1.
Nucleic Acid and Peptide Aptamers: Fundamentals and Bioanalytical Aspects.
Source
Dipartimento di Chimica "Ugo Schiff", Università degli Studi di Firenze, Via della Lastruccia 3, 50019 Sesto Fiorentino (Italy). marco.mascini@unifi.it.
Abstract
In recent years new nucleic acid and protein-based combinatorial molecules have attracted the attention of researchers working in various areas of science, ranging from medicine to analytical chemistry. These molecules, called aptamers, have been proposed as alternatives to antibodies in many different applications. The aim of this Review is to illustrate the peculiarities of these combinatorial molecules which have initially been explored for their importance in molecular medicine, but have enormous potential in other biotechnological fields historically dominated by antibodies, such as bioassays. A description of these molecules is given, and the methods for their selection and production are also summarized. Moreover, critical aspects related to these molecules are discussed.
Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
- PMID:
- 22213382
- [PubMed - as supplied by publisher]
In this issue.
Abstract
COVER IMAGE: The cover shows a histological analysis of an ear section from a C57BL/6 mouse that was first sensitised with 2,4-dinitrofluorobenzene on the belly and then challenged 4 days later with 2,4-dinitrofluorobenzene on the ear. Haematoxylin and eosin staining shows massive skin infiltration by mononuclear cells and granulocytes 24 h after challenge. The image is from Rouzaire et al. (pp. 80-88) in which the authors show that NK cells and T cells induce different types of inflammatory reactions in the skin during recall responses to haptens; of note, NK cell-mediated pathogenesis does not rely on cellular infiltrate. A 'MÉNAGE À TROIS' IN THE GERMINAL CENTRE INVOLVING HUMAN ΓΔ T CELLS: Vγ9/Vδ2 T cells comprise a minor subset of unconventional T cells in the blood which uniquely respond to HMB-PP, a non-peptide metabolite produced by a large range of microbial pathogens. For reasons that are not understood Vγ9/Vδ2 T cells and immune responsiveness to HMB-PP are found only in higher primates and are absent in all other vertebrates including mice. In this issue, Bansal et al. demonstrate that HMB-PP induces upregulation of the IL-21 receptor on human Vγ9/Vδ2 T cells. The authors show that IL-21 plays a co-stimulatory role in the induction of CXCL13, CXCR5 and ICOS expression by HMB-PP-activated Vγ9/Vδ2 T cells and in the potential of these cells to provide B-cell help. These findings suggest that the 'ménage à trois' of Vγ9/Vδ2 T cells, T(FH) cells and B cells in the germinal centres of secondary lymphoid tissues is likely to impact on the generation of high-affinity, class-switched antibodies in microbial infections. pp. 110-119 THE HEART OF HIV-ASSOCIATED TUBERCULOSIS: Compared with those not infected with HIV, HIV-infected individuals are up to 30 times more likely to develop tuberculosis, with an increased risk of extrapulmonary disease. In this issue, Matthews et al. show that in the pericardial form of extrapulmonary tuberculosis, increased HIV replication at the tuberculous disease site leads to the elimination of the CCR5(+) effector memory CD4(+) T-cell population by the virus. This elimination of the memory population is accompanied by the recruitment of less differentiated cells to the disease site. The presence of polyfunctional CD4(+) T cells expressing TNF, IL-2 and IFN-γ is consistent with the less mature phenotype of CD4(+) memory T cells at the disease site of HIV-1-infected patients. Interestingly, and unlike other extrapulmonary tuberculous disease sites, there are instances where the compartmentalisation of antigen-specific T cells is not detectable. As this lack of compartmentalisation might limit diagnostic application in pericardial tuberculosis, the underlying reasons require further elucidation. pp. 147-157 HYBRIDS, NOT JUST FOR CARS: ΓΔ T-CELL TRANSCRIPTOME IS A HYBRID OF ΑΒ T-CELL AND NK-CELL SIGNATURES: Human γδ T cells reactive to non-peptide phosphoantigens represent promising new anti-cancer immunotherapeutics, but they are hitherto poorly characterized. In this issue, Pont et al. perform the first Affymetrix-based transcriptome analysis of human phosphoantigen-specific γδ T cells in resting and activated conditions. In comparison with a broad collection of transcriptomes from both healthy lymphocytes and lymphoma cells, phosphoantigen-specific γδ T cells are found to cluster with - and between - healthy αβ T and NK cells. γδ T cells show gene expression signatures corresponding to both αβ Th1 and NK cell functional