Innovative superparamagnetic iron-oxide nanoparticles coated with silica and conjugated with linoleic acid: Effect on tumor cell growth and viability

One of the goals for the development of more effective cancer therapies with reduced toxic side effects is the optimization of innovative treatments to selectively kill tumor cells. The use of nanovectors loaded with targeted therapeutic payloads is one of the most investigated strategies. In this paper superparamagnetic iron oxide nanoparticles (SPIONs) coated by a silica shell or uncoated, were functionalized with single-layer and bi-layer conjugated linoleic acid (CLA). Silica was used to protect the magnetic core from oxidation, improve the stability of SPIONs and tailor their surface reactivity. CLA was used as novel grafting biomolecule for its anti-tumor activity and to improve particle dispersibility. Mouse breast cancer 4T1 cells were treated with these different SPIONs. SPIONs functionalized with the highest quantity of CLA and coated with silica shell were the most dispersed. Cell viability was reduced by SPIONs functionalized with CLA in comparison with cells which were untreated or treated with SPIONs without CLA. As regards the types of SPIONs functionalized with CLA, the lowest viability was observed in cells treated with uncoated SPIONs with the highest quantity of CLA. In conclusion, the silica shell free SPIONs functionalized with the highest amount of CLA can be suggested as therapeutic carriers because they have the best dispersion and ability to decrease 4T1 cell viability

The effect of elevated atmospheric CO2 on the N2-fixing symbiosis of Trifolium repens L. and Rhizobium leguminosarum biovar trifolii

