Bragg Curve Detection of Low-Energy Protons by Radiophotoluminescence Imaging in Lithium Fluoride Thin Films.

Lithium fluoride (LiF) crystals and thin films are utilized as radiation detectors for energy diagnostics of proton beams. This is achieved by analyzing the Bragg curves in LiF obtained by imaging the radiophotoluminescence of color centers created by protons. In LiF crystals, the Bragg peak depth increases superlinearly with the particle energy. A previous study has shown that, when 35 MeV protons impinge at grazing incidence onto LiF films deposited on Si(100) substrates, the Bragg peak in the films is located at the depth where it would be found in Si rather than in LiF due to multiple Coulomb scattering. In this paper, Monte Carlo simulations of proton irradiations in the 1-8 MeV energy range are performed and compared to experimental Bragg curves in optically transparent LiF films on Si(100) substrates. Our study focuses on this energy range because, as energy increases, the Bragg peak gradually shifts from the depth in LiF to that in Si. The impact of grazing incidence angle, LiF packing density, and film thickness on shaping the Bragg curve in the film is examined. At energies higher than 8 MeV, all these quantities must be considered, although the effect of packing density plays a minor role.

In Vivo Radiobiological Investigations with the TOP-IMPLART Proton Beam on a Medulloblastoma Mouse Model.

Protons are now increasingly used to treat pediatric medulloblastoma (MB) patients. We designed and characterized a setup to deliver proton beams for in vivo radiobiology experiments at a TOP-IMPLART facility, a prototype of a proton-therapy linear accelerator developed at the ENEA Frascati Research Center, with the goal of assessing the feasibility of TOP-IMPLART for small animal proton therapy research. Mice bearing Sonic-Hedgehog (Shh)-dependent MB in the flank were irradiated with protons to test whether irradiation could be restricted to a specific depth in the tumor tissue and to compare apoptosis induced by the same dose of protons or photons. In addition, the brains of neonatal mice at postnatal day 5 (P5), representing a very small target, were irradiated with 6 Gy of protons with two different collimated Spread-Out Bragg Peaks (SOBPs). Apoptosis was visualized by immunohistochemistry for the apoptotic marker caspase-3-activated, and quantified by Western blot. Our findings proved that protons could be delivered to the upper part while sparing the deepest part of MB. In addition, a comparison of the effectiveness of protons and photons revealed a very similar increase in the expression of cleaved caspase-3. Finally, by using a very small target, the brain of P5-neonatal mice, we demonstrated that the proton irradiation field reached the desired depth in brain tissue. Using the TOP-IMPLART accelerator we established setup and procedures for proton irradiation, suitable for translational preclinical studies. This is the first example of in vivo experiments performed with a "full-linac" proton-therapy accelerator.

Evaluation of the impact of vitrification on the actin cytoskeleton of in vitro matured ovine oocytes by means of Raman microspectroscopy.

PURPOSE: Investigation of the changes induced by vitrification on the cortical F-actin of in vitro matured ovine oocytes by Raman microspectroscopy (RMS). METHODS: Cumulus-oocyte complexes, recovered from the ovaries of slaughtered sheep, were matured in vitro and vitrified following the Minimum Essential Volume method using cryotops. The cortical region of metaphase II (MII) oocytes (1) exposed to vitrification solutions but not cryopreserved (CPA-exp), (2) vitrified/warmed (VITRI), and (3) untreated (CTR) was analyzed by RMS. A chemical map of one quadrant of single CPA-exp, VITRI and CTR oocytes was, also, performed. In order to identify the region of Raman spectra representative of the cortical F-actin modification, a group of in vitro matured oocytes were incubated with latrunculin-A (LATA), a specific F-actin destabilizing drug, and processed for RMS analysis. Thereafter, all the oocytes were stained with rhodamine phalloidin and evaluated by fluorescence confocal microscopy. Raman spectra of the oocytes were, statistically, analyzed using Principal Component Analysis (PCA). RESULTS: The PCA score plots showed a marked discrimination between CTR oocytes and CPA-exp/ VITRI groups. The main differences, highlighted by PCA loadings, were referable to proteins (1657, 1440 and 1300 cm(-1)) and, as indicated by LATA experiments, also included the changes of the F-actin. Analysis by confocal microscopy revealed a clear alteration of the cortical F-actin of CPA-exp and VITRI oocytes confirming RMS results. CONCLUSIONS: Raman microspectroscopy may represent an alternative analytical tool for investigating the biochemical modification of the oocyte cortex, including the F-actin cytoskeleton, during vitrification of in vitro matured ovine oocytes.