activities, but not to phagocytic or antigen-presenting cell (APC) bioactivities. Functional studies show that Th1 cytokine, chemokine and cytotoxic activities reflect a high mitotic activity at later time points rather than antigen-presenting functions. The transcriptomes from these phosphoantigen-specific γδ T cells are therefore strikingly distinct from that of NK/T or peripheral T-cell lymphomas of the γδ subtype. pp. 228-240 NATURAL ANTIBODY-PRODUCING B-1 CELLS EXIST IN THE BONE MARROW: A small subset of B cells, B-1 cells, is thought to generate "natural antibodies" (nAbs) - antibodies that are generated without overt immunization and importantly contribute to antimicrobial defenses and tissue homeostasis. However, while nAb production occurs mainly in the spleen and bone marrow (BM), B-1 cells are most prominent in the body cavity of mice and are not thought to exist in BM. In this issue, Choi et al. resolve this apparent conundrum by identifying, for the first time, nAb-secreting B-1 cells in the BM. BM B-1 cells share many characteristics with splenic B-1 cells, but are distinct from their counterparts in body cavities, which despite their relative abundance contribute little to the nAb pools. nAb-producing B-1 cells are also distinct from conventional plasma cells. These data highlight the significant contribution of BM B-1 cells as nAb-producers and supports the view that body cavity B-1 cells function as rapidly inducible cellular reservoirs, but not pools, of nAb-producing cells. pp. 120-129 FROM OUR SISTER JOURNALS - COME ON IN! HOW ANTIBODIES CAN ALSO ACT INSIDE CELLS: It was recently discovered that - besides providing extracellular protection against pathogens - antibodies bound to adenovirus can also enter the cell and lead to the disposal of the virus intracellularly. McEwan et al. describe how this antibody-dependent intracellular neutralisation (ADIN) works: inside the cell the antibodies are recognized by the receptor TRIM21, which binds to IgG with high affinity. The binding of TRIM21 to IgG subsequently recruits the ubiquitin-proteasome system, leading to the degradation of the virus. Furthermore the authors discuss how this discovery alters our view of the way in which antibodies neutralize viral infection and what implications this may have for future research. pp. 803-809.
Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
- PMID:
- 22213041
- [PubMed - in process]
Mitochondrial connexin 43 impacts on respiratory complex I activity and mitochondrial oxygen consumption.
Source
Institut für Pathophysiologie, Universitätsklinikum Essen, Germany.
Abstract
Connexin 43 (Cx43) is present at the sarcolemma and the inner membrane of cardiomyocyte subsarcolemmal mitochondria (SSM). Lack or inhibition of mitochondrial Cx43 is associated with reduced mitochondrial potassium influx, which might affect mitochondrial respiration. Therefore, we analyzed the importance of mitochondrial Cx43 for oxygen consumption. Acute inhibition of Cx43 in rat left ventricular (LV) SSM by 18α glycyrrhetinic acid (GA) or Cx43 mimeticpeptides (Cx43-MP) reduced ADP-stimulated complex I respiration and ATP generation. Chronic reduction of Cx43 in conditional knockout mice (Cx43(Cre-ER(T)/fl) + 4-OHT, 5-10% of Cx43 protein compared to control Cx43(fl/fl) mitochondria) reduced ADP-stimulated complex I respiration of LV SSM to 47.8±2.4 nmol O(2) /min*mg protein (n=8) from 61.9±7.4 nmol O(2) /min*mg protein in Cx43(fl/fl) mitochondria (n=10, p<0.05), while complex II respiration remained unchanged. The LV complex I activities (% of citrate synthase activity) of Cx43(Cre-ER(T)/fl) +4-OHT mice (16.1±0.9%, n=9) were lower than in Cx43(fl/fl) mice (19.8±1.3%, n=8, p<0.05); complex II activities were similar between genotypes. Supporting the importance of Cx43 for respiration, in Cx43-overexpressing HL-1 cardiomyocytes complex I respiration was increased, whereas complex II respiration was unaffected. Taken together, mitochondrial Cx43 is required for optimal complex I activity and respiration and thus mitochondrial ATP-production. © 2011 The Authors Journal of Cellular and Molecular Medicine © 2011 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.
© 2011 The Authors Journal of Cellular and Molecular Medicine © 2011 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.
- PMID:
- 22212640
- [PubMed - as supplied by publisher]
Transgenic sweet potato expressing thionin from barley gives resistance to black rot disease caused by Ceratocystis fimbriata in leaves and storage roots.
Source
Biotechnology Laboratory, Toyota Central R&D Laboratories, Inc., Nagakute, Aichi, 480-1192, Japan, e1130@mosk.tytlabs.co.jp.
