Regulation of the vitellogenin gene B1 promoter after transfer into hepatocytes in primary cultures.

The estrogen-dependent and tissue-specific regulation of the Xenopus laevis vitellogenin gene B1 promoter has been studied by lipid-mediated DNA transfer into Xenopus hepatocytes in primary culture. Hepatocytes achieve an efficient hormonal control of this promoter through a functional interaction between the estrogen responsive elements and a promoter proximal region upstream of the TATA box, which is characterized by a high density of binding sites for the transcription factors CTF/NF-1, C/EBP and HNF3. DNA accessibility to restriction enzymes within the chromosomal copy of the vitellogenin gene B1 promoter shows that the estrogen responsive unit and the promoter proximal region are sensitive to digestion in uninduced and estrogen-induced hepatocytes but not in erythrocyte nuclei. Together, these findings support the notion that chromatin configuration as well as the interplay of promoter elements mediate proper hormone-dependent and tissue-specific expression of the B1 vitellogenin gene.

Glial hyaluronate-binding protein expression in aggregating brain cell cultures.

We analyzed the expression of glial hyaluronate-binding protein (GHAP), an integral component of the extracellular matrix, in aggregating brain cell cultures of fetal rat telencephalon using immunofluorescence. GHAP immunoreactivity appeared after 1 week in culture, simultaneous with the first deposits of myelin basic protein, and showed a development-dependent increase. Comparison of glia-enriched and neuron-enriched cultures showed that only glial cells express GHAP. Three peptide growth factors, epidermal growth factor, fibroblast growth factor and platelet-derived growth factor, which are known to stimulate the differentiation of glial cells, modulated the deposit of GHAP immunoreactivity. The 3-dimensional structure of aggregate cultures promoted GHAP deposition, suggesting that cell-cell interactions are required for extracellular matrix formation. Furthermore GHAP production seemed to depend on the developmental stage of the glial cells.

The lipid composition of rat brain aggregating cell cultures during development.

The lipid and fatty acid composition of rat brain was studied during its development both in vivo and in an aggregating cell culture system. Although the amount of lipid present in the cultures was very low, the increase in glycolipid content corresponded closely to the period of intense myelin formation. Very long chain fatty acids (hydroxylated and unsubstituted) were present in 41-day cultures. In comparison to the in vivo situation, myelination was delayed in vitro and, after 40 days in culture, cholesterol esters were 5-fold higher than in vivo, indicating that demyelination was occurring.

Ochratoxin A at nanomolar concentration perturbs the homeostasis of neural stem cells in highly differentiated but not in immature three-dimensional brain cell cultures.

Ochratoxin A (OTA), a fungal contaminant of basic food commodities, is known to be highly cytotoxic, but the pathways underlying adverse effects at subcytotoxic concentrations remain to be elucidated. Recent reports indicate that OTA affects cell cycle regulation. Therefore, 3D brain cell cultures were used to study OTA effects on mitotically active neural stem/progenitor cells, comparing highly differentiated cultures with their immature counterparts. Changes in the rate of DNA synthesis were related to early changes in the mRNA expression of neural stem/progenitor cell markers. OTA at 10nM, a concentration below the cytotoxic level, was ineffective in immature cultures, whereas in mature cultures it significantly decreased the rate of DNA synthesis together with the mRNA expression of key transcriptional regulators such as Sox2, Mash1, Hes5, and Gli1; the cell cycle activator cyclin D2; the phenotypic markers nestin, doublecortin, and PDGFRα. These effects were largely prevented by Sonic hedgehog (Shh) peptide (500ngml(-1)) administration, indicating that OTA impaired the Shh pathway and the Sox2 regulatory transcription factor critical for stem cell self-renewal. Similar adverse effects of OTA in vivo might perturb the regulation of stem cell proliferation in the adult brain and in other organs exhibiting homeostatic and/or regenerative cell proliferation.

