Molecular identification and antifungal susceptibility testing of Candida species isolated from dental plaques.

OBJECTIVE: The aim of the present study was to provide insight into the prevalence and susceptibility profiles of Candida species isolated from the dental plaque of Iranian immunocompetent patients. As a biofilm, Candida species are responsible for several disorders common to the oral cavity including gingivitis, dental caries, periodontitis, and the less common severe systemic infections specifically in immunosuppressed individuals. METHOD: PCR-RFLP was performed to identify yeasts isolated from the dental plaques of 40 immunocompetent patients. Moreover, antifungal susceptibility testing was performed in according to CLSI guidelines (M27-A3). RESULTS: Among 40 yeasts isolated from the dental plaques of immunocompetent patients, Candida albicans was the most common species (92.5%), followed by P. kudriavzevii (7.5%). It is the first isolation of P. kudriavzevii from dental plaques and the first evaluation of antifungal effect of the new imidazole, luliconazole and echinocandins against these samples worldwide. Luliconazole, voriconazole, amphotericin B and anidulafungin showed the best activity with the lowest geometric mean (GM) 0.03, 0.06, 0.08 and 0.09µg/ml, respectively, followed by miconazole (0.14µg/mL), caspofungin (0.24µg/mL) fluconazole (0.38µg/mL) and itraconazole (0.5µg/mL). CONCLUSION: The current study demonstrated luliconazole and echinocandins displayed excellent activity against all Candida isolates from dental plaques, presenting promising and potent alternative for all oral Candidiasis.

Minocycline attenuates hypoxia-inducible factor-1α expression correlated with modulation of p53 and AKT/mTOR/p70S6K/4E-BP1 pathway in ovarian cancer: in vitro and in vivo studies.

Hypoxia-inducible factor (HIF)-1α is the key cellular survival protein under hypoxia, and is associated with tumor progression and angiogenesis. We have recently shown the inhibitory effects of minocycline on ovarian tumor growth correlated with attenuation of vascular endothelial growth factor (VEGF) and herein report a companion laboratory study to test if these effects were the result of HIF-1α inhibition. In vitro, human ovarian carcinoma cell lines (A2780, OVCAR-3 and SKOV-3) were utilized to examine the effect of minocycline on HIF-1 and its upstream pathway components to elucidate the underlying mechanism of action of minocycline. Mice harboring OVCAR-3 xenografts were treated with minocycline to assess the in vivo efficacy of minocycline in the context of HIF-1. Minocycline negatively regulated HIF-1α protein levels in a concentration-dependent manner and induced its degradation by a mechanism that is independent of prolyl-hydroxylation. The inhibition of HIF-1α was found to be associated with up-regulation of endogenous p53, a tumor suppressor with confirmed role in HIF-1α degradation. Further studies demonstrated that the effect of minocycline was not restricted to proteasomal degradation and that it also caused down-regulation of HIF-1α translation by suppressing the AKT/mTOR/p70S6K/4E-BP1 signaling pathway. Minocycline treatment of mice bearing established ovarian tumors, led to suppression of HIF-1α accompanied by up-regulation of p53 protein levels and inactivation of AKT/mTOR/p70S6K/4E-BP1 pathway. These data reveal the therapeutic potential of minocycline in ovarian cancer as an agent that targets the pro-oncogenic factor HIF-1α through multiple mechanisms.

Production of immune response mediators by HT-29 intestinal cell-lines in the presence of Bifidobacterium-treated infant microbiota.

The colonisation and establishment of the intestinal microbiota starts immediately at birth and is essential for the development of the intestine and the immune system. This microbial community gradually increases in number and diversity until the age of two or three years when it becomes a stable ecosystem resembling that of adults. This period constitutes a unique window of opportunity to modulate it through probiotic action, with a potential impact in later health. In the present work we have investigated how putative bifidobacterial probiotics modify the metabolic profiles and immune-modulatory properties of faecal microbiotas. An in vitro pH-controlled single-stage continuous-culture system (CCS) inoculated with infant faeces was employed to characterise the effects of two Bifidobacterium species on the intestinal microbiotas in three children, together with the effects of these modified microbiotas on cytokine production by HT-29 cells. Intestinal bacterial communities, production of short-chain fatty acids and lactate were determined by quantitative PCR and gas chromatography, respectively. Cytokines production by HT-29 cells was measured by ELISA. The combination of CCS with infant faeces and human intestinal cells provided a suitable model to evaluate the specific modulation of the intestinal microbiota and immune system by probiotics. In the CCS, infant faecal microbiotas were influenced by the addition of bifidobacteria, resulting in changes in their ability to induce the production of immune mediators by HT-29 cells. The different metabolic and immunological responses induced by the bifidobacterial species tested indicate the need to assess potential probiotics in model systems including complex intestinal microbiotas. Potential probiotic bifidobacteria can modulate the infant microbiota and its ability to induce the production of mediators of the immune response by intestinal cells.

p70 Ribosomal protein S6 kinase (Rps6kb1): an update.

