The evolution of the wine microbial ecosystem is generally restricted to Saccharomyces cerevisiae and Oenococcus oeni, which are the two main agents in the transformation of grape must into wine by acting during alcoholic and malolactic fermentation, respectively. But others species like the yeast Brettanomyces bruxellensis and certain ropy strains of Pediococcus parvulus can spoil the wine. The aim of this study was to address the composition of the system more precisely, identifying other components. The advantages of the polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) approach to wine microbial ecology studies are illustrated by bacteria and yeast species identification and their monitoring at each stage of wine production. After direct DNA extraction, PCR-DGGE was used to make the most exhaustive possible inventory of bacteria and yeast species found in a wine environment. Phylogenetic neighbor-joining trees were built to illustrate microbial diversity. PCR-DGGE was also combined with population enumeration in selective media to monitor microbial changes at all stages of production. Moreover, enrichment media helped to detect the appearance of spoilage species. The genetic diversity of the wine microbial community and its dynamics during winemaking were also described. Most importantly, our study provides a better understanding of the complexity and diversity of the wine microbial consortium at all stages of the winemaking process: on grape berries, in must during fermentation, and in wine during aging. On grapes, 52 different yeast species and 40 bacteria could be identified. The diversity was dramatically reduced during winemaking then during aging. Yeast and lactic acid bacteria were also isolated from very old vintages. B. bruxellensis and O. oeni were the most frequent.

This introductory article to the review series entitled "The Cancer Cell's Power Plants as Promising Therapeutic Targets" is written while more than 20 million people suffer from cancer. It summarizes strategies to destroy or prevent cancers by targeting their energy production factories, i.e., "power plants." All nucleated animal/human cells have two types of power plants, i.e., systems that make the "high energy" compound ATP from ADP and P( i ). One type is "glycolysis," the other the "mitochondria." In contrast to most normal cells where the mitochondria are the major ATP producers (>90%) in fueling growth, human cancers detected via Positron Emission Tomography (PET) rely on both types of power plants. In such cancers, glycolysis may contribute nearly half the ATP even in the presence of oxygen ("Warburg effect"). Based solely on cell energetics, this presents a challenge to identify curative agents that destroy only cancer cells as they must destroy both of their power plants causing "necrotic cell death" and leave normal cells alone. One such agent, 3-bromopyruvate (3-BrPA), a lactic acid analog, has been shown to inhibit both glycolytic and mitochondrial ATP production in rapidly growing cancers (Ko et al., Cancer Letts., 173, 83-91, 2001), leave normal cells alone, and eradicate advanced cancers (19 of 19) in a rodent model (Ko et al., Biochem. Biophys. Res. Commun., 324, 269-275, 2004). A second approach is to induce only cancer cells to undergo "apoptotic cell death." Here, mitochondria release cell death inducing factors (e.g., cytochrome c). In a third approach, cancer cells are induced to die by both apoptotic and necrotic events. In summary, much effort is being focused on identifying agents that induce "necrotic," "apoptotic" or apoptotic plus necrotic cell death only in cancer cells. Regardless how death is inflicted, every cancer cell must die, be it fast or slow.

Probiotic microorganisms, especially lactic acid bacteria, are effective in the treatment of infectious diarrhoeal diseases and experimental colitis. Although the mechanisms by which these organisms exert their anti-inflammatory effects are largely unknown, immunomodulating effects are suggested. The objective of this study was to examine the effect of a 5-week oral administration of Lactobacillus rhamnosus subspecies GG (Lb. GG) on the cellular immune response to intestinal microorganisms in ten healthy volunteers. Peripheral blood cells (PB) were stimulated with either 'self' or 'non-self' preparations of faecal samples and isolated Bacteroides fragilis group-organisms (Bfg) or Escherichia coli (Esch. coli), and pro- and anti-inflammatory cytokines (IL-10, IL-4, IL-6, IFN-gamma, TNF-alpha) were measured in the culture supernatant. CD4+ T-lymphocyte activation was determined by measurement of intracellular ATP following lysis of the cells. The activational response of CD4+ T-lymphocytes towards isolated and heat-inactivated intestinal organisms was increased after the probiotic treatment. Additionally, TNF-alpha, IL-6 and in part IFN-gamma cytokine secretion by PB cells following stimulation with whole stool preparations and single members of the flora was significantly decreased, whereas the IL-10 and in part IL-4 cytokine secretion was increased at the end of the study. In contrast, the activational response of CD4+ T-lymphocytes following stimulation with whole 'non-self' intestinal flora was higher than by 'self' intestinal flora, but both responses showed a trend towards a reduction at the end of the study. This study documents a direct effect by Lb. GG on the cellular immune system of healthy volunteers and offers a promising tool to investigate systemic immunomodulation due to oral administration of probiotic microorganisms.

