Toxicity and loss of mitochondrial membrane potential induced by Alkyl Gallates in Trypanosoma cruzi

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Hindawi Publishing Corporation International Scholarly Research Notices Volume 2015, Article ID 924670, 7 pages Research Article Toxicity and Loss of Mitochondrial Membrane Potential Induced by Alkyl Gallates in Trypanosoma cruzi Rogério Andréo,1 Luís Octávio Regasini,1 Maicon Segalla Petrônio,1 Bruna Galdorfini Chiari-Andréo,2 Aline Tansini,2 Dulce Helena Siqueira Silva,1 and Regina Maria Barretto Cicarelli2 1 Instituto de Quı́mica, Universidade Estadual Paulista (UNESP), Rua Professor Franscisco Degni 55, Bairro Quitandinha, CP 355, 14800-900 Araraquara, SP, Brazil 2 Faculdade de Ciências Farmacêuticas, Universidade Estadual Paulista (UNESP), Rodovia Araraquara-Jaú, km 1, CP 355, 14801-902 Araraquara, SP, Brazil Correspondence should be addressed to Regina Maria Barretto Cicarelli; Received 8 September 2014; Revised 18 December 2014; Accepted 22 December 2014 Academic Editor: Tzi Bun Ng Copyright © 2015 Rogério Andréo et al. his is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. American trypanosomiasis or Chagas disease is a debilitating disease representing an important social problem that afects, approximately, 10 million people in the world. he main aggravating factor of this situation is the lack of an efective drug to treat the diferent stages of this disease. In this context, the search for trypanocidal substances isolated from plants, synthetic or semi synthetic molecules, is an important strategy. Here, the trypanocidal potential of gallates was assayed in epimastigotes forms of T. cruzi and also, the interference of these substances on the mitochondrial membrane potential of the parasites was assessed, allowing the study of the mechanism of action of the gallates in the T. cruzi organisms. Regarding the preliminary structure-activity relationships, the side chain length of gallates plays crucial role for activity. Nonyl, decyl, undecyl, and dodecyl gallates showed potent antitrypanosomal efect (IC50 from 1.46 to 2.90 �M) in contrast with benznidazole (IC50 = 34.0 �M). Heptyl gallate showed a strong synergistic activity with benznidazole, reducing by 105 -fold the IC50 of benznidazole. Loss of mitochondrial membrane potential induced by these esters was revealed. Tetradecyl gallate induced a loss of 53% of the mitochondrial membrane potential, at IC50 value. 1. Introduction Chagas disease (or American trypanosomiasis) was recognized by World Health Organization as one of thirteen neglected tropical diseases in the world [1]. It was estimated that over 10 million people are infected, and Latin America was considered the area of the highest prevalence [2]. his parasitic disease is caused by Trypanosoma cruzi (family, Trypanosomatidae and order, Kinetoplastida), a hemolagellate protozoa [3], which can be found on several strains with diferent mechanisms of pathogenesis, immunogenicity, treatment response, and epidemiology [4]. he development of infection includes an acute phase which lasts up to six months ater infection, an indeterminate phase with no symptoms, and a chronic phase in which approximately 30% of patients show clinical evidence of heart disease or megavisceras [5]. he current chemotherapy uses benznidazole, a nitroimidazole derivative, which is orally administered in acute phase and short-term chronic phase [6, 7]. he usefulness of this drug is limited by its narrow therapeutic window and due to serious side efects, such as anorexia, nausea, headache, paresthesia, peripheral neuropathies, dermatitis, central nervous system (CNS) depression, and maniac symptoms [7, 8]. However, several parasite subpopulations with diferent host tissue’s tropism contribute to the low clinical eicacy of this drug [9]. herefore, there are eforts for the discovery and design of new therapeutic compounds to treat Chagas disease, due to the complexity of this disease. 