Since Otto Warburg reported the 'addiction' of cancer cells to fermentative glycolysis, a metabolic pathway that provides energy and building blocks, thousands of studies have shed new light on the molecular mechanisms contributing to altered cancer metabolism. Hypoxia, through hypoxia-inducible factors (HIFs), in addition to oncogenes activation and loss of tumour suppressors constitute major regulators of not only the "Warburg effect" but also many other metabolic pathways such as glutaminolysis. Enhanced glucose and glutamine catabolism has become a recognised feature of cancer cells, leading to accumulation of metabolites in the tumour microenvironment, which offers growth advantages to tumours. Among these metabolites, lactic acid, besides imposing an acidic stress, is emerging as a key signalling molecule that plays a pivotal role in cancer cell migration, angiogenesis, immune escape and metastasis. Although interest in lactate for cancer development only appeared recently, pharmacological molecules blocking its metabolism are already in phase I/II clinical trials. Here, we review the metabolic pathways generating lactate, and we discuss the rationale for targeting lactic acid transporter complexes for the development of efficient and selective anticancer therapies.

The ability to control architectural features in tissue engineering scaffolds is critical to the success of neo-tissue regeneration. In this work, reverse solid freeform fabrication and thermal phase separation of poly(L-lactic acid) (PLLA) solutions were used to create three-dimensional nano-fibrous (NF) scaffolds with complex geometries on the macro- and micro-scales. This approach allows for the fabrication of NF matrices while having precise control of internal pore size and structure, as well as external scaffold shape including architectures generated from computed-tomography scans and histological sections. In vitro cell cultivation experiments with MC3T3-E1 pre-osteoblasts were performed on NF scaffolds and on similarly designed solid-walled (SW) scaffolds that did not have nano-fibers. Proliferation studies showed significantly more cells on NF scaffolds after 7 d. In differentiation studies, the NF scaffolds displayed more uniform matrix and mineral production throughout. Real-time PCR also showed significantly higher expression of osteocalcin and bone sialoprotein mRNAs after 2 and 6 weeks in the NF scaffolds. Expression of type I collagen mRNA was lower in NF scaffolds which possibly indicates quicker differentiation on the NF substrate. In summary, we controlled the geometry of NF PLLA scaffolds at multiple size scales, and the in vitro results showed that these NF scaffolds were advantageous to control scaffolds for bone tissue engineering.

Type B Niemann-Pick disease (NPD) is a multiorgan system disorder caused by a genetic deficiency of acid sphingomyelinase (ASM), for which lung is an important and challenging therapeutic target. In this study, we designed and evaluated new delivery vehicles for enzyme replacement therapy of type B NPD, consisting of polystyrene and poly(lactic-coglycolic) acid polymer nanocarriers targeted to intercellular adhesion molecule (ICAM)-1, an endothelial surface protein up-regulated in many pathologies, including type B NPD. Real-time vascular imaging using intravital microscopy and postmortem imaging of mouse organs showed rapid, uniform, and efficient binding of fluorescently labeled ICAM-1-targeted ASM nanocarriers (anti-ICAM/ASM nanocarriers) to endothelium after i.v. injection in mice. Fluorescence microscopy of lung alveoli actin, tissue histology, and 125I-albumin blood-to-lung transport showed that anti-ICAM nanocarriers cause neither detectable lung injury, nor abnormal vascular permeability in animals. Radioisotope tracing showed rapid disappearance from the circulation and enhanced accumulation of anti-ICAM/125I-ASM nanocarriers over the nontargeted naked enzyme in kidney, heart, liver, spleen, and primarily lung, both in wild-type and ASM knockout mice. These data demonstrate that ICAM-1-targeted nanocarriers may enhance enzyme replacement therapy for type B NPD and perhaps other lysosomal storage disorders.

