Conditioned medium from activated spleen cells supports the survival of rat retinal cells in vitro

Cytokines are a heterogeneous group of molecules that have been associated with several functions in the nervous system, such as survival and differentiation of neuronal and glial cells. In the present study, we demonstrated that conditioned medium from spleen cells activated with concanavalin A increased neuritogenesis and survival of retinal cells, as measured by biochemical and morphological criteria. Our data showed that conditioned medium induced a five-fold increase in the amount of protein after 120 h in vitro. This effect was not inhibited by the blockade of voltage-dependent L-type calcium channels with 5.0 µM nifedipine. However, the use of an intracellular calcium chelator (15.0 µM BAPTA-AM) inhibited this effect. Our results support the idea that factors secreted by activated lymphocytes, such as cytokines, can modulate the maintenance and the differentiation of rat retinal cells in vitro, indicating a possible role of these molecules in the development of retinal cells, as well as in its protection against pathological conditions

Effects of sciatic-conditioned medium on neonatal rat retinal cells in vitro

Schwann cells produce and release trophic factors that induce the regeneration and survival of neurons following lesions in the peripheral nerves. In the present study we examined the in vitro ability of developing rat retinal cells to respond to factors released from fragments of sciatic nerve. Treatment of neonatal rat retinal cells with sciatic-conditioned medium (SCM) for 48 h induced an increase of 92.5 ± 8.8% (N = 7 for each group) in the amount of total protein. SCM increased cell adhesion, neuronal survival and glial cell proliferation as evaluated by morphological criteria. This effect was completely blocked by 2.5 µM chelerythrine chloride, an inhibitor of protein kinase C (PKC). These data indicate that PKC activation is involved in the effect of SCM on retinal cells and demonstrate that fragments of sciatic nerve release trophic factors having a remarkable effect on neonatal rat retinal cells in culture.

Ionic radiocontrast inhibits endothelium-dependent vasodilation of the canine renal artery in vitro: possible mechanism of renal failure following contrast medium infusion

To determine if radiocontrast impairs vascular relaxation of the renal artery, segments (4-5 mm in length) of canine renal artery were suspended in vitro in organ chambers to measure isometric force (95% O2/5% CO2, at 37ºC). Arterial segments with and without endothelium were placed at the optimal point of their length-tension relation and incubated with 10 µM indomethacin to prevent synthesis of endogenous prostanoids. The presence of nonionic radiocontrast (iohexol, Omnipaque 350, 1 ml in 25 ml control solution, 4% (v/v)) did not alter endothelium-dependent relaxation to acetylcholine in rings precontracted with both norepinephrine and prostaglandin F2alpha (N = 6). When the rings were precontracted with prostaglandin F2alpha, the presence of ionic contrast did not inhibit the relaxation of the arteries. However, in canine renal arteries contracted with norepinephrine, the presence of ionic radiocontrast (diatrizoate meglumine and diatrizoate sodium, MD-76, 1 ml in 25 ml control solution, 4% (v/v)) inhibited relaxation in response to acetylcholine, sodium nitroprusside (N = 6 in each group), and isoproterenol (N = 5; P < 0.05). Rings were relaxed less than 50% of norepinephrine contraction. Following removal of the contrast, vascular relaxation in response to the agonists returned to normal. These results indicate that ionic radiocontrast nonspecifically inhibits vasodilation (both cAMP-mediated and cGMP-mediated) of canine renal arteries contracted with norepinephrine. This reversible impairment of vasodilation could inhibit normal renal perfusion and act as a mechanism of renal failure following radiocontrast infusion. In the adopted experimental protocol the isoproterenol-induced relaxation of renal arteries precontracted with norepinephrine was more affected, suggesting a pivotal role of the cAMP system.

