In this study, graphene oxide (GO) and reduced-thiolated GO (rGOSH) were used as 2D substrate to fabricate nanocomposites with nanoparticles of gold nanospheres (AuNS) or nanorods (AuNR), via in situ ...reduction of the metal salt precursor and seed-mediated growth processes. The plasmonic sensing capability of the gold-decorated nanosheets were scrutinized by UV-visible (UV-VIS) spectroscopy. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), thermogravimetric analyses (TGA), and atomic force microscopy (AFM) were performed in order to prove the actual reduction that occurred concomitantly with the thiolation of GO, the increase in the hydrophobic character as well as the size, and preferential gathering of the gold nanoparticles onto the nanosheet substrates, respectively. Moreover, the theoretical electronic and infrared absorption (UV-VIS and IR) spectra were calculated within a time-dependent approach of density functional theory (DFT). Eventually, in vitro cellular experiments on human neuroblastoma cells (SH-SY5Y line) were carried out in order to evaluate the nanotoxicity of the nanocomposites by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide tetrazolium reduction (MTT) colorimetric assay. Results pointed out the promising potential of these hybrids as plasmonic theranostic platforms with different hydrophilic or hydrophobic features as well as cytotoxic effects against cancer cells.
Angiogenin (ANG), an endogenous protein that plays a key role in cell growth and survival, has been scrutinised here as promising nanomedicine tool for the modulation of pro-/anti-angiogenic ...processes in brain cancer therapy. Specifically, peptide fragments from the putative cell membrane binding domain (residues 60-68) of the protein were used in this study to obtain peptide-functionalised spherical gold nanoparticles (AuNPs) of about 10 nm and 30 nm in optical and hydrodynamic size, respectively. Different hybrid biointerfaces were fabricated by peptide physical adsorption (Ang
) or chemisorption (the cysteine analogous Ang
Cys) at the metal nanoparticle surface, and cellular assays were performed in the comparison with ANG-functionalised AuNPs. Cellular treatments were performed both in basal and in copper-supplemented cell culture medium, to scrutinise the synergic effect of the metal, which is another known angiogenic factor. Two brain cell lines were investigated in parallel, namely tumour glioblastoma (A172) and neuron-like differentiated neuroblastoma (d-SH-SY5Y). Results on cell viability/proliferation, cytoskeleton actin, angiogenin translocation and vascular endothelial growth factor (VEGF) release pointed to the promising potentialities of the developed systems as anti-angiogenic tunable nanoplaftforms in cancer cells treatment.
In this work, we aimed to develop a hybrid theranostic nano-formulation based on gold nanoparticles (AuNP)-having a known anti-angiogenic character-and the angiogenin (ANG), in order to tune the ...angiogenesis-related phases involved in the multifaceted process of the wound healing. To this purpose, spherical were surface "decorated" with three variants of the protein, namely, the recombinant (rANG), the wild-type, physiologically present in the human plasma (wtANG) and a new mutant with a cysteine substitution of the serine at the residue 28 (S28CANG). The hybrid biointerface between AuNP and ANG was scrutinized by a multi-technique approach based on dynamic light scattering, spectroscopic (UV-visible, circular dichroism) and microscopic (atomic force and laser scanning confocal) techniques. The analyses of optical features of plasmonic gold nanoparticles allowed for discrimination of different adsorption modes-i.e.; predominant physisorption and/or chemisorption-triggered by the ANG primary sequence. Biophysical experiments with supported lipid bilayers (SLB), an artificial model of cell membrane, were performed by means of quartz crystal microbalance with dissipation monitoring acoustic sensing technique. Cellular experiments on human umbilical vein endothelial cells (HUVEC), in the absence or presence of copper-another co-player of angiogenesis-were carried out to assay the nanotoxicity of the hybrid protein-gold nanoassemblies as well as their effect on cell migration and tubulogenesis. Results pointed to the promising potential of these nanoplatforms, especially the new hybrid Au-S28CANG obtained with the covalent grafting of the mutant on the gold surface, for the modulation of angiogenesis processes in wound care.
Physicochemical properties of the graphene-water interface have been investigated to scrutinize the perturbations with respect to the graphene-air interface, in terms of changes in optical and ...vibrational spectra, as well as in the 3D network of water. Experimental investigations were carried out using Raman spectroscopy and laser scanning confocal microscopy, and integrated with density functional theory (DFT) calculations. Results evidence a substantial orientation of the hydrogen-bonded water molecules at the interfacial region, which, in turn, induces disorder in the water clusters and interfacial charge transfer phenomena.
Water-induced graphene p-doping and preferential orientation of hydrogen-bonded water at the interface with graphene are demonstrated by comparative experimental (Raman spectroscopy and confocal microscopy) and theoretical (DFT) investigations.
The adsorption process of albumin, lysozyme and lactoferrin was investigated onto polymer surfaces, both hydrophobic and hydrophilic treated by oxygen–plasma. In particular thin films of ...polyhydroxymethylsiloxane (about 90° of static water contact angle) were converted by oxygen plasma treatments at reduced pressure into hydrophilic SiO
x
phases (less than 10° of water contact angle). The protein adsorption process was investigated in situ by quartz crystal microbalance with dissipation monitoring in terms of coverage and kinetics mechanism, while chemical structure and topography of the protein adlayers were studied ex situ by angular-resolved XPS and atomic force microscopy, respectively. The plasma surface modification of the polymer film drastically modified the adsorption process of the three proteins, both in terms of kinetics and coverage.