CO2 ist eines der wichtigsten Treibhausgase, das das Klima und terrestrische Ökosysteme nachhaltig beeinflusst. Bis heute ist wenig über den Einfluss von erhöhten atmosphärischen CO2-Kontentrationen auf symbiotische Interaktionen in der Rhizosphäre, insbesondere auf die N2-fixierende Symbiose von Trifolium repens und Rhizobium leguminosarum biovar trifolii bekannt. Erste Ergebnisse eines 10-jährigen Freiland-CO2-Begasungsexperiments (Swiss FACE) zeigten, dass sich nach drei Jahren kontinuierlicher CO2-Begasung die genetische Zusammensetzung der Rhizobien-Population in den Wurzelknöllchen von T. repens unter erhöhter CO2-Konzentration verändert hatte. Im ersten Abschnitt dieser Arbeit sollte der Frage nachgegangen werden, ob nach 10-jähriger CO2-Begasung im Swiss FACE weiterhin genetisch unterschiedliche Rhizobien-Stämme in den Wurzelknöllchen des Weißklees nachgewiesen werden können oder ob eine Anpassung der Knöllchenbakterien an erhöhte CO2-Konzentrationen erfolgt ist. Darüber hinaus sollte diese Arbeit klären, ob erhöhte atmosphärische CO2-Konzentrationen in Abhängigkeit der N-Versorgung bei T. repens zu quantitativen und qualitativen Veränderungen in der Wurzelexsudation führen, insbesondere hinsichtlich der Abgabe von Substanzen mit Signalfunktion während der Nodulation. Im Sommer 2002 wurden mit CO2 begaste und unbegaste Versuchsflächen des FACE beprobt und Rhizobien aus Wurzelknöllchen der geernteten Weißkleepflanzen isoliert. Die Untersuchung von R. l. bv. trifolii mittels BOX-PCR DNA Fingerprinting ergab, dass nach 10-jähriger CO2-Begasung im Swiss FACE keine Veränderung in der genetischen Zusammensetzung der Rhizobien mehr festgestellt werden konnten. Die genetische Verschiebung innerhalb der Rhizobien-Population war somit nur vorübergehend nachweisbar. Dies ist höchstwahrscheinlich auf die Tatsache zurückzuführen, dass sich im Laufe des Freiland-CO2-Experiments ein neues C/N-Gleichgewicht im Graslandökosystem eingestellt hat. Zu Beginn des FACE-Experiments wurde eine Erhöhung des C/N-Verhältnisses im Boden detektiert, das durch die gesteigerte symbiotische N2-Fixierung und den daraus resultierenden vermehrten N-Eintrag in das Ökosystem allmählich ausgeglichen werden konnte. Diese Stabilisierung des Graslandökosystems hatte höchstwahrscheinlich auch einen Rückgang der indirekten CO2-Beinflussung auf die Mikroorganismen zur Folge, so dass nach 10 Jahren im Swiss FACE keine genetische Diversität der Rhizobien-Population mehr nachzuweisen war. Um die Wirkung erhöhter atmosphärischer CO2-Konzentrationen auf die Wurzelexsudation von Signalsubstanzen zu untersuchen, wurden Weißkleepflanzen in zwei unabhängigen Klimakammerexperimenten unter sterilen und unsterilen Versuchsbedingungen bei ambienter und erhöhter CO2-Konzentration (400 und 800 ppm) und unterschiedlicher Stickstoffdüngung hydroponisch kultiviert. Die Gewinnung der Wurzelexsudate erfolgte über einen Zeitraum von sieben Stunden an drei bzw. vier unterschiedlichen Terminen. Die phenolischen Bestandteile der Wurzelexsudate wurden durch eine Festphasenextraktion extrahiert und anschließend mittels HPLC und LC-MS analysiert. Zusätzlich wurden die isolierten Fraktionen, auf ihre Fähigkeit die Nodulationsgene von R. l. bv. trifolii zu aktivieren, anhand eines nod-Gen-Induktionstests überprüft. Die Erhöhung der CO2-Konzentration führte in beiden Versuchsansätzen zu einem gesteigerten Spross- und Wurzelwachstum, aber nicht zu einer Veränderung des C/N-Verhältnisses in der Wurzel. In den Wurzelexsudaten von T. repens konnten, neben der bekannten Signalsubstanz 7,4?-Dihydroxyflavon, neue, bisher in der Literatur nicht beschriebene phenolische Substanzen nachgewiesen werden. Die Fraktionen wurden anhand ihrer Polaritätseigenschaften, Absorptionsverhalten und Molekulargewichte als aglycosidische Flavone identifiziert, die, bis auf eine Fraktion (Fraktion 2) des unsterilen Experiments, alle die Fähigkeit zur nod-Gen-Induktion besaßen. CO2 beeinflusste die Exsudation der Signalsubstanzen quantitativ, nicht aber qualitativ. Die erhöhte Exsudation, insbesondere des 7,4?-Dihydroxyflavons, konnte einerseits auf die höhere Wurzelbiomasse unter erhöhtem CO2 zurückgeführt werden, andererseits aber auch auf eine höhere Abgaberate in Bezug auf die Wurzelfrischmasse. Diese CO2-Reaktion der Weißkleepflanzen, sowohl hinsichtlich der Biomasseproduktion als auch der Wurzelexsudation, war deutlich abhängig von der N-Versorgung und signifikant nur unter sterilen Versuchsbedingungen ausgeprägt. Im Einzelfall überstieg der N-Einfluss sogar den CO2-Effekt: mit zunehmendem N-Mangel exsudierten die Weißkleepflanzen unter sterilen Bedingungen verstärkt die nod-Gen-aktivierende Exsudat-Fraktion C. Diese, als Hydroxyflavon identifizierte Fraktion besitzt folglich bei N-Limitierung eine wichtige Signalfunktion. Neben der CO2-Konzentration und der N-Versorgung beeinflusste das Pflanzenalter maßgeblich die Wurzelexsudation von T. repens, was sich vor allem im unsterilen Versuchsansatz in einem Rückgang der Abgabe von Signalsubstanzen bei älteren Weißkleepflanzen, aber