Maturation of pig oocytes in vitro in a medium with pyruvate

The aim of in vitro maturation oocyte systems is to produce oocytes of comparable quality to those derived in vivo. The present study was designed to examine the surface morphological changes of the cumulus-oocyte complex (COC) and nuclear maturation in a culture system containing pyruvate. Ovaries were obtained from a slaughterhouseand transported to the laboratory within 2 h at 35-39ºC,and rinsed three times in 0.9% NaCl. The COCs were harvested from the ovaries and in vitro maturation was evaluated in San Marcos (SM) medium, a chemically defined culture system containing 22.3 mM sodium pyruvate. Oocytes were cultured in SM, SM + porcine follicular fluid (pFF) and in SM + pFF + gonadotropins (eCG and hCG) for 20-22 h and then without hormonal supplements for an additional 20-22 h. After culture, the degree of cumulus expansion and frequency of nuclear maturation were determined. Oocytes matured in SM (40.9%) and SM + pFF (42.9%) showed moderate cumulus expansion, whereas oocytes matured in SM + pFF + gonadotropins (54.6%) showed high cumulus expansion. The maturation rate of cultured oocytes, measured in function of the presence of the polar corpuscle, did not differ significantly between SM (40.9 ± 3.6%) and SM + pFF (42.9 ± 3.7%). These results indicate that pig oocytes can be successfully matured in a chemically definedmedium and suggest a possible bifunctional role of pyruvate as an energy substrate and as an antioxidant protecting oocytes against the stress of the in vitro environment.

In vitro modulation of alkaline phosphatase activity of Saccharomyces cerevisiae grown in low or high phosphate medium

Our objective was to characterize the modulation of the activity of Saccharomyces cerevisiae alkaline phosphatases (ALPs) by classic inhibitors of ALP activity, cholesterol and steroid hormones, in order to identify catalytic similarities between yeast and mammalian ALPs. S. cerevisiae expresses two ALPs, coded for by the PHO8 and PHO13 genes. The product of the PHO8 gene is repressible by Pi in the medium. ALP activity from yeast (grown in low or high phosphate medium) homogenates was determined with p-nitrophenylphosphate as substrate, pH 10.4 (lPiALP or hPiALP, respectively). Activation of hPiALP was observed with 5 mM L-amino acids (L-homoarginine _ 186%, L-leucine _ 155% and L-phenylalanine - 168%) and with 1 mM levamisole (122%; percentage values, in comparison to control, of recovered activity). EDTA (5 mM) and vanadate (1 mM) distinctly inhibited hPiALP (2 and 20%, respectively). L-homoarginine (5 mM) had a lower activating effect on lPiALP (166%) and was the strongest hPiALP activator. Corticosterone (5 mM) inhibited hPiALP to 90%, but no effect was observed in low phosphate medium. Cholesterol, ß-estradiol and progesterone also had different effects on lPiALP and hPiALP. A concentration-dependent activation of lPiALP minus hPiALP was evident with all three compounds, most especially with ß-estradiol and cholesterol. These results do not allow us to identify similarities of the behavior of S. cerevisiae ALPs and any of the mammalian ALPs but allow us to raise the hypothesis of differential regulation of S. cerevisiae ALPs by L-homoarginine, ß-estradiol and cholesterol and of using these compounds to discriminate between S. cerevisiae lPiALP and hPiALP.

Portfolio as an Indicator of Young Learners’ English Proficiency in Mainstream Language Instruction (EFL) and Bilingual Content Instruction (CLIL)

