in individual ionic current systems to the embryonic development of ventricular myocytes: a simulation study.

Early embryonic rodent ventricular cells exhibit spontaneous action potential (AP), which disappears in later developmental stages. Here, we used 3 mathematical models-the Kyoto, Ten Tusscher-Panfilov, and Luo-Rudy models-to present an overview of the functional landscape of developmental changes in embryonic ventricular cells. We switched the relative current densities of 9 ionic components in the Kyoto model, and 160 of 512 representative combinations were predicted to result in regular spontaneous APs, in which the quantitative changes in Na(+) current (I Na) and funny current (I f) made large contributions to a wide range of basic cycle lengths. In all three models, the increase in inward rectifier current (I K1) before the disappearance of I f was predicted to result in abnormally high intracellular Ca(2+) concentrations. Thus, we demonstrated that the developmental changes in APs were well represented, as I Na increased before the disappearance of I f, followed by a 10-fold increase in I K1.

Prediction of axillary lymph node metastasis in primary breast cancer patients using a decision tree-based model.

BACKGROUND: The aim of this study was to develop a new data-mining model to predict axillary lymph node (AxLN) metastasis in primary breast cancer. To achieve this, we used a decision tree-based prediction method-the alternating decision tree (ADTree). METHODS: Clinical datasets for primary breast cancer patients who underwent sentinel lymph node biopsy or AxLN dissection without prior treatment were collected from three institutes (institute A, n = 148; institute B, n = 143; institute C, n = 174) and were used for variable selection, model training and external validation, respectively. The models were evaluated using area under the receiver operating characteristics (ROC) curve analysis to discriminate node-positive patients from node-negative patients. RESULTS: The ADTree model selected 15 of 24 clinicopathological variables in the variable selection dataset. The resulting area under the ROC curve values were 0.770 [95% confidence interval (CI), 0.689-0.850] for the model training dataset and 0.772 (95% CI: 0.689-0.856) for the validation dataset, demonstrating high accuracy and generalization ability of the model. The bootstrap value of the validation dataset was 0.768 (95% CI: 0.763-0.774). CONCLUSIONS: Our prediction model showed high accuracy for predicting nodal metastasis in patients with breast cancer using commonly recorded clinical variables. Therefore, our model might help oncologists in the decision-making process for primary breast cancer patients before starting treatment.

Biomarine extracts significantly protect from ultraviolet A-induced skin photoaging: an ex vivo study.

We tested the activity of the marine nutraceutical CL-1222 added with a coenzyme Q10 (CoQ10)-lutein-selenium component (Celergen(®), Laboratoires-Dom, Switzerland) to protect human fibroblasts against ultraviolet A (UVA)-induced photoaging. Cells obtained from 22- to 39-year-old healthy donors were pretreated with CL-1222 before UV irradiation, as compared with same quantity of the CoQ10-lutein-selenium component. As compared to untreated control, UVA-irradiated samples exhibited a significant increase of secreted matrix metalloproteinase-1 (MMP-1) (p<0.001) with over four-fold MMP-1 upregulation (p<0.001). Samples treated with CL-1222, but not with the CoQ10-lutein-selenium component, showed a significant decrease of MMP-1 secretion (p<0.01) and expression decrease (>60%, p<0.01) with >54% elastase activity inhibition (p<0.01). This preliminary study shows that such marine nutraceuticals can significantly protect against UV-irradiation irrespective of the CoQ10-lutein-selenium component with a specific protective gene expression modulation amenable to novel clinical applications.

Hepatoprotective activity of a phytotherapeutic formula on thioacetamide--induced liver fibrosis model.

Hepatic fibrosis is a widespread alteration in the liver that primarily consists of increased collagen deposition in the tissue. The aim of the present study was to evaluate the protective effects of poly-phytocompound EH-1501 containing small amounts of silymarin but also other potentially effective substances on thioacetamide (TTA)-induced liver fibrosis and to elucidate the mechanisms underlying these protective effects in rats. Forty rats were randomly divided into four groups. Liver fibrosis was induced by intraperitoneal injection of 200 mg/kg body weight. TAA dissolved in saline was administered thrice a week, for 8 weeks. Groups 1 (normal healthy control) and 2 (liver injury model) received water for 8 weeks or silymarin (50 mg/kg p.o. daily) for 8 weeks (group 3) or a poly-phytocompound EH-1501 (containing grape leaf, wild strawberry, dandelion and milk thistle, EuroHealth, Italy) (200 mg/kg, daily respectively) for 8 weeks (group 4). Biochemistry and serum fibrosis markers were AST, ALT, GGT, bilirubin, thiobarbituric acid reactive substances (TBARs), hyaluronic acid and type IV collagen 7s. Liver tissue was used to assay glutathione peroxidase (GPx), catalase (CAT), superoxide dismutase (SOD), TBARs, hydroxyproline and gene expression of collagen alpha1 (col alpha1) and transforming growth factor-beta1 (TGF-beta1). Silymarine and EH-1501 were equally effective in reducing serum markers of liver damage and fibrosis as well as oxidative stress. However, as compared to silymarine, EH-1501 was significantly more effective in improving tissue level of GPx while decreasing TBARs and hydroproline content (p < 0.05). When looking at gene expression of col alpha1 and TGF-beta1, EH-1501 showed a significantly higher degree of gene down-regulation as compared to silymarine (p < 0.05). Taken altogether, these data suggest that a natural antioxidant-containing phytocompound EH-1501 exerts an effective hepatoprotective property in experimental chronic fibrotizing liver injury to a significantly higher degree than silymarin.

