Acoustic streaming of microparticles using graphene-based interdigital transducers.

Surface acoustic wave (SAW) devices offer many benefits in chemistry and biomedicine, enabling precise manipulation of micro-droplets, mixing of liquids by acoustic streaming and pumping of liquids in enclosed channels, while presenting a cost-effective and easy fabrication and integration with electronic devices. In this work, we present microfluidic devices which use graphene-based interdigital transducers (IDTs) to generate SAWs with a frequency of 100 MHz and an amplitude of up to 200 pm, which allow us to manipulate microparticle solutions by acoustic streaming. Due to the negligible mass loading of the piezoelectric surface by graphene, the SAWs generated by these devices have no frequency shift, typically observed when metal IDTs are used.

Effects of fixatives on myelin molecular order probed with RP-CARS microscopy.

When live imaging is not feasible, sample fixation allows preserving the ultrastructure of biological samples for subsequent microscopy analysis. This process could be performed with various methods, each one affecting differently the biological structure of the sample. While these alterations were well-characterized using traditional microscopy, little information is available about the effects of the fixatives on the spatial molecular orientation of the biological tissue. We tackled this issue by employing rotating-polarization coherent anti-Stokes Raman scattering (RP-CARS) microscopy to study the effects of different fixatives on the myelin sub-micrometric molecular order and micrometric morphology. RP-CARS is a novel technique derived from CARS microscopy that allows probing spatial orientation of molecular bonds while maintaining the intrinsic chemical selectivity of CARS microscopy. By characterizing the effects of the fixation procedures, the present work represents a useful guide for the choice of the best fixation technique(s), in particular for polarization-resolved CARS microscopy. Finally, we show that the combination of paraformaldehyde and glutaraldehyde can be effectively employed as a fixative for RP-CARS microscopy, as long as the effects on the molecular spatial distribution, here characterized, are taken into account.

Workers' Exposure to Nano-Objects with Different Dimensionalities in R&D Laboratories: Measurement Strategy and Field Studies.

With the increasing interest in the potential benefits of nanotechnologies, concern is still growing that they may present emerging risks for workers. Various strategies have been developed to assess the exposure to nano-objects and their agglomerates and aggregates (NOAA) in the workplace, integrating different aerosol measurement instruments and taking into account multiple parameters that may influence NOAA toxicity. The present study proposes a multi-metric approach for measuring and sampling NOAA in the workplace, applied to three case studies in laboratories each dedicated to materials with different shapes and dimensionalities: graphene, nanowires, and nanoparticles. The study is part of a larger project with the aim of improving risk management tools in nanomaterials research laboratories. The harmonized methodology proposed by the Organization for Economic Cooperation and Development (OECD) has been applied, including information gathering about materials and processes, measurements with easy-to-use and hand-held real-time devices, air sampling with personal samplers, and off-line analysis using scanning electron microscopy. Significant values beyond which an emission can be attributed to the NOAA production process were identified by comparison of the particle number concentration (PNC) time series and the corresponding background levels in the three laboratories. We explored the relations between background PNC and microclimatic parameters. Morphological and elemental analysis of sampled filters was done to identify possible emission sources of NOAA during the production processes: rare particles, spherical, with average diameter similar to the produced NOAA were identified in the nanoparticles laboratory, so further investigation is recommended to confirm the potential for worker exposure. In conclusion, the information obtained should provide a valuable basis for improving risk management strategies in the laboratory at work.

Recurrent ETNK1 mutations in atypical chronic myeloid leukemia.

