Extracellular ATP triggers proteolysis and cytosolic Ca2+ rise in Plasmodium berghei and Plasmodium yoelii malaria parasites

Background: Plasmodium has a complex cell biology and it is essential to dissect the cell-signalling pathways underlying its survival within the host. Methods: Using the fluorescence resonance energy transfer (FRET) peptide substrate Abz-AIKFFARQ-EDDnp and Fluo4/AM, the effects of extracellular ATP on triggering proteolysis and Ca2+ signalling in Plasmodium berghei and Plasmodium yoelii malaria parasites were investigated. Results: The protease activity was blocked in the presence of the purinergic receptor blockers suramin (50 mu M) and PPADS (50 mu M) or the extracellular and intracellular calcium chelators EGTA (5 mM) and BAPTA/AM (25, 100, 200 and 500 mu M), respectively for P. yoelii and P. berghei. Addition of ATP (50, 70, 200 and 250 mu M) to isolated parasites previously loaded with Fluo4/AM in a Ca2+-containing medium led to an increase in cytosolic calcium. This rise was blocked by pre-incubating the parasites with either purinergic antagonists PPADS (50 mu M), TNP-ATP (50 mu M) or the purinergic blockers KN-62 (10 mu M) and Ip5I (10 mu M). Incubating P. berghei infected cells with KN-62 (200 mu M) resulted in a changed profile of merozoite surface protein 1 (MSP1) processing as revealed by western blot assays. Moreover incubating P. berghei for 17 h with KN-62 (10 mu M) led to an increase in rings forms (82% +/- 4, n = 11) and a decrease in trophozoite forms (18% +/- 4, n = 11). Conclusions: The data clearly show that purinergic signalling modulates P. berghei protease(s) activity and that MSP1 is one target in this pathway.

Period adding cascades: Experiment and modeling in air bubbling

Period adding cascades have been observed experimentally/numerically in the dynamics of neurons and pancreatic cells, lasers, electric circuits, chemical reactions, oceanic internal waves, and also in air bubbling. We show that the period adding cascades appearing in bubbling from a nozzle submerged in a viscous liquid can be reproduced by a simple model, based on some hydrodynamical principles, dealing with the time evolution of two variables, bubble position and pressure of the air chamber, through a system of differential equations with a rule of detachment based on force balance. The model further reduces to an iterating one-dimensional map giving the pressures at the detachments, where time between bubbles come out as an observable of the dynamics. The model has not only good agreement with experimental data, but is also able to predict the influence of the main parameters involved, like the length of the hose connecting the air supplier with the needle, the needle radius and the needle length. (C) 2012 American Institute of Physics. [http://dx.doi.org/10.1063/1.3695345]

Temperature rise during Er:YAG cavity preparation of primary enamel

This study aimed to assess in vitro thermal alterations taking place during the Er:YAG laser cavity preparation of primary tooth enamel at different energies and pulse repetition rates. Forty healthy human primary molars were bisected in a mesio-distal direction, thus providing 80 fragments. Two small orifices were made on the dentin surface to which type K thermocouples were attached. The fragments were individually fixed with wax in a cylindrical PlexiglassA (R) abutment and randomly assigned to eight groups, according to the laser parameters (n = 10): G1 -aEuro parts per thousand 250 mJ/ 3 Hz, G2 -aEuro parts per thousand 250 mJ/ 4 Hz, G3 -aEuro parts per thousand 250 mJ/ 6 Hz, G4 -aEuro parts per thousand 250 mJ/10 Hz, G5 -aEuro parts per thousand 250 mJ/ 15 Hz, G6 -aEuro parts per thousand 300 mJ/ 3 Hz, G7 -aEuro parts per thousand 300 mJ/ 4 Hz and G8 -aEuro parts per thousand 300 mJ/ 6 Hz. An area of 4 mm(2) was delimited. Cavities were done (2 mm long x 2 mm wide x 1 mm thick) using non-contact (12 mm) and focused mode. Temperature values were registered from the start of laser irradiation until the end of cavity preparation. Data were analyzed by one-way ANOVA and Tukey test (p a parts per thousand currency signaEuro parts per thousand 0.05). Groups G1, G2, G6, and G7 were statistically similar and furnished the lowest mean values of temperature rise. The set 250 mJ/10 and 15 Hz yielded the highest temperature values. The sets 250 and 300 mJ and 6 Hz provided temperatures with mean values below the acceptable critical value, suggesting that these parameters ablate the primary tooth enamel. Moreover, the temperature elevation was directly related to the increase in the employed pulse repetition rates. In addition, there was no direct correlation between temperature rise and energy density. Therefore, it is important to use a lower pulse frequency, such as 300 mJ and 6 Hz, during cavity preparation in pediatric patients.

Pachymetry-Guided Intrastromal Air Injection ("Pachy-Bubble") for Deep Anterior Lamellar Keratoplasty

To evaluate an innovative technique for intrastromal air injection to achieve deep anterior lamellar keratoplasty (DALK) with bare Descemet membrane (DM). Thirty-four eyes with anterior corneal pathology, including 27 with keratoconus, underwent DALK. After 400 mm trephination with a suction trephine, ultrasound pachymetry was performed 0.8 mm internally from the trephination groove in the 11 to 1 o'clock position. In this area, a 2-mm incision was created, parallel to the groove, with a micrometer diamond knife calibrated to 90% depth of the thinnest measurement. A cannula was inserted through the incision and 0.5 mL of air was injected to dissect the DM from the stroma. After peripheral paracentesis, anterior keratectomy was carried out to bare the DM. A 0.25-mm oversized graft was sutured in place. Overall, 94.1% of eyes achieved DALK. Bare DM was achieved in 30 eyes, and a pre-DM dissection was performed in 2 eyes. Air injection was successful in detaching the DM (achieving the big bubble) in 88.2% of the eyes. In keratoconus eyes, the rate was 88.9%. All cases but one required a single air injection to achieve DM detachment. Microperforations occurred in 5 cases: 3 during manual layer-by-layer dissection after air injection failed to detach the DM, 1 during removal of the residual stroma after big-bubble formation, and 1 during the diamond knife incision. Two cases (5.9%) were converted to penetrating keratoplasty because of macroperforations. The technique was reproducible, safe, and highly effective in promoting DALK with bare DM.