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In this paper, we describe the effects of the combination of topographical, mechanical, chemical and intracellular electrical stimuli on a co-culture of fibroblasts and skeletal muscle cells. The co-culture was anisotropically grown onto an engineered micro-grooved (10 µm-wide grooves) polyacrylamide substrate, showing a precisely tuned Young’s modulus (∼ 14 kPa) and a small thickness (∼ 12 µm). We enhanced the co-culture properties through intracellular stimulation produced by piezoelectric nanostructures (i.e., boron nitride nanotubes) activated by ultrasounds, thus exploiting the ability of boron nitride nanotubes to convert outer mechanical waves (such as ultrasounds) in intracellular electrical stimuli, by exploiting the direct piezoelectric effect. We demonstrated that nanotubes were internalized by muscle cells and localized in both early and late endosomes, while they were not internalized by the underneath fibroblast layer. Muscle cell differentiation benefited from the synergic combination of topographical, mechanical, chemical and nanoparticle-based stimuli, showing good myotube development and alignment towards a preferential direction, as well as high expression of genes encoding key proteins for muscle contraction (i.e., actin and myosin). We also clarified the possible role of fibroblasts in this process, highlighting their response to the above mentioned physical stimuli in terms of gene expression and cytokine production. Finally, calcium imaging-based experiments demonstrated a higher functionality of the stimulated co-cultures. [ABSTRACT FROM AUTHOR]

Current medically relevant metals for prosthetic reconstructions enjoy a relatively good success rate, but their performance drops significantly in patients with compromised health status, and post-surgical infections still remain an important challenge. To address these problems, different nanotechnology-based strategies have been exploited to create implantable metals with an enhanced bioactivity and antibacterial capacities. Among these, oxidative nanopatterning has emerged as a very effective approach to engender nanoporous surfaces that stimulate and guide the activity of adhering cells. The resulting nanoporosity is also attractive because it offers nanoconfined volumes that can be exploited to load bioactive compounds and modulate their release over time. Such extended elution is needed since a single exposure to growth factors and/or antibiotics, for instance, may not be adequate to further sustain bone regeneration and/or to counteract bacterial colonization. In this article, we assessed the capacities of nanoporous titanium surfaces generated by oxidative nanopatterning to provide controlled and sustained elution of proteins and antibiotic molecules. To this end, we have selected bovine serum albumin (BSA) and vancomycin to reflect commonly used compounds, and investigated their adsorption and elution by Fourier-transform infrared (FT-IR) and ultraviolet–visible (UV-VIS) spectroscopy. Our results demonstrate that while the elution of albumin is not significantly affected by the nanoporosity, in the case of vancomycin, nanoporous surfaces provided an extended release. These findings were successively correlated to the establishment of interactions with the surface and physical-entrapment effects exerted by the nanopores, ultimately highlighting their synergistic contribution to the release profiles and thus their importance in the design of nanostructured eluting platforms for applications in medicine. [ABSTRACT FROM AUTHOR]

A novel supersaturated self-emulsifying drug delivery system (Super-SEDDS) loaded with scutellarin-phospholipid complex (SPC) was developed. The system aimed to address the limitations presented by conventional SEDDS as delivery carriers for drugs with poor water-solubility, low liposolubility and high dose. As an intermediate, SPC was first prepared based on the response surface design. The presence of amorphous scutellarin was demonstrated through differential scanning calorimetry (DSC) and X-ray diffraction (XRD), while enhanced liposolubility was confirmed through comparison with scutellarin powder via an octanol/water distribution test. On the basis of the solubility study and ternary phase diagram, Super-SEDDS containing SPC of up to 200% equilibrium solubility (Seq) was designed, which composed of ethyl oleate, Cremophor RH40 and Transcutol HP with a ratio of 60∶25∶15 (w/w%). The subsequent in vitro lipolysis study and ex vivo intestinal absorption test indicated that Super-SEDDS enhanced the cumulative dissolution from 70% to 100% and improved the intestinal absorption from 0.04 to 0.12 µg/cm2 compared with scutellarin powder. Furthermore, an in vivo study demonstrated that Super-SEDDS achieved the AUC0-t of scutellarin up to approximate 1.7-fold as scutellarin powder. It was also proved superior to SPC and the conventional SEDDS. Super-SEDDS showed great potential for expanding the usage of SEDDS and could act as an alternative to conventional SEDDS. [ABSTRACT FROM AUTHOR]

The Hedgehog (Hh) pathway inhibitors have shown great promise in cancer therapeutics. SANT75, a novel compound we previously designed to specially inhibit the Smoothened (SMO) protein in the Hh pathway, has greater inhibitory potency than many of commonly used Hh inhibitors. However, preclinical studies of SANT75 revealed water insolubility and acute toxicity. To overcome these limitations, we developed a liposomal formulation of SANT75 and investigated its antitumor efficacy in vitro and in vivo. We encapsulated SANT75 into PEGylated liposome and the mean particle size distribution and zeta-potential (ZP) of liposomes were optimized. Using the Shh-light2 cell and Gli-GFP or Flk-GFP transgenic reporter zebrafish, we confirmed that liposome-encapsulated SANT75 inhibited Hh activity with similar potency as the original SANT75. SANT75 encapsulated into liposome exerted strong tumor growth-inhibiting effects in vitro and in vivo. In addition, the liposomal SANT75 therapy efficiently improved the survival time of tumor-bearing mice without obvious systemic toxicity. The pathological morphology and immunohistochemistry staining revealed that liposomal SANT75 induced tumor cell apoptosis, inhibited tumor angiogenesis as assessed by CD31 and down-regulated the expression of Hh target protein Gli-1 in tumor tissues. Our findings suggest that liposomal formulated SANT75 has improved solubility and bioavailability and should be further developed as a drug candidate for treating tumors with abnormally high Hh activity. [ABSTRACT FROM AUTHOR]

Nanostructured three-dimensional constructs combining layer-by-layer technology (LbL) and template leaching were processed and evaluated as possible support structures for cartilage tissue engineering. Multilayered constructs were formed by depositing the polyelectrolytes chitosan (CHT) and chondroitin sulphate (CS) on either bidimensional glass surfaces or 3D packet of paraffin spheres. 2D CHT/CS multi-layered constructs proved to support the attachment and proliferation of bovine chondrocytes (BCH). The technology was transposed to 3D level and CHT/CS multi-layered hierarchical scaffolds were retrieved after paraffin leaching. The obtained nanostructured 3D constructs had a high porosity and water uptake capacity of about 300%. Dynamical mechanical analysis (DMA) showed the viscoelastic nature of the scaffolds. Cellular tests were performed with the culture of BCH and multipotent bone marrow derived stromal cells (hMSCs) up to 21 days in chondrogenic differentiation media. Together with scanning electronic microscopy analysis, viability tests and DNA quantification, our results clearly showed that cells attached, proliferated and were metabolically active over the entire scaffold. Cartilaginous extracellular matrix (ECM) formation was further assessed and results showed that GAG secretion occurred indicating the maintenance of the chondrogenic phenotype and the chondrogenic differentiation of hMSCs. [ABSTRACT FROM AUTHOR]