Concentrations of organic acids in cerebrospinal fluid (CSF) appear to be directly dependent upon their rate of production in the brain. There is evidence that the net release of short-chain monocarboxylic acids from the brain is a major route for removing these products of cerebral metabolism. Concentrations of organic acids in blood and CSF are largely independent of each other. Quantitative reference values for the concentrations of organic acids in CSF and plasma as well as ratios of individual organic acids between CSF and plasma were determined in 35 pairs of samples from paediatric patients. Over 25 organic acids were quantifiable in all or in the majority of CSF and/or plasma specimens (limit of detection 1 mumol/L). There were substantial differences in the CSF/plasma ratios between subgroups of organic acids. Metabolites related to fatty-acid oxidation were present in CSF in substantially less amounts than in plasma. Organic acids related to carbohydrate and energy metabolism and to amino acid degradation were present in CSF in the same amounts as or slightly smaller amounts than in plasma. Finally, some organic acids were found in substantially higher amounts in CSF than in plasma, e.g. glycolate, glycerate, 2,4-dihydroxybutyrate, citrate and isocitrate. Studies of organic acids in CSF and plasma samples are presented from patients with 'cerebral' lactic acidosis, disorders of propionate and methylmalonate metabolism, glutaryl-CoA dehydrogenase deficiency and L-2-hydroxy-glutaric aciduria. It became apparent that derangements of organic acids in the CSF may occur independently of the systemic metabolism. Quantitative organic acid analysis in CSF will yield new information on the pathophysiology in the central nervous system (CNS) of these disorders and may prove necessary for successful monitoring of treatment of organoacidopathies, which present mainly with neurological disease. For example, in glutaryl-CoA dehydrogenase deficiency the urinary excretion of glutarate appears to be an inadequate parameter for monitoring the effect of dietary therapy, without plasma and CSF determinations. In L-2-hydroxyglutaric aciduria the elevation of L-2-hydroxyglutarate was found to be greater in CSF than in plasma. In addition, some other organic acids, glycolate, glycerate, 2,4-dihydroxybutyrate, citrate and isocitrate, were also elevated in the CSF of the patients out of proportion to normal levels in plasma and urine. High concentrations of an unknown compound, which was tentatively identified as 2,4-dihydroxyglutarate, were found in the CSF of patients with L-2-hydroxyglutaric aciduria.(ABSTRACT TRUNCATED AT 400 WORDS)

The extracellular pH in malignant tumors is known to be lower than in normal tissues and may therefore facilitate extracellular activation of secreted lysosomal cathepsins. We have tested the capability of human mammary cells (continuous cell lines and primary culture) to acidify their extracellular environment, using two techniques. By measuring pH changes through alterations of phenolsulfone phthaleine absorbance, we found that the more aggressive MDA-MB-231 human breast cancer cells were more active in acidifying a non-buffered balanced salt solution than the estrogen receptor positive MCF7 and ZR75 cell lines and than normal mammary epithelial cells in primary culture. Metastatic breast cancer cells from pleural effusions were up to 200-fold more active in acidifying their extracellular milieu than non-malignant mammary cells cultured in the same conditions, strongly suggesting that this difference also occurs in vivo. The use of inhibitors in the presence or absence of glucose showed that both lactate and an ATP-driven proton pump sharing some characteristics of the vacuolar H+ pump were involved. Bafilomycin A1, a specific inhibitor of the vacuolar (V-type) ATP-H+ pump inhibited part of the acidification by MCF7 cells, but not by MDA-MB-231 cells. We also used microelectrodes to measure extracellular pH, in close contact to the MCF7 breast cancer cells. The pH at the free surface of MCF7 cells was lower by 0.33 +/- 0.14 unit than that of the surrounding medium, while insertion of the microelectrode tip beneath the attached surface of the cells showed a greater lowering of pH from 0.3 to 1.7 pH unit as long as cell attachment on the substrate prevented H+ diffusion. We conclude that breast carcinoma cells have a higher capacity for acidifying their extracellular milieu than normal mammary cells, and that both a plasma membrane H(+)-ATPase, and lactic acid production are involved in this acidification. It is therefore possible that the aspartyl and cysteinyl pro-cathepsins secreted in excess by tumor cells may be activated extracellularly in vivo close to the basement membrane.