OBJECTIVE

Because the muscle response to incremental exercise is not well documented in patients suffering from chronic fatigue syndrome (CFS), we combined electrophysiological (compound-evoked muscle action potential, M wave), and biochemical (lactic acid production, oxidative stress) measurements to assess any muscle dysfunction in response to a routine cycling exercise.

METHODS

This case-control study compared 15 CFS patients to a gender-, age- and weight-matched control group (n=11) of healthy subjects.

METHODS

All subjects performed an incremental cycling exercise continued until exhaustion.

METHODS

We measured the oxygen uptake (VO2), heart rate (HR), systemic blood pressure, percutaneous O2 saturation (SpO2), M-wave recording from vastus lateralis, and venous blood sampling allowing measurements of pH (pHv), PO2 (PvO2), lactic acid (LA), and three markers of the oxidative stress (thiobarbituric acid-reactive substances, TBARS, reduced glutathione, GSH, and ascorbic acid, RAA).

RESULTS

Compared with control, in CFS patients (i) the slope of VO2 versus work load relationship did not differ from control subjects and there was a tendency for an accentuated PvO2 fall at the same exercise intensity, indicating an increased oxygen uptake by the exercising muscles; (ii) the HR and blood pressure responses to exercise did not vary; (iii) the anaerobic pathways were not accentuated; (iv) the exercise-induced oxidative stress was enhanced with early changes in TBARS and RAA and enhanced maximal RAA consumption; and (v) the M-wave duration markedly increased during the recovery period.

CONCLUSIONS

The response of CFS patients to incremental exercise associates a lengthened and accentuated oxidative stress together with marked alterations of the muscle membrane excitability. These two objective signs of muscle dysfunction are sufficient to explain muscle pain and postexertional malaise reported by our patients.

OBJECTIVE

When Lactobacillus spp. dominate the vaginal microbiota of women of reproductive age they acidify the vagina to pH <4.0 by producing ∼1% lactic acid in a nearly racemic mixture of d- and l-isomers. We determined the HIV virucidal activity of racemic lactic acid, and its d- and l-isomers, compared with acetic acid and acidity alone (by the addition of HCl).

METHODS

HIV-1 and HIV-2 were transiently treated with acids in the absence or presence of human genital secretions at 37°C for different time intervals, then immediately neutralized and residual infectivity determined in the TZM-bl reporter cell line.

RESULTS

l-lactic acid at 0.3% (w/w) was 17-fold more potent than d-lactic acid in inactivating HIVBa-L. Complete inactivation of different HIV-1 subtypes and HIV-2 was achieved with ≥0.4% (w/w) l-lactic acid. At a typical vaginal pH of 3.8, l-lactic acid at 1% (w/w) more potently and rapidly inactivated HIVBa-L and HIV-1 transmitter/founder strains compared with 1% (w/w) acetic acid and with acidity alone, all adjusted to pH 3.8. A final concentration of 1% (w/w) l-lactic acid maximally inactivated HIVBa-L in the presence of cervicovaginal secretions and seminal plasma. The anti-HIV activity of l-lactic acid was pH dependent, being abrogated at neutral pH, indicating that its virucidal activity is mediated by protonated lactic acid and not the lactate anion.

CONCLUSIONS

l-lactic acid at physiological concentrations demonstrates potent HIV virucidal activity distinct from acidity alone and greater than acetic acid, suggesting a protective role in the sexual transmission of HIV.

