Enhanced laser-driven proton acceleration via improved fast electron heating in a controlled pre-plasma.

The interaction of ultraintense laser pulses with solids is largely affected by the plasma gradient at the vacuum-solid interface, which modifies the absorption and ultimately, controls the energy distribution function of heated electrons. A micrometer scale-length plasma has been predicted to yield a significant enhancement of the energy and weight of the fast electron population and to play a major role in laser-driven proton acceleration with thin foils. We report on recent experimental results on proton acceleration from laser interaction with foil targets at ultra-relativistic intensities. We show a threefold increase of the proton cut-off energy when a micrometer scale-length pre-plasma is introduced by irradiation with a low energy femtosecond pre-pulse. Our realistic numerical simulations agree with the observed gain of the proton cut-off energy and confirm the role of stochastic heating of fast electrons in the enhancement of the accelerating sheath field.

Toward an effective use of laser-driven very high energy electrons for radiotherapy: Feasibility assessment of multi-field and intensity modulation irradiation schemes.

Radiotherapy with very high energy electrons has been investigated for a couple of decades as an effective approach to improve dose distribution compared to conventional photon-based radiotherapy, with the recent intriguing potential of high dose-rate irradiation. Its practical application to treatment has been hindered by the lack of hospital-scale accelerators. High-gradient laser-plasma accelerators (LPA) have been proposed as a possible platform, but no experiments so far have explored the feasibility of a clinical use of this concept. We show the results of an experimental study aimed at assessing dose deposition for deep seated tumours using advanced irradiation schemes with an existing LPA source. Measurements show control of localized dose deposition and modulation, suitable to target a volume at depths in the range from 5 to 10 cm with mm resolution. The dose delivered to the target was up to 1.6 Gy, delivered with few hundreds of shots, limited by secondary components of the LPA accelerator. Measurements suggest that therapeutic doses within localized volumes can already be obtained with existing LPA technology, calling for dedicated pre-clinical studies.

Effects of small misalignments on the intensity and Strehl ratio for a laser beam focused by an off-axis parabola.

A general procedure is described to calculate the intensity and Strehl ratio, at a generic plane in the focal region, of a beam focused by an off-axis parabolic mirror in the presence of small misalignments. The general theoretical framework is first developed, which allows a full vector diffraction treatment in the case of general misalignments. Then, a parametric numerical study is reported, aimed at highlighting the tolerances of both the intensity and Strehl ratio for small misalignments, for different focusing and off-axis parabola parameters. A set of experimental measurements aimed at validating the theoretical model is also discussed.

Radiobiological Effectiveness of Ultrashort Laser-Driven Electron Bunches: Micronucleus Frequency, Telomere Shortening and Cell Viability.

Laser-driven electron accelerators are capable of producing high-energy electron bunches in shorter distances than conventional radiofrequency accelerators. To date, our knowledge of the radiobiological effects in cells exposed to electrons using a laser-plasma accelerator is still very limited. In this study, we compared the dose-response curves for micronucleus (MN) frequency and telomere length in peripheral blood lymphocytes exposed to laser-driven electron pulse and X-ray radiations. Additionally, we evaluated the effects on cell survival of in vitro tumor cells after exposure to laser-driven electron pulse compared to electron beams produced by a conventional radiofrequency accelerator used for intraoperative radiation therapy. Blood samples from two different donors were exposed to six radiation doses ranging from 0 to 2 Gy. Relative biological effectiveness (RBE) for micronucleus induction was calculated from the alpha coefficients for electrons compared to X rays (RBE = alpha laser/alpha X rays). Cell viability was monitored in the OVCAR-3 ovarian cancer cell line using trypan blue exclusion assay at day 3, 5 and 7 postirradiation (2, 4, 6, 8 and 10 Gy). The RBE values obtained by comparing the alpha values were 1.3 and 1.2 for the two donors. Mean telomere length was also found to be reduced in a significant dose-dependent manner after irradiation with both electrons and X rays in both donors studied. Our findings showed a radiobiological response as mirrored by the induction of micronuclei and shortening of telomere as well as by the reduction of cell survival in blood samples and cancer cells exposed in vitro to laser-generated electron bunches. Additional studies are needed to improve preclinical validation of the radiobiological characteristics and efficacy of laser-driven electron accelerators in the future.

UV Radiation and Visible Light Induce hsp70 Gene Expression in the Antarctic Psychrophilic Ciliate Euplotes focardii.