2 Gallic acid (3,4,5-trihydroxybenzoic acid) is a precursor of hydrolysable tannins in the biosynthesis that occurs in plants [10] and its natural and semisynthetic derivatives have been associated with a wide variety of biological activities, such as antiproliferative [11–13], chemopreventive [14], antihemolytic [15], antioxidant [16], and anti-inlammatory [17] activities. However, the main interest in gallic acid and in its derivatives has been associated with their antimicrobial properties. It has been shown that n-octyl gallate possesses fungicidal activity against Saccharomyces cerevisiae and Zygosaccharomyces bailii in any stage of their growth [18, 19]. Leal and coworkers reported the potent fungitoxicity of n-nonyl gallate against yeasts, dermatophytes, and hialohyphomycetes [20]. Lauryl gallate showed antibacterial activity against Gram-positive bacteria, including methicillinresistant Staphylococcus aureus (MRSA) [21, 22]. hus, the aim of this study was to investigate the antitrypanosomal activity of gallic acid and its esters against epimastigote forms of Trypanosoma cruzi. Further analysis involving interactions generated by combinations between alkyl gallates and benznidazole was also carried out. In addition, the apoptosis-inducing activity by alkyl gallates was studied through the parasite mitochondrial membrane potential assay. 2. Material and Methods 2.1. Compounds. he compounds tested in this study were synthesized as described previously [14, 15]. 2.2. Parasites. Epimastigote forms of Trypanosoma cruzi of Y strain, described by Silva and Nussenzweig (1953) [23], which is considered a standard strain of type I, were grown in LIT media (liver infusion tryptose media) at 28∘ C [24]. 2.3. Cytotoxicity Assay Using MTT. he cytotoxicity assay with epimastigotes T. cruzi was performed using the MTT colorimetric method as described by Muelas-Serrano et al. (2000) [25] with modiications described in Cotinguiba et al. (2009) [26]. he parasites were treated with diferent concentrations of the substances for 72 hours and then, the parasite viability was estimated by measuring the absorbance at 595 nm. he concentration versus response curve was constructed to enable the determination of IC50 . An equation that describes the curve was obtained using Origin 7.0 sotware. Means and standard deviations were calculated. ANOVA and Tukey’s tests were carried out when necessary. Diferences in means were treated as signiicant when � < 0.05. Benznidazole was used as positive control (IC50 = 34.0 �M). 2.4. Mitochondrial Membrane Potential Assay. he inluence of the treatments with the gallates in the mitochondrial membrane potential was assessed. he parasites were treated with the substances during 72 hours using, as standard concentration, the IC50 previously determined by the MTT assay. he percentage of parasite cells which sufered loss in the mitochondrial membrane potential due to the treatment International Scholarly Research Notices with the tested compounds was measured by low cytometry using JC-1 dye (5,5� ,6,6� -tetrachloro-1,1� ,3,3� -tetraethylbenzimidazolcarbocyanine iodide, BD MitoScreen kit) according to the manufacturer’s instruction [27, 28]. A FacsCanto I cytometer was used, and the data were recorded and analyzed on the sotware of the equipment. Pentamidine was used as positive control, at 58.7 �M for 24-hour treatment. 3. Results and Discussion he antitrypanosomal activity of a homologous series of alkyl gallates, gallic acid, and other analogues was evaluated against epimastigote forms of Trypanosoma cruzi. his study was carried out in order to correlate chemical characteristics of the molecules with the observed activity, including the importance of free hydroxyl groups and side chain length. Table 1 summarizes the IC50 values of these compounds. Gallic acid (1) was found to be totally inactive against T. cruzi (IC50 > 100 �M) and its derivatives up to hexyl gallate (2–7) exhibited similar IC50 values (IC50 > 100 �M). Heptyl gallate (8, IC50 = 37.3 ± 0.9) showed weak activity which increased in the case of longer carbon chain derivatives up to undecyl gallate (12, IC50 = 1.46±0.0). However, the increase in the carbon chains, becoming bigger than undecyl gallate, led to lower activity, as shown for compounds 13 and 14 (Table 1). Such results indicate that the 3,4,5-trihydroxyphenyl moiety appeared to be necessary but not suicient for antiprotozoal activity. On the other hand, Kubo et al. (2001) [18] veriied that the length of the alkyl chain is not a major contributor but plays an important role in eliciting the activity of the gallates. Among the thirteen tested gallates (2–14) esteriied with linear alcohols from C1 to C14, best results were observed for esters with chain lengths ranging from 9 to 12, which present � log � values of 4.11−5.30, as reported previously by Rosso et