The survival and the physiology of lactococcal cells in the different compartments of the digestive tracts of rats were studied in order to know better the fate of ingested lactic acid bacteria after oral administration. For this purpose, we used strains marked with reporter genes, the luxA-luxB gene of Vibrio harveyi and the gfp gene of Aequora victoria, that allowed us to differentiate the inoculated bacteria from food and the other intestinal bacteria. Luciferase was chosen to measure the metabolic activity of Lactococcus lactis in the digestive tract because it requires NADH, which is available only in metabolically active cells. The green fluorescent protein was used to assess the bacterial lysis independently of death. We report not only that specific factors affect the cell viability and integrity in some digestive tract compartments but also that the way bacteria are administrated has a dramatic impact. Lactococci which transit with the diet are quite resistant to gastric acidity (90 to 98% survival). In contrast, only 10 to 30% of bacteria survive in the duodenum. Viable cells are metabolically active in each compartment of the digestive tract, whereas most dead cells appear to be subject to rapid lysis. This property suggests that lactococci could be used as a vector to deliver specifically into the duodenum the proteins produced in the cytoplasm. This type of delivery vector would be particularly appropriate for targeting digestive enzymes such as lipase to treat pancreatic deficiencies.

Mammalian cells grown in culture excrete lactic acid and ammonium ions in quantities that may limit growth and reduce product synthesis. Frequent replenishment of the culture medium is often necessary to prevent waste product accumulation which could inhibit cell growth. Since increased medium replenishment results in increased usage of animal serum, the most expensive raw material, excessive production of waste products lowers the cell and product yield on serum, and hence increases production costs. Strategies for reducing the production of lactic acid and ammonium bymammalian cells via controlled addition of glucose and glutamine will be demonstrated. Mathematical relations coupling ammonium and glutamine kinetics will be described. Additionally, a method for automatic on-line estimation of the cell concentration was developed. This method involves calculating the ATP production rate from the oxygen uptake rate and the lactic acid production rate. Automatic online estimation of the cell concentration is critical if nutrient levels in large-scale mammaliancell cultures are to be accurately maintained via process control.

OBJECTIVE

The pathophysiology of sports-related concussion (SRC) is incompletely understood. Human adult and experimental animal investigations have revealed structural axonal injuries, decreases in the neuronal metabolite N-acetyl aspartate, and reduced cerebral blood flow (CBF) after SRC and minor traumatic brain injury. The authors of this investigation explore these possibilities after pediatric SRC.

METHODS

Twelve children, ages 11 to 15 years, who experienced SRC were evaluated by ImPACT neurocognitive testing, T1 and susceptibility weighted MRI, diffusion tensor imaging, proton magnetic resonance spectroscopy, and phase contrast angiography at <72 hours, 14 days, and 30 days or greater after concussion. A similar number of age- and gender-matched controls were evaluated at a single time point.

RESULTS

ImPACT results confirmed statistically significant differences in initial total symptom score and reaction time between the SRC and control groups, resolving by 14 days for total symptom score and 30 days for reaction time. No evidence of structural injury was found on qualitative review of MRI. No decreases in neuronal metabolite N-acetyl aspartate or elevation of lactic acid were detected by proton magnetic resonance spectroscopy. Statistically significant alterations in CBF were documented in the SRC group, with reduction in CBF predominating (38 vs 48 mL/100 g per minute; P = .027). Improvement toward control values occurred in only 27% of the participants at 14 days and 64% at >30 days after SRC.

CONCLUSIONS

Pediatric SRC is primarily a physiologic injury, affecting CBF significantly without evidence of measurable structural, metabolic neuronal or axonal injury. Further study of CBF mechanisms is needed to explain patterns of recovery.

Many substances are used in the production of biomaterials: metals (titanium), ceramics (alumina), synthetic polymers (polyurethanes, silicones, polyglycolic acid (PGA), polylactic acid (PLA), copolymers of lactic and glycolic acids (PLGA), polyanhydrides, polyorthoesters) and natural polymers (chitosan, glycosaminoglycans, collagen). With the rapid development in tissue engineering, these different biomaterials have been used as three-dimensional scaffolds and cell transplant devices. The principal biochemical and biological characteristics of the collagen-based biomaterials are presented, including their interactions with cells (fibroblasts), distinct from those of synthetic polymers, and their potential use in gene therapy through the formation of neo-organs or organoids.

