Antibacterial activity of lactoferrin and a pepsin-derived lactoferrin peptide fragment

Although the antimicrobial activity of lactoferrin has been well described, its mechanism of action has been poorly characterized. Recent work has indicated that in addition to binding iron, human lactoferrin damages the outer membrane of gram-negative bacteria. In this study, we determined whether bovine lactoferrin and a pepsin-derived bovine lactoferrin peptide (lactoferricin) fragment have similar activities. We found that both 20 microM bovine lactoferrin and 20 microM lactoferricin release intrinsically labeled [3H]lipopolysaccharide ([3H]LPS) from three bacterial strains, Escherichia coli CL99 1-2, Salmonella typhimurium SL696, and Salmonella montevideo SL5222. Under most conditions, more LPS is released by the peptide fragment than by whole bovine lactoferrin. In the presence of either lactoferrin or lactoferricin there is increased killing of E. coli CL99 1-2 by lysozyme. Like human lactoferrin, bovine lactoferrin and lactoferricin have the ability to bind to free intrinsically labeled [3H]LPS molecules. In addition to these effects, whereas bovine lactoferrin was at most bacteriostatic, lactoferricin demonstrated consistent bactericidal activity against gram-negative bacteria. This bactericidal effect is modulated by the cations Ca2+, Mg2+, and Fe3+ but is independent of the osmolarity of the medium. Transmission electron microscopy of bacterial cells exposed to lactoferricin show the immediate development of electron-dense membrane blisters. These experiments offer evidence that bovine lactoferrin and lactoferricin damage the outer membrane of gram-negative bacteria. Moreover, the peptide fragment lactoferricin has direct bactericidal activity. As lactoferrin is exposed to proteolytic factors in vivo which could cleave the lactoferricin fragment, the effects of this peptide are of both mechanistic and physiologic relevance

Hereditary palmoplantar keratoderma - phenotypes and mutations in 64 patients

Background Hereditary palmoplantar keratodermas (PPK) represent a heterogeneous group of rare skin disorders with epidermal hyperkeratosis of the palms and soles, with occasional additional manifestations in other tissues. Mutations in at least 69 genes have been implicated in PPK, but further novel candidate genes and mutations are still to be found. Objectives To identify mutations underlying PPK in a cohort of 64 patients. Methods DNA of 48 patients was analysed on a custom-designed in-house panel for 35 PPK genes, and 16 patients were investigated by a diagnostic genetic laboratory either by whole-exome sequencing, gene panels or targeted single-gene sequencing. Results Of the 64 PPK patients, 32 had diffuse (50%), 19 focal (30%) and 13 punctate (20%) PPK. None had striate PPK. Pathogenic mutations in altogether five genes were identified in 31 of 64 (48%) patients, the majority (22/31) with diffuse PPK. Of them, 11 had a mutation in AQP5, five in SERPINB7, four in KRT9 and two in SLURP1. AAGAB mutations were found in nine punctate PPK patients. New mutations were identified in KRT9 and AAGAB. No pathogenic mutations were detected in focal PPK. Variants of uncertain significance (VUS) in PPK-associated and other genes were observed in 21 patients that might explain their PPK. No suggestive pathogenic variants were found for 12 patients. Conclusions Diffuse PPK was the most common (50%) and striate PPK was not observed. We identified pathogenic mutations in 48% of our PPK patients, mainly in five genes: AQP5, AAGAB, KRT9, SERPINB7 and SLURP1.Peer reviewe

Four-dimensional distribution of the 2010 Eyjafjallajökull volcanic cloud over Europe observed by EARLINET

© Author(s) 2013. This work is distributed under the Creative Commons Attribution 3.0 License.The eruption of the Icelandic volcano Eyjafjallaj ökull in April-May 2010 represents a "natural experiment" to study the impact of volcanic emissions on a continental scale. For the first time, quantitative data about the presence, altitude, and layering of the volcanic cloud, in conjunction with optical information, are available for most parts of Europe derived from the observations by the European Aerosol Research Lidar NETwork (EARLINET). Based on multi-wavelength Raman lidar systems, EARLINET is the only instrument worldwide that is able to provide dense time series of high-quality optical data to be used for aerosol typing and for the retrieval of particle microphysical properties as a function of altitude. In this work we show the four-dimensional (4-D) distribution of the Eyjafjallajökull volcanic cloud in the troposphere over Europe as observed by EARLINET during the entire volcanic event (15 April-26 May 2010). All optical properties directly measured (backscatter, extinction, and particle linear depolarization ratio) are stored in the EARLINET database available at www.earlinet.org. A specific relational database providing the volcanic mask over Europe, realized ad hoc for this specific event, has been developed and is available on request at www.earlinet.org. During the first days after the eruption, volcanic particles were detected over Central Europe within a wide range of altitudes, from the upper troposphere down to the local planetary boundary layer (PBL). After 19 April 2010, volcanic particles were detected over southern and south-eastern Europe. During the first half of May (5-15 May), material emitted by the Eyjafjallajökull volcano was detected over Spain and Portugal and then over the Mediterranean and the Balkans. The last observations of the event were recorded until 25 May in Central Europe and in the Eastern Mediterranean area. The 4-D distribution of volcanic aerosol layering and optical properties on European scale reported here provides an unprecedented data set for evaluating satellite data and aerosol dispersion models for this kind of volcanic events.Peer reviewe