Despite the recent identification of recurrent SETBP1 mutations in atypical chronic myeloid leukemia (aCML), a complete description of the somatic lesions responsible for the onset of this disorder is still lacking. To find additional somatic abnormalities in aCML, we performed whole-exome sequencing on 15 aCML cases. In 2 cases (13.3%), we identified somatic missense mutations in the ETNK1 gene. Targeted resequencing on 515 hematological clonal disorders revealed the presence of ETNK1 variants in 6 (8.8%) of 68 aCML and 2 (2.6%) of 77 chronic myelomonocytic leukemia samples. These mutations clustered in a small region of the kinase domain, encoding for H243Y and N244S (1/8 H243Y; 7/8 N244S). They were all heterozygous and present in the dominant clone. The intracellular phosphoethanolamine/phosphocholine ratio was, on average, 5.2-fold lower in ETNK1-mutated samples (P < .05). Similar results were obtained using myeloid TF1 cells transduced with ETNK1 wild type, ETNK1-N244S, and ETNK1-H243Y, where the intracellular phosphoethanolamine/phosphocholine ratio was significantly lower in ETNK1-N244S (0.76 ± 0.07) and ETNK1-H243Y (0.37 ± 0.02) than in ETNK1-WT (1.37 ± 0.32; P = .01 and P = .0008, respectively), suggesting that ETNK1 mutations may inhibit the catalytic activity of the enzyme. In summary, our study shows for the first time the evidence of recurrent somatic ETNK1 mutations in the context of myeloproliferative/myelodysplastic disorders.

Effect of scattering on coherent anti-Stokes Raman scattering (CARS) signals.

We develop a computational framework to examine the factors responsible for scattering-induced distortions of coherent anti-Stokes Raman scattering (CARS) signals in turbid samples. We apply the Huygens-Fresnel wave-based electric field superposition (HF-WEFS) method combined with the radiating dipole approximation to compute the effects of scattering-induced distortions of focal excitation fields on the far-field CARS signal. We analyze the effect of spherical scatterers, placed in the vicinity of the focal volume, on the CARS signal emitted by different objects (2µm diameter solid sphere, 2µm diameter myelin cylinder and 2µm diameter myelin tube). We find that distortions in the CARS signals arise not only from attenuation of the focal field but also from scattering-induced changes in the spatial phase that modifies the angular distribution of the CARS emission. Our simulations further show that CARS signal attenuation can be minimized by using a high numerical aperture condenser. Moreover, unlike the CARS intensity image, CARS images formed by taking the ratio of CARS signals obtained using x- and y-polarized input fields is relatively insensitive to the effects of spherical scatterers. Our computational framework provide a mechanistic approach to characterizing scattering-induced distortions in coherent imaging of turbid media and may inspire bottom-up approaches for adaptive optical methods for image correction.

Local anodic oxidation on hydrogen-intercalated graphene layers: oxide composition analysis and role of the silicon carbide substrate.

We investigate nanoscale local anodic oxidation (LAO) on hydrogen-intercalated graphene grown by controlled sublimation of silicon carbide (SiC). Scanning probe microscopy was used as a lithographic and characterization tool in order to investigate the local properties of the nanofabricated structures. The anomalous thickness observed after the graphene oxidation process is linked to the impact of LAO on the substrate. Micro-Raman (µ-Raman) spectroscopy was employed to demonstrate the presence of two oxidation regimes depending on the applied bias. We show that partial and total etching of monolayer graphene can be achieved by tuning the bias voltage during LAO. Finally, a complete compositional characterization was achieved by scanning electron microscopy and energy dispersive spectroscopy.

GHz Electroluminescence Modulation in Nanoscale Subwavelength Emitters.

We investigate light emission from nanoscale point-sources obtained in hybrid metal-GaAs nanowires embedding two sharp axial Schottky barriers. Devices are obtained via the formation of Ni-rich metallic alloy regions in the nanostructure body thanks to a technique of controlled thermal annealing of Ni/Au electrodes. In agreement with recent findings, visible-light electroluminescence can be observed upon suitable voltage biasing of the junctions. We investigate the time-resolved emission properties of our devices and demonstrate an electrical modulation of light generation up to 1 GHz. We explore different drive configurations and discuss the intrinsic bottlenecks of the present device architecture. Our results demonstrate a novel technique for the realization of fast subwavelength light sources with possible applications in sensing and microscopy beyond the diffraction limit.

Immune response in peripheral axons delays disease progression in SOD1G93A mice.

