Études de la réponse du métabolisme énergétique à la carence en fer dans les cultures cellulaires de Solanum tuberosum

Le fer est un micronutriment important pour la croissance et le développement des plantes. Il agit comme cofacteur pour plusieurs enzymes et il est important pour des processus tels que la photosynthèse et la respiration. Souvent, le Fe dans le sol n’est pas bio-disponible pour la plante. Les plantes ont développé des stratégies pour solubiliser le Fe du sol pour le rendre disponible et assimilable pour elles. Il y a deux stratégies, la première est caractéristique des dicotylédones et la seconde est caractéristique des monocotylédones. Le modèle utilisé dans cette étude est une culture cellulaire de Solanum tuberosum. Une partie de la recherche effectuée a permis la mesure d’activité et d’expression relative de certaines enzymes impliquées dans le métabolisme énergétique et la fourniture de précurseurs pour la synthèse d’ADN : la Nucléoside diphosphate kinase, la Ribonucléotide reductase, la Glucose 6-phosphate déshydrogénase et la 6-Phosphogluconate déshydrogénase dans les cellules en présence ou en absence de Fe. Chez certains organismes, la déficience en Fe est associée à une perte de croissance qui est souvent liée à une diminution de la synthèse d’ADN. Chez les cultures de cellules de S. tuberosum, les résultats indiquent que la différence de biomasse observée entre les traitements n’est pas due à une variation de l’activité ou l’expression relative d’une de ces enzymes. En effet, aucune variation significative n’a été détectée entre les traitements (+/- Fe) pour l’activité ni l’expression relative de ces enzymes. Une autre partie de la recherche a permis d’évaluer l’activité des voies métaboliques impliquées dans la stratégie 1 utilisée par S. tuberosum. Cette stratégie consomme des métabolites énergétiques: de l’ATP pour solubiliser le Fe et du pouvoir réducteur (NAD(P)H), pour réduire le Fe3+ en Fe2+. Des études de flux métaboliques ont été faites afin d’étudier les remaniements du métabolisme carboné en déficience en Fe chez S. tuberosum. Ces études ont démontré une baisse du régime dans les différentes voies du métabolisme énergétique dans les cellules déficientes en Fe, notamment dans le flux glycolytique et le flux de C à travers la phosphoenolpyruvate carboxylase. En déficience de Fe il y aurait donc une dépression du métabolisme chez S. tuberosum qui permettrait à la cellule de ralentir son métabolisme pour maintenir sa vitalité. En plus des flux, les niveaux de pyridines nucléotides ont été mesurés puisque ceux-ci servent à réduire le Fe dans la stratégie 1. Les résultats démontrent des niveaux élevés des formes réduites de ces métabolites en déficience de Fe. L’ensemble des résultats obtenus indiquent qu’en déficience de Fe, il y a une baisse du métabolisme permettant à la cellule de s’adapter et survivre au stress.

Études de la réponse du métabolisme énergétique à la carence en fer dans les cultures cellulaires de Solanum tuberosum