Tämä soveltavan kielitieteen ja kielitaidon arvioinnin toimintatutkimus tarkasteli kieliportfolion ominaisuuksia ja mahdollisuuksia nuorten oppijoiden englannin kielen arvioinnissa kahdessa eri oppimiskontekstissa: englanti oppiaineena (EFL) ja kaksikielinen sisällönopetus (CLIL). Tutkielman itsenäiset, kahteen eri englannin kielen rekisteriin (arkikieli ja akateeminen kieli) kohdistuneet portfoliokokeilut olivat erillisiä tapaustutkimuksia. Molemmat portfoliot perustuivat väljästi Eurooppalaiseen kielisalkkumalliin, ja ne olivat osa tutkielmantekijän luokkaopetusta ja -toimintaa. EFL -portfoliokokeilu 9-10-vuotiaille kolmasluokkalaisille toteutettiin marraskuun 2011 ja toukokuun 2012 välisenä aikana, kun CLIL -portfoliokokeilu n. 7-9-vuotiallle ensimmäisen ja toisen luokan oppilaille kesti kaksi lukuvuotta 2012–2014. Molemmissa kokeiluissa myös oppilaiden vanhemmat kuuluivat tutkimusjoukkoon, samoin CLIL -portfolion toteutuksessa avustaneet ja opettajanäkökulmaa edustaneet opettajaopiskelijat. Portfoliokokeilun aloitti myös kaksi muuta CLIL -opettajaa, mutta kumpikin kokeilu päättyi alkuvaiheeseensa. Tarkemman tarkastelun kohteina olivat tutkimuksen osallistujien kokemukset ja mielipiteet portfoliokokeiluista. Erityisesti tavoitteena oli selvittää, miten informatiivisena englannin kielitaidon indikaattorina kieliportfoliota pidettiin. Myös kehitysehdotuksia kerättiin. Trianguloitu aineisto koottiin sekä puolistrukturoiduin kyselyin että vapaaehtoisin teemahaastatteluin, jotka äänitettiin. EFL -aineisto koostui 18 oppilaskyselystä, 17 huoltajakyselystä ja 7 oppilashaastattelusta. CLIL -aineistoon sisältyi 19 oppilaskyselyä, 18 huoltajakyselyä, 7 oppilashaastattelua ja yksi opettajaopiskelijoiden (N=3) ryhmähaastattelu. Aineisto analysoitiin pääosin kvalitatiivisin menetelmin temaattisen sisältöanalyysin keinoin, mutta myös laskien frekvenssejä ja prosenttisosuuksia. Osallistujien mielipiteet ja kokemukset olivat hyvin samankaltaiset ja positiiviset kummassakin portfoliokokeilussa. Merkittävä enemmistö sekä oppilaista että huoltajista koki, että portfolion avulla on mahdollista osoittaa englannin kielitaitoa ja sen kehittymistä. Oppilaat kuvailivat portfoliotyötä hauskaksi ja kivaksi, ja heidän mielestään portfoliotehtävien pitäisi olla tarpeeksi haastavia, sisältää taiteellisia ja luovia elementtejä sekä kohdistua tuttuihin, mielenkiintoisiin aiheisiin. He totesivat, että portfolion avulla voi oppia lisää kieltä. Vanhempien mielestä portfolio kertoo koulun vieraiksi jääneistä oppisisällöistä, auttaa ymmärtämään lapsen ajatusmaailmaa ja motivaatiotasoa sekä paljastaa heidän kielitaidostaan uusia ulottuvuuksia. Opettajaopiskelijat havaitsivat, että portfolion avulla voi tutustua oppilaiden kieli- ja kulttuuritaustoihin sekä kartoittaa heidän kielellisiä tarpeitaan. Tämän tutkielman teoreettisen tarkastelun ja tulosten mukaan kieliportfolio tukee erinomaisesti uuden Perusopetuksen Opetussuunnitelman (NCC 2014) tavoitteita ja arvioinnin uudistuspyrkimyksiä sekä lainsäädännön arvioinnille asettamia edellytyksiä. Portfolio on erittäin suositeltava nuorten oppijoiden kielitaidon arviointimenetelmä perinteisten rinnalle.

Prevalence of BRCA1 and BRCA2 gene mutations in families with medium and high risk of breast and ovarian cancer in Brazil

Of all malignant neoplasias affecting women, breast cancer has the highest incidence rate in Brazil. The objective of the present study was to determine the frequency of genetic modifications in families with medium and high risk for breast and ovarian cancer from different regions of Brazil. An exploratory, descriptive study was carried out on the prevalence of the BRCA1 and BRCA2 mutations in case series of high-risk families for breast and/or ovarian cancer. After heredogram construction, a blood sample was taken and DNA extraction was performed in all index cases. The protein truncation test was used to screen for truncated mutations in exon 11 of the BRCA1 gene and in exons 10 and 11 of the BRCA2 gene. Of the 612 individuals submitted to genetic testing, 21 (3.4%), 19 women and 2 men, had mutations in the BRCA1 or BRCA2 genes. Of the 19 BRCA1 mutations found in the 18 participants, 7 consisted of ins6kb mutations, 4 were 5382insC, 3 were 2156delGinsCC, 2 were 185delAG, 1 was C1201G, 1 was C3522T, and 1 was 3450del4. With respect to the BRCA2 gene, 3 mutations were found: 5878del10, 5036delA and 4232insA (one case each). The prevalence of germline mutations in the BRCA1 and BRCA2 genes found in the present study was lower than reported by other studies on high-risk Brazilian populations. The inclusion of individuals with medium risk may have contributed to the lower prevalence observed.