al. (2006) [16]. n-Undecyl gallate (12), the most active derivative, with IC50 of 1.46 �M (� log � = 4.90), was twenty-three times more potent than benznidazole, used as positive control (IC50 = 34.0 �M). Such results indicate that esteriication led to appearance of antitrypanosomal activity, suggesting that a free carboxyl group was not crucial for protozoal death. In contrast, esters with � log � values lower than 3.32 or higher than 0.92 exhibited lower potency than the positive control. Altogether, there is a clear and positive correlation among IC50 values, alkyl chain length and its contribution to the lipophilicity, which are in agreement with previous studies of this homologous series [19, 29]. Furthermore, the trypanocidal potential increased with the increase in the number of carbons found on the side chain until reaching the maximum activity, in this case at n-undecyl gallate (12), and longer carbon chains showed lower trypanocidal activity, reaching a cutof at n-tetradecyl gallate (14). If the homologs longer than 14 might show antitrypanosomal activity, their IC50 values will be superior at 17.6 �M, representing low potency, not corroborating for further chemical or biological investments. In previous studies, the cutof phenomenon was observed for alkanols, which was correlated with their amphipathic properties. In the case of alkyl gallates, their amphiphilicity appeared to be dependent on the presence of two International Scholarly Research Notices 3 Table 1: Antitrypanosomal activity of gallic acid (1) and n-alkyl gallates (2–14) against epimastigote forms of Trypanosoma cruzi. IC50 values were expressed in �M. Compound Gallic acid (1) Methyl gallate (2) Ethyl gallate (3) Propyl gallate (4) Butyl gallate (5) Pentyl gallate (6) Hexyl gallate (7) Heptyl gallate (8) Octyl gallate (9) Nonyl gallate (10) Decyl gallate (11) Undecyl gallate (12) Dodecyl gallate (13) Tetradecyl gallate (14) Benznidazolec R H CH3 CH2 CH3 (CH2 )2 CH3 (CH2 )3 CH3 (CH2 )4 CH3 (CH2 )5 CH3 (CH2 )6 CH3 (CH2 )7 CH3 (CH2 )8 CH3 (CH2 )9 CH3 (CH2 )10 CH3 (CH2 )11 CH3 (CH2 )13 CH3 — � log �a 0.89 0.92 1.27 1.73 2.13 2.53 2.92 3.32 3.72 4.11 4.51 4.90 5.30 6.09 — IC50 >100b >100b >100b >100b >100b >100b >100b 37.3 ± 0.9 23.0 ± 5.3 2.90 ± 0.1 1.50 ± 0.3 1.46 ± 0.0 2.13 ± 0.2 17.6 ± 1.8 34.0 a Values described by Rosso and coworkers (2006) [16]. Percentage of inhibition at 100 �M. c Positive control. b features: hydrophilic phenolic hydroxyls and hydrophobic alkyl chain. Fujita and Kubo (2002) [19] reported the antifungal activity of gallates, suggesting that these compounds possessed amphiphilic capacity and surfactant properties and they might act disrupting the luidic bilayer membrane, leading to fungal death. he synergistic efects of the combination using gallic acid and its esters with benznidazole, the antichagasic drug, were also evaluated. he deinitions and mathematical determination of the formula for synergism were used as described by Hellmann et al. (2010) [30]. Briely, the fractional inhibitory concentration (FIC) index was calculated by using the formula FIC index = �/� + �/�. In this formula, �, �, and � were IC50 of gallate with benznidazole, IC50 of gallate alone, and IC50 of benznidazole alone, respectively. From this calculation, a FIC index ≤ 0.5 was considered evidence of synergistic efect, as this value can be generated only if the concentrations of both compounds in combination do not exceed one-quarter of the IC50 of either drug tested alone. A FIC index > 4.0 indicates antagonism. When tested alone, some gallates (2–7) have not led to a signiicant number of epimastigotes toxicity, even at the highest concentration tested (100 �M). herefore, to calculated FIC index for these compounds, the IC50 was arbitrarily deined as 50 �M (Table 2). he interactions between compounds 1–14 and benznidazole were highly variable, indicating dependence on the gallates chain length. Association between gallic acid (1) and short esters (2–4) with benznidazole was indiferent. Additive interaction was observed for inactive short esters (5 and 6) and most potent long esters, which exhibited FIC index values ranging from 0.560 to 0.765. Interestingly, medium chain length gallates (7–9) and the long chain length ester 14 with FIC index values bellow 0.257 exhibited synergism. he most signiicant synergistic efect was observed for heptyl gallate (8, FIC = 0.084), which reduced by 105 -fold the concentration