Cardiovascular diseases are one of the major causes of deaths in adults in the western world. Elevated levels of certain blood lipids have been reported to be the principal cause of cardiovascular disease and other disabilities in developed countries. Several animal and clinical trials have shown a positive association between cholesterol levels and the risks of coronary heart disease. Current dietary strategies for the prevention of cardiovascular disease advocate adherence to low-fat/low-saturated-fat diets. Although there is no doubt that, in experimental conditions, low-fat diets offer an effective means of reducing blood cholesterol concentrations on a population basis, these appear to be less effective, largely due to poor compliance, attributed to low palatability and acceptability of these diets to the consumers. Due to the low consumer compliance, attempts have been made to identify other dietary components that can reduce blood cholesterol levels. Supplementation of diet with fermented dairy products or lactic acid bacteria containing dairy products has shown the potential to reduce serum cholesterol levels. Various approaches have been used to alleviate this issue, including the use of probiotics, especially Bifidobacterium spp. and Lactobacillus spp.. Probiotics, the living microorganisms that confer health benefits on the host when administered in adequate amounts, have received much attention on their proclaimed health benefits which include improvement in lactose intolerance, increase in natural resistance to infectious disease in gastrointestinal tract, suppression of cancer, antidiabetic, reduction in serum cholesterol level, and improved digestion. In addition, there are numerous reports on cholesterol removal ability of probiotics and their hypocholesterolemic effects. Several possible mechanisms for cholesterol removal by probiotics are assimilation of cholesterol by growing cells, binding of cholesterol to cellular surface, incorporation of cholesterol into the cellular membrane, deconjugation of bile via bile salt hydrolase, coprecipitation of cholesterol with deconjugated bile, binding action of bile by fibre, and production of short-chain fatty acids by oligosaccharides. The present paper reviews the mechanisms of action of anti-cholesterolemic potential of probiotic microorganisms and probiotic food products, with the aim of lowering the risks of cardiovascular and coronary heart diseases.

Ribosomally synthesized peptides with antimicrobial properties (antimicrobial peptides-AMPs) are produced by eukaryotes and prokaryotes and represent crucial components of their defense systems against microorganisms. Although they differ in structure, they are nearly all cationic and very often amphiphilic, which reflects the fact that many of them attack their target cells by permeabilizing the cell membrane. They can be roughly categorized into those that have a high content of a certain amino acid, most often proline, those that contain intramolecular disulfide bridges, and those with an amphiphilic region in their molecule if they assume an alpha-helical structure. Most of the known ribosomally synthesized peptides with antimicrobial functions have been identified and studied during the last 20 years. As a result of these studies, new knowledge has been acquired into biology and biochemistry. It has become evident that these peptides may be developed into useful antimicrobial additives and drugs. The use of two-peptide antimicrobial peptides as replacement for clinical antibiotics is promising, though their applications in preservation of foods (safe and effective for use in meat, vegetables, and dairy products), in veterinary medicine, and in dentistry are more immediate. This review focuses on the current status of some of the main types of ribosomally synthesized AMPs produced by eucaryotes and procaryotes and discusses the novel antimicrobial functions, new developments, e.g. heterologous production of bacteriocins by lactic acid bacteria, or construction of multibacteriocinogenic strains, novel applications related to these peptides, and future research paradigms.