Competition between microbial species is a product of, yet can lead to a reduction in, the microbial diversity of specific habitats. Microbial habitats can resemble ecological battlefields where microbial cells struggle to dominate and/or annihilate each other and we explore the hypothesis that (like plant weeds) some microbes are genetically hard-wired to behave in a vigorous and ecologically aggressive manner. These 'microbial weeds' are able to dominate the communities that develop in fertile but uncolonized--or at least partially vacant--habitats via traits enabling them to out-grow competitors; robust tolerances to habitat-relevant stress parameters and highly efficient energy-generation systems; avoidance of or resistance to viral infection, predation and grazers; potent antimicrobial systems; and exceptional abilities to sequester and store resources. In addition, those associated with nutritionally complex habitats are extraordinarily versatile in their utilization of diverse substrates. Weed species typically deploy multiple types of antimicrobial including toxins; volatile organic compounds that act as either hydrophobic or highly chaotropic stressors; biosurfactants; organic acids; and moderately chaotropic solutes that are produced in bulk quantities (e.g. acetone, ethanol). Whereas ability to dominate communities is habitat-specific we suggest that some microbial species are archetypal weeds including generalists such as: Pichia anomala, Acinetobacter spp. and Pseudomonas putida; specialists such as Dunaliella salina, Saccharomyces cerevisiae, Lactobacillus spp. and other lactic acid bacteria; freshwater autotrophs Gonyostomum semen and Microcystis aeruginosa; obligate anaerobes such as Clostridium acetobutylicum; facultative pathogens such as Rhodotorula mucilaginosa, Pantoea ananatis and Pseudomonas aeruginosa; and other extremotolerant and extremophilic microbes such as Aspergillus spp., Salinibacter ruber and Haloquadratum walsbyi. Some microbes, such as Escherichia coli, Mycobacterium smegmatis and Pseudoxylaria spp., exhibit characteristics of both weed and non-weed species. We propose that the concept of nonweeds represents a 'dustbin' group that includes species such as Synodropsis spp., Polypaecilum pisce, Metschnikowia orientalis, Salmonella spp., and Caulobacter crescentus. We show that microbial weeds are conceptually distinct from plant weeds, microbial copiotrophs, r-strategists, and other ecophysiological groups of microorganism. Microbial weed species are unlikely to emerge from stationary-phase or other types of closed communities; it is open habitats that select for weed phenotypes. Specific characteristics that are common to diverse types of open habitat are identified, and implications of weed biology and open-habitat ecology are discussed in the context of further studies needed in the fields of environmental and applied microbiology.

Adrenocorticotropic hormone (ACTH) levels were compared before and after submaximal and exhaustive isotonic exercise in six normally active college students (3 men and 3 women). Each subject participated in three experiments conducted at the same morning hour. Venous plasma was obtained before and immediately after 20-min runs at 65 and 80% of maximal O2 consumption (VO2 max) and after a run of gradually increasing work intensity which resulted in exhaustion (100% VO2 max) in 12.6 +/- 1.3 min. ACTH (mean +/- SE) was 48 +/- 15, 57 +/- 12, and 61 +/- 11 pg/ml before the 65, 80, and 100% VO2 max runs, and increased to 61 +/- 15 (NS), 128 +/- 18 (P less than 0.05), and 292 +/- 72 (P less than 0.05) pg/ml, respectively. Plasma glucose, growth hormone, cortisol, and lactic acid concentrations increased in a similar fashion. Cortisol and ACTH levels were significantly correlated at the higher levels of exertion: r = 0.18 (NS) for the 65% VO2 max run, r = 0.65 (P less than 0.05) for the 80% VO2 max run, and r = 0.64 (P less than 0.05) for the run to exhaustion. Both the change in ACTH with exercise and its postrun concentration were significantly related to the change in plasma lactic acid (r = 0.65, P less than 0.05) and the postrun plasma lactic acid (r = 0.64, P less than 0.05). We conclude that exercise-induced increases in plasma ACTH and their correlation with circulating cortisol depend on the intensity of isotonic exercise. Our observations also suggest that plasma lactic acid may influence ACTH release during exercise.

The relative impermeability of the blood-brain barrier (BBB) results from tight junctions and efflux transport systems limits drug delivery to the central nervous system (CNS), and thus severely restricts the therapy of many central nervous system diseases. In order to enhance the brain-specific drug delivery, we employed a 12-mer phage display peptide library to isolate peptides that could target the drug delivery system to the brain. A 12-amino-acid-peptide (denoted as Pep TGN) which was displayed by bacteriophage Clone 12-2 was finally selected by rounds of in vivo screening. Pep TGN was covalently conjugated onto the surface of poly (ethyleneglycol)-poly (lactic-co-glycolic acid) (PEG-PLGA) based nanoparticles (NPs). The cellular uptake of Pep TGN decorated nanoparticles was significantly higher than that of unmodified nanoparticles when incubated with bEnd.3 cells. Enhanced brain accumulation efficiency together with lower accumulation in liver and spleen was observed in the nude mice intravenously injected with Pep TGN conjugated nanoparticles compared with those injected with plain nanoparticles, showing powerful brain selectivity of Pep TGN. Coumarin 6 was used as a fluorescent probe for the evaluation of brain delivery properties. The brain Drug Targeting Index (DTI) of coumarin 6 incorporated in targeted nanoparticles was significantly higher than that of coumarin 6 incorporated in plain nanoparticles. In conclusion, the Pep TGN is a motif never been reported before and Pep TGN modified nanoparticles showed great potential in targeted drug delivery across the blood brain barrier.