The Rps6kb1 gene encodes the 70 kDa ribosomal protein S6 kinase (p70S6K), which is a serine/threonine kinase regulated by phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) pathway. p70S6K plays a crucial role in controlling cell cycle, growth and survival. The PI3K/mTOR signalling pathway is one of the major mechanisms for controlling cell survival, proliferation and metabolism and is the central regulator of translation of some components of protein synthesis system. Upon activation, this kinase phosphorylates S6 protein of ribosomal subunit 40S resulting in selective translation of unique family of mRNAs that contain oligopyrimidine tract on 5' transcriptional site (5'TOP). 5'TOP mRNAs are coding the components of translational apparatus including ribosomal proteins and elongation factors. Due to the role of p70S6K in protein synthesis and also its involvement in a variety of human diseases ranging from diabetes and obesity to cancer, p70S6K is now being considered as a new therapeutic target for drug development. Furthermore, p70S6K acts as a biomarker for response to immunosuppressant as well as anticancer effects of inhibitors of the mTOR. Because of the narrow therapeutic index of mTOR inhibitors, drug monitoring is essential, and this is usually done by measuring blood drug levels, therapeutic response and drug-induced adverse effects. Recent studies have suggested that plasma p70S6K is a reliable index for the monitoring of patient response to mTOR inhibitors. Therefore, a better understanding of p70S6K and its role in various pathological conditions could enable the development of strategies to aid diagnosis, prognosis and treatment schedules.

Synbiotic consumption changes the metabolism and composition of the gut microbiota in older people and modifies inflammatory processes: a randomised, double-blind, placebo-controlled crossover study.

BACKGROUND: Ageing can result in major changes in the composition and metabolic activities of bacterial populations in the large gut and an impaired immune system. AIM: To investigate the effects of synbiotic consumption on the colonic microbiota, immune function and health status in older people. METHODS: A randomised, double-blind placebo-controlled, 4-week crossover study was carried out, involving 43 older volunteers, using a synbiotic comprising the probiotic Bifidobacterium longum and an inulin-based prebiotic Synergy 1 (SudZucker, Mannheim, Germany). Faecal and blood samples were collected, and clinical status scored at the start, and at 2- and 4-week intervals, with a 4-week washout between each feeding period. Faecal bacteria were determined by fluorescent in situ hybridisation. Short-chain fatty acid concentrations, cytokine production, bowel habit and a range of clinical parameters were measured. RESULTS: The synbiotic increased bifidobacterial numbers by 1.4 log units (P < 0.0001) and also increased members of the phyla Actinobacteria and Firmicutes (P = 0.0004, P < 0.0001). Proteobacteria were reduced by 1.0 log units (P < 0.0001). Synbiotic feeding was associated with increased butyrate production (P = 0.0399). The pro-inflammatory response was modified by the synbiotic, with significantly reduced pro-inflammatory cytokine TNF-α in peripheral blood after 2 and 4 weeks of synbiotic consumption (P = 0.02, P = 0.0406). The synbiotic had no effect on bowel habit or any clinical parameters. CONCLUSION: Short-term synbiotic use can be effective in improving the composition and metabolic activities of colonic bacterial communities and immune parameters in older people. This study was registered at clinicaltrials.gov as NCT01226212.

Online social network size is reflected in human brain structure.

The increasing ubiquity of web-based social networking services is a striking feature of modern human society. The degree to which individuals participate in these networks varies substantially for reasons that are unclear. Here, we show a biological basis for such variability by demonstrating that quantitative variation in the number of friends an individual declares on a web-based social networking service reliably predicted grey matter density in the right superior temporal sulcus, left middle temporal gyrus and entorhinal cortex. Such regions have been previously implicated in social perception and associative memory, respectively. We further show that variability in the size of such online friendship networks was significantly correlated with the size of more intimate real-world social groups. However, the brain regions we identified were specifically associated with online social network size, whereas the grey matter density of the amygdala was correlated both with online and real-world social network sizes. Taken together, our findings demonstrate that the size of an individual's online social network is closely linked to focal brain structure implicated in social cognition.

Induction of cytokine formation by human intestinal bacteria in gut epithelial cell lines.

