Evaluation of direct light processing for the fabrication of bioactive ceramic scaffolds: Effect of pore/strut size on manufacturability and mechanical performance

Bioactive ceramic scaffolds for bone regeneration consisting of a three-dimensional mesh of interpenetrating struts with square section were fabricated via Digital Light Processing (DLP). The ability of the technique to manufacture 3D porous structures from β-tricalcium phosphate (β-TCP) powders with different dimensions of struts and pores was evaluated, identifying the possibilities and limitations of the manufacturing process. Small pore sizes were found to seriously complicate the elimination of excess slurry from the scaffold’s innermost pores. The effect of the strut/pore size on the mechanical performance of the scaffolds under compressive stresses was also evaluated, but no significant influence was found. Under compressive stresses, the structures resulted weaker when tested perpendicularly to the printing plane due to interlayer shear failure. Interlayer superficial grooves are proposed as potential failure-controlling defects, which could also explain the lack of a Weibull size effect on the mechanical strength of the fabricated DLP scaffolds.     

The Age-Sensitive Efficacy of Calorie Restriction on Mitochondrial Biogenesis and mtDNA Damage in Rat Liver

Calorie restriction (CR) is the most efficacious treatment to delay the onset of age-related changes such as mitochondrial dysfunction. However, the sensitivity of mitochondrial markers to CR and the age-related boundaries of CR efficacy are not fully elucidated. We used liver samples from ad libitum-fed (AL) rats divided in: 18-month-old (AL-18), 28-month-old (AL-28), and 32-month-old (AL-32) groups, and from CR-treated (CR) 28-month-old (CR-28) and 32-month-old (CR-32) counterparts to assay the effect of CR on several mitochondrial markers. The age-related decreases in citrate synthase activity, in TFAM, MFN2, and DRP1 protein amounts and in the mtDNA content in the AL-28 group were prevented in CR-28 counterparts. Accordingly, CR reduced oxidative mtDNA damage assessed through the incidence of oxidized purines at specific mtDNA regions in CR-28 animals. These findings support the anti-aging effect of CR up to 28 months. Conversely, the protein amounts of LonP1, Cyt c, OGG1, and APE1 and the 4.8 Kb mtDNA deletion content were not affected in CR-28 rats. The absence of significant differences between the AL-32 values and the CR-32 counterparts suggests an age-related boundary of CR efficacy at this age. However, this only partially curtails the CR benefits in counteracting the generalized aging decline and the related mitochondrial involvement.     

Implication of the NLRP3 Inflammasome in Bovine Age-Related Sarcopenia

Sarcopenia is defined as the age-related loss of skeletal muscle mass, quality, and strength. The pathophysiological mechanisms underlying sarcopenia are still not completely understood. The aim of this work was to evaluate, for the first time, the expression of NLRP3 inflammasome in bovine skeletal muscle in order to investigate the hypothesis that inflammasome activation may trigger and sustain a pro-inflammatory environment leading to sarcopenia. Samples of skeletal muscle were collected from 60 cattle belonging to three age-based groups. Morphologic, immunohistochemical and molecular analysis were performed to assess the presence of age-related pathologic changes and chronic inflammation, the expression of NLRP3 inflammasome and to determine the levels of interleukin-1β, interleukin-18 and tumor necrosis factor alpha in muscle tissue. Our results revealed the presence of morphologic sarcopenia hallmark, chronic lymphocytic inflammation and a type II fibers-selective NLRP3 expression associated to a significant decreased number of immunolabeled-fibers in aged animals. Moreover, we found a statistically significant age-related increase of pro-inflammatory cytokines such as interleukin-1β and interleukin-18 suggesting the activation of NLRP3 inflammasome. Taken together, our data suggest that NLRP3 inflammasome components may be normally expressed in skeletal muscle, but its priming and activation during aging may contribute to enhance a pro-inflammatory environment altering normal muscular anabolism and metabolism.     

Maskless Preparation of Spatially-Resolved Plasmonic Nanoparticles on Polydimethylsiloxane via In Situ Fluoride-Assisted Synthesis

Here, a fluoride-assisted route for the controlled in-situ synthesis of metal nanoparticles (NPs) (i.e., AgNPs, AuNPs) on polydimethylsiloxane (PDMS) is reported. The size and coverage of the NPs on the PDMS surface are modulated with time and over space during the synthetic process, leveraging the improved yield (10×) and faster kinetics (100×) of NP formation in the presence of F⁻ ions, compared to fluoride-free approaches. This enables the maskless preparation of both linear and step gradients and patterns of NPs in 1D and 2D on the PDMS surface. As an application in flexible plasmonics/photonics, continuous and step-wise spatial modulations of the plasmonic features of PDMS slabs with 1D and 2D AgNP gradients on the surface are demonstrated. An excellent spatially resolved tuning of key optical parameters, namely, optical density from zero to 5 and extinction ratio up to 100 dB, is achieved with AgNP gradients prepared in AgF solution for 12 minutes; the performance are comparable to those of commercial dielectric/interference filters. When used as a rejection filter in optical fluorescence microscopy, the AgNP-PDMS slabs are able to reject the excitation laser at 405 nm and retain the green fluorescence of microbeads (100 µm) used as test cases.     

Recent Biophysical Issues About the Preparation of Solute-Filled Lipid Vesicles

Here we report some recent biophysical issues on the preparation of solute-filled lipid vesicles and their relevance to the construction of “synthetic cells.” First, we introduce the “semi-synthetic minimal cells” as the liposome-based cell-like systems, which contain a minimal number of biomolecules required to display simple and complex biological functions. Next, we focus on recent aspects related to the construction of synthetic cells. Emphasis is given to the interplay between the methods of synthetic cell preparation and the physics of solute encapsulation. We briefly introduce the notion of structural and compositional “diversity” in synthetic cell populations.     