Enrichment cultures should be performed in the detection of bacterial oral human pathogens in DUWLs

Water delivered by dental units during routine dental practice is densely contaminated by bacteria. The aim of this study was to determine actual isolation of the microorganisms sprayed from Dental Unit Water Lines (DUWLs) when enrichment cultures are performed and to compare frequencies with those obtained without enrichment cultures. Moreover, the antimicrobial susceptibilities of the microorganisms isolated were also studied. Water samples were collected from one hundred dental equipments in use at Dental Hospital of our University in order to evaluate the presence/absence of microorganisms and to perform their presumptive identification. Aliquots from all of the samples were inoculated in eight different media including both enrichment and selective media. Minimal inhibitory concentrations (MIC) were determined by the broth dilution method. The results herein reported demonstrate that most of the DUWLs were colonized by bacteria from human oral cavity; when enrichment procedures were applied the percentage of DUWLs with detectable human bacteria was one hundred percent. The results showed that in order to evaluate the actual risk of infections spread by DUWLs the inclusion of a step of pre-enrichment should be performed. The need for devices preventing bacterial contamination of DUWLs is a goal to be achieved in the near future that would contribute to maintain safety in dental medical assistance

Cultures en contacte a Catalunya

En el actual escenario mundial, interconectado en el circuito de la mundialización cultural, con poblaciones cada vez más heterogéneas a causa de los desplazamientos internos y extracomunitarios, el contacto de civilizaciones provoca tensiones y efectos múltiples y conectados en cadena en el ámbito económico, social, político y cultural. En este sentido, la cuestión que centra los debates es la problemática resultante de la vida, en común o en paralelo, en un mismo territorio, de las culturas que historicamente están ya asentadas con las que van llegando, que son (muy) diferentes. En nuestro estado multinacional, Cataluña tiene unas condiciones óptimas para ser observada como un paradigma, porque forma parte de este circuito mundializado; porque es un punto de llegada de una parte importante de la actual inmigración comunitaria y extracomunitaria; porque, además, tiene ya como bagaje una experiencia, no tan lejana, de recepción de emigrantes culturalmente distantes, de la que se desprenden tendencias interesantes para orientar el presente y consolidar en el futuro el escenario multicultural que ya se empieza a vertebrar.

Cultures familiars, mitjans digitals i joves. Restriccions explícites i implícites en la reproducció de l'avantatge generalitzat

La irrupció dels mitjans digitals està generant en les famílies amb adolescents noves formes d'estar junts i de relacionar-se amb l'exterior, que són viscuts sovint amb ansietat per part dels pares. Per estudiar les normes que tant els pares com els seus fills estan generant per regular-les, i com aquestes regulacions es relacionen amb la reproducció de l'avantatge i el desavantatge generalitzat, es proposa superar l'orientació restringida dels riscos i oportunitats i fixar-se en canvi en la manera comles cultures i identitats familiars generen equilibris particulars en les tensions normatives del treball, el consum i l'autenticitat. Mitjançant una recerca empírica qualitativa amb 23 famílies de dos centres d'educació secundària de l'àrea metropolitana de Barcelona, es constata que enlloc de buscar com normes o estils parentals concrets es relacionen amb diferents posicions socials, és preferible analitzar com les diferents cultures i identitats familiars regulen l'adquisició d'autonomia per part dels fills. Aspectes com la familiaritat i reflexivitat vers la tecnologia, l'establiment de rutines i horaris familiars o els contextos relacionals esdevenen així claus per entendre la reproducció de l'avantatge generalitzat. Aquests elements permeten contextualitzar dinàmicament la complexa combinació de les regulacions implícites i explícites en el procés de maduració dels fills.

Identification of neuronal network properties from the spectral analysis of calcium imaging signals in neuronal cultures

Neuronal networks in vitro are prominent systems to study the development of connections in living neuronal networks and the interplay between connectivity, activity and function. These cultured networks show a rich spontaneous activity that evolves concurrently with the connectivity of the underlying network. In this work we monitor the development of neuronal cultures, and record their activity using calcium fluorescence imaging. We use spectral analysis to characterize global dynamical and structural traits of the neuronal cultures. We first observe that the power spectrum can be used as a signature of the state of the network, for instance when inhibition is active or silent, as well as a measure of the network's connectivity strength. Second, the power spectrum identifies prominent developmental changes in the network such as GABAA switch. And third, the analysis of the spatial distribution of the spectral density, in experiments with a controlled disintegration of the network through CNQX, an AMPA-glutamate receptor antagonist in excitatory neurons, reveals the existence of communities of strongly connected, highly active neurons that display synchronous oscillations. Our work illustrates the interest of spectral analysis for the study of in vitro networks, and its potential use as a network-state indicator, for instance to compare healthy and diseased neuronal networks.