Le fer est un micronutriment important pour la croissance et le développement des plantes. Il agit comme cofacteur pour plusieurs enzymes et il est important pour des processus tels que la photosynthèse et la respiration. Souvent, le Fe dans le sol n’est pas bio-disponible pour la plante. Les plantes ont développé des stratégies pour solubiliser le Fe du sol pour le rendre disponible et assimilable pour elles. Il y a deux stratégies, la première est caractéristique des dicotylédones et la seconde est caractéristique des monocotylédones. Le modèle utilisé dans cette étude est une culture cellulaire de Solanum tuberosum. Une partie de la recherche effectuée a permis la mesure d’activité et d’expression relative de certaines enzymes impliquées dans le métabolisme énergétique et la fourniture de précurseurs pour la synthèse d’ADN : la Nucléoside diphosphate kinase, la Ribonucléotide reductase, la Glucose 6-phosphate déshydrogénase et la 6-Phosphogluconate déshydrogénase dans les cellules en présence ou en absence de Fe. Chez certains organismes, la déficience en Fe est associée à une perte de croissance qui est souvent liée à une diminution de la synthèse d’ADN. Chez les cultures de cellules de S. tuberosum, les résultats indiquent que la différence de biomasse observée entre les traitements n’est pas due à une variation de l’activité ou l’expression relative d’une de ces enzymes. En effet, aucune variation significative n’a été détectée entre les traitements (+/- Fe) pour l’activité ni l’expression relative de ces enzymes. Une autre partie de la recherche a permis d’évaluer l’activité des voies métaboliques impliquées dans la stratégie 1 utilisée par S. tuberosum. Cette stratégie consomme des métabolites énergétiques: de l’ATP pour solubiliser le Fe et du pouvoir réducteur (NAD(P)H), pour réduire le Fe3+ en Fe2+. Des études de flux métaboliques ont été faites afin d’étudier les remaniements du métabolisme carboné en déficience en Fe chez S. tuberosum. Ces études ont démontré une baisse du régime dans les différentes voies du métabolisme énergétique dans les cellules déficientes en Fe, notamment dans le flux glycolytique et le flux de C à travers la phosphoenolpyruvate carboxylase. En déficience de Fe il y aurait donc une dépression du métabolisme chez S. tuberosum qui permettrait à la cellule de ralentir son métabolisme pour maintenir sa vitalité. En plus des flux, les niveaux de pyridines nucléotides ont été mesurés puisque ceux-ci servent à réduire le Fe dans la stratégie 1. Les résultats démontrent des niveaux élevés des formes réduites de ces métabolites en déficience de Fe. L’ensemble des résultats obtenus indiquent qu’en déficience de Fe, il y a une baisse du métabolisme permettant à la cellule de s’adapter et survivre au stress.

Characterisation Of The Membrane Transport Of Pilocarpine In Cell Suspension Cultures Of Pilocarpus Microphyllus.

Pilocarpine is an alkaloid obtained from the leaves of Pilocarpus genus, with important pharmaceutical applications. Previous reports have investigated the production of pilocarpine by Pilocarpus microphyllus cell cultures and tried to establish the alkaloid biosynthetic route. However, the site of pilocarpine accumulation inside of the cell and its exchange to the medium culture is still unknown. Therefore, the aim of this study was to determine the intracellular accumulation of pilocarpine and characterise its transport across membranes in cell suspension cultures of P. microphyllus. Histochemical analysis and toxicity assays indicated that pilocarpine is most likely stored in the vacuoles probably to avoid cell toxicity. Assays with exogenous pilocarpine supplementation to the culture medium showed that the alkaloid is promptly uptaken but it is rapidly metabolised. Treatment with specific ABC protein transporter inhibitors and substances that disturb the activity of secondary active transporters suppressed pilocarpine uptake and release suggesting that both proteins may participate in the traffic of pilocarpine to inside and outside of the cells. As bafilomicin A1, a specific V-type ATPase inhibitor, had little effect and NH4Cl (induces membrane proton gradient dissipation) had moderate effect, while cyclosporin A and nifedipine (ABC proteins inhibitors) strongly inhibited the transport of pilocarpine, it is believed that ABC proteins play a major role in the alkaloid transport across membranes but it is not the exclusive one. Kinetic studies supported these results.

Reproductive biology of Davilla kunthii A. St-Hil. (Dilleniaceae) in Central Amazonia

This survey aimed at describing the interactions of floral visitors and Davilla kunthii A. St.-Hil. as well as characteristics of its reproductive biology in Itacoatiara, state of Amazonas, Brazil. Tests of the breeding system were performed. The guild of visitors was described according to richness, abundance, relative frequency and constancy. The breeding system tests indicated that D. kunthii is self-compatible. The pollination system was characterized as generalist, with 39 visitor species, from three different orders. Bees were the main group of pollinators, thus some behavioural aspects were described. Th e period of highest foraging activity was between 7 and 10 am. Some species presented agonistic and monopolistic behaviour. Given the behaviour and destructive potential, the Curculionidae seem to have a greater impact as seed predators than pollinators.

Antioxidative responses of cell suspension cultures of two Coffea arabica varieties to low aluminum levels at pH 5.8

The effects of aluminum (Al) on the activities of antioxidant enzymes and ferritin expression were studied in cell suspension cultures of two varieties of Coffea arabica, Mundo Novo and Icatu, in medium with pH at 5.8. The cells were incubated with 300 µM Al3+, and the Al speciation as Al3+ was 1.45% of the mole fraction. The activities of superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferase (GST) were increased in Mundo Novo, whereas glutathione reductase (GR) and guaiacol peroxidase (GPOX) activities remained unchanged. SOD, GR, and GST activities were increased in Icatu, while CAT activity was not changed, and GPOX activity decreased. The expression of two ferritin genes (CaFer1 and CaFer2) were analyzed by Real-Time PCR. Al caused a downregulation of CaFER1 expression and no changes of CaFER2 expression in both varieties. The Western blot showed no alteration in ferritin protein levels in Mundo Novo and a decrease in Icatu. The differential enzymes responses indicate that the response to Al is variety-dependent.