BACKGROUND: Increasing evidence suggests that the immune system has a beneficial role in the progression of amyotrophic lateral sclerosis (ALS) although the mechanism remains unclear. Recently, we demonstrated that motor neurons (MNs) of C57SOD1G93A mice with slow disease progression activate molecules classically involved in the cross-talk with the immune system. This happens a lot less in 129SvSOD1G93A mice which, while expressing the same amount of transgene, had faster disease progression and earlier axonal damage. The present study investigated whether and how the immune response is involved in the preservation of motor axons in the mouse model of familial ALS with a more benign disease course. METHODS: First, the extent of axonal damage, Schwann cell proliferation, and neuromuscular junction (NMJ) denervation were compared between the two ALS mouse models at the disease onset. Then, we compared the expression levels of different immune molecules, the morphology of myelin sheaths, and the presence of blood-derived immune cell infiltrates in the sciatic nerve of the two SOD1G93A mouse strains using immunohistochemical, immunoblot, quantitative reverse transcription PCR, and rotating-polarization Coherent Anti-Stokes Raman Scattering techniques. RESULTS: Muscle denervation, axonal dysregulation, and myelin disruption together with reduced Schwann cell proliferation are prominent in 129SvSOD1G93A compared to C57SOD1G93A mice at the disease onset, and this correlates with a faster disease progression in the first strain. On the contrary, a striking increase of immune molecules such as CCL2, MHCI, and C3 was seen in sciatic nerves of slow progressor C57SOD1G93A mice and this was accompanied by heavy infiltration of CD8+ T lymphocytes and macrophages. These phenomena were not detectable in the peripheral nervous system of fast-progressing mice. CONCLUSIONS: These data show for the first time that damaged MNs in SOD1-related ALS actively recruit immune cells in the peripheral nervous system to delay muscle denervation and prolong the lifespan. On the contrary, the lack of this response has a negative impact on the disease course.

RP-CARS: label-free optical readout of the myelin intrinsic healthiness.

Here we present a method based on Rotating-Polarization Coherent Anti-Stokes Raman Scattering (RP-CARS) imaging to assess the myelin health status in mouse sciatic nerves. Differently from the existing techniques, our method is based on the readout of intrinsic molecular architecture rather than on the image analysis, relying on the fact that healthy myelin is characterized by a high degree of molecular order. We exploit RP-CARS imaging to demonstrate that the degree of spatial anisotropy of the CARS signal displays a strong correlation with the g-ratio (a well-known image-based index of myelin damage) in a chemical-damage model and therefore that the former is a good indicator for the local myelin health status.

Rotating-polarization CARS microscopy: combining chemical and molecular orientation sensitivity.

Coherent Anti-Stokes Raman Spectroscopy (CARS) is a non-linear process in which the energy difference of a pair of incoming photons matches the energy of the vibrational mode of a molecular bond of interest. This phonon population is coherently probed by a third photon and anti-Stokes radiation is emitted. Here a novel approach to CARS microscopy is presented yielding the intensity of the anti-Stokes emission, the directionality the molecular bonds of interest, and their average orientation. Myelinated axons in fixed mouse-brain slices have been imaged by RP-CARS. We were able to detect the local average direction of the acylic chains of membrane phospholipids and their spatial anisotropy. This novel method may impact the study of healthy brain circuitry as well as demyelinating diseases or other pathological states associated with altered neural connectivity.

Total mesorectal excision laparoscopic versus transanal approach for rectal cancer: A systematic review and meta-analysis.

INTRODUCTION: Total mesorectal excision (TME) performed for the first time by Held through an open approach, it has become the standard technique for the surgical treatment of rectal cancer. The aim the of this meta-analysis is to compare the outcomes provided by TaTME than LaTME. MATERIAL AND METHODS: In this meta-analysis, we included all comparative studies, prospective and retrospective, which addressed in low and middle rectal cancer, a comparison between TaTME and LaTME. A search was performed through MEDLINE and Cochrane Database. 846 records were identified. RESULTS: Eight relevant studies have been included in this meta-analysis. The studies were from France, Russia, USA, Netherlands, Taiwan, Egypt. The eight studies including 471 patients with middle or low rectal cancer. CONCLUSION: The meta-analysis confirmed safety of TaTME for low and middle rectal cancer. TaTME can lead to a high quality of rectal cancer resection specimen.

Short-term outcomes after laparoscopic right hemicolectomy for colon cancer: intracorporeal versus extracorporeal anastomosis.