Abstract
Black rot of sweet potato caused by pathogenic fungus Ceratocystis fimbriata severely deteriorates both growth of plants and post-harvest storage. Antimicrobial peptides from various organisms have broad range activities of killing bacteria, mycobacteria, and fungi. Plant thionin peptide exhibited anti-fungal activity against C. fimbriata. A gene for barley α-hordothionin (αHT) was placed downstream of a strong constitutive promoter of E12Ω or the promoter of a sweet potato gene for β-amylase of storage roots, and introduced into sweet potato commercial cultivar Kokei No. 14. Transgenic E12Ω:αHT plants showed high-level expression of αHT mRNA in both leaves and storage roots. Transgenic β-Amy:αHT plants showed sucrose-inducible expression of αHT mRNA in leaves, in addition to expression in storage roots. Leaves of E12Ω:αHT plants exhibited reduced yellowing upon infection by C. fimbriata compared to leaves of non-transgenic Kokei No. 14, although the level of resistance was weaker than resistance cultivar Tamayutaka. Storage roots of both E12Ω:αHT and β-Amy:αHT plants exhibited reduced lesion areas around the site inoculated with C. fimbriata spores compared to Kokei No. 14, and some of the transgenic lines showed resistance level similar to Tamayutaka. Growth of plants and production of storage roots of these transgenic plants were not significantly different from non-transgenic plants. These results highlight the usefulness of transgenic sweet potato expressing antimicrobialpeptide to reduce damages of sweet potato from the black rot disease and to reduce the use of agricultural chemicals.
- PMID:
- 22212462
- [PubMed - as supplied by publisher]
Effect of aracnotoxin from Latrodectus mactans on bovine sperm function: modulatory action of bovine oviduct cells and their secretions.
Source
Center of Neurosciences and Peptides Biology (CEBIOR), BIOREN, Universidad de La Frontera, Temuco, Chile;; PhD in Sciences Applied Cellular and Molecular Biology, University of La Frontera, Temuco, Chile;
Abstract
Latrodectus mactans' aracnotoxin (Atx) induces changes in sperm function that could be used as a co-adjuvant in male contraceptive barrier methods. This effect includes the suppression of intracellular reactive oxygen species (ROS), an event necessary for capacitation, chemotaxis and acrosome reaction (AR). The sperm that are not trapped by the barrier method can reach the oviduct before fertilisation and be exposed to the secretions of the oviducts. This study evaluated the effect of bovine tubal explants (TU) and conditioned media (CM) from the ampullar and isthmal regions on spermatozoa exposed to Atx. Thawed bovine sperm were incubated with Atx, TU and CM from the ampullar and isthmal regions for 4 h and then DNA integrity, intracellular ROS and lysophosphatidylcholine-induced AR were determined. Spermatozoa exposed to Atx and co-incubated with TU and CM for 4 h produced an increase in sperm DNA damage, a decrease in ROS production and a decrease in %AR, compared with the control. A similar result was obtained from the co-incubation of spermatozoa with Atx. In conclusion, the effect of Atx is not modified by tubal cells or their secretions and this opens the door to future studies to evaluate the application of synthetic peptides obtained from Atx as a co-adjuvant of contraceptive barrier methods.
© 2011 Blackwell Verlag GmbH.
- PMID:
- 22211875
- [PubMed - as supplied by publisher]
Interleukin-17- and protease-activated receptor 2-mediated production of CXCL1 and CXCL8 modulated by cyclosporine A, vitamin D(3) and glucocorticoids in human keratinocytes.
Source
Department of Dermatology, Hyogo College of Medicine, Hyogo, Japan Department of Physiology and Pharmacology, and Department of Medicine, University of Calgary Faculty of Medicine, Calgary, Alberta, Canada.
Abstract
Protease-activated receptor 2 (PAR2) is a G protein-coupled receptor which mediates a variety of functions in the skin including cutaneous inflammation. SLIGKV-NH(2) , an agonist peptide for PAR2, enhanced the interleukin (IL)-17-induced production of two CXC chemokines, CXCL1 (GRO-α) and CXCL8 (IL-8), in normal human epidermal keratinocytes (NHEK) in a concentration-dependent manner. The enhanced production of those chemokines was suppressed by a PAR2-specific siRNA. The SLIGKV-NH(2) -induced production of both CXCL1 and CXCL8 was markedly reduced by cyclosporine A. The enhanced production of CXCL1 was suppressed by 1α, 24R-dihydroxyvitamin D(3) , an active form of vitamin D(3) , and weakly by glucocorticoids, dexamethasone and clobetasol propionate, whereas production of CXCL8 was not altered by any of those receptor agonists. In psoriatic skin, the thickened upper spinous layer of the epidermis was positive for PAR2 protein and the expression of the IL17A mRNA was increased. These results suggest that the IL-17-induced pro-inflammatory reaction is enhanced by the activation of PAR2 in keratinocytes, and that the effect of PAR2 is differentially modulated by cyclosporine A, the active form of vitamin D(3) and glucocorticoids.