The development and maintenance of immune homeostasis indispensably depend on signals from the gut flora. Lactic acid bacteria (LAB), which are gram-positive (G+) organisms, are plausible significant players and have received much attention. Gram-negative (G-) commensals, such as members of the family Enterobacteriaceae, may, however, be immunomodulators that are as important as G+ organisms but tend to be overlooked. Dendritic cells (DCs) are crucial immune regulators, and therefore, the present study aimed at investigating differences among human gut flora-derived LAB and G- bacteria in their patterns of DC polarization. Human monocyte-derived DCs were exposed to UV-killed bacteria, and cytokine secretion and surface marker expression were analyzed. Profound differences in the DC polarization patterns were found among the strains. While strains of LAB varied greatly in their capacity to induce interleukin-12 (IL-12) and tumor necrosis factor alpha (TNF-alpha), G- strains were consistently weak IL-12 and TNF-alpha inducers. All strains induced significant amounts of IL-10, but G- bacteria were far more potent IL-10 inducers than LAB. Interestingly, we found that when weakly IL-12- and TNF-alpha-inducing LAB and strong IL-12- and TNF-alpha-inducing LAB were mixed, the weakly IL-12- and TNF-alpha-inducing LAB efficiently inhibited otherwise strong IL-12- and TNF-alpha-inducing LAB, yet when weakly IL-12- and TNF-alpha-inducing LAB were mixed with G- bacteria, they synergistically induced IL-12 and TNF-alpha. Furthermore, strong IL-12- and TNF-alpha-inducing LAB efficiently up-regulated surface markers (CD40, CD83, CD86, and HLA-DR), which were inhibited by weakly IL-12- and TNF-alpha-inducing LAB. All G- bacteria potently up-regulated surface markers; however, these markers were not inhibited by weakly IL-12- and TNF-alpha-inducing LAB. These much divergent DC stimulation patterns among intestinal bacteria, which encompass both antagonistic and synergistic relationships, support the growing evidence that the composition of the gut flora affects immune regulation and that compositional imbalances may be involved in disease etiology.

Class IIa bacteriocins may be used as natural food preservatives, yet resistance development in the target organisms is still poorly understood. In this study, the understanding of class IIa resistance development in Listeria monocytogenes is extended, linking the seemingly diverging results previously reported. Eight resistant mutants having a high resistance level (at least a 10(3)-fold increase in MIC), originating from five wild-type listerial strains, were independently isolated following exposure to four different class IIa bacteriocin-producing lactic acid bacteria (including pediocin PA-1 and leucocin A producers). Two of the mutants were isolated from food model systems (a saveloy-type sausage at 10 degrees C, and salmon juice at 5 degrees C). Northern blot analysis showed that the eight mutants all had increased expression of EII(Bgl) and a phospho-beta-glucosidase homologue, both originating from putative beta-glucoside-specific phosphoenolpyruvate-dependent phosphotransferase systems (PTSs). However, disruption of these genes in a resistant mutant did not confer pediocin sensitivity. Comparative two-dimensional gel analysis of proteins isolated from mutant and wild-type strains showed that one spot was consistently missing in the gels from mutant strains. This spot corresponded to the MptA subunit of the mannose-specific PTS, found only in the gels of wild-type strains. The mptACD operon was recently shown to be regulated by the sigma(54) transcription factor in conjunction with the activator ManR. Class IIa bacteriocin-resistant mutants having defined mutations in mpt or manR also exhibited the two diverging PTS expression changes. It is suggested here that high-level class IIa resistance in L. monocytogenes and at least some other Gram-positive bacteria is developed by one prevalent mechanism, irrespective of wild-type strain, class IIa bacteriocin, or the tested environmental conditions. The changes in expression of the beta-glucoside-specific and the mannose-specific PTS are both influenced by this mechanism. The current understanding of the actual cause of class IIa resistance is discussed.

Streptococcus pyogenes is a human-specific pathogen that relies on its host for metabolic substrates. Rgg-like proteins constitute a family of transcriptional regulators present in several gram-positive bacteria. In S. pyogenes, Rgg influences the expression of several virulence-associated proteins localized to the cell wall and extracellular environment. Secreted enzymes may degrade host macromolecules, thereby liberating metabolic substrates. To determine if Rgg regulation of exoprotein expression is associated with altered metabolism, the catabolic activities of S. pyogenes strain NZ131 (serotype M49) and an isogenic rgg mutant strain were analyzed during growth with complex and defined media. As expected, the wild-type strain preferentially used glucose and produced lactic acid during the exponential phase of growth. In contrast, the rgg mutant fermented arginine in the exponential phase of growth, even in the presence of glucose. Arginine degradation was associated with a neutral culture pH and excretion of NH(3) and ornithine. Arginine, serine, and asparagine were depleted from mutant cultures during growth. The addition of arginine and serine to culture media increased the growth yield and NH(3) production of mutant but not wild-type cultures. Addition of asparagine had no effect on the growth yield of either strain. Altered metabolism of arginine and serine in the mutant was associated with increased transcript levels of genes encoding arginine deiminase and a putative serine dehydratase. Thus, Rgg coordinates virulence factor synthesis and catabolic activity and may be important in the pathogen's adaptation to changes in the availability of metabolic substrates.