AIMS: To investigate the effects of human gut micro-organisms on cytokine production by human intestinal cell lines. METHODS AND RESULTS: Quantitative real-time PCR assays were developed to measure the production of pro-inflammatory (IL-1α, IL-6, IL-18 and TNFα) and anti-inflammatory (TGF-ß1, TGF-ß2, TGF-ß3, IL-4 and IL-10) cytokines in HT-29 and Caco-2 cell lines. They were co-cultured with a range of mucosal bacteria isolated from ulcerative colitis patients, together with lactobacilli and bifidobacteria obtained from healthy people. HT-29 cells were also co-cultured with Campylobacter jejuni, enterotoxigenic Escherichia coli (ETEC), enteropathogenic E. coli and Salmonella typhimurium. The majority of commensal bacteria tested suppressed the expression of anti-inflammatory cytokine mRNA, increased IL-18, reduced IL-1α, and with the exception of nonpathogenic E. coli, reduced TNF-α. All overtly pathogenic species increased both pro-inflammatory and anti-inflammatory cytokine mRNA. CONCLUSION: Commensal and pathogenic species induced fundamentally different cytokine responses in human intestinal epithelial cell lines. SIGNIFICANCE AND IMPACT OF THE STUDY: Interactions between commensal bacteria tested in this study and the innate immune system were shown to be anti-inflammatory in nature, in contrast to the pathogenic organisms investigated. These data contribute towards our understanding of how potential probiotic species can be used to suppress the pro-inflammatory response in inflammatory bowel disease.

Effect of low fluency dentin conditioning on tensile bond strength of composite bonded to Er:YAG laser-prepared dentin: a preliminary study.

Several studies in the literature have previously shown that the bond strength of a composite bonded to dentin is almost equivalent as when dentin is prepared by either bur or Er:YAG laser. The aim of this preliminary study is to assess the hypothesis that dentin conditioning at low fluency by means of Er:YAG laser can improve the value of adhesion of composites resin to dentin. Sixty surfaces of caries-free human third molars extracted for orthodontic purposes were randomly divided into five groups of 12 teeth. The bur group was the control, prepared using bur, group L was prepared using Er:YAG 200 mJ, SSP (50 µs), 20 Hz, 15 seconds of sweeping, for groups L80, L100, L120, they were prepared first, with the same parameters of the group L 200, and then they received a conditioning, which is, respectively, 15 s of irradiations at: 80 mJ (SSP, 10 Hz), 100 mJ (SSP, 10 Hz), and 120 mJ (SSP, 10 Hz). All samples were restored in a single-component adhesive system: Xenon (DENTSPLY), and ceramX (DENTSPLY) as the resin composite. The specimens were submitted to tensile bond strength test using a universal testing machine. Data were submitted to statistical analysis using ANOVA coupled to a Tukey-Kramer test at the 95% level. The mean values in MPa were 33.3 for group B, 36.73 for group L 200, 41.7 for group L80, 37.9 for group L100, and 39.1 for group L120. Our results showed that dentin conditioning at a low fluency of 12.58 J/cm(2) per pulse, with 80 mJ output energy and 50-µs pulse duration can significantly improve tensile bond strength of a composite bonded to Er:YAG laser-prepared dentine.

Clinical trial: the microbiological and immunological effects of synbiotic consumption - a randomized double-blind placebo-controlled study in active Crohn's disease.

BACKGROUND: Crohn's disease is an inflammatory illness in which the immune response against gut microorganisms is believed to drive an abnormal immune response. Consequently, modification of mucosal bacterial communities, and the immune effects they elicit, might be used to modify the disease state. AIM: To investigate the effects of synbiotic consumption on disease processes in patients with Crohn's disease. METHODS: A randomized, double-blind placebo-controlled trial was conducted involving 35 patients with active Crohn's disease, using a synbiotic comprising Bifidobacterium longum and Synergy 1. Clinical status was scored and rectal biopsies were collected at the start, and at 3- and 6-month intervals. Transcription levels of immune markers and mucosal bacterial 16S rRNA gene copy numbers were quantified using real-time PCR. RESULTS: Significant improvements in clinical outcomes occurred with synbiotic consumption, with reductions in both Crohn's disease activity indices (P = 0.020) and histological scores (P = 0.018). The synbiotic had little effect on mucosal IL-18, INF-gamma and IL-1beta; however, significant reductions occurred in TNF-alpha expression in synbiotic patients at 3 months (P = 0.041), although not at 6 months. Mucosal bifidobacteria proliferated in synbiotic patients. CONCLUSION: Synbiotic consumption was effective in improving clinical symptoms in patients with active Crohn's disease.

Crystals of an ATPase fragment of bovine clathrin uncoating ATPase.

A 44,000 Mr amino-terminal, clathrin-independent ATPase fragment of the bovine clathrin uncoating ATPase has been crystallized in a form suitable for X-ray diffraction studies. The crystals are orthorhombic, space group P2(1)2(1)2(1), a = 145.3 A, b = 65.0 A, c = 46.9 A, with one protein molecule per asymmetric unit (1 A = 0.1 nm).

Development of a camel kidney cell strain and its use in virology.

A strain of camel kidney cells was developed and carried in serial passages. The subcultures were slow-growing in the early passages and were composed of heterogeneous cell population. By the 35th passage, the growth rate increased, and more homogeneous cells, mostly of the epithelioid type, were seen. The cell strain was highly susceptible to West Nile, Sindbis, vesicular stomatitis, adeno, and vaccinia viruses, and also was susceptible to herpes simplex, rinderpest, measles, and canine distemper viruses.