Histone deacetylase 1 and 2 differentially regulate apoptosis by opposing effects on extracellular signal-regulated kinase 1/2

Histone deacetylases (HDACs) are epigenetic regulators that are important for the control of various pathophysiological events. We found that HDAC inhibitors completely abolished transforming growth factor-β1 (TGF-β1)-induced apoptosis in AML-12 and primary mouse hepatocytes. Expression of a dominant-negative mutant of HDAC1 or downregulation of HDAC1 by RNAi both suppressed TGF-β1-induced apoptosis. In addition, overexpression of HDAC1 enhanced TGF-β1-induced apoptosis, and the rescue of HDAC1 expression in HDAC1 RNAi cells restored the apoptotic response of cells to TGF-β1. These data indicate that HDAC1 functions as a proapoptotic factor in TGF-β1-induced apoptosis. In contrast, downregulation of HDAC2 by RNAi increased spontaneous apoptosis and markedly enhanced TGF-β1-induced apoptosis, suggesting that HDAC2 has a reciprocal role in controlling cell survival. Furthermore, inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) by MEK1 inhibitor PD98059 or expression of a kinase-dead mutant of MEK1 restored the apoptotic response to TGF-β1 in HDAC1 RNAi cells. Strikingly, HDAC2 RNAi caused an inhibition of ERK1/2, and the spontaneous apoptosis can be abolished by reactivation of ERK1/2. Taken together, our data demonstrate that HDAC1 and 2 reciprocally affect cell viability by differential regulation of ERK1/2; these observations provide insight into the roles and potential mechanisms of HDAC1 and 2 in apoptosis

Decreased Cerebrovascular Brain-Derived Neurotrophic Factor–Mediated Neuroprotection in the Diabetic Brain

Objective: Diabetes is an independent risk factor for stroke. However, the underlying mechanism of how diabetes confers that this risk is not fully understood. We hypothesize that secretion of neurotrophic factors by the cerebral endothelium, such as brain-derived neurotrophic factor (BDNF), is suppressed in diabetes. Consequently, such accrued neuroprotective deficits make neurons more vulnerable to injury. Research Design and Methods: We examined BDNF protein levels in a streptozotocin-induced rat model of diabetes by Western blotting and immunohistochemistry. Levels of total and secreted BDNF protein were quantified in human brain microvascular endothelial cells after exposure to advanced glycation end product (AGE)-BSA by enzyme-linked immunosorbent assay and immunocytochemistry. In media transfer experiments, the neuroprotective efficacy of conditioned media from normal healthy endothelial cells was compared with AGE-treated endothelial cells in an in vitro hypoxic injury model. Results: Cerebrovascular BDNF protein was reduced in the cortical endothelium in 6-month diabetic rats. Immunohistochemical analysis of 6-week diabetic brain sections showed that the reduction of BDNF occurs early after induction of diabetes. Treatment of brain microvascular endothelial cells with AGE caused a similar reduction in BDNF protein and secretion in an extracellular signal–related kinase-dependent manner. In media transfer experiments, conditioned media from AGE-treated endothelial cells were less neuroprotective against hypoxic injury because of a decrease in secreted BDNF. Conclusions: Taken together, our findings suggest that a progressive depletion of microvascular neuroprotection in diabetes elevates the risk of neuronal injury for a variety of central nervous system diseases, including stroke and neurodegeneration