Computational assessment of insulin secretion and insulin sensitivity from 2-h oral glucose tolerance tests for clinical use for type 2 diabetes.

In type 2 diabetes mellitus, glucose homeostasis is tightly maintained through insulin secretion and insulin sensitivity. Therefore, finding an accurate method to assess insulin secretion and sensitivity using clinically available data would enhance the quality of diabetic medical care. In an effort to find such a method, we developed a computational approach to derive indices of these factors using a 2-h oral glucose tolerance test (OGTT). To evaluate our method, clinical data from subjects who received an OGTT and a glucose clamp test were examined. Our insulin secretion index was significantly correlated with an analogous index obtained from a hyperglycemic clamp test (r = 0.90, n = 46, p < 0.001). Our insulin sensitivity index sensitivity was also significantly correlated with an analogous index obtained from a hyperinsulinemic-euglycemic clamp test (r = 0.56, n = 79, p < 0.001). These results suggest that our method can potentially provide an accurate and convenient tool toward improving the management of diabetes in clinical practice by assessing insulin secretion and insulin sensitivity.

Predictive value of CD24 and CD44 for neoadjuvant chemotherapy response and prognosis in primary breast cancer patients.

PURPOSE: We investigated the significance of CD24 and CD44 expression for predicting responses to chemotherapy and prognosis in primary breast cancer patients. PATIENTS AND METHODS: Diagnosis of breast cancer was confirmed by core needle biopsy, and immunohistochemical studies were performed. Preoperatively, patients received anthracycline-containing chemotherapy. Expression of CD44 and CD24 was assessed immunohistochemically and the relationship with chemotherapy response and with prognosis was analyzed. RESULTS: Between 2001 and 2004, 139 women were enrolled in this study. In the correlation analysis, CD24 expression was negatively associated with pathological response to chemotherapy (p = 0.0003). A machine learning technique with an alternating decision tree (ADTree) showed that four logical rules are involved in predicting the response depending on the combination of CD24, HER2, tumor stage, CD44, progesterone receptor, and patient age. In the survival analysis, patients having CD44 (++) showed a significantly favorable prognosis as compared with others (p = 0.0002). A multivariate analysis showed that CD44 expression had an independent prognostic value (p < .0001). CONCLUSION: We found a significant correlation between CD44 expression and prognosis and between CD24 expression and response to chemotherapy. CD24 and CD44 expressions would be useful predictive markers, although further studies are needed.

Anti-allergic effects of probiotic Dahi through modulation of the gut immune system.

BACKGROUND/AIMS: The alarming increase in allergy in the last few decades demands the development of new anti-allergic prevention strategies, and consumption of functional foods (i.e. probiotic Dahi, which has already been proven to enhance immunity by modulation of the gut mucosal immune system) may be one of them. In the present study, we evaluated anti-allergic effects of a Dahi (yogurt) containing probiotic Lactobacillus acidophilus, L. casei and normal Dahi culture Lactococcus lactis biovar diacetylactis (named probiotic Dahi) on ovalbumin induced allergy in mice. METHODS: Allergy was induced by injecting (i.p.) ovalbumin at 0 and 14 days. Animals were fed with standard diet (control), milk, control Dahi or probiotic Dahi for 21 days. Total and ovalbumin-specific IgE, cytokines and lymphocyte proliferation index were examined after 7, 14 and 21 days. RESULTS: Feeding of probiotic Dahi completely suppressed the elevation of total and ovalbumin-specific IgE in the serum of ovalbumin-injected mice. Similarly, splenocytes collected from mice fed with probiotic Dahi entirely lost the total and ovalbumin-specific IgE production property during in-vitro culture. Production of T helper (Th)-1 cell-specific cytokines, i.e. interferon -γ and interleukin (IL)-2, increased, while Th2-specific cytokines, i.e. IL-4 and IL-6, decreased in the supernatant of cultured splenocytes collected from mice fed with probiotic Dahi compared to the other groups. Moreover, ovalbumin-stimulated lymphocyte proliferation was strongly suppressed by feeding of probiotic Dahi in comparison to milk and control Dahi. CONCLUSIONS: Results of the present study indicate that probiotic Dahi suppressed ovalbumin-induced allergic consequences characterized by decreasing levels of total and ovalbumin-specific IgE and lymphocyte proliferation and skewed ovalbumin-induced Th2-specific immune response towards Th1-specific response.