© 2011 Japanese Dermatological Association.
- PMID:
- 22211698
- [PubMed - as supplied by publisher]
[What's new in dermatological research?].
Source
Université de Franche Comté, EA3181, IFR133 et Service de Dermatologie, CHU de Besançon, 2 place Saint-Jacques 25030 Besançon cedex, France.
Abstract
Dermatological research has been very active this year. Most of the numerous fields investigated involve the mechanisms of cutaneous regeneration and barrier function. A novel target of early ultraviolet-induced skin photodamage, the Syk kinase, has been recently identified. Synergistic relationship between telomere damage and cutaneous progerin production during cell senescence may also participate in the natural skin aging process. Interestingly, ultraviolet radiation induces an inhibitory effect on subcutaneous lipogenesis. Androgenetic alopecia or common baldness is not characterized by loss of hair follicle stem cells but by a defect in the conversion of hair follicle stem cells into active progenitor cells. It has been shown that the cornified envelope functions not only as a physicomechanical barrier, but also as both a biochemical line of antoxidant defense and an immunological line of defense. Like human papillomaviruses, Merckel cell polyomavirus belongs to the skin microbiome and different studies have demonstrated the protective role of epidermal resident microflora through the activation of innate immunity.Production of antimicrobial peptides and the activation of inflammasome and plasmacytoid dendritic cells are involved in the modulation of the cutaneous barrier function. Results from different studies suggest that IL-22 and IL-36 may be common mediators of both innate and adaptive immune responses. All these pathways interact not only to maintain cutaneous homeostasis and integrity (wound healing) but also to regulate autoinflammatory and autoimmune dermatoses (psoriasis, lupus, rosacea, atopic dermatitis, etc…). In addition, molecular mechanisms that regulate T helper type 2 differentiation and the retention at the site of inflammation of Th2 cells have been identified. New promising therapeutic targets for different chronic dermatosis are thus suggested. Mechanobiology and mechanotransduction are also emerging fields that investigate mechanical interactions between living cells and their environment and the conversion of mechanical cues into biochemical signals. Electronic second skin is now a current concept through bio-integrated epidermal electronics platforms used for different monitoring and stimulations of body functions.
Copyright © 2011 Elsevier Masson SAS. All rights reserved.
- PMID:
- 22202644
- [PubMed - in process]
Pathophysiology of rosacea: redness, telangiectasia, and rosacea.
Source
Clinique Dermatologique, Faculté de Médecine, Université de Strasbourg et Hôpitaux, Universitaires de Strasbourg, 1, place de l'Hôpital, 67091 Strasbourg, France.
Abstract
The pathophysiology of rosacea involves a large number of factors that are at times difficult to correlate. There is not a single physiopathological model. Nevertheless, today it seems to have been established that two essential factors are involved: vascular and inflammatory. The disease occurs in individuals with a predisposition, mainly a light phototype subjected to substantial variations in climate. On a background of primary vascular anomaly, external factors (climate, exposure to ultraviolet rays, cutaneous flora, etc.) contribute to the development of abnormal superficial blood vessels, with a low permeability. The edema that results undoubtedly favors the colonization and multiplication of Demodex folliculorum. This parasite creates inflammation, directly and indirectly, which is seen in the papules and pustules as well as granulomas. Inflammation from rosacea is also characterized by innate immune system anomalies, with an increase in the expression of epidermal proteases and production of pro-inflammatory cathelicidin peptides. In addition, facial hypersensitivity exists, even though the cutaneous barrier is not altered. Finally, rhinophyma remains poorly explained; the vascular abnormalities induce local production of transforming growth factor β1 (TGF-β1) capable of creating fibrosis and therefore cutaneous thickening.
Copyright © 2011 Elsevier Masson SAS. All rights reserved.
- PMID:
- 22183097
- [PubMed - in process]
Conserved microRNA miR-8 in fat body regulates innate immune homeostasis in Drosophila.
Source
School of Biological Sciences, Chung-Ang University, Seoul 156-756, Republic of Korea.