Influence of a Solid Surface on PNIPAM Microgel Films.

Stimuli-responsive microgels have attracted great interest in recent years as building blocks for fabricating smart surfaces with many technological applications. In particular, PNIPAM microgels are promising candidates for creating thermo-responsive scaffolds to control cell growth and detachment via temperature stimuli. In this framework, understanding the influence of the solid substrate is critical for tailoring microgel coatings to specific applications. The surface modification of the substrate is a winning strategy used to manage microgel-substrate interactions. To control the spreading of microgel particles on a solid surface, glass substrates are coated with a PEI or an APTES layer to improve surface hydrophobicity and add positive charges on the interface. A systematic investigation of PNIPAM microgels spin-coated through a double-step deposition protocol on pristine glass and on functionalised glasses was performed by combining wettability measurements and Atomic Force Microscopy. The greater flattening of microgel particles on less hydrophilic substrates can be explained as a consequence of the reduced shielding of the water-substrate interactions that favors electrostatic interactions between microgels and the substrate. This approach allows the yielding of effective control on microgel coatings that will help to unlock new possibilities for their application in biomedical devices, sensors, or responsive surfaces.

Raman spectroscopy-based approach to detect aging-related oxidative damage in the mouse oocyte.

PURPOSE: Detection of chemical modifications induced by aging-related oxidative damage in mouse metaphase II (MII) oocytes by Raman microspectroscopy. METHODS: CD-1 mice at the age of 4-8 weeks (young mice) and 48-52 weeks (old mice), were superovulated and oocytes at metaphase II stage were recovered from oviducts. MII oocytes from young animals were divided into three groups: A) young oocytes, processed immediately after collection; B) in vitro aged oocytes, cultured in vitro for 10 h before processing; C) oxidative-stressed oocytes, exposed to 10 mM hydrogen peroxide for 2 min before processing. Oocytes from reproductively old mice were referred to as old oocytes (D). All the oocytes were analyzed by confocal Raman microspectroscopy. The spectra were statistically analyzed using Principal Component Analysis (PCA). RESULTS: PCA evidenced that spectra from young oocytes (A) were clearly distinguishable from those obtained from in vitro-aged, oxidative-damaged and old oocytes (B, C, D) and presented significant differences in the bands attributable to lipid components (C = C stretching, 1,659 cm⁻¹; CH2 bending, 1,450 cm⁻¹; CH3 deformation,1,345 cm⁻¹; OH bending, C-N stretching, 1,211 cm⁻¹) and protein components (amide I band,1,659 cm⁻¹; CH2 bending modes and CH3 deformation, 1,450 cm⁻¹; C-N and C-C stretching vibrations, 1,132 cm⁻¹; phenylalanine's vibration, 1,035 cm⁻¹) CONCLUSIONS: Raman spectroscopy is a valuable non-invasive tool for the identification of biochemical markers of oxidative damage and could represent a highly informative method of investigation to evaluate the oocyte quality.

Hybrid materials with an increased resistance to hard X-rays using fullerenes as radical sponges.

The protection of organic and hybrid organic-inorganic materials from X-ray damage is a fundamental technological issue for broadening the range of applications of these materials. In the present article it is shown that doping hybrid films with fullerenes C(60) gives a significant reduction of damage upon exposure to hard X-rays generated by a synchrotron source. At low X-ray dose the fullerene molecules act as `radical scavengers', considerably reducing the degradation of organic species triggered by radical formation. At higher doses the gradual hydroxylation of the fullerenes converts C(60) into fullerol and a bleaching of the radical sinking properties is observed.

Raman microspectroscopy as a non-invasive tool to assess the vitrification-induced changes of ovine oocyte zona pellucida.

