Orazio Vittorio

Physicochemical properties affecting cellular uptake of carbon nanotubes

Carbon nanotubes (CNTs) are widely used for biomedical applications as intracellular transporters of biomolecules owing to their ability to cross cell membranes. In this article, we survey the reported literature and results of our published work in an attempt to provide a rational view of the various CNT internalization mechanisms. Essentially three uptake mechanisms (phagocytosis, diffusion and endocytosis) have been reported in the literature. In addressing the subject of cellular internalization of CNTs, the unique physicochemical characteristics of CNTs that influence and drive the cell uptake pathway are considered. According to available evidence, the degree of dispersion, the formation of supramolecular complexes and the nanotube length are crucial factors in determining the exact mechanism of cellular uptake. In conclusion, phagocytosis appears to be the internalization pathway for CNT aggregates, bundles, cluster or single dispersed nanotubes 1 microm or more in length; endocytosis is the internalization mechanism for nanotubes forming supramolecular structures; and diffusion is the internalization mechanism for submicron CNTs that do not form supramolecular complexes. This information may be relevant to the rational design of CNT-based carriers for cell therapy.

Influence of purity and surface oxidation on cytotoxicity of multiwalled carbon nanotubes with human neuroblastoma cells

UNLABELLED There are conflicting data concerning the safety and biocompatibility of carbon nanotubes (CNTs). In some reports CNTs have been used for gene delivery without significant toxicity, whereas in others various cytotoxic effects were observed, including induction of intracellular reactive oxygen species (ROS), DNA damage, and apoptosis. Although it is clear that CNT production methods, purity, and functionalization treatments impact on biocompatibility, most of the published reports lack detailed characterization of the CNT samples used. We investigated the effect of various physicochemical features of multiwalled carbon nanotubes (MWCNTs) on toxicity and biocompatibility with cultured human neuroblastoma cells by using MTT, WST-1, Hoechst, and oxidative stress assays. In vitro experiments confirm that after 3 days of incubation with three different types of CNTs dispersed in Pluronic F127 solution, 0.01% cell viability is not affected and apoptosis and ROS are not induced in the SH-SY5Y cells. With prolonged cultures and continued propagation in the presence of MWCNTs, the loss of cell viability was minimal for pure MWCNTs (99% purity), but cell proliferation decreased significantly for 97% purity MWCNTs and acid-treated MWCNTs (97% purity, surface oxidation 8%); no intracellular ROS were detected. When the concentration of CNTs increases, purity and surface oxidation seem to affect cell viability (ED(25) is 48, 34.4, and 18.4 mug/mL, respectively, for 99% purity MWCNTs, 97% purity MWCNTs, and acid-treated 97% purity MWCNTs. Our results indicate that concentrations of 5-10 mug/mL MWCNTs seem ideal for studies on the design and development of artificial MWCNT nanovectors for gene and drug therapy against cancer. FROM THE CLINICAL EDITOR With prolonged cultures, loss of cell viability was minimal for preparations with 99% purity, but significant adverse effects were detected with 97% purity and with acid-treated preparations. A concentrations of 5-10 mug/mL of MWCNTs seems ideal for gene and drug therapy against cancer.

Dextran-Catechin Conjugate: A Potential Treatment Against the Pancreatic Ductal Adenocarcinoma

ABSTRACTPurposeA polysaccharide-flavonoid conjugate was developend and proposed for the treatment of pancreatic ductal adenocarcinoma (PDAC).MethodsThe conjugate was synthesized by free radical grafting reaction between catechin and dextran. The chemical characterization of the conjugate was obtained by UV-Vis, 1H-NMR, FT-IR and GPC analyses, while the functionalization degree was determined by the Folin-Ciocalteu assay. The biological activity of the catechin-dextran conjugate was tested on two different cell lines derived from human pancreatic cancer (MIA PaCa-2 and PL45 cells), and the toxicity towards human pancreatic nestin-expressing cells evaluated.ResultsBoth the cancer cell lines are killed when exposed to the conjugate, and undergo apoptosis after the incubation with catechin-dextran which resulted more effective in killing pancreatic tumor cells compared to the catechin alone. Moreover, our experimental data indicate that the conjugate was less cytotoxic to human pancreatic nestin-expressing cells which are considered a good model of non-neoplastic pancreatic cells.ConclusionThe suitability of newly synthesized Dextran-Catechin conjugate in the treatment of PDAC was proved confirming the high potential application of the proposed macromolecula system in the cancer therapy.