Abstract
Antimicrobial peptides (AMPs) constitute a major arm of the innate immune system across diverse organisms. In Drosophila, septic injury by microbial pathogens rapidly induces the production of the AMPs in fat body via well elucidated pathways such as Toll and IMD. However, several epithelial tissues were reported to locally express AMPs without septic injury via poorly characterized ways. Here, we report that microRNA miR-8 regulates the levels of AMPs basally expressed in Drosophila. The levels of AMPs such as Drosomycin and Diptericin are significantly increased in miR-8 null animals in non-pathogen stimulated conditions. Analysis of various larval tissues revealed that the increase of Drosomycin is fat body specific. Supporting this observation, re-introduction of miR-8 only in the fat body restored the altered AMP expression in miR-8 null flies. Although loss of miR-8 impedes PI3K in the fat body, inhibition of PI3K does not phenocopy the AMP expression of miR-8 null flies, indicating that miR-8 regulates AMP independently of PI3K. Together, our findings suggest a role of miR-8 in systemic immune homeostasis in generally non-pathogenic conditions in flies.
Copyright © 2011 Elsevier Ltd. All rights reserved.
Telmisartan acts through the modulation of ACE-2/ANG 1-7/mas receptor in rats with dilated cardiomyopathy induced by experimental autoimmune myocarditis.
Source
Department of Clinical Pharmacology, Faculty of Pharmaceutical Sciences, Niigata University of Pharmacy and Applied Life Sciences, Niigata, Japan.
Abstract
AIM:
Recent findings have suggested that a therapeutic approach to amplify or stimulate the angiotensin-converting enzyme-2 [ACE-2]-angiotensin 1-7 [ANG 1-7] mas axis could provide protection against the development of cardiovascular diseases. We investigated the cardioprotective effects of telmisartan in rats with dilated cardiomyopathy [DCM] after experimental autoimmune myocarditis [EAM].
MAIN METHODS:
DCM was elicited in Lewis rats by immunization with cardiac myosin, and twenty-eight days after immunization, the surviving Lewis rats were divided into two groups and treated with either telmisartan (10mg/kg/day) or vehicle.
KEY FINDINGS:
Telmisartan treatment effectively suppressed myocardial protein and mRNA expressions of inflammatory markers [CD68, iNOS, NF-kB, interleukin-1β, interferon-γ, monocyte chemotactic protein-1] in comparison to vehicle-treated rats. In contrast, myocardial protein levels of ACE-2 and ANG 1-7 mas receptor were upregulated in the telmisartan-treated group compared with vehicle-treated rats. Telmisartan treatment significantly reduced fibrosis and hypertrophy and their marker molecules [OPN, CTGF, TGF-β1 and collagens I and III and atrial natriuretic peptide and GATA-4, respectively] compared with those of vehicle-treated rats. In addition, telmisartan treatment significantly lowered the protein expressions of NADPH oxidase subunits p47phox, p67phox, and superoxide production when compared with vehicle-treated rats. Telmisartan treatment significantly decreased the expression levels of mitogen-activated protein kinase (MAPK) signaling molecules than with those of vehicle-treated rats. Also, telmisartan treatment significantly improved LV systolic and diastolic function.
SIGNIFICANCE:
These results indicate that telmisartan treatment significantly improved LV function and ameliorated the progression of cardiac remodeling through modulation of the ACE-2/ANG 1-7/Mas receptor axis in rats with DCM after EAM.
Copyright © 2011. Published by Elsevier Inc.
Peptide vaccination is superior to genetic vaccination using a recombineered bacteriophage λ subunit vaccine.
Source
Southern Alberta Cancer Research Institute, Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Calgary. Calgary, AB, Canada; Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
Abstract
Genetic immunization holds promise as a vaccination method, but has so far proven ineffective in large primate and human trials. Herein, we examined the relative merits of genetic immunization and peptide immunization using bacteriophage λ. Bacteriophage λ has proven effective in immune challenge models using both immunization methods, but there has never been a direct comparison of efficacy and of the quality of immune response. In the current study, this vector was produced using a combination of cis and trans phage display. When antibody titers were measured from immunized animals together with IL-2, IL-4 and IFNγ production from splenocytes in vitro, we found that proteins displayed on λ were superior at eliciting an immune response in comparison to genetic immunization with λ. We also found that the antibodies produced in response to immunization with λ displayed proteins bound more epitopes than those produced in response to genetic immunization. Finally, the general immune response to λ inoculation, whetherpeptide or genetic, was dominated by a Th1 response, as determined by IFNγ and IL-4 concentration, or by a higher concentration of IgG2a antibodies.
Copyright © 2011. Published by Elsevier Ltd.
Suppression of EAE and prevention of blood-brain barrier breakdown after vaccination with novel bifunctional peptide inhibitor.