Construction of a biological tissue model based on a single-cell model: a computer simulation of metabolic heterogeneity in the liver lobule.

An enormous body of information has been obtained by molecular and cellular biology in the last half century. However, even these powerful approaches are not adequate when it comes to higher-level biological structures, such as tissues, organs, and individual organisms, because of the complexities involved. Thus, accumulation of data at the higher levels supports and broadens the context for that obtained on the molecular and cellular levels. Under such auspices, an attempt to elucidate mesoscopic and macroscopic subjects based on plentiful nanoscopic and microscopic data is of great potential value. On the other hand, fully realistic simulation is impracticable because of the extensive cost entailed and enormous amount of data required. Abstraction and modeling that balance the dual requirements of prediction accuracy and manageable calculation cost are of great importance for systems biology. We have constructed an ammonia metabolism model of the hepatic lobule, a histological component of the liver, based on a single-hepatocyte model that consists of the biochemical kinetics of enzymes and transporters. To bring the calculation cost within reason, the porto-central axis, which is an elemental structure of the lobule, is defined as the systems biological unit of the liver, and is accordingly modeled. A model including both histological structure and position-specific gene expression of major enzymes largely represents the physiological dynamics of the hepatic lobule in nature. In addition, heterogeneous gene expression is suggested to have evolved to optimize the energy efficiency of ammonia detoxification at the macroscopic level, implying that approaches like this may elucidate how properties at the molecular and cellular levels, such as regulated gene expression, modify higher-level phenomena of multicellular tissue, organs, and organisms.

Oxidative-inflammatory damage in cirrhosis: effect of vitamin E and a fermented papaya preparation.

BACKGROUND AND AIM: Oxidative DNA damage occurs as an early event in hepatitis C virus (HCV) infection and is an indication of the potential for carcinogenesis. The aim of this study was to test a novel antioxidant/immunomodulator in patients with HCV-related cirrhosis. METHODS: The study group consisted of 50 patients with HCV-related cirrhosis with transaminase values less than twofold increased (alanine aminotransferase [ALT] < 80 IU/L). Patients underwent a standardized food-vitamin composition assessment and were assessed for dietary intake, nutritional status and iron level. Patients were randomly allocated into two groups and then given either alpha-tocopherol 900 IU/day or 9 g/day of a fermented papaya preparation (FPP, Immun-Age, Osato Research Institute, Gifu, Japan) at bedtime for 6 months. Ten healthy subjects served as controls. Patients were checked monthly for: routine tests, redox status (reduced glutathione, glutathione peroxidase, oxidized glutathione, malondialdehyde), plasma alpha-tocopherol, 8-hydroxy-deoxy-guanidine (8-OHdG) level in circulating leukocyte DNA and serum levels of cytokines. RESULTS: Patients with cirrhosis showed a significant imbalance of redox status (low antioxidants/high oxidative stress markers) (P < 0.005 vs controls). Neither treatment regimen affected transaminases as a whole. However, vitamin E supplementation almost normalized ALT only in the limited vitamin-E-deficient subgroup. A significant improvement of redox status was obtained by both regimens. However, only FPP significantly decreased 8-OHdG and the improvement of cytokine balance with FPP was significantly better than with vitamin E treatment (P < 0.05). CONCLUSIONS: Although the present data seem to suggest a potential supportive role of antioxidants/immunomodulators as FPP in HCV patients, more studies are needed to substantiate their effect on the natural history of the disease.

Simulation of developmental changes in action potentials with ventricular cell models.

During cardiomyocyte development, early embryonic ventricular cells show spontaneous activity that disappears at a later stage. Dramatic changes in action potential are mediated by developmental changes in individual ionic currents. Hence, reconstruction of the individual ionic currents into an integrated mathematical model would lead to a better understanding of cardiomyocyte development. To simulate the action potential of the rodent ventricular cell at three representative developmental stages, quantitative changes in the ionic currents, pumps, exchangers, and sarcoplasmic reticulum (SR) Ca(2+) kinetics were represented as relative activities, which were multiplied by conductance or conversion factors for individual ionic systems. The simulated action potential of the early embryonic ventricular cell model exhibited spontaneous activity, which ceased in the simulated action potential of the late embryonic and neonatal ventricular cell models. The simulations with our models were able to reproduce action potentials that were consistent with the reported characteristics of the cells in vitro. The action potential of rodent ventricular cells at different developmental stages can be reproduced with common sets of mathematical equations by multiplying conductance or conversion factors for ionic currents, pumps, exchangers, and SR Ca(2+) kinetics by relative activities.