Effects of a novel calcium aluminate cement on the early events of the progression of osteogenic cell cultures

The present study evaluated the progression of osteogenic cell cultures exposed to a novel calcium aluminate cement (CAC+) in comparison with the gold standard mineral trioxide aggregate (MTA). Cells were enzimatically isolated from newborn rat calvarial bone, plated on glass coverslips containing either CAC+ or a control MTA samples in the center, and grown under standard osteogenic conditions. Over the 10-day culture period, roundening of sample edges was clearly noticed only for MTA group. Although both cements supported osteogenic cell adhesion, spreading, and proliferation, CAC+-exposed cultures showed significantly higher values in terms of total cell number at days 3 and 7, and total protein content and alkaline phosphatase activity at day 10. The present in vitro results indicate that the exposure to CAC+ supports a higher differentiation of osteogenic cells compared with the ones exposed to MTA. Further experimental studies should consider CAC+ as a potential alternative to MTA when the repair of mineralized tissues is one of the desired outcomes in endodontic therapy.

Neolignans and sesquiterpenes from leaves and embryogenic cultures of Ocotea Catharinensis (Lauraceae)

The extracts from leaves of Ocotea catharinensis Mez (Lauraceae) were found to contain fourteen neolignans and two sesquiterpenes: nine benzofuran types (including three new compounds 1e, 2f and 4b), one new seco-benzofuran type (3b), two bicyclo[3.2.1]octane types (including the new compound 5c), two new dimers of bicyclo[3.2.1]octane type (7a and 7b) and two sesquiterpenes (including a new humulanol 9). In addition, seven neolignans were also showed to occur in somatic embryos of O. catharinensis including one new bicyclo[3.2.1]octane type (4a).

Array-CGH testing in spontaneous abortions with normal karyotypes

In about 50% of first trimester spontaneous abortion the cause remains undetermined after standard cytogenetic investigation. We evaluated the usefulness of array-CGH in diagnosing chromosome abnormalities in products of conception from first trimester spontaneous abortions. Cell culture was carried out in short- and long-term cultures of 54 specimens and cytogenetic analysis was successful in 49 of them. Cytogenetic abnormalities (numerical and structural) were detected in 22 (44.89%) specimens. Subsequent, array-CGH based on large insert clones spaced at ~1 Mb intervals over the whole genome was used in 17 cases with normal G-banding karyotype. This revealed chromosome aneuplodies in three additional cases, giving a final total of 51% cases in which an abnormal karyotype was detected. In keeping with other recently published works, this study shows that array-CGH detects abnormalities in a further ~10% of spontaneous abortion specimens considered to be normal using standard cytogenetic methods. As such, array-CGH technique may present a suitable complementary test to cytogenetic analysis in cases with a normal karyotype.

Complex networks: the key to systems biology

Though introduced recently, complex networks research has grown steadily because of its potential to represent, characterize and model a wide range of intricate natural systems and phenomena. Because of the intrinsic complexity and systemic organization of life, complex networks provide a specially promising framework for systems biology investigation. The current article is an up-to-date review of the major developments related to the application of complex networks in biology, with special attention focused on the more recent literature. The main concepts and models of complex networks are presented and illustrated in an accessible fashion. Three main types of networks are covered: transcriptional regulatory networks, protein-protein interaction networks and metabolic networks. The key role of complex networks for systems biology is extensively illustrated by several of the papers reviewed.