Enhancing the therapeutic effects of polyphenols with macromolecules

Well-known for their antioxidant properties, polyphenols are naturally occurring compounds containing one or more phenol groups. A high dietary intake of polyphenols has been linked to a reduced incidence of a number of diseases, including cancer, cardiovascular diseases, diabetes, osteoporosis and neurodegenerative diseases. Furthermore, in vitro and in vivo studies show the potential for polyphenols as therapeutic agents with cardioprotective, antimicrobial, anticancer, neuroprotective, and antidiabetic effects demonstrated. Despite their impressive therapeutic effects, polyphenols suffer from a number of drawbacks, including instability when exposed to light, heat and basic conditions; poor bioavailability; rapid metabolism; and poor membrane permeability. These drawbacks limit the clinical applications of polyphenols. Polymers and other macromolecules are well-known for their ability to stabilise and improve the bioavailability of therapeutic agents. A number of macromolecular systems have been developed that stabilise polyphenols whilst enhancing their therapeutic effects, including direct polymerisation of polyphenol monomers via step-growth, free radical and enzyme catalysed reactions; and conjugation with macromolecules via enzyme grafting, free-radical grafting, esterification and amidation. In this review, we will detail the key techniques employed to stabilise polyphenols with macromolecules and provide examples of each technique.

Progress in nanotechnology for healthcare

Abstract This review based on the Wickham lecture given by AC at the 2009 SMIT meeting in Sinaia outlines the progress made in nano-technology for healthcare. It describes in brief the nature of nano-materials and their unique properties which accounts for the significant research both in scientific institutions and industry for translation into new therapies embodied in the emerging field of nano-medicine. It stresses that the potential of nano-medicine to make significant inroads for more effective therapies both for life-threatening and life-disabling disorders will only be achieved by high-quality life science research. The first generation of passive nano-diagnostics based on nanoparticle contrast agents for magnetic resonance imaging is well established in clinical practice and new such contrast agents are undergoing early clinical evaluation. Likewise active (second generation) nano-therapies, exemplified by targeted control drug release systems are undergoing early clinical evaluation. The situation concerning other nano-materials such as carbon nanotubes (CNTs) and boron nitride nanotubes (BNNTs) is less advanced although considerable progress has been made on their coating for aqueous dispersion and functionalisation to enable carriage of drugs, genes and fluorescent markers. The main problem related to the clinical use of these nanotubes is that there is no consent among scientists on the fate of such nano-materials following injection or implantation in humans. Provided carbon nanotubes are manufactured to certain medical criteria (length around 1 μm, purity of 97–99% and low Fe content) they exhibit no cytotoxicity on cell cultures and demonstrate full bio-compatibility on in vivo animal studies. The results of recent experimental studies have demonstrated the potential of technologies based on CNTs for low voltage wireless electro-chemotherapy of tumours and for electro-stimulation therapies for cardiac, neurodegenerative and skeletal and visceral muscle disorders.

Incorporation of carbon nanotubes into a gelatin–catechin conjugate: Innovative approach for the preparation of anticancer materials

A new hybrid material made of gelatin, catechin and carbon nanotubes was prepared by the non-covalent incorporation of carbon nanotubes (CNTs) into a gelatin-catechin covalent conjugate. The composite materials was tested by means of determination of the dispersion stability in water and the functionalization degree was assessed by the Folin-Ciocalteu method, finding a 0.9 mg of CT/g of protein conjugate. Subsequently, the complete retention of the antioxidant properties of the flavonoid after incorporation into the composite was proved by DPPH and ABTS assays and IC50 values of 5.74 mg mL(-1) and 0.39 mg mL(-1) were recorded. The presence of CNT into the materials did not interfere with the scavenging activities of the catechin. Finally, the anticancer activity on HeLa cancer cells was evaluated and a considerable increase in the therapeutic activity of the flavonoid was recorded moving from the free to the conjugated form in the presence of CNT, while in absence of CNT a reduction of the efficiency was observed.