Source
Department of Pharmaceutical Chemistry, University of Kansas, 2095 Constant Avenue, Lawrence, KS 66047, USA.
Abstract
The efficacy of bifunctional peptide inhibitor (BPI) in preventing blood-brain barrier (BBB) breakdown during onset of experimental autoimmune encephalomyelitis (EAE) and suppression of the disease was evaluated in mice. The mechanism that defines how BPI prevents the disease was investigated by measuring the in vitro cytokine production of splenocytes. Peptides were injected 5-11 days prior to induction of EAE, and the severity of the disease was monitored by a standard clinical scoring protocol and change in body weight. The BBB breakdown in diseased and treated mice was compared to that in normal control mice by determining deposition of gadolinium diethylenetriaminepentaacetate (Gd-DTPA) in the brain using magnetic resonance imaging (MRI). Mice treated with PLP-BPI showed no or low indication of EAE as well as normal increase in body weight. In contrast, mice treated with the control peptide or PBS showed a decrease in body weight and a high disease score. The diseased mice had high deposition of Gd-DTPA in the brain, indicating breakdown in the BBB. However, the deposition of Gd-DTPA in PLP-BPI-treated mice was similar to that in normal control mice. Thus, PLP-BPI can suppress EAE when administered as a peptide vaccine and maintain the integrity of the BBB.
Copyright © 2011 Elsevier Ltd. All rights reserved.
Altered regulation of nitric oxide and natriuretic peptide system in cisplatin-induced nephropathy.
Source
Department of Internal Medicine, Chonnam National University Medical School, Gwangju, Korea.
Abstract
Cisplatin is a chemotherapeutic agent used for treating solid tumors. However, nephrotoxicity is the dose-limiting factor in its clinical use. The present study was aimed to determine whether altered regulation of the local nitric oxide (NO) and natriuretic peptide (NP) systems is involved in the pathogenesis of cisplatin-induced nephropathy. Cisplatin (6mg/kg) was injected intraperitoneally into male Sprague-Dawley rats. The control group was not treated with cisplatin. Expression levels of nitric oxide synthase (NOS), nitrotyrosine, soluble guanylyl cyclase and neutral endopeptidase (NEP) in the kidneys were determined 4days after treatment by semiquantitative immunoblotting. mRNA expression of NPs and natriuretic peptide receptors (NPRs) was determined by real-time polymerase chain reaction. The activities of soluble and particulate guanylyl cyclase were determined by measuring the amount of cyclic 3',5'-guanosine monophosphate (cGMP) generated in responses to sodium nitroprusside and atrial natriuretic peptide (ANP), respectively. In the test rats, creatinine clearance was decreased, while sodium and water excretion were increased. The expression of inducible NOS (iNOS) and nitrotyrosine was increased in the cortex/outer stripe of outer medullar and inner medullar, while that of endothelial and neuronal NOS was decreased in the inner medullar. Excretion of NO metabolites was increased in these rats. The catalytic activity of soluble guanyly cyclase was blunted in the papilla after cisplatin was administered. The mRNA expression of ANP, brain natriuretic peptide, and C-type natriuretic peptide was increased, while that of NPR-A and NPR-C were decreased in the test rats. The catalytic activity of soluble and particulate guanylyl cyclase in the papilla was blunted after cisplatin was administered. In conclusion, increasedproduction of NO by iNOS may contribute to cytotoxic injury, resulting in cisplatin-induced nephropathy, while the up-regulation of renal natriuretic peptide synthesis together with the down-regulation of NEP and NPR-C may contribute to the natriuresis and diuresis seen in cisplatin-induced nephropathy.
Copyright © 2011. Published by Elsevier B.V.
NO-dependent protective effect of VEGF against excitotoxicity on layer VI of the developing cerebral cortex.
Source
EA "NeoVasc" 4309, Laboratory of Microvascular Endothelium and Neonate Brain Lesions, Rouen Institute for Biomedical Research, European Institute for Peptide Research (IFR 23), University of Rouen, Rouen, France.