BACKGROUND: The aim of this study was to compare the short-term outcomes of laparoscopic right hemicolectomy between intracorporeal (IA) and extracorporeal (EA) anastomosis techniques. METHODS: A retrospective chart review of 77 consecutive patients who underwent laparoscopic right hemicolectomy from January 2016 to June 2018 was performed. The intracorporeal group included 36 patients and the extracorporeal group 41 patients. RESULTS: Patient demographics and disease-related characteristics were similar. Mean operative time was 152±38.3 minutes in IA vs. 148±34.8 minutes in EA (P=0.664). IA was not associated with less overall postoperative complications and decreased rate of surgical site infections. There was no statistically significant difference in the incidence of postoperative leak, length of stay, mortality in both groups. There was no perioperative difference in median number of lymph node harvested. There was statistically significant difference in the length of larger incision (50 [50-70] mm in IA vs. 80 [70-100] mm in EA; P<0.001), length of the specimen (323±52.7 mm in IA vs. 295±64.7 mm in EA; P<0.05), free tumor margin (113.6±47.4 in IA vs. 75.6±37.4 mm in EA; P<0.001). There was a statistically significant difference in IA in the less postoperative use of analgesics (P<0.032) and return of bowel function (P<0.021). CONCLUSIONS: IA seems feasible and safe; it does not significantly affect the length of surgery, and it guarantees maintenance of radical oncological standards. Furthermore, IA significantly improves cosmesis and patient comfort postoperatively, reducing the rates of emesis, which leads to higher rates of early regular diet tolerance. Therefore, laparoscopic right hemicolectomy with intracorporeal anastomosis is associated with improved short-term outcomes.

RP-CARS reveals molecular spatial order anomalies in myelin of an animal model of Krabbe disease.

Krabbe disease (KD) is a rare demyelinating sphingolipidosis, often fatal in the first years of life. It is caused by the inactivation of the galactocerebrosidase (GALC) enzyme that causes an increase in the cellular levels of psychosine considered to be at the origin of the tissue-level effects. GALC is inactivated also in the Twitcher (TWI) mouse: a genetic model of KD that is providing important insights into the understating of the pathogenetic process and the development of possible treatments. In this article an innovative optical technique, RP-CARS, is proposed as a tool to study the degree of order of the CH2 bonds inside the myelin sheaths of TWI-mice sciatic-nerve fibres. RP-CARS, a recently developed variation of CARS microscopy, is able to combine the intrinsic chemical selectivity of CARS microscopy with molecular-bond-spatial-orientation sensibility. This is the first time RP-CARS is applied to the study of a genetic model of a pathology, leading to the demonstration of a post-onset progressive spatial disorganization of the myelin CH2 bonds. The presented result could be of great interest for a deeper understanding of the pathogenic mechanisms underlying the human KD and, moreover, it is an additional proof of the experimental validity of this microscopy technique. RP-CARS image (2850 cm-1 , CH2 bonds) of a sciatic-nerve optical longitudinal section from a Twitcher P23 (symptomatic) mouse. Scale bar: 10 microns. The image was constructed by colour-mapping the degree of molecular order of the CH2 bonds inside the myelin walls, as displayed in the colour bar on the right.

Ionic Strength Responsive Sulfonated Polystyrene Opals.

Stimuli-responsive photonic crystals (PCs) represent an intriguing class of smart materials very promising for sensing applications. Here, selective ionic strength responsive polymeric PCs are reported. They are easily fabricated by partial sulfonation of polystyrene opals, without using toxic or expensive monomers and etching steps. The color of the resulting hydrogel-like ordered structures can be continuously shifted over the entire visible range (405-760 nm) by changing the content of ions over an extremely wide range of concentration (from about 70 µM to 4 M). The optical response is completely independent from pH and temperature, and the initial color can be fully recovered by washing the sulfonated opals with pure water. These new smart photonic materials could find important applications as ionic strength sensors for environmental monitoring as well as for healthcare screening.

Femtosecond-Laser-Pulse Characterization and Optimization for CARS Microscopy.

We present a simple method and its experimental implementation to determine the pulse durations and linear chirps of the pump-and-probe pulse and the Stokes pulse in a coherent anti-Stokes Raman scattering microscope at sample level without additional autocorrelators. Our approach exploits the delay line, ubiquitous in such microscopes, to perform a convolution of the pump-and-probe and Stokes pulses as a function of their relative delay and it is based on the detection of the photons emitted from an appropriate non-linear sample. The analysis of the non-resonant four-wave-mixing and sum-frequency-generation signals allows for the direct retrieval of the pulse duration on the sample and the linear chirp of each pulse. This knowledge is crucial in maximizing the spectral-resolution and contrast in CARS imaging.

