Roberta Ruffilli

Lipid Droplets: A New Player in Colorectal Cancer Stem Cells Unveiled by Spectroscopic Imaging

The cancer stem cell (CSC) model is describing tumors as a hierarchical organized system and CSCs are suggested to be responsible for cancer recurrence after therapy. The identification of specific markers of CSCs is therefore of paramount importance. Here, we show that high levels of lipid droplets (LDs) are a distinctive mark of CSCs in colorectal (CR) cancer. This increased lipid content was clearly revealed by label‐free Raman spectroscopy and it directly correlates with well‐accepted CR‐CSC markers as CD133 and Wnt pathway activity. By xenotransplantation experiments, we have finally demonstrated that CR‐CSCs overexpressing LDs retain most tumorigenic potential. A relevant conceptual advance in this work is the demonstration that a cellular organelle, the LD, is a signature of CSCs, in addition to molecular markers. A further functional characterization of LDs could lead soon to design new target therapies against CR‐CSCs. Stem Cells 2015;33:35–44

Surface morphology and mechanical properties of new-generation flowable resin composites for dental restoration

OBJECTIVES The purpose of this study was to characterize the surface morphology and the elastic properties of four dental restorative flowable composites currently on the market (Venus Diamond Flow, Vertise Flow, Filtex Supreme XT Flow, Surefil SDR Flow). Additionally, one adhesive system (Adhese One F) and one non-flowable composite (Venus Diamond) have also been characterized as the control materials. METHODS Surface morphology was studied by both scanning electron and atomic force microscopy, and the elastic modulus and the hardness measured by instrumented indentation. Grain analysis was performed on the microscopic images, and statistical analysis was carried out on the results of the nanoindentation measurements. RESULTS It was observed that Vertise, Filtek XT and Surefil SDR exhibit stiffness similar to the non-flowable Venus Diamond, whereas Venus Diamond Flow presents itself as the more compliant flowable composite, with Adhese showing intermediate stiffness. Grain analysis of the images confirmed the general rule that the mechanical properties improve with increasing filler loading, with the notable exception of Vertise Flow that shows modulus and hardness as high as 9.1±0.6 and 0.43±0.03GPa, respectively, for an estimated loading of only ∼40% by volume. SIGNIFICANCE Whereas generally flowable composites are confirmed not to possess sufficiently strong mechanical properties for bulk restorations, exceptions can eventually be found upon appropriate laboratory screening, as presently seems to be the case for Vertise Flow. However, real practice in actual restorations and respective clinical evaluation are required for final assessment of the suggested results.

Nanostructured superhydrophobic substrates trigger the development of 3D neuronal networks

The generation of 3D networks of primary neurons is a big challenge in neuroscience. Here, a novel method is presented for a 3D neuronal culture on superhydrophobic (SH) substrates. How nano-patterned SH devices stimulate neurons to build 3D networks is investigated. Scanning electron microscopy and confocal imaging show that soon after plating neurites adhere to the nanopatterned pillar sidewalls and they are subsequently pulled between pillars in a suspended position. These neurons display an enhanced survival rate compared to standard cultures and develop mature networks with physiological excitability. These findings underline the importance of using nanostructured SH surfaces for directing 3D neuronal growth, as well as for the design of biomaterials for neuronal regeneration.

Superhydrophobic nanocomposites from biodegradable thermoplastic starch composites (Mater-Bi®), hydrophobic nano-silica and lycopodium spores