Antibiosis and dark-pigments secretion by the phytopathogenic and environmental fungal species after interaction in vitro with a Bacillus subtilis isolate

In this work, different reactions in vitro between an environmental bacterial isolate and fungal species were related. The Gram-positive bacteria had terminal and subterminal endospores, presented metabolic characteristics of mesophilic and acidophilic growth, halotolerance, positive to nitrate reduction and enzyme production, as caseinase and catalase. The analysis of partial sequences containing 400 to 700 bases of the 16S ribosomal RNA gene showed identity with the genus Bacillus. However, its identity as B. subtilis was confirmed after analyses of the rpoB, gyrA, and 16S rRNA near-full-length sequences. Strong inhibitory activity of environmental microorganisms, such as Penicillium sp, Aspergillus flavus, A. niger, and phytopathogens, such as Colletotrichum sp, Alternaria alternata, Fusarium solani and F. oxysporum f.sp vasinfectum, was shown on co-cultures with B. subtilis strain, particularly on Sabouraud dextrose agar (SDA) and DNase media. Red and red-ochre color pigments, probably phaeomelanins, were secreted by A. alternata and A. niger respectively after seven days of co-culture.

Merluccius hubbsi (Teleostei: Merlucciidae): stock identification based on reproductive biology in the south-southeast brazilian region

The Argentine hake, Merluccius hubbsi, a demersal-pelagic species found from Rio de Janeiro, Brazil to the Tierra del Fuego, Argentina, has become an important target of the Brazilian bottom-trawler fleet since 2001. Earlier studies focusing on the species have suggested that more than one stock might occur off the Brazilian coast, in accordance with environmental features. In order to evaluate this hypothesis, fish were collected from four different areas in the Brazilian waters in which the hake is distributed, during the summers and winters of 1996-2001 and 2004, the females being used to analyze and compare spatial-temporal variations in ovarian maturation. Gonad indexes were also applied for the same purpose. Results indicate a north-south spawning gradient occurring as from summer at around 21°S to winter near 34°S, leading to the identification of two distinct stocks: one located between 21°S and 29°S (Southeastern stock) and the other between 29°S and 34°S (Southern stock), this latter shared with Uruguay and Argentina. Brazilian stocks present clear signs of overexploitation, the situation calling for an urgent solution.

The influence of the 1992-1993 el niño on the reproductive biology of Scomber japonicus peruanus (Jordán & Hubb, 1925)

The influence of the 1992-1993 El Niño events on the reproductive behavior of the Scomber japonicus peruanus (Chub mackerel) was studied from samples collected monthly, along the Peruvian coast (3º23'S-14º00'S), from January 1990 to December 1993. The monthly variation of the gonadosomatic index and the frequency of the periods of gonad maturation evidenced that the spawning of the species occurred all year long, being more intense in summer. The values of the gonadosomatic index were higher during the occurrence of the 1992-1993 El Niño, while the body weight and gonad weight decreased. Regarding the condition factor, its values decreased in females over 35 cm in fork length.

Embryonic, Larval, and Juvenile Development of the Sea Biscuit Clypeaster subdepressus (Echinodermata: Clypeasteroida)

Sea biscuits and sand dollars diverged from other irregular echinoids approximately 55 million years ago and rapidly dispersed to oceans worldwide. A series of morphological changes were associated with the occupation of sand beds such as flattening of the body, shortening of primary spines, multiplication of podia, and retention of the lantern of Aristotle into adulthood. To investigate the developmental basis of such morphological changes we documented the ontogeny of Clypeaster subdepressus. We obtained gametes from adult specimens by KCl injection and raised the embryos at 26 degrees C. Ciliated blastulae hatched 7.5 h after sperm entry. During gastrulation the archenteron elongated continuously while ectodermal red-pigmented cells migrated synchronously to the apical plate. Pluteus larvae began to feed in 3 d and were similar to 20 d old at metamorphosis; starved larvae died 17 d after fertilization. Postlarval juveniles had neither mouth nor anus nor plates on the aboral side, except for the remnants of larval spicules, but their bilateral symmetry became evident after the resorption of larval tissues. Ossicles of the lantern were present and organized in 5 groups. Each group had 1 tooth, 2 demipyramids, and 2 epiphyses with a rotula in between. Early appendages consisted of 15 spines, 15 podia (2 types), and 5 sphaeridia. Podial types were distributed in accordance to Loven's rule and the first podium of each ambulacrum was not encircled by the skeleton. Seven days after metamorphosis juveniles began to feed by rasping sand grains with the lantern. Juveniles survived in laboratory cultures for similar to 9 months and died with <500 mu m wide, a single open sphaeridium per ambulacrum, aboral anus, and no differentiated food grooves or petaloids. Tracking the morphogenesis of early juveniles is a necessary step to elucidate the developmental mechanisms of echinoid growth and important groundwork to clarify homologies between irregular urchins.