It is a longstanding debate whether cancer is one disease or a set of very diverse diseases. The goal of this paper is to suggest strongly that most of (if not all) the hallmarks of cancer could be the consequence of the Warburg's effect. As a result of the metabolic impairment of the oxidative phosphorylation, there is a decrease in ATP concentration. To compensate the reduced energy yield, there is massive glucose uptake, anaerobic glycolysis, with an up-regulation of the Pentose Phosphate Pathway resulting in increased biosynthesis leading to increased cell division and local pressure. This increased pressure is responsible for the fractal shape of the tumor, the secretion of collagen by the fibroblasts and plays a critical role in metastatic spread. The massive extrusion of lactic acid contributes to the extracellular acidity and the activation of the immune system. The decreased oxidative phosphorylation leads to impairment in CO2 levels inside and outside the cell, with increased intracellular alkalosis and contribution of carbonic acid to extracellular acidosis-mediated by at least two cancer-associated carbonic anhydrase isoforms. The increased intracellular alkalosis is a strong mitogenic signal, which bypasses most inhibitory signals. Mitochondrial disappearance (such as seen in very aggressive tumors) is a consequence of mitochondrial swelling, itself a result of decreased ATP concentration. The transmembrane pumps, which extrude, from the mitochondria, ions, and water, are ATP-dependant. Therapy aiming at increasing both the number and the efficacy of mitochondria could be very useful.

A review is presented on the present status of biological preservation of foods. Recent developments are discussed with respect to underlying mechanisms of inhibition by 'protective' cultures, and special reference is made to lactic acid bacteria (LAB) and their 'food-grade' safety. The role of bacteriocins, their limitations and potentiating role in biological systems, is also addressed. The use of enzymes (e.g. lysozyme) for food preservation is mainly restricted by economic factors, their inactivation by endogenous food components and their limited activity spectrum. Practical applications of protective cultures refer to particular food commodities that either constitute novel systems with respect to packaging and/or composition, or represent special hygienic risks. It is concluded that biological preservation cannot substitute GMP; it, however, offers an additional (and acceptable) processing parameter for improving the safety and assuring the quality of a given food.

This study has three main objectives: Study (1) test the reproducibility and accuracy of the ICDAS I and ICDAS II caries detection systems; Study (2) validate a new impression material (Clinpro, 3M ESPE), which is said to detect lactic acid in plaque fermenting sucrose; Study (3) devise and test a scoring system for the assessment of caries activity of coronal lesions. Study (1): 141 extracted teeth were examined by two examiners using the ICDAS I and ICDAS II caries detection systems and validated against a histological classification system. Study (2): The accuracy of the impression material in predicting plaque with pH lower/higher than 5.5 was determined in an in situ study of 45 root dentin specimens by comparing the color change in the impression with the actual pH of the plaque, determined with a pH meter. Study (3): A scoring system to assess lesion activity was devised based on the predictive power of the visual appearance of the lesion (ICDAS II system), location of the lesion in a plaque stagnation area and, finally, the tactile feeling, rough/soft or smooth/hard, when running a perio-probe over the lesion. The accuracy was tested in a clinical study of 35 children with 225 lesions/sound surfaces and was validated using the Clinpro impression material for construct validity. Study (1): Intra- and inter-examiner reproducibility was found to be excellent (Kappa-values>> 0.82) and the associations strong (Spearmans correlation coefficients>> 0.90). Study (2): The Clinpro impression material was found to be acceptable as compared to the results of a pH meter, the combined sensitivity and specificity was 1.63. Study (3): ROC analysis showed that the devised classification system for determining lesion activity had acceptable accuracy (area under curve = 0.84 and the highest combined sum of specificity and sensitivity was 1.67). Thus, it is possible to predict lesion depth and assess the activity of primary coronal caries lesions accurately by using the combined knowledge obtained from visual appearance, location of the lesion and tactile sensation during probing.