of benznidazole necessary to inhibit cell growth in 50% (IC50 of 37.3 ± 0.93 �M). Synergism was observed at �M level, evidencing a strong antitrypanosomal activity and the potential of medium and long chain alkyl gallates as prototypes for further studies and development of novel therapeutic agents. It is also worth mentioning that the signiicant results from the association of benznidazole and alkyl gallates, which enabled the use of smaller doses of benznidazole, extended dosing intervals between consecutive administrations and short-term treatment leading to fewer and less intense side efects. Few reports have been found on the antimicrobial synergistic efect against trypanosomatidae protozoa. Urbina and coauthors (1988, 1995, 2002, 2009) [31–34] described systematic studies involving antifungal drugs and their associations as trypanocidal combined agents for treatment of Chagas disease, such as azoles (ketoconazole and posaconazole) with mevinolin, terbinaine, aspirin, and amiodarone. Recently, the interaction between benznidazole and parthenolide was investigated against T. cruzi, which evidenced synergistic and additive activities against epimastigotes and trypomastigotes, respectively. Additionally, trypanocidal gallates (8–14) were investigated for their inluence on the mitochondrial membrane potential. Considering that the loss of mitochondrial membrane potential is a typical characteristic of apoptotic cells [35], it was used to elucidate a possible mode of parasite death. In living cells, JC-1 dye crosses the plasmatic membrane as monomers, penetrating into the mitochondria. his process is controlled by the membrane potential of this organelle. he membrane of healthy mitochondria is polarized and JC-1 is rapidly internalized, increasing the concentration gradient, which may lead to its aggregation. In the low cytometer, the JC-1 monomers were detected in the FL-1 channel (FITCS), while its aggregates were observed in the red channel FL2 (PE). On the other hand, injured mitochondria exhibits depolarized membrane and JC-1 remains in the cytoplasm as monomers. hen, we suggest that the cells which loss their mitochondrial membrane potential, which could be apoptotic cells, lose their luorescence and could be captured in Q2 quadrant, whereas normal cells are detected in Q4 quadrant. Figure 1 presents the low cytometry graphs. Figure 1 presents the low cytometry analysis of n-undecyl gallate (12) on T. cruzi. Among the tested gallates, compounds 8, 9, and 14 promoted important loss in the mitochondrial membrane potential, which maybe an apoptosis-inducing activity, with percentage of cells with reduction of mitochondrial membrane potential of 46.6, 43.6, and 53.2, respectively. he results are presented on Table 3. Figure 1(a) shows that untreated epimastigotes (control) did not lose their mitochondrial membrane potential which can be observed due to the high luorescence detected in FL-2 channel (PE). Similarly, it is observed that n-undecyl gallate (12) (Figure 1(b)) did not stimulate this inluence in the treated cells since it did not induce a reduction 4 International Scholarly Research Notices 105 104 104 PE-A PE-A 105 103 103 102 102 102 103 105 104 102 103 104 105 FITC-A FITC-A (a) Negative control (b) n-Undecyl gallate (12) 105 PE-A 104 103 102 102 103 104 105 FITC-A (c) Pentamidine Figure 1: Flow cytometry analysis of epimastigotes forms of T. cruzi Y strain, without any treatment (a), treated with 1.46 �M of n-Undecyl gallate (12) for 72 h (b), and treated with 58.7 �M of pentamidine for 24 h (c). of luorescence as shown by the control, suggesting the maintenance of the membrane potential. In contrast, the cells treated with pentamidine (Figure 1(c)) show intense decrease in the luorescence represented by PE, suggesting a loss of mitochondrial membrane potential which may indicate apoptosis of the parasites analyzed. he results of low cytometry for n-undecyl gallate (12) are presented on Table 3. he results showed that the gallate esters used in this study induced loss of the mitochondrial membrane potential and maybe apoptosis in the parasites. his indicates that one of the mechanisms of cell death induced by the gallates activity could be similar to the apoptosis. his can be visualized by comparing the results with pentamidine, an apoptosisinducing drug [36], which led to diferent percentages of mitochondrial membrane potential relatively to