Manipulation of the extracellular and/or intracellular pH of tumors may have considerable potential in cancer therapy. The extracellular space of most tumors is mildly acidic, owing to exuberant production of lactic acid. Aerobic glycolysis - attributable largely to chronic activation of hypoxia-inducible factor-1 (HIF-1) - as well as tumor hypoxia, are chiefly responsible for this phenomenon. Tumor acidity tends to correlate with cancer aggressiveness; in part, this reflects the ability of HIF-1 to promote invasiveness and angiogenesis. But there is growing evidence that extracellular acidity per se boosts the invasiveness and metastatic capacity of cancer cells; moreover, this acidity renders cancer cells relatively resistant to the high proportion of chemotherapeutic drugs that are mildly basic, and may impede immune rejection of tumors. Thus, practical strategies for raising the extracellular pH of tumors may have therapeutic utility. In rodents, oral administration of sodium bicarbonate can raise the extracellular pH of tumors, an effect associated with inhibition of metastasis and improved responsiveness to certain cytotoxic agents; clinical application of this strategy appears feasible. As an alternative approach, drugs that inhibit proton pumps in cancer cells may alleviate extracellular tumor acidity while lowering the intracellular pH of cancer cells; reduction of intracellular pH slows proliferation and promotes apoptosis in various cancer cell lines. Well-tolerated doses of the proton pump inhibitor esomeprazole have markedly impeded tumor growth and prolonged survival in nude mice implanted with a human melanoma. Finally, it may prove feasible to exploit the aerobic glycolysis of cancers in hyperacidification therapies; intense intracellular acidification of cancer cells achieved by induced hyperglycemia, concurrent administration of proton pump inhibitor drugs, and possibly dinitrophenol, may have the potential to kill cancer cells directly, or to potentiate their responsiveness to adjunctive measures. A similar strategy, but without proton pump inhibition, could be employed to maximize extracellular tumor acidity, enabling tumor-selective release of cytotoxic drugs encased in pH-sensitive nanoparticles.

The rate of DNA synthesis in cultures of chicken embryo fibroblasts is reduced by deprivation of serum, high population density, and other "physiological" effectors, through a reduction in the number of cells in the S-period of the cell cycle. The same effect can be produced by drastically reducing the concentration of Mg++ added to the medium. This effect is erratic, however, and better control of [Mg++] can be achieved with phosphorylated compounds which preferentially bind Mg++. Both ATP and ADP, at concentrations in the medium less than or equal to [Mg++], stimulate DNA synthesis in cultures, and at greater concentrations inhibit DNA synthesis by affe-ting the proportion of cells in the S-period. Sodium pyrophosphate, which strongly complexes Mg++, causes little stimulation of DNA synthesis at low concentrations, but causes a striking decrease at concentrations exceeding [Mg++] of the medium. The inhibition can be fully reversed by adding an excess of Mg++, and the kinetics of increase in DNA synthesis resemble those which follow the restoration of serum to serum-deprived cultures. Limitation of [Mg++] by pyrophosphate also reduces the rates of RNA and protein synthesis, 2-deoxy-D-glucose uptake, and lactic acid production to an extent comparable to the reduction caused by the removal of serum from the medium. A model for the coordinate control of metabolism, differentiated function, and growth through the activity of divalent cations is described. The compartmentalization of Mg++ within the cell serves as the key element in this coordinate control by regulating those metabolic pathways in which the rate-limiting steps are transphosphorylation reactions.