Abstract
In industrialized countries, cerebral palsy affects 2.5‰ of preterm and term infants. At a neurochemical level, the massive release of glutamate constitutes a major process leading to excitotoxicity and neonatal brain lesions. Previous studies, conducted in the laboratory, revealed that, in (δ/δ)VEGF(A) transgenic mice, glutamate-induced brain lesions are exacerbated suggesting that VEGF(A) could play a protective action against excitotoxicity. Using a model of cultured cortical brain slices, the aim of the study was to characterize the central effects of VEGF against glutamate-induced excitotoxicity in neonates. Exposure of brain slices to glutamate induced a strong increase of necrotic cell death in the deep cortical layer VI and a decrease of apoptotic death in superficial layers II-IV. When administered alone, a 6-h treatment with VEGF(A) had no effect on both apoptotic and necrotic deaths. In contrast, VEGF(A) abolished the glutamate-induced necrosis observed in layer VI. While MEK and PI3-K inhibitors had no effect on the protective action of VEGF(A), L-NAME, a pan inhibitor of NOS, abrogated the effect of VEGF(A) and exacerbated the excitotoxic action of glutamate. Calcimetry experiments performed on brain slices revealed that VEGF(A) reduced the massive calcium influx induced by glutamate in layer VI and this effect was blocked by L-NAME. Neuroprotective effect of VEGF(A) was also blocked by LNIO and NPLA, two inhibitors of constitutive NOS, while AGH, an iNOS inhibitor, had no effect. Nitrite measurements, electron paramagnetic resonance spectroscopy and immunohistochemistry indicated that glutamate was a potent inducer of NO production via activation of nNOS in the cortical layer VI. In vivo administration of nNOS siRNA promoted excitotoxicity and mimicked the effects of L-NAME, LNIO and NPLA. A short-term glutamate treatment increased nNOS Ser1412 phosphorylation, while a long-term exposure inhibited nNOS/NR2B protein-protein interactions. Altogether, these findings indicate that, in deep cortical layers of mice neonates, glutamate stimulates nNOS activity. Contrasting with mature brain, NO production induced by high concentrations of glutamate is neuroprotective and is required for the anti-necrotic effect of VEGF(A).
Copyright © 2011. Published by Elsevier Inc.
Trans fatty acids enhance amyloidogenic processing of the Alzheimer amyloid precursor protein (APP).
Source
Deutsches Institut für DemenzPrävention (DIDP), Neurodegeneration and Neurobiology, 66421 Homburg, Germany.
Abstract
Hydrogenation of oils and diary products of ruminant animals leads to an increasing amount of trans fatty acids in the human diet. Trans fatty acids are incorporated in several lipids and accumulate in the membrane of cells. Here we systematically investigate whether the regulated intramembrane proteolysis of the amyloid precursor protein (APP) is affected by trans fatty acids compared to the cis conformation. Our experiments clearly show that trans fatty acids compared to cis fatty acids increase amyloidogenic and decrease nonamyloidogenic processing of APP, resulting in an increased production of amyloid beta (Aβ) peptides, main components of senile plaques, which are a characteristic neuropathological hallmark for Alzheimer's disease (AD). Moreover, our results show that oligomerization and aggregation of Aβ are increased by trans fatty acids. The mechanisms identified by this in vitro study suggest that the intake of trans fatty acids potentially increases the AD risk or causes an earlier onset of the disease.
Copyright © 2012 Elsevier Inc. All rights reserved.
Oral enzyme therapy for celiac sprue.
Source
Division of Biology, California Institute of Technology, Pasadena, California, USA.
Abstract
Celiac sprue is an inflammatory disease of the small intestine caused by dietary gluten and treated by adherence to a life-long gluten-free diet. The recent identification of immunodominant gluten peptides, the discovery of their cogent properties, and the elucidation of the mechanisms by which they engender immunopathology in genetically susceptible individuals have advanced our understanding of the molecular pathogenesis of this complex disease, enabling the rational design of new therapeutic strategies. The most clinically advanced of these is oral enzyme therapy, in which enzymes capable of proteolyzing gluten (i.e., glutenases) are delivered to the alimentary tract of a celiac sprue patient to detoxify ingested gluten in situ. In this chapter, we discuss the key challenges for discovery and preclinical development of oral enzyme therapies for celiac sprue. Methods for lead identification, assay development, gram-scaleproduction and formulation, and lead optimization for next-generation proteases are described and critically assessed.
Copyright © 2012 Elsevier Inc. All rights reserved.
- PMID:
- 22208988
- [PubMed - in process]
Vasoactive Intestinal Peptide Enhances TNF-α-Induced IL-6 and IL-8 Synthesis in Human Proximal Renal Tubular Epithelial Cells by NF-κB-Dependent Mechanism.
Source
Division of Nephrology, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China.