the control. he treatment with some gallates, such as the substances 8, 9, and 14, showed a equilibrium value of MeCpG steps (,+14 deg.) [31,44]. In comparison, methylation has a significantly lower stability cost when happening at major groove positions, such as 211 and 21 base pair from dyad (mutations 9 and 12), where the roll of the nucleosome bound conformation (+10 deg.) is more compatible with the equilibrium geometry of MeCpG steps. The nucleosome destabilizing effect of cytosine methylation increases with the number of methylated cytosines, following the same position dependence as the single methylations. The multiple-methylation case reveals that each major groove meth- PLOS Computational Biology | 3 November 2013 | Volume 9 | Issue 11 | e1003354 DNA Methylation and Nucleosome Positioning ylation destabilizes the nucleosome by around 1 kJ/mol (close to the average estimate of 2 kJ/mol obtained for from individual methylation studies), while each minor groove methylation destabilizes it by up to 5 kJ/mol (average free energy as single mutation is around 6 kJ/mol). This energetic position-dependence is the reverse of what was observed in a recent FRET/SAXS study [30]. The differences can be attributed to the use of different ionic conditions and different sequences: a modified Widom-601 sequence of 157 bp, which already contains multiple CpG steps in mixed orientations, and which could assume different positioning due to the introduction of new CpG steps and by effect of the methylation. The analysis of our trajectories reveals a larger root mean square deviation (RMSD) and fluctuation (RMSF; see Figures S2– S3 in Text S1) for the methylated nucleosomes, but failed to detect any systematic change in DNA geometry or in intermolecular DNA-histone energy related to methylation (Fig. S1B, S1C, S4–S6 in Text S1). The hydrophobic effect should favor orientation of the methyl group out from the solvent but this effect alone is not likely to justify the positional dependent stability changes in Figure 2, as the differential solvation of the methyl groups in the bound and unbound states is only in the order of a fraction of a water molecule (Figure S5 in Text S1). We find however, a reasonable correlation between methylation-induced changes in hydrogen bond and stacking interactions of the bases and the change in nucleosome stability (see Figure S6 in Text S1). This finding suggests that methylation-induced nucleosome destabilization is related to the poorer ability of methylated DNA to fit into the required conformation for DNA in a nucleosome. Changes in the elastic deformation energy between methylated and un-methylated DNA correlate with nucleosomal differential binding free energies To further analyze the idea that methylation-induced nucleosome destabilization is connected to a worse fit of methylated DNA into the required nucleosome-bound conformation, we computed the elastic energy of the nucleosomal DNA using a harmonic deformation method [36,37,44]. This method provides a rough estimate of the energy required to deform a DNA fiber to adopt the super helical conformation in the nucleosome (full details in Suppl. Information Text S1). As shown in Figure 2, there is an evident correlation between the increase that methylation produces in the elastic deformation energy (DDE def.) and the free energy variation (DDG bind.) computed from MD/TI calculations. Clearly, methylation increases the stiffness of the CpG step [31], raising the energy cost required to wrap DNA around the histone octamers. This extra energy cost will be smaller in regions of high positive roll (naked DNA MeCpG steps have a higher roll than CpG steps [31]) than in regions of high negative roll. Thus, simple elastic considerations explain why methylation is better tolerated when the DNA faces the histones through the major groove (where positive roll is required) that when it faces histones through the minor groove (where negative roll is required). Nucleosome methylation can give rise to nucleosome repositioning We have established that methylation affects the wrapping of DNA in nucleosomes, but how does this translate into chromatin structure? As noted above, accumulation of minor groove methylations strongly destabilizes the nucleosome, and could trigger nucleosome unfolding, or notable changes in positioning or phasing of DNA around the histone core. While accumulation of methylations