Lactic acid bacteria (LAB) constitute a diverse group of Gram positive obligately fermentative microorganisms which include both beneficial and pathogenic strains. LAB generally have complex nutritional requirements and therefore they are usually associated with nutrient-rich environments such as animal bodies, plants and foodstuffs. Amino acids represent an important resource for LAB and their utilization serves a number of physiological roles such as intracellular pH control, generation of metabolic energy or redox power, and resistance to stress. As a consequence, the regulation of amino acid catabolism involves a wide set of both general and specific regulators and shows significant differences among LAB. Moreover, due to their fermentative metabolism, LAB amino acid catabolic pathways in some cases differ significantly from those described in best studied prokaryotic model organisms such as Escherichia coli or Bacillus subtilis. Thus, LAB amino acid catabolism constitutes an interesting case for the study of metabolic pathways. Furthermore, LAB are involved in the production of a great variety of fermented products so that the products of amino acid catabolism are also relevant for the safety and the quality of fermented products.

Nanotechnology is an innovative approach that has potential applications in nutraceutical research. Phytochemicals have promising potential for maintaining and promoting health, as well as preventing and potentially treating some diseases. However, the generally low solubility, stability, bioavailability and target specificity, together with the side effects seen when used at high levels, have limited their application. Indeed, nanoparticles can increase solubility and stability of phytochemicals, enhance their absorption, protect them from premature degradation in the body and prolong their circulation time. Moreover, these nanoparticles exhibit high differential uptake efficiency in the target cells (or tissue) over normal cells (or tissue) through preventing them from prematurely interacting with the biological environment, enhanced permeation and retention effect in disease tissues and improving their cellular uptake, resulting in decreased toxicity, In this review, we outline the commonly used biocompatible and biodegradable nanoparticles including liposomes, emulsions, solid lipid nanoparticles, nanostructured lipid carriers, micelles and poly(lactic-co-glycolic acid) nanoparticles. We then summarize studies that have used these nanoparticles as carriers for epigallocatechin gallate, quercetin, resveratrol and curcumin administration to enhance their aqueous solubility, stability, bioavailability, target specificity and bioactivities.

Exosomes, nanoscale extracellular vesicles functioning as cell-to-cell communicators, are an emerging promising therapeutic in the field of bone tissue engineering. Here, we report the construction and evaluation of a novel cell-free tissue-engineered bone that successfully accelerated the restoration of critical-sized mouse calvarial defects through combining exosomes derived from human adipose-derived stem cells (hASCs) with poly(lactic-co-glycolic acid) (PLGA) scaffolds. The exosomes were immobilized on the polydopamine-coating PLGA (PLGA/pDA) scaffolds under mild chemical conditions. Specifically, we investigated the effects of hASC-derived exosomes on the osteogenic, proliferation, and migration capabilities of human bone marrow-derived mesenchymal stem cells in vitro and optimized their osteoinductive effects through osteogenic induction. Furthermore, an in vitro assay showed exosomes could release from PLGA/pDA scaffold slowly and consistently and in vivo results showed this cell-free system enhanced bone regeneration significantly, at least partially through its osteoinductive effects and capacities of promoting mesenchymal stem cells migration and homing in the newly formed bone tissue. Therefore, overall results demonstrated that our novel cell-free system comprised of hASC-derived exosomes and PLGA/pDA scaffold provides a new therapeutic paradigm for bone tissue engineering and showed promising potential in repairing bone defects.

Succinic acid is a four-carbon dicarboxylic acid produced as one of the fermentation products of anaerobic metabolism. Based on the complete genome sequence of a capnophilic succinic acid-producing rumen bacterium, Mannheimia succiniciproducens, gene knockout studies were carried out to understand its anaerobic fermentative metabolism and consequently to develop a metabolically engineered strain capable of producing succinic acid without by-product formation. Among three different CO2-fixing metabolic reactions catalyzed by phosphoenolpyruvate (PEP) carboxykinase, PEP carboxylase, and malic enzyme, PEP carboxykinase was the most important for the anaerobic growth of M. succiniciproducens and succinic acid production. Oxaloacetate formed by carboxylation of PEP was found to be converted to succinic acid by three sequential reactions catalyzed by malate dehydrogenase, fumarase, and fumarate reductase. Major metabolic pathways leading to by-product formation were successfully removed by disrupting the ldhA, pflB, pta, and ackA genes. This metabolically engineered LPK7 strain was able to produce 13.4 g/liter of succinic acid from 20 g/liter glucose with little or no formation of acetic, formic, and lactic acids, resulting in a succinic acid yield of 0.97 mol succinic acid per mol glucose. Fed-batch culture of M. succiniciproducens LPK7 with intermittent glucose feeding allowed the production of 52.4 g/liter of succinic acid, with a succinic acid yield of 1.16 mol succinic acid per mol glucose and a succinic acid productivity of 1.8 g/liter/h, which should be useful for industrial production of succinic acid.