auch in der Qualität der exsudierten Phenole äußerte. Die anfänglich im Swiss FACE beobachtete Veränderung in der genetischen Zusammensetzung von R. l. bv. trifolii war mit sehr großer Wahrscheinlichkeit die Folge einer gesteigerten Exsudation phenolischer Signalsubstanzen von T. repens unter erhöhter atmosphärischer CO2-Konzentration.CO2 is one of the main greenhouse gases strongly influencing the climate and the terrestrial ecosystem. Up to know little is known about the impact of elevated atmospheric CO2 on symbiotic interactions in the rhizosphere, especially on the N2-fixing symbiosis between Trifolium repens and Rhizobium leguminosarum biovar trifolii. First results of a ten-year Free-Air CO2 enrichment experiment (Swiss FACE) showed that after three years of CO2 fumigation the genetic composition of the Rhizobium population in the root nodules of T. repens had changed. The first part of this thesis set out to clarify the question whether a genetic difference in the Rhizobium population of root nodules of white clover could still be detected after ten years of CO2 fumigation or if an adaptation of the nodule bacteria to elevated CO2 concentrations had occurred. Furthermore the thesis addressed the question whether elevated atmospheric CO2 leads to quantitative and qualitative changes in the root exudation of T. repens particularly with regard to exudation of signal substances during the nodulation process. In summer 2002 white clover plants were collected from plots fumigated with CO2 and control plots of the Swiss FACE. Rhizobium strains were isolated from the clover root nodules and used for rep-PCR DNA fingerprinting. Results clearly showed that after ten years of CO2 enrichment changes in the genetic composition of the R. l. bv. trifolii could no longer be observed. Thus, CO2-induced changes in the population structure of rhizobia seemed to be transient. This can be traced back to the possibility that over the experimental period a new C/N equilibrium in the grassland ecosystem has been established. At the beginning of the FACE experiment an increase in the C/N ratio of the soil was detected, which could be balanced in the course of time through enhanced symbiotic N2 fixation and consequently a higher N input into the ecosystem. The observed stabilisation of the grassland ecosystem most likely caused a reduction of the indirect CO2 impact on the microorganisms. This might explain why a change in the genetic composition of Rhizobium strains was not longer detected after ten years in the Swiss FACE. To investigate an influence of elevated atmospheric CO2 concentration on the release of signalling compounds clover plants were cultivated hydroponically in two independent climate chamber trials under axenic and non-axenic conditions at ambient and elevated CO2 concentrations (400 and 800 ppm) and different levels of N supply. Root exudates were collected over a period of seven hours and at three and four different plant ages, respectively. Phenolic compounds were extracted by solid-phase extraction and afterwards analysed with HPLC and LC-MS. Additionally, the isolated fractions were tested for their ability to induce the nodulation genes of R. l. bv. trifolii using a nod-gene induction test. The CO2 enrichment caused an increase in shoot and root growth in both experimental setups, but did not provoke a change in the C/N ratio of the roots. Besides the known signal compound 7,4?-dihydroxyflavone new phenolic substances could be detected, which have not yet been described in literature. The fractions were identified by their polarity, light absorption and molecular weight as aglyca and flavones. All of these had the ability for nod-gene induction except one fraction (fraction 2). CO2 influenced the exudation of signalling compounds quantitatively but not qualitatively. The enhanced exudation, especially of 7,4?-dihydroxyflavone, could be attributed to the higher root mass under elevated CO2 but also to a higher release rate on a root fresh weight basis. The CO2 reaction of the clover plants, for the biomass production as well as for the root exudation, was clearly dependent on the N supply and only significant under axenic conditions. In individual cases the N impact was more pronounced than the CO2 effect: with increasing N demand axenic clover plants enhanced the exudation of the nod-gene inducing fraction C. It is concluded that this fraction, identified as a hydroxyflavone, has therefore an important signal function under N limitation. Besides the CO2 concentration and N supply, root exudation by T. repens was considerably influenced by the plant age, which caused a reduction of the signal exudation in older plants and qualitative changes of the released phenols, especially under non-axenic conditions. The present study suggests that the genetic shift of R. l. bv. trifolii detected at the beginning of the Swiss FACE experiment was most likely a consequence of the enhanced exudation of phenolic signal compounds of T. repens under elevated atmospheric CO2 concentrations