might be well tolerated if placed in favorable positions, accumulation in unfavorable positions would destabilize the nucleosome, which might trigger changes in chromatin structure. Chromatin could in fact react in two different ways in response to significant levels of methylation in unfavorable positions: i) the DNA could either detach from the histone core, leading to nucleosome eviction or nucleosome repositioning, or ii) the DNA could rotate around the histone core, changing its phase to place MeCpG steps in favorable positions. Both effects are anticipated to alter DNA accessibility and impact gene expression regulation. The sub-microsecond time scale of our MD trajectories of methylated DNAs bound to nucleosomes is not large enough to capture these effects, but clear trends are visible in cases of multiple mutations occurring in unfavorable positions, where unmethylated and methylated DNA sequences are out of phase by around 28 degrees (Figure S7 in Text S1). Due to this repositioning, large or small, DNA could move and the nucleosome structure could assume a more compact and distorted conformation, as detected by Lee and Lee [29], or a slightly open conformation as found in Jimenez-Useche et al. [30]. Using the harmonic deformation method, we additionally predicted the change in stability induced by cytosine methylation for millions of different nucleosomal DNA sequences. Consistently with our calculations, we used two extreme scenarios to prepare our DNA sequences (see Fig. 3): i) all positions where the minor grooves contact the histone core are occupied by CpG steps, and ii) all positions where the major grooves contact the histone core are occupied by CpG steps. We then computed the elastic energy required to wrap the DNA around the histone proteins in unmethylated and methylated states, and, as expected, observed that methylation disfavors DNA wrapping (Figure 3A). We have rescaled the elastic energy differences with a factor of 0.23 to match the DDG prediction in figure 2B. In agreement with the rest of our results, our analysis confirms that the effect of methylation is position-dependent. In fact, the overall difference between the two extreme methylation scenarios (all-in-minor vs all-in-major) is larger than 60 kJ/mol, the average difference being around 15 kJ/ mol. We have also computed the elastic energy differences for a million sequences with CpG/MeCpG steps positioned at all possible intermediate locations with respect to the position (figure 3B). The large differences between the extreme cases can induce rotations of DNA around the histone core, shifting its phase to allow the placement of the methylated CpG steps facing the histones through the major groove. It is illustrative to compare the magnitude of CpG methylation penalty with sequence dependent differences. Since there are roughly 1.5e88 possible 147 base pairs long sequence combinations (i.e., (4n+4(n/2))/2, n = 147), it is unfeasible to calculate all the possible sequence effects. However, using our elastic model we can provide a range of values based on a reasonably large number of samples. If we consider all possible nucleosomal sequences in the yeast genome (around 12 Mbp), the energy difference between the best and the worst sequence that could form a nucleosome is 0.7 kj/mol per base (a minimum of 1 kJ/mol and maximum of around 1.7 kJ/mol per base, the first best and the last worst sequences are displayed in Table S3 in Text S1). We repeated the same calculation for one million random sequences and we obtained equivalent results. Placing one CpG step every helical turn gives an average energetic difference between minor groove and major groove methylation of 15 kJ/ mol, which translates into ,0.5 kJ/mol per methyl group, 2 kJ/ mol per base for the largest effects. Considering that not all nucleosome base pair steps are likely to be CpG steps, we can conclude that the balance between the destabilization due to CpG methylation and sequence repositioning will depend on the PLOS Computational Biology | 4 November 2013 | Volume 9 | Issue 11 | e1003354 DNA Methylation and Nucleosome Positioning Figure 3. Methylated and non-methylated DNA elastic deformation energies. (A) Distribution of deformation energies for 147 bplong random DNA sequences with CpG steps positioned every 10 base steps (one helical turn) in minor (red and dark red) and major (light and dark blue) grooves respectively. The