We exploit Surface-Enhanced Raman Scattering (SERS) to investigate aqueous droplets of genomic DNA deposited onto silver-coated silicon nanowires, and we show that it is possible to efficiently discriminate between spectra of tumoral and healthy cells. To assess the robustness of the proposed technique, we develop two different statistical approaches, one based on the Principal Components Analysis of spectral data and one based on the computation of the `2 distance between spectra. Both methods prove to be highly efficient, and we test their accuracy via the Cohen's ? statis- tics. We show that the synergistic combination of the SERS spectroscopy and the statistical analysis methods leads to efficient and fast cancer diagnostic applications allowing rapid and unexpansive discrimination between healthy and tumoral genomic DNA alternative to the more complex and expensive DNA sequencing

This article demonstrates the possibility to use a novel powerful approach based on Raman mapping of analyte solutions drop casted on a disordered array of Ag covered silicon nanowires (Ag/SiNWs), to identify the characteristic spectral signal of the four DNA bases, adenine (A), thymine (T), cytosine (C), and guanine (G), at concentration as low as 10 ng/?L, and to study their specific way of interacting with the nanostructured substrate. The results show a distinctive and amplified interaction of guanine, the base that is most susceptible to oxidation, with the nanostructured surface. Our findings explain the recently revealed diverse behaviour of cancer and normal DNA deposited on the same Ag/SiNWs, which is ascribed to mechanical deformation and base lesions present on the oxidised DNA molecule backbone and causes detectable variation in the Raman signal, usable for diagnostic purposes. The notable bio-analytical capability of the presented platform, and its sensitivity to the molecule mechanical conformation at the single-base level, thus provides a new reliable, rapid, label-free DNA

Genomic deoxyribonucleic acid (DNA) stores and carries the information required to maintain and replicate cellular life. While much efforts have been devoted in decoding the sequence of DNA basis to detect the genetic mutations related to cancer disease, it is becoming clear that physical properties, like structural conformation, stiffness and shape, can play an important role to recognize DNA modifications. Here, silver-coated silicon nanowires (Ag/SiNWs) are exploited as Raman spectroscopic platform to easily discriminate healthy and cancer genomic DNA, extracted from human normal skin and malignant melanoma cells, respectively. In particular, aqueous DNA droplets are directly deposited onto a forest of Ag/SiNWs and Raman maps are acquired after sample dehydration. By applying principal component analysis (PCA) to the Raman spectra collected within the droplets, healthy and cancer cell DNA can be distinguished without false negative identifications and with few false positive results (< 2%). The discrimination occurs regardless the analysis of specific DNA sequencing, but through Raman bands strictly related to the interfacing of the DNA and the NWs. The observed phenomenon can be ascribed to conformational differences and/or diverse charge properties between healthy and cancer cell DNA determining a different arrangement of the molecules adsorbed onto the NWs upon water evaporation. The unique interaction with DNA and facile fabrication technology make Ag/SiNWs an effective platform for a robust, rapid and label-free cancer diagnosis, as well as a potential tool to investigate physical properties of DNA.

Ultrasmall superparamagnetic iron oxide nanoparticles with a size <5 nm are emerging nanomaterials for their excellent biocompatibility, chemical stability, and tunable surface modifications. The applications explored include dual-modal or multi-modal imaging, drug delivery, theranostics and, more recently, magnetic resonance angiography. Good biocompatibility and biosafety are regarded as the preliminary requirements for their biomedical applications and further exploration in this field is still required. We previously synthesized and characterized ultrafine (average core size of 3 nm) silica-coated superparamagnetic iron oxide fluorescent nanoparticles, named sub-5 SIO-Fl, uniform in size, shape, chemical properties and composition. The cellular uptake and in vitro biocompatibility of the as-synthesized nanoparticles were demonstrated in a human colon cancer cellular model. Here, we investigated the biocompatibility of sub-5 SIO-Fl nanoparticles in human Amniotic Mesenchymal Stromal/Stem Cells (hAMSCs). Kinetic analysis of cellular uptake showed a quick nanoparticle internalization in the first hour, increasing over time and after long exposure (48 h), the uptake rate gradually slowed down. We demonstrated that after internalization, sub-5 SIO-Fl nanoparticles neither affect hAMSC growth, viability, morphology, cytoskeletal organization, cell cycle progression, immunophenotype, and the expression of pro-angiogenic and immunoregulatory paracrine factors nor the osteogenic and myogenic differentiation markers. Furthermore, sub-5 SIO-Fl nanoparticles were intravenously injected into mice to investigate the in vivo biodistribution and toxicity profile for a time period of 7 weeks. Our findings showed an immediate transient accumulation of nanoparticles in the kidney, followed by the liver and lungs, where iron contents increased over a 7-week period. Histopathology, hematology, serum pro-inflammatory response, body weight and mortality studies demonstrated a short- and long-term biocompatibility and biosafety profile with no apparent acute and chronic toxicity caused by these nanoparticles in mice. Overall, these results suggest the feasibility of using sub-5 SIO-Fl nanoparticles as a promising agent for stem cell magnetic targeting as well as for diagnostic and therapeutic applications in oncology.