BACKGROUND

Stem cell-based bone tissue engineering with adipose-derived stromal cells (ASCs) has shown great promise for revolutionizing treatment of large bone deficits. However, there is still a lack of consensus on cell surface markers identifying osteoprogenitors. Fluorescence-activated cell sorting has identified a subpopulation of CD105(low) cells with enhanced osteogenic differentiation. The purpose of the present study was to compare the ability of CD90 (Thy-1) to identify osteoprogenitors relative to CD(105).

METHODS

Unsorted cells, CD90(+), CD90(-), CD105(high), and CD105(low) cells were treated with an osteogenic differentiation medium. For evaluation of in vitro osteogenesis, alkaline phosphatase (ALP) staining and alizarin red staining were performed at 7 days and 14 days, respectively. RNA was harvested after 7 and 14 days of differentiation, and osteogenic gene expression was examined by quantitative real-time polymerase chain reaction. For evaluation of in vivo osteogenesis, critical-sized (4-mm) calvarial defects in nude mice were treated with the hydroxyapatite-poly(lactic-co-glycolic acid) scaffold seeded with the above-mentioned subpopulations. Healing was followed using micro-CT scans for 8 weeks. Calvaria were harvested at 8 weeks postoperatively, and sections were stained with Movat's Pentachrome.

RESULTS

Transcriptional analysis revealed that the CD90(+) subpopulation was enriched for a more osteogenic subtype relative to the CD105(low) subpopulation. Staining at day 7 for ALP was greatest in the CD90(+) cells, followed by the CD105(low) cells. Staining at day 14 for alizarin red demonstrated the greatest amount of mineralized extracellular matrix in the CD90(+) cells, again followed by the CD105(low) cells. Quantification of in vivo healing at 2, 4, 6, and 8weeks postoperatively demonstrated increased bone formation in defects treated with CD90(+) ASCs relative to all other groups. On Movat's Pentachrome-stained sections, defects treated with CD90(+) cells showed the most robust bony regeneration. Defects treated with CD90(-) cells, CD105(high) cells, and CD105(low) cells demonstrated some bone formation, but to a lesser degree when compared with the CD90(+) group.

CONCLUSIONS

While CD105(low) cells have previously been shown to possess an enhanced osteogenic potential, we found that CD90(+) cells are more capable of forming bone both in vitro and in vivo. These data therefore suggest that CD90 may be a more effective marker than CD105 to isolate a highly osteogenic subpopulation for bone tissue engineering.

Ten men and one woman were studied before, during, and following five 1-min cycling bouts (100% VO2 max) 1 h after consuming either 0.2 g/kg B.W. of NaHCO3 or a placebo drink (NaCl). The fifth exercise bout was performed until exhaustion and the time recorded to judge the effects of the NaHCO3 drink on performance. Blood samples taken from a forearm vein revealed that the intake of NaHCO3 increased resting pH (7.34 to 7.41), HCO3 (27.5 to 31.0 mM), and base excess (1.1 to 4.6 mM). During and following the exercise, blood pH and HCO3 were always higher (P less than 0.05) in the NaHCO3 than placebo trial. Performance times during the fifth cycling bout averaged 113.5 (SE +/- 12.4) and 160.8 (SE +/- 19.1) s in the NaCl and NaHCO3 trials, respectively. This 42% difference between the means was significant at the 0.01 level. The blood hydrogen ion to lactic acid ratios (nM/mM) measured between each exercise bout and during recovery from the exhaustive fifth bout suggests that the enhanced performance during the NaHCO3 trial was the result of greater buffer capacity. Thigh muscle (vastus lateralis) pH measured immediately before the fifth cycling bout in four of the subjects revealed that the working muscles were less acid in the NaHCO3 trial (pH = 6.81) than during the NaCl treatment (pH = 6.73). Thus, the alkalizing influence of oral HCO3 supports the concept that the hydrogen ion concentration in blood and muscle has a direct influence on performance during repeated, supramaximal exercise.

Every biotechnology process that relies on the use of bacteria to make a product or to overproduce a molecule may, at some time, struggle with the presence of virulent phages. For example, phages are the primary cause of fermentation failure in the milk transformation industry. This review focuses on the recent scientific advances in the field of lactic acid bacteria phage research. Three specific topics, namely, the sources of contamination, the detection methods and the control procedures will be discussed.

Poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles with anionic surface charge were surface coated with cationic di-block copolymer, poly(L-lysine)-poly(ethylene glycol)-folate (PLL-PEG-FOL) conjugate, for enhancing their site-specific intracellular delivery against folate receptor overexpressing cancer cells. The PLGA nanoparticles coated with the conjugate were characterized in terms of size, surface charge, and change in surface composition by XPS. By employing the flow cytometry method and confocal image analysis, the extent of cellular uptake was comparatively evaluated under various conditions. PLL-PEG-FOL coated PLGA nanoparticles demonstrated far greater extent of cellular uptake to KB cells, suggesting that they were mainly taken up by folate receptor-mediated endocytosis. The enhanced cellular uptake was also observed even in the presence of serum proteins, possibly due to the densely seeded PEG chains. The PLL-PEG-FOL coated PLGA nanoparticles could be potentially applied for cancer cell targeted delivery of various therapeutic agents.

The pattern of decrease in arterial bicarbonate concentration ([HCO3-]) during progressive incremental exercise was compared with that of the rise in arterial lactate ([La-]) to determine the degree of buffering of lactic acid by bicarbonate. A mathematical model was derived for the change in [HCO3-] beyond the lactate threshold. This was based on a log-log transformation of the data, a model previously found to provide a very good fit to the [La-]-O2 consumption (VO2) relationship. The results of the analysis of incremental exercise data from 10 subjects show that the decrease in [HCO3-] very nearly matches the increase in [La-]. However, it was found by comparing regression models that the correspondence between [HCO3-] and [La-] could be improved by assuming that the [HCO3-] decrease was delayed until the arterial lactate level had increased by approximately 0.4 meq/l. This result is compatible with the existence of buffering mechanisms in the cell which buffer the initial increase of lactic acid. Beyond this initial buffering, lactic acid appears to be buffered almost entirely by the bicarbonate buffer system.

Dental composites do not hinder bacteria colonization and plaque formation. Caries at the restoration margins is a frequent reason for replacement of existing restorations, which accounts for 50 to 70% of all restorations. The objectives of this study were to examine the filler level effect on nanocomposite containing nanoparticles of amorphous calcium phosphate (NACP) and investigate the load-bearing and acid-neutralizing properties and bacteria inhibition. NACP with 116-nm particle size were synthesized via a spray-drying technique and incorporated into a resin. Flexural strength of nanocomposite with 10 to 30% NACP fillers matched the strength of a commercial hybrid composite (p>> 0.1). Nanocomposite with 40% NACP matched the strength of a microfill composite, which was 2-fold that of a resin-modified glass ionomer. Nanocomposite with 40% NACP neutralized a lactic acid solution of pH 4 by rapidly increasing the pH to 5.69 in 10 min. In contrast, the commercial controls had pH staying at near 4. Using Streptoccocus mutans, an agar disk-diffusion test showed no inhibition zone for commercial controls. In contrast, the inhibition zone was (2.5 ± 0.7) mm for nanocomposite with 40% NACP. Crystal violet staining showed that S. mutans coverage on nanocomposite was 1/4 that on commercial composite. In conclusion, novel calcium-phosphate nanocomposite matched the mechanical properties of commercial composite and rapidly neutralized lactic acid of pH 4. The nanocomposite appeared to moderately reduce the S. mutans growth, and further study is needed to obtain strong antimicrobial properties. The new nanocomposite may have potential to reduce secondary caries and restoration fracture, two main challenges facing tooth cavity restorations.

Four sourdoughs (A to D) were produced under practical conditions by using a starter mixture of three commercially available sourdough starters and a baker's yeast constitutively containing various species of lactic acid bacteria (LAB). The sourdoughs were continuously propagated until the composition of the LAB flora remained stable. Two LAB-specific PCR-denaturing gradient gel electrophoresis (DGGE) systems were established and used to monitor the development of the microflora. Depending on the prevailing ecological conditions in the different sourdough fermentations, only a few Lactobacillus species were found to be competitive and became dominant. In sourdough A (traditional process with rye flour), Lactobacillus sanfranciscensis and a new species, L. mindensis, were detected. In rye flour sourdoughs B and C, which differed in the process temperature, exclusively L. crispatus and L. pontis became the predominant species in sourdough B and L. crispatus, L. panis, and L. frumenti became the predominant species in sourdough C. On the other hand, in sourdough D (corresponding to sourdough C but produced with rye bran), L. johnsonii and L. reuteri were found. The results of PCR-DGGE were consistent with those obtained by culturing, except for sourdough B, in which L. fermentum was also detected. Isolates of the species L. sanfranciscensis and L. fermentum were shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively.