The psychrophilic ciliate Euplotes focardii inhabits the shallow marine coastal sediments of Antarctica, where, over millions of years of evolution, it has reached a strict molecular adaptation to such a constant-temperature environment (about -2 °C). This long evolution at sub-zero temperatures has made E. focardii unable to respond to heat stress with the activation of its heat shock protein (hsp) 70 genes. These genes can, however, be expressed in response to other stresses, like the oxidative one, thus indicating that the molecular adaptation has exclusively altered the heat stress signaling pathways, while it has preserved hsp70 gene activation in response to other environmental stressors. Since radiative stress has proved to be affine to oxidative stress in several organisms, we investigated the capability of UV radiation to induce hsp70 transcription. E. focardii cell cultures were exposed to several different irradiation regimes, ranging from visible only to a mixture of visible, UV-A and UV-B. The irradiation values of each spectral band have been set to be comparable with those recorded in a typical Antarctic spring. Using Northern blot analysis, we measured the expression level of hsp70 immediately after irradiation (0-h-labeled samples), 1 h, and 2 h from the end of the irradiation. Surprisingly, our results showed that besides UV radiation, the visible light was also able to induce hsp70 expression in E. focardii. Moreover, spectrophotometric measurements have revealed no detectable endogenous pigments in E. focardii, making it difficult to propose a possible explanation for the visible light induction of its hsp70 genes. Further research is needed to conclusively clarify this point.

ß-Amyloid amorphous aggregates induced by the small natural molecule ferulic acid.

There is an emerging interest in small natural molecules for their potential therapeutic use in neurodegenerative disorders like Alzheimer's disease (AD). Ferulic acid (FA), an antioxidant phenolic compound present in fruit and vegetables, has been proposed as an inhibitor of beta amyloid (Aß) pathological aggregation. Using fluorescence and Fourier transform infrared spectroscopy, electrophoresis techniques, chromatographic analysis, and confocal microscopy, we investigated the effects of FA in the early stages of Aß fibrillogenesis in vitro. Our results show that FA interacts promptly with Aß monomers/oligomers, interfering since the beginning with its self-assembly and finally forming amorphous aggregates more prone to destabilization. These findings highlight the molecular basis underlying FA antiamyloidogenic activity in AD.

Understanding UV-driven metabolism in the hypersaline ciliate Fabrea salina.

By using NMR spectroscopy, a non-invasive investigation technique, we performed in vivo experiments aimed at uncovering the metabolic pathways involved in the early response of Fabrea salina cells to ultraviolet (UV) radiation. This hypersaline ciliate was chosen as a model organism because of its well-known high resistance to UV radiation. Identical cell samples were exposed to visible radiation only (control samples, CS) and to UV-B + UV-A + visible radiation (treated samples, TS), and NMR spectra of in vivo cells were collected at different exposure times. Resonances were identified through one- and two-dimensional experiments. To compare experiments performed at variable irradiation times on different culture batches, metabolite signals affected by the UV exposure were normalized to corresponding intensity at τ = 0, the zero exposure time. The most affected metabolites are all osmoprotectants, namely, choline, glycine-betaine, betaines, ectoine, proline, α-trehalose and sucrose. The time course of these signals presents qualitative differences between CS and TS, and most of these osmoprotectants tend to accumulate significantly in TS in a UV dose-dependent manner. A picture of the immediate stress response of F. salina against UV radiation in terms of osmoprotection, water retention and salting-out prevention is described.

In vivo NMR metabolic profiling of Fabrea salina reveals sequential defense mechanisms against ultraviolet radiation.

Fabrea salina is a hypersaline ciliate that is known to be among the strongest ultraviolet (UV)-resistant microorganisms; however, the molecular mechanisms of this resistance are almost unknown. By means of in vivo NMR spectroscopy, we determined the metabolic profile of living F. salina cells exposed to visible light and to polychromatic UV-B + UV-A + Vis radiation for several different exposure times. We used unsupervised pattern-recognition analysis to compare these profiles and discovered some metabolites whose concentration changed specifically upon UV exposure and in a dose-dependent manner. This variation was interpreted in terms of a two-phase cell reaction involving at least two different pathways: an early response consisting of degradation processes, followed by a late response activating osmoprotection mechanisms. The first step alters the concentration of formate, acetate, and saturated fatty-acid metabolites, whereas the osmoprotection modifies the activity of betaine moieties and other functionally related metabolites. In the latter pathway, alanine, proline, and sugars suggest a possible incipient protein synthesis as defense and/or degeneration mechanisms. We conclude that NMR spectroscopy on in vivo cells is an optimal approach for investigating the effect of UV-induced stress on the whole metabolome of F. salina because it minimizes the invasiveness of the measurement.

Optimizing soluble protein extraction and two-dimensional polyacrylamide gel electrophoresis quality for extremophile ciliates.

An efficient protein extraction methodology is quite important for sample preparation and subsequent 2-D PAGE and MS analysis. Cell lysis is the first step in protein extraction and purification. Many techniques are available for cell disruption, including physical and detergent-based methods. Here, we report on a very fast and efficient detergent-free Tris-based method to extract the soluble fraction proteins of extremophile ciliates, comparing it with a detergent-based protocol. This comparison has been carried out by means of 2-D PAGE and subsequent MALDI-compatible silver staining of protein samples obtained from the intensely pigmented hypersaline ciliate Fabrea salina and the Antarctic hypotrich ciliate Euplotes focardii. Our results indicate that this fast and easy extraction method allows to obtain more clear crude extracts and more spot-abundant polyacrylamide gels.