Due to increasing environmental concerns related to bio-persistence of petroleum based polymers, research efforts have intensified towards developing biodegradable materials with equivalent performance properties. Among these properties liquid repellence is one of the most challenging to obtain due to the fact that most biopolymers such as polysaccharides or bio-polyesters are hygroscopic or hydrophilic. One possible approach is to formulate biopolymer nanocomposites with water repellent properties. Here, we demonstrate such an approach using a commercial thermoplastic starch composite, Materi-Bi®, hydrophobic fumed silica nanoparticles and lycopodium spores. We found that both hydrophobic fumed silica and Materi-Bi® have excellent colloidal stability and solubility in chloroform and they can be spray coated onto various surfaces like paper, metals and semiconductors. By varying the concentration ratio of biopolymer to hydrophobic fumed silica, the wetting properties of the nanocomposites could be tuned. Superhydrophobic nanocomposites displayed raspberry-like surface roughness with water contact angles exceeding 160° with very low water droplet roll-off angles (∼1°). On the other hand, composites of Materi-Bi® and lycopodium spores displayed sticky superhydrophobicity (rose petal effect). Superhydrophobic nanocomposites were found to withstand thermal aging at 250 °C without loss of properties. Due to the resultant micro-morphology of the nanocomposites, certain coatings were rendered superoleophobic by functionalizing with a dilute fluoroacrylic polymer solution in acetone. Oil droplet contact angles reached 166° with droplet roll-off angles of approximately 15°.

Self-Cleaning Organic/Inorganic Photo-Sensors

We present the fabrication of a multifunctional, hybrid organic-inorganic micropatterned device, which is capable to act as a stable photosensor and, at the same time, displaying inherent superhydrophobic self-cleaning wetting characteristics. In this framework several arrays of epoxy photoresist square micropillars have been fabricated on n-doped crystalline silicon substrates and subsequently coated with a poly(3-hexylthiophene-2,5-diyl) (P3HT) layer, giving rise to an array of organic/inorganic p-n junctions. Their photoconductivity has been measured under a solar light simulator at different illumination intensities. The current-voltage (I-V) curves show high rectifying characteristics, which are found to be directly correlated with the illumination intensity. The photoresponse occurs in extremely short times (within few tens of milliseconds range). The influence of the interpillar distance on the I-V characteristics of the sensors is also discussed. Moreover, the static and dynamic wetting properties of these organic/inorganic photosensors can be easily tuned by changing the pattern geometry. Measured static water contact angles range from 125° to 164°, as the distance between the pillars is increased from 14 to 120 μm while the contact angle hysteresis decreases from 36° down to 2°.

Biomimetic Approach for Liquid Encapsulation with Nanofibrillar Cloaks

Technologies that are able to handle microvolumes of liquids, such as microfluidics and liquid marbles, are attractive for applications that include miniaturized biological and chemical reactors, sensors, microactuators, and drug delivery systems. Inspired from natural fibrous envelopes, here, we present an innovative approach for liquid encapsulation and manipulation using electrospun nanofibers. We demonstrated the realization of non-wetting soft solids consisting of a liquid core wrapped in a hydrophobic fibrillar cloak of a fluoroacrylic copolymer and cellulose acetate. By properly controlling the wetting and mechanical properties of the fibers, we created final architectures with tunable mechanical robustness that were stable on a wide range of substrates (from paper to glass) and floated on liquid surfaces. Remarkably, the realized fiber-coated drops endured vortex mixing in a continuous oil phase at high stirring speed without bursting or water losses, favoring mixing processes inside the entrapped liquid volume. Moreover, the produced cloak can be easily functionalized by incorporating functional particles, active molecules, or drugs inside the nanofibers.

Mucuna pruriens (Velvet bean) Rescues Motor, Olfactory, Mitochondrial and Synaptic Impairment in PINK1B9 Drosophila melanogaster Genetic Model of Parkinson’s Disease