energy values were rescaled by the slope of a best-fit straight line of figure 2, which is 0.23, to por la lectura a través de la lectura de la prensa. La educación en los medios las fuerzas dispersas en función de los soportes mediáticos y orientarse más hacia la educación en medios que al dominio adquiere pleno derecho y entidad en la sección sexta titulada «competencias sociales y cívi- técnico de los aparatos. cas» que indica que «los alum- nos deberán ser capaces de juz- gar y tendrán espíritu crítico, lo que supone ser educados en los las programaciones oficiales, ya que, a lo largo de un medios y tener conciencia de su lugar y de su influencia estudio de los textos, los documentalistas del CLEMI en la sociedad». han podido señalar más de una centena de referencias a la educación de los medios en el seno de disciplinas 4. Un entorno positivo como el francés, la historia, la geografía, las lenguas, Si nos atenemos a las cifras, el panorama de la las artes plásticas : trabajos sobre las portadas de educación en medios es muy positivo. Una gran ope- prensa, reflexiones sobre temas mediáticos, análisis de ración de visibilidad como la «Semana de la prensa y publicidad, análisis de imágenes desde todos los ángu- de los medios en la escuela», coordinada por el CLE- los, reflexión sobre las noticias en los países europeos, MI, confirma año tras año, después de 17 convocato- información y opinión rias, el atractivo que ejerce sobre los profesores y los Esta presencia se constata desde la escuela mater- alumnos. Concebida como una gran operación de nal (2 a 6 años) donde, por ejemplo, se le pregunta a complementariedad entre la escuela y los profesiona- los niños más pequeños si saben diferenciar entre un les de los medios, alrededor del aprendizaje ciudada- periódico, un libro, un catálogo, a través de activida- no de la comunicación mediática, este evento moviliza des sensoriales, si saben para qué sirve un cartel, un durante toda una semana un porcentaje elevado de periódico, un cuaderno, un ordenador si son capa- centros escolares que representan un potencial de 4,3 ces de reconocer y distinguir imágenes de origen y de millones de alumnos (cifras de 2006). Basada en el naturaleza distintas. Podríamos continuar con más voluntariado, la semana permite desarrollar activida- ejemplos en todos los niveles de enseñanza y práctica- des más o menos ambiciosas centradas en la introduc- Páginas 43-48 ción de los medios en la vida de la escuela a través de la instalación de kioscos, organización de debates con profesionales y la confección por parte de los alumnos de documentos difundidos en los medios profesionales. Es la ocasión de dar un empujón a la educación en medios y de disfrutarlos. Los medios –un millar en 2006– se asocian de maneras diversas ofreciendo ejemplares de periódicos, acceso a noticias o a imágenes, proponiendo encuentros, permitiendo intervenir a los jóvenes en sus ondas o en sus columnas Esta operación da luz al trabajo de la educación en medios y moviliza a los diferentes participantes en el proyecto. 5. La formación de los docentes La formación es uno de los pilares principales de la educación en los medios. Su función es indispensable ya que no se trata de una disciplina, sino de una enseñanza que se hace sobre la base del voluntariado y del compromiso personal. Se trata de convencer, de mostrar, de interactuar. En primer lugar es necesario incluirla en la formación continua de los docentes, cuyo volumen se ha incrementado desde 1981 con la aparición de una verdadera política de formación continua de personal. Es difícil dar una imagen completa del volumen y del público, pero si nos atenemos a las cifras del CLEMI, hay más de 24.000 profesores que han asistido y se han involucrado durante 2004-05. 