Ultrasmall superparamagnetic iron oxide nanoparticles with a size <5 nm are emerging nanomaterials for their excellent biocompatibility, chemical stability, and tunable surface modifications. The applications explored include dual-modal or multi-modal imaging, drug delivery, theranostics and, more recently, magnetic resonance angiography. Good biocompatibility and biosafety are regarded as the preliminary requirements for their biomedical applications and further exploration in this field is still required. We previously synthesized and characterized ultrafine (average core size of 3 nm) silica-coated superparamagnetic iron oxide fluorescent nanoparticles, named sub-5 SIO-Fl, uniform in size, shape, chemical properties and composition. The cellular uptake and in vitro biocompatibility of the as-synthesized nanoparticles were demonstrated in a human colon cancer cellular model. Here, we investigated the biocompatibility of sub-5 SIO-Fl nanoparticles in human Amniotic Mesenchymal Stromal/Stem Cells (hAMSCs). Kinetic analysis of cellular uptake showed a quick nanoparticle internalization in the first hour, increasing over time and after long exposure (48 h), the uptake rate gradually slowed down. We demonstrated that after internalization, sub-5 SIO-Fl nanoparticles neither affect hAMSC growth, viability, morphology, cytoskeletal organization, cell cycle progression, immunophenotype, and the expression of pro-angiogenic and immunoregulatory paracrine factors nor the osteogenic and myogenic differentiation markers. Furthermore, sub-5 SIO-Fl nanoparticles were intravenously injected into mice to investigate the in vivo biodistribution and toxicity profile for a time period of 7 weeks. Our findings showed an immediate transient accumulation of nanoparticles in the kidney, followed by the liver and lungs, where iron contents increased over a 7-week period. Histopathology, hematology, serum pro-inflammatory response, body weight and mortality studies demonstrated a short- and long-term biocompatibility and biosafety profile with no apparent acute and chronic toxicity caused by these nanoparticles in mice. Overall, these results suggest the feasibility of using sub-5 SIO-Fl nanoparticles as a promising agent for stem cell magnetic targeting as well as for diagnostic and therapeutic applications in oncology.

Cell therapy represents the promising alternative strategy for end-stage liver disease, and hepatic progenitors are the best candidates. The possibility to maximize the paracrine effects of transplanted cells represents a great potential benefit for cell therapy success. We studied how cell type and microenvironment modulate the Wnt/?-catenin signaling in vitro and in vivo. In vitro, the onset of hepatocyte commitment was characterized by the presence of nuclear truncated ?-catenin. In vivo, we analyzed the effect of human hepatic progenitors on damage recovery and functional regeneration in a mouse model of acute liver injury, either in combination or in absence of a selected mix of hepatogenic factors. Animals injected with human hepatic progenitors and hepatogenic factors showed improved engraftment triggering the Wnt/?-catenin signaling cascade. Human hepatic progenitors expressing the human oval cell marker OV6 displayed a consistent co-localization with ?-catenin and co-localized with Wnt1 main ligand of the canonical pathway. Wnt5a, on the contrary, was expressed in distinct liver cell populations. Epithelial mesenchymal transition-related markers showed enhanced expression and wider distribution and the hepato-mesenchymal population Thy1+CK19- was also present. Control animals injected with hepatogenic factors alone exhibited higher ?-catenin and decreased Wnt5a levels, and persistent proliferation of the hepato-mesenchymal population. In conclusion, the combination of human hepatic progenitors with selected hepatogenic factors creates a positive synergy with local microenvironment, ameliorates cell engraftment, stimulates and accelerates regenerative process and improves the rescue of hepatic function, by modulating the Wnt/?catenin signaling and activating hepato-mesenchymal population.