The relationships of organizational cultures, trust and innovativeness

Sekä organisaatiokulttuuria, luottamusta että innovatiivisuutta on tutkittu paljon, mutta toistaiseksi nämä käsitteet yhdistävää kokonaisvaltaista tutkimusta ei juuri ole tehty tai ainakaan raportoitu tieteellisissä julkaisuissa. Tätä tutkimus käsitteli organisaatiokulttuurin, luottamuksen ja innovatiivisuuden suhteita erilaisissa organisaatiokulttuureissa. Tutkimuksen tavoitteena oli tutkia organisaatiokulttuurin vaikutusta luottamukseen (sekä kompetenssiin, hyväntahtoisuuteen että rehellisyyteen perustuvaan lateraaliin, vertikaaliin ja institutionaaliseen luottamukseen), innovaatioilmastoon ja innovaatiotoiminnan tuloksellisuuteen. Organisaatiokulttuurin, luottamuksen ja innovatiivisuuden yhteyttä tarkasteltiin neljässä erityyppisessä organisaatiokulttuurissa (klaani-, adhokratia-, hierarkia- ja markkinakulttuurit), jotka pohjautuvat kilpailevien arvojen malliin. Tutkimuksen empiirinen osa toteutettiin posti ja Internet -pohjaisena kyselytutkimuksena 40 organisaatioyksikössä tilastollisen analyysin menetelmin. Yleisellä tasolla työssä saatiin selville, että luottamuksen ja innovatiivisuuden tasot vaihtelevat erityyppisissä organisaatio-kulttuureissa. Tarkemmin sanottuna klaani- ja adhokratiakulttuureissa luottamus ja innovatiivisuus olivat korkeita, ja näillä kulttuureilla oli myös positiivinen vaikutus innovaatiotoiminnan tuloksellisuuteen. Erityisesti institutionaalisen luottamuksen ja innovaatiotuen merkitykset olivat tärkeitä, sillä ne toimivat mediaattoreina organisaatiokulttuurin ja innovatiivisuuden välisessä suhteessa. Luottamuksella ja innovatiivisuudelle ei sitä vastoin ollut vaikutusta hierarkia- ja markkinakulttuureissa, tai vaikutus oli negatiivinen. Tässä työssä osoitettiin myös aiemmin hyvin vähän tutkitun institutionaalisen organisatorisen luottamuksen merkitys organisaatioiden innovatiivisuudessa.

Microglial cells in astroglial cultures: a cautionary note

Primary rodent astroglial-enriched cultures are the most popular model to study astroglial biology in vitro. From the original methods described in the 1970's a great number of minor modifications have been incorporated into these protocols by different laboratories. These protocols result in cultures in which the astrocyte is the predominant cell type, but astrocytes are never 100% of cells in these preparations. The aim of this review is to bring attention to the presence of microglia in astroglial cultures because, in my opinion, the proportion of and the role that microglial cells play in astroglial cultures are often underestimated. The main problem with ignoring microglia in these cultures is that relatively minor amounts of microglia can be responsible for effects observed on cultures in which the astrocyte is the most abundant cell type. If the relative contributions of astrocytes and microglia are not properly assessed an observed effect can be erroneously attributed to the astrocytes. In order to illustrate this point the case of NO production in activated astroglial-enriched cultures is examined. Lipopolysaccharide (LPS) induces nitric oxide (NO) production in astroglial-enriched cultures and this effect is very often attributed to astrocytes. However, a careful review of the published data suggests that LPS-induced NO production in rodent astroglial-enriched cultures is likely to be mainly microglial in origin. This review considers cell culture protocol factors that can affect the proportion of microglial cells in astroglial cultures, strategies to minimize the proportion of microglia in these cultures, and specific markers that allow the determination of such microglial proportions.

Cell type-specific expression and localization of cytochrome P450 isoforms in tridimensional aggregating rat brain cell cultures.