Abstract
Vasoactive intestinal polypeptide (VIP) is a 28-amino acid neuropeptide with vasodilator, bronchodilator, and anti-inflammatory effects. But little is known about its pro-inflammatory effects. We investigated the effect of VIP on the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8), two pro-inflammatory cytokines, in TNF-α-activated proximal renal tubular epithelial cell line (HK-2 cells). Cultured HK-2 cells were treated with TNF-α in the presence or absence of VIP with a dose range from 1 to 100 nM, followed by analysis of pro-inflammatory cytokines (IL-6 and IL-8) induction and their signal events including activation of the NF-κB pathway. We report here that tumor necrosis factor-α (TNF-α) increased IL-6 and IL-8 production, and that these effects were potentiated by VIP at 10 nM in HK-2 cells. However, VIP at 1 and 100 nM did not display this function. Consistent with these observations, we were able to show that VIP at 10 nM upregulated TNF-α-induced phosphorylation of IκB-α, leading to IκB-α degradation and the subsequent nuclear translocation of NF-κB. Furthermore, VIP-enhanced activation of NF-κB transcription activity was demonstrated using a NF-κB reporter construct upon transient transfection into HK-2 cells. These results strongly suggest that VIP synergistically enhances TNF-α-stimulated IL-6 and IL-8 synthesis via activating the NF-κB pathway in HK-2 cells.
Mechanisms by which pesticides affect insect immunity.
Source
USDA-ARS, Pollinating Insects Research Unit, Dept. Biology UMC 5310, Utah State University, Logan, UT 84322-5310, USA.
Abstract
The current state of knowledge regarding the effect of pesticides on insect immunity is reviewed here. A basic understanding of these interactions is needed for several reasons, including to improve methods for controlling pest insects in agricultural settings, for controlling insect vectors of human diseases, and for reducing mortality in beneficial insects. Bees are particularly vulnerable to sublethal pesticide exposures because they gather nectar and pollen, concentrating environmental toxins in their nests in the process. Pesticides do have effects on immunity. Organophosphates and some botanicals have been found to impact hemocyte number, differentiation, and thus affect phagocytosis. The phenoloxidase cascade and malanization have also been shown to be affected by several insecticides. Many synthetic insecticides increase oxidative stress, and this could have severe impacts on theproduction of some antimicrobial peptides in insects, but research is needed to determine the actual effects. Pesticides can also affect grooming behaviors, rendering insects more susceptible to disease. Despite laboratory data documenting pesticide/pathogen interactions, little field data is available at the population level.
Copyright © 2011. Published by Elsevier Inc.
Fibrillar Amyloid-β1-42 Modifies Actin Organization Affecting the Cofilin Phosphorylation State: A Role for Rac1/cdc42 Effector Proteins and the Slingshot Phosphatase.
Source
Laboratory of Cellular and Molecular Neurosciences, University of Chile and International Center for Biomedicine (ICC), Santiago, Chile.
Abstract
The neuronal cytoskeleton regulates numerous processes that occur in normal homeostasis. Under pathological conditions such as those of Alzheimer's disease (AD), major alterations in cytoskeleton organization have been observed and changes in both microtubules and actin filaments have been reported. Many neurodegenerative consequences of AD are linked to the production and accumulation of amyloid peptides (Aβ) and their oligomers, produced from the internal cleavage of the amyloid-β protein precursor. We previously reported that fibrillar Aβ1-42 (fAβ) treatment of hippocampal neurons induced an increase in Rac1 and Cdc42 activities linking fAβ effects with changes in actin dynamics. Here we show fAβ-induces increased activity of PAK1 and cyclin-dependent kinase 5, and that p21-activated kinase (PAK1) activation targets the LIMK1-cofilin signaling pathway. Increased cofilin dephosphorylation under conditions of enhanced LIM-Kinase 1 (LIMK1) activity suggests that fAβ co-stimulates bifurcating pathways impacting cofilin phosphorylation. Overexpression of slingshot (SSH) prevents the augment of F-actin induced by fAβ after 24 h, suggesting that fAβ-induced changes in actin assembly involve both LIMK1 and SSH. These results suggest that fAb may alter the PAK1/LIMK1/cofilin axis and therefore actin organization in AD.
The multifaceted activities of Mammalian defensins.
Source
Departments of Microbiology and Internal Medicine, Center for Microbial Interface Biology, Ohio State University, Columbus, OH. seveau.1@osu.edu.
Abstract
Defensins are an important family of cationic and cysteine-rich host defense peptides that are widely distributed in plants, fungi, and animals. In mammals, defensins exert potent antimicrobial and immunomodulatory activities linking the innate and adaptive immune responses. These peptides play critical roles in health and disease as defects in theirproduction are associated with abnormal host responses to infection, chronic inflammatory diseases, and cancer. There is much interest in elucidating the structure-function relation and modes of action of the defensins to better understand how these peptides kill microbes and regulate the host immune responses. Such knowledge is expected to help in the design of novel defensin-based therapeutics. This review focuses on the multifaceted antimicrobial and immunomodulatory activities of human and murine defensins.
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