In situ observation of synthesized nanoparticles in ultra-dilute aerosols via X-ray scattering

In-air epitaxy of nanostructures (Aerotaxy) has recently emerged as a viable route for fast, large-scale production. In this study, we use small-angle X-ray scattering to perform direct in-flight characterizations of the first step of this process, i.e., the engineered formation of Au and Pt aerosol nanoparticles by spark generation in a flow of N₂ gas. This represents a particular challenge for characterization because the particle density can be extremely low in controlled production. The particles produced are examined during production at operational pressures close to atmospheric conditions and exhibit a lognormal size distribution ranging from 5–100 nm. The Au and Pt particle production and detection are compared. We observe and characterize the nanoparticles at different stages of synthesis and extract the corresponding dominant physical properties, including the average particle diameter and sphericity, as influenced by particle sintering and the presence of aggregates. We observe highly sorted and sintered spherical Au nanoparticles at ultra-dilute concentrations (< 5 × 10⁵ particles/cm³) corresponding to a volume fraction below 3 × 10^–10, which is orders of magnitude below that of previously measured aerosols. We independently confirm an average particle radius of 25 nm via Guinier and Kratky plot analysis. Our study indicates that with high-intensity synchrotron beams and careful consideration of background removal, size and shape information can be obtained for extremely low particle concentrations with industrially relevant narrow size distributions.

     

2-Phenyloxazole-4-carboxamide as a Scaffold for Selective Inhibition of Human Monoamine Oxidase B

A series of 2-phenyloxazoles bearing an amide group at position 4 were designed and synthesized for evaluation as potential inhibitors of human recombinant monoamine oxidases (hrMAOs). Results of kinetics experiments demonstrated that all compounds behave as competitive MAO inhibitors, with good selectivity toward the MAO-B isoform. The most potent and selective derivatives are characterized by inhibition constant (K_i) values in the sub-micromolar range and a good selectivity index (K_{i MAO-A}/K_{i MAO-B}>50). Some derivatives were also found to be able to inhibit MAO activity in nerve growth factor (NGF)-differentiated PC12 cells, taken as a model of neuronal cells. In particular, 2-(2-hydroxyphenyl)-N-phenyloxazole-4-carboxamide (compound 4 a ) may be a promising new scaffold, exerting the highest selectivity and inhibitory effect toward MAOs in NGF-differentiated PC12 cell lysates, without compromising cell viability. Molecular docking analysis allowed a rationalization of the experimentally observed binding affinity and selectivity.     

Dihydrotanshinone,a Natural Diterpenoid,Preserves Blood-Retinal Barrier Integrity via P2X7 Receptor

Activation of P2X7 signaling, due to high glucose levels, leads to blood retinal barrier (BRB) breakdown, which is a hallmark of diabetic retinopathy (DR). Furthermore, several studies report that high glucose (HG) conditions and the related activation of the P2X7 receptor (P2X7R) lead to the over-expression of pro-inflammatory markers. In order to identify novel P2X7R antagonists, we carried out virtual screening on a focused compound dataset, including indole derivatives and natural compounds such as caffeic acid phenethyl ester derivatives, flavonoids, and diterpenoids. Molecular Mechanics/Generalized Born Surface Area (MM/GBSA) rescoring and structural fingerprint clustering of docking poses from virtual screening highlighted that the diterpenoid dihydrotanshinone (DHTS) clustered with the well-known P2X7R antagonist JNJ47965567. A human-based in vitro BRB model made of retinal pericytes, astrocytes, and endothelial cells was used to assess the potential protective effect of DHTS against HG and 2′(3′)-O-(4-Benzoylbenzoyl)adenosine-5′-triphosphate (BzATP), a P2X7R agonist, insult. We found that HG/BzATP exposure generated BRB breakdown by enhancing barrier permeability (trans-endothelial electrical resistance (TEER)) and reducing the levels of ZO-1 and VE-cadherin junction proteins as well as of the Cx-43 mRNA expression levels. Furthermore, HG levels and P2X7R agonist treatment led to increased expression of pro-inflammatory mediators (TLR-4, IL-1β, IL-6, TNF-α, and IL-8) and other molecular markers (P2X7R, VEGF-A, and ICAM-1), along with enhanced production of reactive oxygen species. Treatment with DHTS preserved the BRB integrity from HG/BzATP damage. The protective effects of DHTS were also compared to the validated P2X7R antagonist, JNJ47965567. In conclusion, we provided new findings pointing out the therapeutic potential of DHTS, which is an inhibitor of P2X7R, in terms of preventing and/or counteracting the BRB dysfunctions elicited by HG conditions.     

Evaluation of the effect of processing on cocoa polyphenols: antiradical activity,anthocyanins and procyanidins profiling from raw beans to chocolate

The content and composition of anthocyanins and procyanidins in fermented cocoa beans (from different geographic origins: Ecuador, Cameroon, Ivory Coast, Ghana and Nigeria), roasted nibs, cocoa mass and chocolate were determined, beside the determination of the total antiradical capacity. Concerning geographic origin, cocoa beans and processed products from Ecuador showed the highest levels of anthocyanins, followed by Nigeria and Cameroon. Generally, as cocoa beans were further processed, the levels of anthocyanins and flavan‐3‐ols decreased. The largest observed losses of phenolics occurred during roasting. A progressive decreasing trend in polyphenol concentration was observed in the other processed samples as well. Despite the original content of polyphenols in raw cocoa beans, technological processes imply a significant impact on cocoa quality, confirming the need of specific optimisation to obtain high value chocolate.