The Role of Water in the Preparation and Stabilization of High-Quality Phosphorene Flakes.

The stability of phosphorene is shown to be degraded by water. However, the presence of a small amount of water allows the synthesis of high-quality material in liquid exfoliation of black phosphorus using dimethylsulfoxide as solvent. A phosphorus/water molar ratio between 1.5 and 14 maximizes the quality of the phosphorene flakes and their stability.

Ultrastructural Characterization of the Lower Motor System in a Mouse Model of Krabbe Disease.

Krabbe disease (KD) is a neurodegenerative disorder caused by the lack of ß- galactosylceramidase enzymatic activity and by widespread accumulation of the cytotoxic galactosyl-sphingosine in neuronal, myelinating and endothelial cells. Despite the wide use of Twitcher mice as experimental model for KD, the ultrastructure of this model is partial and mainly addressing peripheral nerves. More details are requested to elucidate the basis of the motor defects, which are the first to appear during KD onset. Here we use transmission electron microscopy (TEM) to focus on the alterations produced by KD in the lower motor system at postnatal day 15 (P15), a nearly asymptomatic stage, and in the juvenile P30 mouse. We find mild effects on motorneuron soma, severe ones on sciatic nerves and very severe effects on nerve terminals and neuromuscular junctions at P30, with peripheral damage being already detectable at P15. Finally, we find that the gastrocnemius muscle undergoes atrophy and structural changes that are independent of denervation at P15. Our data further characterize the ultrastructural analysis of the KD mouse model, and support recent theories of a dying-back mechanism for neuronal degeneration, which is independent of demyelination.

Nanostructured ultra-thin patches for ultrasound-modulated delivery of anti-restenotic drug.

This work aims to demonstrate the possibility to fabricate ultra-thin polymeric films loaded with an anti-restenotic drug and capable of tunable drug release kinetics for the local treatment of restenosis. Vascular nanopatches are composed of a poly(lactic acid) supporting membrane (thickness: ~250 nm) on which 20 polyelectrolyte bilayers (overall thickness: ~70 nm) are alternatively deposited. The anti-restenotic drug is embedded in the middle of the polyelectrolyte structure, and released by diffusion mechanisms. Nanofilm fabrication procedure and detailed morphological characterization are reported here. Barium titanate nanoparticles (showing piezoelectric properties) are included in the polymeric support and their role is investigated in terms of influence on nanofilm morphology, drug release kinetics, and cell response. Results show an efficient drug release from the polyelectrolyte structure in phosphate-buffered saline, and a clear antiproliferative effect on human smooth muscle cells, which are responsible for restenosis. In addition, preliminary evidences of ultrasound-mediated modulation of drug release kinetics are reported, thus evaluating the influence of barium titanate nanoparticles on the release mechanism. Such data were integrated with quantitative piezoelectric and thermal measurements. These results open new avenues for a fine control of local therapies based on smart responsive materials.

Age-related changes in the function and structure of the peripheral sensory pathway in mice.

This study is aimed at describing the changes occurring in the entire peripheral nervous system sensory pathway along a 2-year observation period in a cohort of C57BL/6 mice. The neurophysiological studies evidenced significant differences in the selected time points corresponding to childhood, young adulthood, adulthood, and aging (i.e., 1, 7, 15, and 25 months of age), with a parabolic course as function of time. The pathological assessment allowed to demonstrate signs of age-related changes since the age of 7 months, with a remarkable increase in both peripheral nerves and dorsal root ganglia at the subsequent time points. These changes were mainly in the myelin sheaths, as also confirmed by the Rotating-Polarization Coherent-Anti-stokes-Raman-scattering microscopy analysis. Evident changes were also present at the morphometric analysis performed on the peripheral nerves, dorsal root ganglia neurons, and skin biopsies. This extensive, multimodal characterization of the peripheral nervous system changes in aging provides the background for future mechanistic studies allowing the selection of the most appropriate time points and readouts according to the investigation aims.