Nutraceutical supplementation: effect of a fermented papaya preparation on redox status and DNA damage in healthy elderly individuals and relationship with GSTM1 genotype: a randomized, placebo-controlled, cross-over study.

Our study group consisted of 54 elderly patients without major invalidating diseases who were randomly divided into two fully matched groups. Group A was given a certified fermented papaya preparation 9 g/day by mouth, while group B received placebo. Treatment was carried out in a cross-over manner with a 3-month supplementation followed by a 6-week washout period. Blood samples were drawn at entry and on a monthly basis to check routine parameters, redox status, and 8-OHdG in circulating leukocyte DNA. Polymorphism analysis of GSTM1 was carried out as well. The glutathune-S transferase M1 (GSTM1) genotype was null (-) in 40% and 46% of groups A and B, respectively. GSTM1 (-) smokers had a significantly higher level of plasma DNA adducts and leukocytes level of 8-OHdG than their GSTM1 (+) counterparts (P < 0.01). There was a weak correlation between cigarettes smoked/day and DNA adduct (r: 0.61, P < 0.05), which also correlated with antioxidant concentrations, but only in GSTM1 (-) smokers (P < 0.01). The fermented papaya preparation (FPP)-supplemented group showed a significant enhancement of the antioxidant protection (P < 0.01 vs. A) within the subgroups with GSTM1 (-) and of plasma DNA adduct, irrespective of the GSTM1 genotype. Only the GSTM1 (-) subgroup was the one that, under FPP treatment, increased lymphocyte 8-OHdG (P < 0.01). Such preliminary data show that FPP is a promising nutraceutical for improving antioxidant-defense in elderly patients even without any overt antioxidant-deficiency state while helping explain some inconsistent results of prior interventional studies.

A trial of somatic gene targeting in vivo with an adenovirus vector.

BACKGROUND: Gene targeting in vivo provides a potentially powerful method for gene analysis and gene therapy. In order to sensitively detect and accurately measure designed sequence changes, we have used a transgenic mouse system, MutaMouse, which has been developed for detection of mutation in vivo. It carries bacteriophage lambda genome with lacZ+ gene, whose change to lacZ-negative allele is detected after in vitro packaging into bacteriophage particles. We have also demonstrated that gene transfer with a replication-defective adenovirus vector can achieve efficient and accurate gene targeting in vitro. METHODS: An 8 kb long DNA corresponding to the bacteriophage lambda transgene with one of two lacZ-negative single-base-pair-substitution mutant allele was inserted into a replication-defective adenovirus vector. This recombinant adenovirus was injected to the transgenic mice via tail-vein. Twenty-four hours later, genomic DNA was extracted from the liver tissue and the lambda::lacZ were recovered by in vitro packaging. The lacZ-negative phage was detected as a plaque former on agar with phenyl-beta-D-galactoside. RESULTS: The mutant frequency of the lacZ-negative recombinant adenovirus injected mice was at the same level with the control mouse (approximately 1/10000). Our further restriction analysis did not detect any designed recombinant. CONCLUSION: The frequency of gene targeting in the mouse liver by these recombinant adenoviruses was shown to be less than 1/20000 in our assay. However, these results will aid the development of a sensitive, reliable and PCR-independent assay for gene targeting in vivo mediated by virus vectors and other means.

A DNA methyltransferase can protect the genome from postdisturbance attack by a restriction-modification gene complex.

In prokaryotic genomes, some DNA methyltransferases form a restriction-modification gene complex, but some others are present by themselves. Dcm gene product, one of these orphan methyltransferases found in Escherichia coli and related bacteria, methylates DNA to generate 5'-C(m)CWGG just as some of its eukaryotic homologues do. Vsr mismatch repair function of an adjacent gene prevents C-to-T mutagenesis enhanced by this methylation but promotes other types of mutation and likely has affected genome evolution. The reason for the existence of the dcm-vsr gene pair has been unclear. Earlier we found that several restriction-modification gene complexes behave selfishly in that their loss from a cell leads to cell killing through restriction attack on the genome. There is also increasing evidence for their potential mobility. EcoRII restriction-modification gene complex recognizes the same sequence as Dcm, and its methyltransferase is phylogenetically related to Dcm. In the present work, we found that stabilization of maintenance of a plasmid by linkage of EcoRII gene complex, likely through postsegregational cell killing, is diminished by dcm function. Disturbance of EcoRII restriction-modification gene complex led to extensive chromosome degradation and severe loss of cell viability. This cell killing was partially suppressed by chromosomal dcm and completely abolished by dcm expressed from a plasmid. Dcm, therefore, can play the role of a "molecular vaccine" by defending the genome against parasitism by a restriction-modification gene complex.