Multiwall carbon nanotubes as MRI contrast agents for tracking stem cells

In this study we investigate the potential of multiwall carbon nanotubes (MWCNTs) with low metal impurities (2.57% iron) as magnetic resonance imaging (MRI) contrast agents. Taking into account probable aggregation at high MWCNTs concentration analysis shows that the r(2) relaxivity of MWCNTs in 1% agarose gels at 19 °C is 564 ± 41 s(-1) mM(-1); this is attributed to both the presence of iron oxide impurities and also to the carbon MWCNT structure itself. Stem cells were labelled with MWCNTs to demonstrate the effectiveness of MWCNTs as MRI contrast agents for cellular MRI. The MWCNTs did not impair cell viability or proliferation. These results suggest that the MRI contrast agent properties of the MWCNTs could be used in vivo for stem cell tracking/imaging and during MWCNT-mediated targeted electro-chemotherapy of tumours.

Polyphenol Conjugates and Human Health: A Perspective Review

In recent years, antioxidants have gained great importance because of their potential use in food, pharmaceutical, and cosmetic industries. This interest is rooted in the cumulative evidence connecting active oxygen and free radicals with numerous human degenerative disorders, such as cardiovascular diseases, cancer, aging, and atherosclerosis. Polyphenols are the major class of antioxidant able to reduce the oxidative damages of lipids, proteins, enzymes, carbohydrates, and DNA in living cells and tissues. Among the realm of polyphenol compounds, polyphenol conjugates have been proposed as innovative materials which, by combining the advantageous properties of both the components, can increase the efficiency of antioxidants and their range of application in nutritional and biomedical fields. This work is an overview of the different class of polyphenol conjugates, which will be analyzed in terms of nutritional and biological properties, showing how these bio-conjugates will positively affect the human health.

Magnetic carbon nanotubes: a new tool for shepherding mesenchymal stem cells by magnetic fields.

AIMS We investigated the interaction between magnetic carbon nanotubes (CNTs) and mesenchymal stem cells (MSCs), and their ability to guide these intravenously injected cells in living rats by using an external magnetic field. MATERIALS & METHODS Multiwalled CNTs were used to treat MSCs derived from rat bone marrow. Cytotoxicity induced by nanotubes was studied using the WST-1 proliferation and Hoechest 33258 apoptosis assays. The effects of nanotubes on MSCs were evaluated by monitoring the effects on cellular growth rates, immunophenotyping and differentiation, and on the arrangement of cytoskeletal actin. MSCs loaded with nanotubes were injected in vivo in the portal vein of rats driving their localization in the liver by magnetic field. An histological analysis was performed on the liver, lungs and kidneys of all animals. RESULTS CNTs did not affect cell viability and their ability to differentiate in osteocytes and adipocytes. Both the CNTs and the magnetic field did not alter the cell growth rate, phenotype and cytoskeletal conformation. CNTs, when exposed to magnetic fields, are able to shepherd MSCs towards the magnetic source in vitro. Moreover, the application of a magnetic field alters the biodistribution of CNT-labelled MSCs after intravenous injection into rats, increasing the accumulation of cells into the target organ (liver). CONCLUSION Multiwalled CNTs hold the potential for use as nanodevices to improve therapeutic protocols for transplantation and homing of stem cells in vivo. This could pave the way for the development of new strategies for the manipulation/guidance of MSCs in regenerative medicine and cell transplantation.

Cancer phototherapy in living cells by multiphoton release of doxorubicin from gold nanospheres

Doxorubicin is a widely used but toxic cancer chemotherapeutic agent. In order to localize its therapeutic action and minimize side effects, it was covalently conjugated to peptide-encapsulated gold nanospheres by click-chemistry and then photo-released in a controlled fashion by a multiphoton process. Selective treatment of a chosen region in a 2D layer of U2Os cancer cells is shown by driving photorelease with 561 nm irradiation at μW power. These results show promising directions for the development of practical applications based on nanocarriers that can ensure drug delivery with high spatial and temporal control.