Targeted delivery of therapeutics possesses the potential to localize therapeutic agents to a specific tissue as a mechanism to enhance treatment efficacy and abrogate side effects. Antibodies have been used clinically as therapeutic agents and are currently being explored for targeting drug-loaded nanoparticles. Peptides such as RGD peptides are also being developed as an inexpensive and stable alternative to antibodies. In this study, cyclo(1,12)PenITDGEATDSGC (cLABL) peptide was used to target nanoparticles to human umbilical cord vascular endothelial cell (HUVEC) monolayers that have upregulated intercellular cell-adhesion molecule-1 (ICAM-1) expression. The cLABL peptide has been previously demonstrated to possess high avidity for ICAM-1 receptors on the cell surface. Poly( dl-lactic-coglycolic acid) nanoparticles conjugated with polyethylene glycol and cLABL demonstrated rapid binding to HUVEC with upregulated ICAM-1, which was induced by treating cells with the proinflammatory cytokine, interferon-gamma. Binding of the nanoparticles could be efficiently blocked by preincubating cells with free peptide suggesting that the binding of the nanoparticles is specifically mediated by surface peptide binding to ICAM-1 on HUVEC. The targeted nanoparticles were rapidly endocytosed and trafficked to lysosomes to a greater extent than the untargeted PLGA-PEG nanoparticles. Verification of peptide-mediated nanoparticle targeting to ICAM-1 may ultimately lead to targeting therapeutic agents to inflammatory sites expressing upregulated ICAM-1.

Bacteriophage-resistant dairy streptococci were obtained following conjugal transfer of pTR2030 from a lactose-negative donor, Streptococcus lactis TEK12, to lactose-positive recipient strains, Streptococcus cremoris LMA13 and 924 and S. lactis LMA12. Fast acid-producing, phage-resistant transconjugants were selected by challenge with homologous phage on fast-slow differential agar or lactose indicator agar. Acquisition of pTR2030 by the transconjugants was confirmed by DNA-DNA hybridization. Resistance of transconjugants to homologous phage was complete. Curing or deletion of pTR2030 in the transconjugants confirmed that phage resistance was due to pTR2030 acquisition and not to coincident background mutation. Phage-sensitive pTR2030 deletion derivatives of LMA12 transconjugants were isolated in vivo. The HindIII fragment B of pTR2030 was subcloned into pBR322 to yield a recombinant plasmid, pMET2, useful as a source of pTR2030 DNA. A specific, chemically synthesized oligomer useful as a pTR2030 probe was derived from the sequence of a small portion of pTR2030. The conjugal strategy presented here was effective in yielding fast acid-producing, phage-resistant S. cremoris and S. lactis strains without the use of antibiotic resistance markers and without interfering with the acid-producing ability of the recipient strain.

Mutations in the POLG gene have emerged as one of the most common causes of inherited mitochondrial diseases in children and adults. This study sequenced the exons and flanking intronic regions of the POLG gene from 2697 unrelated patients with clinical presentations suggestive of POLG deficiency. Informative mutations have been identified in 136 unrelated individuals (5%), including 92 patients with two recessive pathogenic alleles and three patients harbouring a dominant mutation. Twenty-four novel recessive mutations and a novel possible dominant mutation, p.Y951N, were identified. All missense mutations occurred at evolutionarily conserved amino acids within functionally important regions identified by molecular modelling analyses. Oligonucleotide array comparative genomic hybridisation analyses performed on DNA samples from 81 patients with one mutant POLG allele identified a large intragenic deletion in only one patient, suggesting that large deletions in POLG are rare. The 92 patients with two mutant alleles exhibited a broad spectrum of disease. Almost all patients in all age groups had some degree of neuropathy. Seizures, hepatopathy, and lactic acidaemia were predominant in younger patients. By comparison, patients who developed symptoms in adulthood had a higher percentage of myopathy, sensory ataxia, and chronic progressive external ophthalmoplegia (CPEO)/ptosis. In conclusion, POLG mutations account for a broad clinical spectrum of mitochondrial disorders. Sequence analysis of the POLG gene should be considered as a part of routine screening for mitochondrial disorders, even in the absence of apparent mitochondrial DNA abnormalities.