Cryopreservation-induced modifications of zona pellucida (ZP) have been explored to a lesser extent compared to other oocyte compartments. Different methods have been applied to identify ZP changes, but most of them are invasive and measure only few properties of ZP. Raman microspectroscopy (RMS) is a powerful technique for studying the molecular composition of cells but to date few studies have been performed on the oocytes using this method. The aim of the present study is to investigate the structural modifications of ZP of vitrified/warmed in vitro matured ovine oocytes by means of RMS. Cumulus-oocyte complexes were recovered from the ovaries of slaughtered adult sheep, matured in vitro and vitrified following the Minimum Essential Volume method using cryotops. ZPs of vitrified/warmed oocytes (VITRI), were exposed to vitrification solutions but not cryopreserved (CPA-exp) and untreated oocytes (CTR) were analyzed by RMS. We focused our analysis on the ZP protein and carbohydrate components by analyzing the 1230-1300 cm(-1) amide III region and the 1020-1140 cm(-1) spectral range in RMS spectra, respectively. The spectral profiles in the ranges of proteins and carbohydrates were comparable between CTR and CPA-exp ZPs, whereas VITRI ZPs showed a significantly altered protein secondary structure characterized by an increase in ß-sheet content and a decrease in the α-helix content. A significant modification of the carbohydrate components was also observed. This study demonstrates that vitrification of ovine oocytes induces biochemical changes of ZP related to the secondary structure of proteins and carbohydrate residues. Cryoprotectants do not strongly alter the molecular composition of ZP which is affected mainly by cooling. Raman technology offers a powerful and non-invasive tool to assess molecular modifications induced by cryopreservation in oocytes.

Structural evolution during evaporation of a 3-glycidoxypropyltrimethoxysilane film studied in situ by time resolved infrared spectroscopy.

Time resolved infrared spectroscopy has been applied to study in situ the evaporation process of a 3-glycidoxypropyltrimethoxysilane hybrid sol by casting a droplet on a ZnSe substrate; the analysis has been performed in the middle-infrared range and in the near-infrared range. The experiment has allowed following the structural changes induced by water evaporation and the formation of ordered structures within the cast film; the CH(2) scissoring bands have been used as a fingerprint for the disorder to order transition of the hybrid. The experiment has been done using both a fresh sol and an aged sol which produce respectively an amorphous material and a crystalline hybrid material. The analysis has shown that the epoxy groups do not react during the evaporation while the silica structure shows only a slight condensation and an increase in open cage-like species. At the end of evaporation the hybrid has a "soft-like" state which allows structural rearrangements to self-order.

Open Issues and Practical Suggestions for Telemedicine in Chronic Pain.

Telemedicine represents a major opportunity to facilitate continued assistance for patients with chronic pain and improve their access to care. Preliminary data show that an improvement can be expected of the monitoring, treatment adherence, assessment of treatment effect including the emotional distress associated with pain. Moreover, this approach seems to be convenient and cost-effective, and particularly suitable for personalized treatment. Nevertheless, several open issues must be highlighted such as identification of assessment tools, implementation of monitoring instruments, and ability to evaluate personal needs and expectations. Open questions exist, such as how to evaluate the need for medical intervention and interventional procedures, and how to define when a clinical examination is required for certain conditions. In this context, it is necessary to establish dynamic protocols that provide the right balance between face-to-face visits and telemedicine. Useful tips are provided to start an efficient experience. More data are needed to develop precise operating procedures. In the meantime, the first experiences from such settings can pave the way to initiate effective care pathways in chronic pain.

Facing the challenge of biosample imaging by FTIR with a synchrotron radiation source.

Fourier-transform infrared (FTIR) synchrotron radiation (SR) microspectroscopy is a powerful molecular probe of biological samples at cellular resolution (<10 microm). As the brilliance of SR is 100-1000 times higher than that of a conventional Globar source, FTIR microscopes are now available in almost all advanced SR facilities around the world. However, in spite of this superior performance, the expected advances in IR SR microscopy have not yet been realised, particularly with regard to bio-analytical studies of single cells and soft tissues. In recent decades solid-state array detectors have revolutionized the fields of molecular spectroscopy and chemical imaging, and now new IR focal plane array detectors implemented at ultra-bright SR facilities will extend the performance and overcome the existing limitations, possibly allowing IR SR instrumentation to achieve the highest sensitivity and resolution of molecular imaging. The impact of IR imaging on large tissue area and the complexity of the analysis are discussed. In view of the high brilliance of SR sources, a comparison of published microscope images is given. Finally, it is briefly outlined how an optimized combination of IR instrumentation and SR optical systems could reach the expected advantages of a SR-based FTIR imaging system.

Tracking infrared signatures of drugs in cancer cells by Fourier transform microspectroscopy.