Within the Predict-IV FP7 project a strategy for measurement of in vitro biokinetics was developed, requiring the characterization of the cellular model used, especially regarding biotransformation, which frequently depends on cytochrome P450 (CYP) activity. The extrahepatic in situ CYP-mediated metabolism is especially relevant in target organ toxicity. In this study, the constitutive mRNA levels and protein localization of different CYP isoforms were investigated in 3D aggregating brain cell cultures. CYP1A1, CYP2B1/B2, CYP2D2/4, CYP2E1 and CYP3A were expressed; CYP1A1 and 2B1 represented almost 80% of the total mRNA content. Double-immunolabeling revealed their presence in astrocytes, in neurons, and to a minor extent in oligodendrocytes, confirming the cell-specific localization of CYPs in the brain. These results together with the recently reported formation of an amiodarone metabolite following repeated exposure suggest that this cell culture system possesses some metabolic potential, most likely contributing to its high performance in neurotoxicological studies and support the use of this model in studying brain neurotoxicity involving mechanisms of toxication/detoxication.

Identitat diversa. Conflicte identitari en persones vinculades a diferents cultures

Aquest treball de Final de Grau es centra en investigar un fenomen personal i social experimentat per persones d’origen immigrat al plantejar-se la seva identitat vinculada a dues o més cultures. Aquesta recerca pretén apropar-se a les vivències, pensaments, sentiments, etc. de persones que han viscut un conflicte identitari i esbrinar si han experimentat una certa pressió social per definir la seva identitat en una o altra cultura i com ho han viscut. La investigació compta amb la participació de sis persones d’origen marroquí, algunes nascudes al territori i altres no, les quals expliquen las seva experiència i perspectiva sobre aquest fenomen.

Cyclosporine A kinetics in brain cell cultures and its potential of crossing the blood-brain barrier.

There is an increasing need to develop improved systems for predicting the safety of xenobiotics. However, to move beyond hazard identification the available concentration of the test compounds needs to be incorporated. In this study cyclosporine A (CsA) was used as a model compound to assess the kinetic profiles in two rodent brain cell cultures after single and repeated exposures. CsA induced-cyclophilin B (Cyp-B) secretion was also determined as CsA-specific pharmacodynamic endpoint. Since CsA is a potent p-glycoprotein substrate, the ability of this compound to cross the blood-brain barrier (BBB) was also investigated using an in vitro bovine model with repeated exposures up to 14days. Finally, CsA uptake mechanisms were studied using a parallel artificial membrane assay (PAMPA) in combination with a Caco-2 model. Kinetic results indicate a low intracellular CsA uptake, with no marked bioaccumulation or biotransformation. In addition, only low CsA amounts crossed the BBB. PAMPA and Caco-2 experiments revealed that CsA is mostly trapped to lipophilic compartments and exits the cell apically via active transport. Thus, although CsA is unlikely to enter the brain at cytotoxic concentrations, it may cause alterations in electrical activity and is likely to increase the CNS concentration of other compounds by occupying the BBBs extrusion capacity. Such an integrated testing system, incorporating BBB, brain culture models and kinetics could be applied for assessing neurotoxicity potential of compounds.

Amiodarone biokinetics, the formation of its major oxidative metabolite and neurotoxicity after acute and repeated exposure of brain cell cultures.

The difficulty in mimicking nervous system complexity and cell-cell interactions as well as the lack of kinetics information has limited the use of in vitro neurotoxicity data. Here, we assessed the biokinetic profile as well as the neurotoxicity of Amiodarone after acute and repeated exposure in two advanced rodent brain cell culture models, consisting of both neurons and glial cells organized in 2 or 3 dimensions to mimic the brain histiotypic structure and function. A strategy was applied to evidence the abiotic processes possibly affecting Amiodarone in vitro bioavailability, showing its ability to adsorb to the plastic devices. At clinically relevant Amiodarone concentrations, known to induce neurotoxicity in some patients during therapeutic treatment, a complete uptake was observed in both models in 24h, after single exposure. After repeated treatments, bioaccumulation was observed, especially in the 3D cell model, together with a greater alteration of neurotoxicity markers. After 14days, Amiodarone major oxidative metabolite (mono-N-desethylamiodarone) was detected at limited levels, indicating the presence of active drug metabolism enzymes (i.e. cytochrome P450) in both models. The assessment of biokinetics provides useful information on the relevance of in vitro toxicity data and should be considered in the design of an Integrated Testing Strategy aimed to identify specific neurotoxic alerts, and to improve the neurotoxicity assay predictivity for human acute and repeated exposure.