Graphene oxide (GO) nanosheets with different content in the defective carbon species bound to oxygen sp3 were functionalized with the angiogenin (ANG) protein, to create a novel nanomedicine for ...modulating angiogenic processes in cancer therapies. The GO@ANG nanocomposite was scrutinized utilizing UV-visible and fluorescence spectroscopies. GO exhibits pro- or antiangiogenic effects, mostly attributed to the disturbance of ROS concentration, depending both on the total concentration (i.e., >100 ng/mL) as well as on the number of carbon species oxidized, that is, the C/O ratio. ANG is considered one of the most effective angiogenic factors that plays a vital role in the angiogenic process, often in a synergic role with copper ions. Based on these starting hypotheses, the GO@ANG nanotoxicity was assessed with the MTT colorimetric assay, both in the absence and in the presence of copper ions, by in vitro cellular experiments on human prostatic cancer cells (PC-3 line). Laser confocal microscopy (LSM) cell imaging evidenced an enhanced internationalization of GO@ANG than bare GO nanosheets, as well as significant changes in cell cytoskeleton organization and mitochondrial staining compared to the cell treatments with free ANG.
Inhibitors of PARP-1(Poly(ADP-ribose) polymerase-1) act by competing with NAD+, the enzyme physiological substrate, which play a protective role in many pathological conditions characterized by ...PARP-1 overactivation. It has been shown that PARP-1 also promotes tumor growth and progression through its DNA repair activity. Since angiogenesis is an essential requirement for these activities, we sought to determine whether PARP inhibition might affect rat brain microvascular endothelial cells (GP8.3) migration, stimulated by C6-glioma conditioned medium (CM). Through wound-healing experiments and MTT analysis, we demonstrated that PARP-1 inhibitor PJ-34 N-(6-Oxo-5,6-dihydrophenanthridin-2-yl)-N,N-dimethylacetamide abolishes the migratory response of GP8.3 cells and reduces their viability. PARP-1 also acts in a DNA independent way within the Extracellular-Regulated-Kinase (ERK) signaling cascade, which regulates cell proliferation and differentiation. By western analysis and confocal laser scanning microscopy (LSM), we analyzed the effects of PJ-34 on PARP-1 expression, phospho-ERK and phospho-Elk-1 activation. The effect of MEK (mitogen-activated-protein-kinase-kinase) inhibitor PD98059 (2-(2-Amino-3-methoxyphenyl)-4 H-1-benzopyran-4-one) on PARP-1 expression in unstimulated and in CM-stimulated GP8.3 cells was analyzed by RT-PCR. PARP-1 expression and phospho-ERK activation were significantly reduced by treatment of GP8.3 cells with PJ-34 or PD98059. By LSM, we further demonstrated that PARP-1 and phospho-ERK are coexpressed and share the same subcellular localization in GP8.3 cells, in the cytoplasm as well as in nucleoplasm. Based on these data, we propose that PARP-1 and phospho-ERK interact in the cytosol and then translocate to the nucleus, where they trigger a proliferative response. We also propose that PARP-1 inhibition blocks CM-induced endothelial migration by interfering with ERK signal-transduction pathway.
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In this work, nanobiohybrids of plasmonic gold nanoparticles (AuNP, anti-angiogenic) and a peptide mimicking the vascular endothelial growth factor (VEGF, pro-angiogenic) were assembled and ...scrutinized in terms of physicochemical characterization, including optical properties, surface charge, surface chemical structure and morphology of the bioengineered metal nanoparticles, for their potential application as multifunctional theranostic (i.e., therapy + sensing) nanoplatform (AuNP/VEGF). Specifically, a peptide sequence encompassing the VEGF cellular receptor domain 73–101 (VEGF73–101) and its single point cysteine mutated were immobilized onto AuNP by physi- and chemi-sorption, respectively. The new hybrid systems were characterized by means of a multitechnique approach, including dynamic light scattering (DLS) analyses, zeta potential (ZP), spectroscopic (UV-Vis, FT-IR, XPS), spectrometric (TOF-SIMS) and microscopic (AFM, SEM) techniques. Proof-of-work cellular experiments in human umbilical vein endothelial cells (HUVEC) upon the treatment with AuNP/VEGF samples, demonstrated no toxicity up to 24 h (MTT assay) as well an effective internalization (laser confocal microscopy, LSM).
Nerve growth factor (NGF) is a protein necessary for development and maintenance of the sympathetic and sensory nervous systems. We have previously shown that the NGF N-terminus peptide NGF(1-14) is ...sufficient to activate TrkA signaling pathways essential for neuronal survival and to induce an increase in brain-derived neurotrophic factor (BDNF) expression. Cu
ions played a critical role in the modulation of the biological activity of NGF(1-14). Using computational, spectroscopic, and biochemical techniques, here we report on the ability of a newly synthesized peptide named d-NGF(1-15), which is the dimeric form of NGF(1-14), to interact with TrkA. We found that d-NGF(1-15) interacts with the TrkA-D5 domain and induces the activation of its signaling pathways. Copper binding to d-NGF(1-15) stabilizes the secondary structure of the peptides, suggesting a strengthening of the noncovalent interactions that allow for the molecular recognition of D5 domain of TrkA and the activation of the signaling pathways. Intriguingly, the signaling cascade induced by the NGF peptides ultimately involves cAMP response element-binding protein (CREB) activation and an increase in BDNF protein level, in keeping with our previous result showing an increase of BDNF mRNA. All these promising connections can pave the way for developing interesting novel drugs for neurodegenerative diseases.
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