Pluronic-silica (PluS) nanoparticles doped with multiple dyes featuring complete energy transfer

We report here the design of two sets Of multifltiorophoric silica nanoparticles, observing unprecedented efficiencies in the energy-transfer processes among the doping dyes. These nanomaterials show a very high overall sensitization, allowing under a single wavelength excitation to obtain many different colors (one per nanoparticle) in emission with negligible crosstalk. Moreover, each particle can present very large and tunable pseudo-Stokes shifts (up to 435 nm), a very high brightness even exciting the bluest donor, and a negligible residual emission intensity from all donor dyes. All these features, combined with colloidal stability and synthetic method reliability, make these multicomponent nanoparticles very promising for multiplex analysis and for all the diagnostic techniques requiring high sensitivity associated with a large Stokes shift

Anti‐dopamine D2 receptor antibodies in chronic tic disorders

Aim: To investigate the association between circulating anti-dopamine D2 receptor (D2R) autoantibodies and the exacerbation of tics in children with chronic tic disorders (CTDs). Method: One hundred and thirty-seven children with CTDs (108 males, 29 females; mean age [SD] 10y 0mo [2y 7mo], range 4-16y) were recruited over 18 months. Patients were assessed at baseline, at tic exacerbation, and at 2 months after exacerbation. Serum anti-D2R antibodies were evaluated using a cell-based assay and blinded immunofluorescence microscopy scoring was performed by two raters. The association between visit type and presence of anti-D2R antibodies was measured with McNemar's test and repeated-measure logistic regression models, adjusting for potential demographic and clinical confounders. Results: At exacerbation, 11 (8%) participants became anti-D2R-positive ('early peri-exacerbation seroconverters'), and nine (6.6%) became anti-D2R-positive at post-exacerbation ('late peri-exacerbation seroconverters'). The anti-D2R antibodies were significantly associated with exacerbations when compared to baseline (McNemar's odds ratio=11, p=0.003) and conditional logistic regression confirmed this association (Z=3.49, p<0.001) after adjustment for demographic and clinical data and use of psychotropic drugs. Interpretation: There is a potential association between immune mechanisms and the severity course of tics in adolescents with CTDs

Antibodies to neuronal surface proteins in Tourette Syndrome: Lack of evidence in a European paediatric cohort

In Tourette Syndrome (TS) a role for autoantibodies directed against neuronal proteins has long been suspected, but so far results are still inconsistent. The aim of this study was to look for antibodies to specific or undefined neuronal proteins that could be involved in the aetiology of the disease. Sera from children with Tourette Syndrome or another chronic tic disorder (TS/TD), collected as part of the longitudinal European Multicenter Tics in Children Study, were investigated. Participants included 30 siblings of patients with TS/TD prior to developing tics (preclinical stage) and the same children after the first tic onset (onset), and 158 patients in the chronic phase undergoing an acute relapse (exacerbation). Presence of antibodies binding to rodent brain tissue was assessed by immunohistology on rat brain sections and by immunofluorescent staining of live hippocampal neurons. Live cell-based assays were used to screen for antibodies to NMDAR, CASPR2, LGI1, AMPAR and GABA(A)R. Immunohistology indicated evidence of antibodies reactive with brain tissue, binding mainly to the hippocampus, the basal ganglia or the cerebellum in 26/218 (12%), with 8% of the preclinical or onset sera binding to the dentate gyrus/CA3 region or cerebellum. Only two individuals (one pre-clinical, one chronic) had antibodies binding the NMDAR and the binding was only weakly positive. No other specific antibodies were detected. Despite some immunoreactivity towards neuronal antigens on brain tissue, this was not mirrored by antibodies binding to live neurons, suggesting the presence of non-specific antibodies or those that bind non-pathogenic intracellular epitopes. NMDAR or the other neuronal surface antibodies tested were very infrequent in these patients. The evidence for pathogenic antibodies that could be causative of TS is weak