5.1. La formación continua En la mayoría de los casos, los profesores reciben su formación en contextos cercanos a su centro de trabajo, o incluso en este mismo. Después de una política centrada en la oferta que hacían los formadores, se valora más positivamente la demanda por parte del profesorado, ya que sólo así será verdaderamente fructífera. Los cursos de formación se repartieron en varias categorías: desde los formatos más tradicionales (cursos, debates, animaciones), hasta actividades de asesoramiento y de acompañamiento, y por supuesto los coloquios que permiten un trabajo en profundidad ya que van acompañados de expertos investigadores y profesionales. Citemos, por ejemplo en 2005, los coloquios del CLEMI-Toulouse sobre el cine documental o el del CLEMI-Dijon sobre «Políticos y medios: ¿connivencia?». Estos coloquios, que forman parte de un trabajo pedagógico regular, reagrupan a los diferentes participantes regionales y nacionales alrededor de grandes temas de la educación en medios y permiten generar nuevos conocimientos de aproximación y una profundización. Páginas 43-48 Hay otro tipo de formación original que se viene desarrollando desde hace menos tiempo, a través de cursos profesionales, como por ejemplo, en el Festival Internacional de Foto-periodismo «Visa para la imagen», en Perpignan. La formación se consolida en el curso, da acceso a las exposiciones, a las conferencias de profesionales y a los grandes debates, pero añade además propuestas pedagógicas y reflexiones didácticas destinadas a los docentes. Estas nuevas modalidades de formación son también consecuencia del agotamiento de la formación tradicional en las regiones. Los contenidos más frecuentes en formación continua conciernen tanto a los temas más clásicos como a los cambios que se están llevando a cabo en las prácticas mediáticas. Así encontramos distintas tendencias para 2004-05: La imagen desde el ángulo de la producción de imágenes animadas, el análisis de la imagen de la información o las imágenes del J.T. La prensa escrita y el periódico escolar. Internet y la información en línea. Medios y educación de los medios. 5.2 La formación inicial La formación inicial está aun en un grado muy ini- cial. El hecho de que la educación en medios no sea una disciplina impide su presencia en los IUFM (Institutos Universitarios de Formación de Maestros) que dan una prioridad absoluta a la didáctica de las disciplinas. En 2003, alrededor de 1.400 cursillistas sobre un total de 30.000 participaron en un momento u otro de un módulo de educación en medios. Estos módulos se ofrecen en función del interés que ese formador encuentra puntualmente y forman parte a menudo de varias disciplinas: documentación, letras, historia-geografía Estamos aún lejos de una política concertada en este dominio. La optativa «Cine-audiovisual» ha entrado desde hace muy poco tiempo en algunos IUFM destinada a obtener un certificado de enseñanza de la opción audiovisual y cine. Internet tiene cabida también en los cursos de formación inicial, recientemente con la aparición de un certificado informático y de Internet para los docentes, dirigido más a constatar competencias personales que a valorar una aptitud para enseñarlos. 6. ¿Y el futuro? El problema del futuro se plantea una vez más por la irrupción de nuevas técnicas y nuevos soportes. La difusión acelerada de lo digital replantea hoy muchas cuestiones relativas a prácticas mediáticas. Muchos Comunicar, 28, 2007 47 Comunicar, 28, 2007 Enrique Martínez-Salanova '2007 para Comunicar 48 trabajos que llevan el rótulo de la educación en medios solicitan una revisión ya que los conceptos cambian. La metodología elaborada en el marco de la educación en medios parece incluso permitir la inclinación de la sociedad de la información hacia una sociedad del conocimiento, como defiende la UNESCO. En Francia, se necesitaría unir las fuerzas dispersas en función de los soportes mediáticos y orientarse más hacia la educación en medios que al dominio técnico de los aparatos. Los avances recientes en el reconocimiento de estos contenidos y las competencias que supondrían podrían permitirlo. Referencias CLEMI/ACADEMIE DE BORDEAUX (Ed.) (2003): Parcours médias au collège: approches disciplinaires et transdisciplinaires. Aquitaine, Sceren-CRDP. GONNET, J. (2001): Education aux médias. Les controverses fécondes. Paris, Hachette Education/CNDP. SAVINO, J.; MARMIESSE, C. et BENSA, F. (2005): L’éducation aux médias de la maternelle au lycée. Direction de l’Enseignement Scolaire. Paris, Ministère de l’Education Nationale, Sceren/CNDP, Témoigner. BEVORT, E. et FREMONT, P. (2001): Médias, violence et education. Paris, CNDP, Actes et rapports pour l’éducation. – fiches pédagogiques, rapports et liens avec les pages régionales/académiques. – Le site «Quai des images» est dédié à l’enseignement du cinéma et de l’audiovisuel. – la rubrique «Côté profs» a une entrée «education aux médias». – Programme européen d’éducation aux risques liés à Internet. dResedfeleexliobnuetsacón Páginas 43-48
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