Cellular changes in boric acid-treated DU-145 prostate cancer cells

Epidemiological, animal, and cell culture studies have identified boron as a chemopreventative agent in prostate cancer. The present objective was to identify boron-induced changes in the DU-145 human prostate cancer cell line. We show that prolonged exposure to pharmacologically-relevant levels of boric acid, the naturally occurring form of boron circulating in human plasma, induces the following morphological changes in cells: increases in granularity and intracellular vesicle content, enhanced cell spreading and decreased cell volume. Documented increases in β-galactosidase activity suggest that boric acid induces conversion to a senescent-like cellular phenotype. Boric acid also causes a dose-dependent reduction in cyclins A–E, as well as MAPK proteins, suggesting their contribution to proliferative inhibition. Furthermore, treated cells display reduced adhesion, migration and invasion potential, along with F-actin changes indicative of reduced metastatic potential. Finally, the observation of media acidosis in treated cells correlated with an accumulation of lysosome-associated membrane protein type 2 (LAMP-2)-negative acidic compartments. The challenge of future studies will be to identify the underlying mechanism responsible for the observed cellular responses to this natural blood constituent

SFRP1 reduction results in an increased sensitivity to TGF-β signaling

Background Transforming growth factor (TGF)-β plays a dual role during mammary gland development and tumorigenesis and has been shown to stimulate epithelial-mesenchymal transition (EMT) as well as cellular migration. The Wnt/β-catenin pathway is also implicated in EMT and inappropriate activation of the Wnt/β-catenin signaling pathway leads to the development of several human cancers, including breast cancer. Secreted frizzled-related protein 1 (SFRP1) antagonizes this pathway and loss of SFRP1 expression is frequently observed in breast tumors and breast cancer cell lines. We previously showed that when SFRP1 is knocked down in immortalized non-malignant mammary epithelial cells, the cells (TERT-siSFRP1) acquire characteristics associated with breast tumor initiating cells. The phenotypic and genotypic changes that occur in response to SFRP1 loss are consistent with EMT, including a substantial increase in the expression of ZEB2. Considering that ZEB2 has been shown to interact with mediators of TGF-β signaling, we sought to determine whether TGF-β signaling is altered in TERT-siSFRP1 cells. Methods Luciferase reporter assays and real-time PCR analysis were employed to measure TGF-β transcriptional targets. Western blot analysis was used to evaluate TGF-β-mediated ERK1/2 phosphorylation. Migration chamber assays were utilized to quantify cellular migration. TERT-siSFRP1 cells were transfected with Stealth RNAi™ siRNA in order to knock-down the expression of ZEB2. Results TERT-siSFRP1 cells exhibit a significant increase in both TGF-β-mediated luciferase activity as well as TGF-β transcriptional targets, including Integrin β3 and PAI-1. Phosphorylation of ERK1/2 is increased in TERT-siSFRP1 cells in response to enhanced TGF-β signaling. Furthermore, when the TGF-β pathway is blocked with a TGF-βR antagonist (LY364947), cellular migration is significantly hindered. Finally, we found that when ZEB2 is knocked-down, there is a significant reduction in the expression of exogeneous and endogenous TGF-β transcriptional targets and cellular migration is impeded. Conclusions We demonstrate that down-regulation of SFRP1 renders mammary epithelial cells more sensitive to TGF-β signaling which can be partially ameliorated by blocking the expression of ZEB2

Search for the glueball candidates f0(1500) and fJ(1710) in gamma gamma collisions

Data taken with the ALEPH detector at LEP1 have been used to search for gamma gamma production of the glueball candidates f0(1500) and fJ(1710) via their decay to pi+pi-. No signal is observed and upper limits to the product of gamma gamma width and pi+pi- branching ratio of the f0(1500) and the fJ(1710) have been measured to be Gamma_(gamma gamma -> f0(1500)). BR(f0(1500)->pi+pi-) < 0.31 keV and Gamma_(gamma gamma -> fJ(1710)). BR(fJ(1710)->pi+pi-) < 0.55 keV at 95% confidence level.Comment: 10 pages, 3 figure

Search for supersymmetry with a dominant R-parity violating LQDbar couplings in e+e- collisions at centre-of-mass energies of 130GeV to 172 GeV

A search for pair-production of supersymmetric particles under the assumption that R-parity is violated via a dominant LQDbar coupling has been performed using the data collected by ALEPH at centre-of-mass energies of 130-172 GeV. The observed candidate events in the data are in agreement with the Standard Model expectation. This result is translated into lower limits on the masses of charginos, neutralinos, sleptons, sneutrinos and squarks. For instance, for m_0=500 GeV/c^2 and tan(beta)=sqrt(2) charginos with masses smaller than 81 GeV/c^2 and neutralinos with masses smaller than 29 GeV/c^2 are excluded at the 95% confidence level for any generation structure of the LQDbar coupling.Comment: 32 pages, 30 figure