We investigated the blood-pressure-lowering effects of gamma-aminobutyric acid (GABA) and a GABA-enriched fermented milk product (FMG) by low-dose oral administration to spontaneously hypertensive (SHR/Izm) and normotensive Wistar-Kyoto (WKY/Izm) rats. FMG was a non-fat fermented milk product produced by lactic acid bacteria, and the GABA contained in FMG was made from the protein of the milk during fermentation. A single oral dose of GABA or FMG (5 ml/kg; 0.5 mg GABA/kg) significantly (P<0.05) decreased the blood pressure of SHR/Izm from 4 to 8 h after administration, but did not increase that of WKY/Izm rats. The hypotensive activity of GABA was dose-dependent from 0.05 to 5.00 mg/kg in SHR/Izm. During the chronic administration of experimental diets to SHR/Izm, a significantly slower increase in blood pressure with respect to the control group was observed at 1 or 2 weeks after the start of feeding with the GABA or FMG diet respectively (P<0.05) and this difference was maintained throughout the period of feeding. The time profile of blood-pressure change due to administration of FMG was similar to that of GABA. FMG did not inhibit angiotensin 1-converting enzyme. Furthermore, an FMG peptide-containing fraction from reverse-phase chromatography lacked a hypotensive effect in SHR/Izm rats. The present results suggest that low-dose oral GABA has a hypotensive effect in SHR/Izm and that the hypotensive effect of FMG is due to GABA.

Stress-induced abdominal dysfunction is an attractive target for probiotics. To investigate the effects of the probiotic Lactobacillus casei strain Shirota on abdominal dysfunction, a double-blind, placebo-controlled trial was conducted with healthy medical students undertaking an authorized nationwide examination for academic advancement. For 8 weeks, until the day before the examination, 23 and 24 subjects consumed an L. casei strain Shirota-fermented milk and a placebo milk daily, respectively. In addition to assessments of abdominal symptoms, psychophysical state, and salivary stress markers, gene expression changes in peripheral blood leukocytes and composition of the gut microbiota were analyzed using DNA microarray analysis and 16S rRNA gene amplicon sequence analysis, respectively, before and after the intervention. Stress-induced increases in a visual analog scale measuring feelings of stress, the total score of abdominal dysfunction, and the number of genes with changes in expression of more than 2-fold in leukocytes were significantly suppressed in the L. casei strain Shirota group compared with those in the placebo group. A significant increase in salivary cortisol levels before the examination was observed only in the placebo group. The administration of L. casei strain Shirota, but not placebo, significantly reduced gastrointestinal symptoms. Moreover, 16S rRNA gene amplicon sequencing demonstrated that the L. casei strain Shirota group had significantly higher numbers of species, a marker of the alpha-diversity index, in their gut microbiota and a significantly lower percentage of Bacteroidaceae than the placebo group. Our findings indicate that the daily consumption of probiotics, such as L. casei strain Shirota, preserves the diversity of the gut microbiota and may relieve stress-associated responses of abdominal dysfunction in healthy subjects exposed to stressful situations.

A novel clinical trial was conducted with healthy medical students under examination stress conditions. It was demonstrated that the daily consumption of lactic acid bacteria provided health benefits to prevent the onset of stress-associated abdominal symptoms and a good change of gut microbiota in healthy medical students.

The innate and adaptive immune systems are important mechanisms for resistance to pathogens in the female lower genital tract. Lactobacilli at this site help maintain a healthy vagina by producing several factors including lactic acid. Indeed, bacterial vaginosis, a condition in which the genital microbiota is altered, is strongly associated with increased rates of a number of infections including HIV. However, the precise factors that contribute to increased rates of microbial and viral infections in bacterial vaginosis remain to be elucidated. We have studied the effects of bacterial microbiota in the lower genital tract on innate immunity and have found that Toll-like receptor ligands and short chain fatty acids, produced by bacterial microbiota, have dramatic effects on immune function. In this review, we will discuss these results, in addition to some recent articles that we believe will enhance our understanding of how microbes might interact with the immune system.