The fruit fly Drosophila melanogaster (Dm) mutant for PTEN-induced putative kinase 1 (PINK1B9) gene is a powerful tool to investigate physiopathology of Parkinsons disease (PD). Using PINK1B9 mutant Dm we sought to explore the effects of Mucuna pruriens methanolic extract (Mpe), a L-Dopa-containing herbal remedy of PD. The effects of Mpe on PINK1B9 mutants, supplied with standard diet to larvae and adults, were assayed on 3–6 (I), 10–15 (II) and 20–25 (III) days old flies. Mpe 0.1% significantly extended lifespan of PINK1B9 and fully rescued olfactory response to 1-hexanol and improved climbing behavior of PINK1B9 of all ages; in contrast, L-Dopa (0.01%, percentage at which it is present in Mpe 0.1%) ameliorated climbing of only PINK1B9 flies of age step II. Transmission electron microscopy analysis of antennal lobes and thoracic ganglia of PINK1B9 revealed that Mpe restored to wild type (WT) levels both T-bars and damaged mitochondria. Western blot analysis of whole brain showed that Mpe, but not L-Dopa on its own, restored bruchpilot (BRP) and tyrosine hydroxylase (TH) expression to age-matched WT control levels. These results highlight multiple sites of action of Mpe, suggesting that its effects cannot only depend upon its L-Dopa content and support the clinical observation of Mpe as an effective medication with intrinsic ability of delaying the onset of chronic L-Dopa-induced long-term motor complications. Overall, this study strengthens the relevance of using PINK1B9 Dm as a translational model to study the properties of Mucuna pruriens for PD treatment.

Complex architectures formed by alginate drops floating on liquid surfaces

We demonstrate the generation of natural polymeric structures of complex shapes and controlled composition, starting from the collision of aqueous drops of alginate with the surface of a calcium ion-based liquid. We prove that by tuning the impact velocity of the alginate drops on the target surface one can control the floating state of the drops inducing the formation of mushroom-like structures, upon alginate gelation. Besides the geometric peculiarity, the presented approach allows us to provide dual functionality to the polymeric objects, attaching different kinds of functional molecules onto their surface areas, which are immersed or not in the liquid, making such architectures attractive for the development of a novel class of bionanocomposites.

Impaired sense of smell in a Drosophila Parkinson's model.

Parkinson’s disease (PD) is one of the most common neurodegenerative disease characterized by the clinical triad: tremor, akinesia and rigidity. Several studies have suggested that PD patients show disturbances in olfaction at the earliest onset of the disease. The fruit fly Drosophila melanogaster is becoming a powerful model organism to study neurodegenerative diseases. We sought to use this system to explore olfactory dysfunction, if any, in PINK1 mutants, which is a model for PD. PINK1 mutants display many important diagnostic symptoms of the disease such as akinetic motor behavior. In the present study, we describe for the first time, to the best of our knowledge, neurophysiological and neuroanatomical results concerning the olfactory function in PINK1 mutant flies. Electroantennograms were recorded in response to synthetic and natural volatiles (essential oils) from groups of PINK1 mutant adults at three different time points in their life cycle: one from 3–5 day-old flies, from 15–20 and from 27–30 days. The results obtained were compared with the same age-groups of wild type flies. We found that mutant adults showed a decrease in the olfactory response to 1-hexanol, α-pinene and essential oil volatiles. This olfactory response in mutant adults decreased even more as the flies aged. Immunohistological analysis of the antennal lobes in these mutants revealed structural abnormalities, especially in the expression of Bruchpilot protein, a marker for synaptic active zones. The combination of electrophysiological and morphological results suggests that the altered synaptic organization may be due to a neurodegenerative process. Our results indicate that this model can be used as a tool for understanding PD pathogensis and pathophysiology. These results help to explore the potential of using olfaction as a means of monitoring PD progression and developing new treatments.

Zwitterionic Nanofibers of Super-Glue for Transparent and Biocompatible Multi-Purpose Coatings

Here we show that macrozwitterions of poly(ethyl 2-cyanoacrylate), commonly called Super Glue, can easily assemble into long and well defined fibers by electrospinning. The resulting fibrous networks are thermally treated on glass in order to create transparent coatings whose superficial morphology recalls the organization of the initial electrospun mats. These textured coatings are characterized by low liquid adhesion and anti-staining performance. Furthermore, the low friction coefficient and excellent scratch resistance make them attractive as solid lubricants. The inherent texture of the coatings positively affects their biocompatibility. In fact, they are able to promote the proliferation and differentiation of myoblast stem cells. Optically-transparent and biocompatible coatings that simultaneously possess characteristics of low water contact angle hysteresis, low friction and mechanical robustness can find application in a wide range of technological sectors, from the construction and automotive industries to electronic and biomedical devices.