Aimed at developing accurate, reliable and cost-saving analytical techniques for drugs screening we evaluated the potential of Fourier Transform (FT) InfraRed (IR) microspectroscopy (microFTIR) as a quantitative pre-diagnostic approach for the rapid identification of IR signatures of drugs targeting specific molecular pathways causing Chronic Myeloid Leukemia (CML). To obtain reproducible FTIR absorbance spectra at the necessary spatial resolution we optimized sample preparation and acquisition parameters on a single channel Mercury-Cadmium-Telluride (MCT) detector in the spectral interval of frequencies from 4000 to 800 cm(-1). Single K562 cells were illuminated by Synchrotron Radiation (SR) and a number of ~15 K562 cells spread in monolayer were illuminated by a conventional IR source (Globar), respectively. Combining IR spectral data with the results of complementary biochemical investigations carried out in samples by different analytical methods we identified and cross-validated IR signatures of drugs targeting the oncogenic protein BCR/ABL and its associated abnormal tyrosine kinase activity in K562 cell line. Unsupervised pattern recognition performed by Hierarchical Cluster Analysis (HCA) clustered the spectra of single K562 cells in two distinct groups roughly corresponding to living and to apoptotic cells, respectively. The corresponding IR spectral profiles were assumed to represent drug-resistant and drug-sensitive cells. Significant variations with increasing percentages of apoptotic cells were observed after the treatment of K562 cells with drugs that directly or indirectly target BCR/ABL. In conclusion, we suggest that microFTIR associated with multivariate data analysis may be useful to assess drug compounds in ex vivo cancer cell models and possibly peripheral blast cells from CML patients.

Evaporation-induced crystallization of pluronic F127 studied in situ by time-resolved infrared spectroscopy.

Rapid scan time-resolved infrared spectroscopy has been used to study in situ the crystallization induced by evaporation in an aqueous solution of a triblock copolymer, Pluronic F127. A droplet of the solution was cast on a silicon substrate and the evaporation followed by an infrared microscope in transmission mode. The evaporation rate of water, in the last stage of the process, has been shown to be correlated to the changes in the block copolymer; four different stages can be distinguished. The block copolymer passes from an amorphous micellar state in water to a partially crystallized phase in well-defined stages of the evaporation; the complete change from amorphous to crystalline state of Pluronic F127 is observed only after all water is evaporated.

Stain effects studied by time-resolved infrared imaging.

Pattern formation in evaporating colloidal droplets is an important phenomenon that is commonly observed in several solute-solvent systems; this is also an emerging technique for obtaining fine patterning through controlled conditions of drying. After evaporation of the solvent a ring-like pattern remains on the solid substrate under the condition of contact line pinning. We have used a new analytical technique, time-resolved infrared imaging, to investigate the formation of patterned structures with droplet drying, which is a typical time-dependent phenomenon. We have coupled the technique with optical imaging to follow the evolution of the droplet shape and dimension in correspondence with the chemical images. The main advantage of the technique is represented by the possibility to have simultaneous spatial and time-resolved information; we have applied the method to a water-methylene blue system that has been studied during drying. We have monitored the droplet profile change, in terms of water and methylene blue variation with time and space, at the droplet edge. The analysis has allowed a detailed reconstruction of the evaporating droplet profile with a micrometer scale resolution and of the change in concentration of the dye as a function of evaporating time. A uniform ring-like pattern, after evaporation in controlled relative humidity, is observed. The data are consistent with a constant evaporation model, whose conditions are realized when a constant evaporation is achieved along the entire droplet. The technique has allowed elucidating the evaporation phenomenon close to the contact line in a dye solute-solvent system, which is very difficult to study with other techniques.

Water evaporation studied by in situ time-resolved infrared spectroscopy.

Evaporation of water is a fundamental and ubiquitous process that is on the ground of different types of nanoscience phenomena such as evaporation induced self-assembly. Even if water evaporation is a very basic phenomenon, there is still a lack of experiments that give a direct insight of the process. In situ application of rapid scan time-resolved infrared spectroscopy to an evaporating droplet has allowed monitoring the process at different relative humidity conditions. The experiments have been performed in the near-infrared range using water and deuterated water. The water evaporation appears as a continuous process that is not affected by changes of relative humidity in the external environment. This result, however, is affected by the impossibility to discriminate the contribution of the adsorbed water. The same experiment repeated with a deuterated water droplet has allowed, instead, a direct observation of the contribution during the evaporation process from water in the external environment. The time-resolved analysis has shown that at higher relative humidity the water adsorption is enhanced and that this process is time delayed with respect to the beginning of the evaporation process.

Application of terahertz spectroscopy to time-dependent chemical-physical phenomena.