Association of sulfur dioxide exposure with circulatory system deaths in a medium-sized city in Brazil

There is a demonstrable association between exposure to air pollutants and deaths due to cardiovascular diseases. The objective of this study was to estimate the effects of exposure to sulfur dioxide on mortality due to circulatory diseases in individuals 50 years of age or older residing in São José dos Campos, SP. This was a time-series ecological study for the years 2003 to 2007 using information on deaths due to circulatory disease obtained from Datasus reports. Data on daily levels of pollutants, particulate matter, sulfur dioxide (SO2), ozone, temperature, and humidity were obtained from the São Paulo State Environmental Agency. Moving average models for 2 to 7 days were calculated by Poisson regression using the R software. Exposure to SO2 was analyzed using a unipollutant, bipollutant or multipollutant model adjusted for mean temperature and humidity. The relative risks with 95%CI were obtained and the percent decrease in risk was calculated. There were 1928 deaths with a daily mean (± SD) of 1.05 ± 1.03 (range: 0-6). Exposure to SO2 was significantly associated with mortality due to circulatory disease: RR = 1.04 (95%CI = 1.01 to 1.06) in the 7-day moving average, after adjusting for ozone. There was an 8.5% decrease in risk in the multipollutant model, proportional to a decrease of SO2 concentrations. The results of this study suggest that residents of medium-sized Brazilian cities with characteristics similar to those of São José dos Campos probably have health problems due to exposure to air pollutants.

Induction of chagasic-like arrhythmias in the isolated beating hearts of healthy rats perfused with Trypanosoma cruzi-conditioned medium

Chagas' myocardiopathy, caused by the intracellular protozoan Trypanosoma cruzi, is characterized by microvascular alterations, heart failure and arrhythmias. Ischemia and arrythmogenesis have been attributed to proteins shed by the parasite, although this has not been fully demonstrated. The aim of the present investigation was to study the effect of substances shed by T. cruzi on ischemia/reperfusion-induced arrhythmias. We performed a triple ischemia-reperfusion (I/R) protocol whereby the isolated beating rat hearts were perfused with either Vero-control or Vero T. cruzi-infected conditioned medium during the different stages of ischemia and subsequently reperfused with Tyrode's solution. ECG and heart rate were recorded during the entire experiment. We observed that triple I/R-induced bradycardia was associated with the generation of auricular-ventricular blockade during ischemia and non-sustained nodal and ventricular tachycardia during reperfusion. Interestingly, perfusion with Vero-infected medium produced a delay in the reperfusion-induced recovery of heart rate, increased the frequency of tachycardic events and induced ventricular fibrillation. These results suggest that the presence of parasite-shed substances in conditioned media enhances the arrhythmogenic effects that occur during the I/R protocol.

A Web-Based Environmental Toolkit to Support Small and Medium-Sized Enterprises in the Implementation of Their Own Environmental Management System

With small and medium sized-enterprises (SMEs) taking up the majority of the global businesses, it is important they act in an environmentally responsible manner. Environmental management systems (EMS) help companies evaluate and improve their environmental impact but they often require human, financial, and temporary resources that not all SMEs can provide. This research encompasses interviews with representatives of two small enterprises in Germany to provide insights into their understanding, and knowledge of an EMS and how they perceive their responsibility towards the environment. Furthermore, it presents a toolkit created especially for small and medium-sized enterprises that serves as a simplified version of an EMS based on the ISO 14001 standard and is evaluated by the representatives of the SMEs. Some of the findings are: while being open to the idea of improving their environmental impact, SMEs do not always feel it is their responsibility to do so; they seem to lack the means to fully implement an EMS. The developed toolkit is considered useful and usable and recommendations are drawn for its future enhancement.

Biotransformation of citronellol in rose-oxide using cassava wastewater as a medium

The use of liquid cassava waste (manipueira) as the medium for the biotransformation of citronellol using a Penicillium sp strain was studied. The strain was able to grow in the waste and production of cellular mass reaching 25 g/L over three days of contact of the spores with the medium. Submerged cultures of Penicillium sp grown in manipueira were able to convert the substrate into cis- and trans-rose oxides when the cells were transferred into a mineral medium for the biotransformation experiments. The production of rose oxide increased by more than 2.4 times using this 2 media process as compared to processes using only a manipueira medium (cassava medium). Auto-oxidation products were not detected in the control experiments.