Diverse microorganisms are able to grow on food matrixes and along food industry infrastructures. This growth may give rise to biofilms. This review summarizes, on the one hand, the current knowledge regarding the main bacterial species responsible for initial colonization, maturation and dispersal of food industry biofilms, as well as their associated health issues in dairy products, ready-to-eat foods and other food matrixes. These human pathogens include Bacillus cereus (which secretes toxins that can cause diarrhea and vomiting symptoms), Escherichia coli (which may include enterotoxigenic and even enterohemorrhagic strains), Listeria monocytogenes (a ubiquitous species in soil and water that can lead to abortion in pregnant women and other serious complications in children and the elderly), Salmonella enterica (which, when contaminating a food pipeline biofilm, may induce massive outbreaks and even death in children and elderly), and Staphylococcus aureus (known for its numerous enteric toxins). On the other hand, this review describes the currently available biofilm prevention and disruption methods in food factories, including steel surface modifications (such as nanoparticles with different metal oxides, nanocomposites, antimicrobial polymers, hydrogels or liposomes), cell-signaling inhibition strategies (such as lactic and citric acids), chemical treatments (such as ozone, quaternary ammonium compounds, NaOCl and other sanitizers), enzymatic disruption strategies (such as cellulases, proteases, glycosidases and DNAses), non-thermal plasma treatments, the use of bacteriophages (such as P100), bacteriocins (such us nisin), biosurfactants (such as lichenysin or surfactin) and plant essential oils (such as citral- or carvacrol-containing oils).

Inflammatory bowel disease (IBD), which includes Crohn's disease (CD) and ulcerative colitis (UC), is a chronic inflammation of the small intestine and colon caused by a dysregulated immune response to host intestinal microbiota in genetically susceptible subjects. A number of fermented dairy products contain lactic acid bacteria (LAB) and bifidobacteria, some of which have been characterized as probiotics that can modify the gut microbiota and may be beneficial for the treatment and the prevention of IBD. The objective of this review was to carry out a systematic search of LAB and bifidobacteria probiotics and IBD, using the PubMed and Scopus databases, defined by a specific equation using MeSH terms and limited to human clinical trials. The use of probiotics and/or synbiotics has positive effects in the treatment and maintenance of UC, whereas in CD clear effectiveness has only been shown for synbiotics. Furthermore, in other associated IBD pathologies, such as pouchitis and cholangitis, LAB and bifidobacteria probiotics can provide a benefit through the improvement of clinical symptoms. However, more studies are needed to understand their mechanisms of action and in this way to understand the effect of probiotics prior to their use as coadjuvants in the therapy and prevention of IBD conditions.

A cell adhesive peptide, Arg-Gly-Asp (RGD), was immobilized onto the surface of electrospun poly(D,L-lactic-co-glycolic acid) PLGA nanofiber mesh in an attempt to mimic an extracellular matrix structure. A blend mixture of PLGA and PLGA-b-PEG-NH(2) di-block copolymer dissolved in a 1:1 volume mixture of dimethylformamide and tetrahydrofuran was electrospun to produce a nanofiber mesh with functional primary amino groups on the surface. Various electrospinning parameters, such as polymer concentration and the blend ratio, were optimized to produce a nanofiber mesh with desirable morphology, surface characteristics, and fiber diameter. A cell adhesive peptide, GRGDY, was covalently grafted onto the aminated surface of the electrospun mesh under a hydrating condition. The amounts of surface primary amino groups and grafted RGD peptides were quantitatively determined. Cell attachment, spreading, and proliferation were greatly enhanced in the RGD modified electrospun PLGA nanofiber mesh compared with that of the unmodified one.

The lactic acid bacteria are a functionally related group of organisms known primarily for their bioprocessing roles in food and beverages. More recently, selected members of the lactic acid bacteria have been implicated in a number of probiotic roles that impact general health and well-being. Genomic analyses of multiple members of the lactic acid bacteria, at the genus, species, and strain level, have now elucidated many genetic features that direct their fermentative and probiotic roles. This information is providing an important platform for understanding core mechanisms that control and regulate bacterial growth, survival, signaling, and fermentative processes and, in some cases, potentially underlying probiotic activities within complex microbial and host ecosystems.