We present here a "proof of concepts" experiment that has been realized to show that time-dependent phenomena can be successfully studied in the terahertz region in a non pump-probe configuration. We have built-up an original analytical setup that has allowed following the evaporation of a deuterated water droplet cast on a CVD diamond substrate simultaneously in the near-middle infrared region and in the terahertz range. We have used a synchrotron light source in the terahertz region and a conventional thermal source in the infrared range. The results demonstrate that it is possible to study time-dependent phenomena simultaneously in the middle and terahertz ranges monitoring the entire chemical-physical process that occurs in the time domain of minutes.

Evaporation of ethanol and ethanol-water mixtures studied by time-resolved infrared spectroscopy.

The knowledge of the physics and the chemistry behind the evaporation of solvents is very important for the development of several technologies, especially in the fabrication of thin films from liquid phase and the organization of nanostructures by evaporation-induced self-assembly. Ethanol, in particular, is one of the most common solvents in sol-gel and evaporation-induced self-assembly processing of thin films, and a detailed understanding of its role during these processes is of fundamental importance. Rapid scan time-resolved infrared spectroscopy has been applied to study in situ the evaporation of ethanol and ethanol-water droplets on a ZnSe substrate. Whereas the evaporation rate of ethanol remains constant during the process, water is adsorbed by the ethanol droplet from the external environment and evaporates in three stages that are characterized by different evaporation rates. The adsorption and evaporation process of water in an ethanol droplet has been observed to follow a complex behavior: due to this reason, it has been analyzed by two-dimensional infrared correlation. Three different components in the water bending band have been resolved.

Time-resolved infrared spectroscopy as an in situ tool to study the kinetics during self-assembly of mesostructured films.

Rapid scan time-resolved infrared spectroscopy has been used to investigate in situ the kinetics of the chemical processes involved in the formation of self-assembled mesostructured films. The experiments have been done in transmission mode on films cast on a diamond disk using an infrared microscope. Two specific materials have been studied: silica and titania mesoporous films templated by a triblock copolymer surfactant (Pluronic F-127). The time dependence of solvent evaporation and condensation of the chemical species have been clearly observed. Different stages in the film formation have been identified, which support well the general theory of self-assembly. The in situ FTIR spectroscopy using time-resolved rapid scan has proven to be a very effective tool for in situ analysis of film formation from a liquid phase.

Kinetics of polycondensation reactions during self-assembly of mesostructured films studied by in situ infrared spectroscopy.

In situ synchrotron FTIR experiments have been performed during evaporation-induced self-assembly (EISA) of mesoporous films and the role of silica polycondensation in obtaining highly organized mesostructures has been illuminated.

Octreotide in the prevention of intra-abdominal complications following elective pancreatic resection: a prospective, multicenter randomized controlled trial.

HYPOTHESIS: Prophylactic administration of octreotide acetate decreases the rate of postoperative intra-abdominal complications (IACs) after elective pancreatic resection. DESIGN: Single-blind, controlled, randomized trial. SETTING: Multicenter (N = 20) trial in France. PATIENTS: Of 230 randomized patients undergoing pancreatoduodenectomy and pancreatic enteric anastomosis or distal pancreatectomy for either malignant or benign tumor or chronic pancreatitis, 122 were allotted intraoperatively to receive octreotide; 108 served as controls. RESULTS: All 230 patients were analyzed. Both groups were comparable except that significantly more patients in the octreotide group had biological glue injected into the main pancreatic duct alone (P<.001) or reinforcing the pancreatic enteric anastomosis (68% [83/122] vs 39% [42/108]; P =.002). Fewer patients (P =.08) in the octreotide group sustained 1 or more IACs (22% vs 32%). In subgroup analysis, octreotide significantly reduced the rate of patients sustaining 1 or more IACs when the main pancreatic duct diameter was less than 3 mm (P<.02), when pancreatojejunostomy was performed (P<.02), or both (P<.02). No significant differences were found regarding IAC severity. Twenty-three patients (10%) died postoperatively, 16 (70% of deaths) of whom had 1 or more IACs. The only independent risk factor for IACs found on multivariate analysis was pancreatoduodenectomy compared with distal pancreatectomy (P<.01) (odds ratio, 3.54 [95% confidence interval, 1.44-8.65]). CONCLUSIONS: Our results suggest that octreotide is not necessary for all patients undergoing pancreatic resection; it could be useful when the main pancreatic duct is less than 3 mm in diameter and when pancreatoduodenectomy is completed by pancreatojejunostomy.