Cell therapy is an innovative strategy for tissue repair, since adult stem cells could have limited regenerative ability as in the case of myocardial damage. This leads to a local contractile dysfunction due to scar formation. For these reasons, refining strategy approaches for "in vitro" stem cell commitment, preparatory to the "in vivo" stem cell differentiation, is imperative. In this work, we isolated and characterized at molecular and cellular level, human Amniotic Mesenchymal Stromal Cells (hAMSCs) and exposed them to a physical Extremely Low Frequency Electromagnetic Field (ELF-EMF) stimulus and to a chemical Nitric Oxide treatment. Physically exposed cells showed a decrease of cell proliferation and no change in metabolic activity, cell vitality and apoptotic rate. An increase in the mRNA expression of cardiac and angiogenic differentiation markers, confirmed at the translational level, was also highlighted in exposed cells. Our data, for the first time, provide evidence that physical ELF-EMF stimulus (7 Hz, 2.5 µT), similarly to the chemical treatment, is able to trigger hAMSC cardiac commitment. More importantly, we also observed that only the physical stimulus is able to induce both types of commitments contemporarily (cardiac and angiogenic), suggesting its potential use to obtain a better regenerative response in cell-therapy protocols

Stem cell therapy is a regenerative medicine technique consisting in introducing new cells into a damaged tissue in order to restore it. In this paper we present our multidisciplinary approach about the possible use of the Human Mesenchymal Stem Cells in treating heart failure. A multidisciplinary team composed by biologists, mathematicians, cardiologists and bioengineers was selected in order to investigate the feasibility of repairing the necrotic tissue damaged by myocardial infarction by means of Mesenchymal Stem Cells and to implement a new software simulator able to reproduce cells implantation, migration and proliferation. In the first step, biologists and cardiologists studied the Human Mesenchymal Stem Cells and measured selected parameters. Stem cells were isolated and cultured in order to study their growth and characterization. In the second step, mathematicians and bioengineers developed a new numerical model based on the studies and the data measured during in vitro experiments. The first version of the software simulator named MiStTher is able to give a first qualitative description of the stem cell therapy in some simplified schemes.

Photothermal therapy (PTT) assisted by nanomaterials is a promising minimally invasive technique for cancer treatment. Here, we explore the PTT properties of a silicon- and gold-based nanostructured platform suitable for being directly integrated in fibre laser systems rather than injected into the human body, which occurs for the most commonly unreported PTT nanoagents. In particular, the photothermal properties of an array of disordered silicon nanowires coated by a thin gold film (Au/SiNWs) were tested on a monolayer of human colon adenocarcinoma cells (Caco-2) irradiated with a 785 nm laser. Au/SiNWs allowed an efficient photothermal action and simultaneous monitoring of the process evolution through the Raman signal coming from the irradiated cellular zone. Strong near infra-red (NIR) absorption, overlapping three biological windows, cell-friendly properties and effective fabrication technology make Au/SiNWs suitable both to be integrated in surgical laser tools and as an in vitro platform to develop novel PTT protocols using different cancer types and NIR sources.

Cell therapy is an innovative strategy for tissue repair, since adult stem cells could have limited regenerative ability as in the case of myocardial damage. This leads to a local contractile dysfunction due to scar formation. For these reasons, refining strategy approaches for "in vitro" stem cell commitment, preparatory to the "in vivo" stem cell differentiation, is imperative. In this work, we isolated and characterized at molecular and cellular level, human Amniotic Mesenchymal Stromal Cells (hAMSCs) and exposed them to a physical Extremely Low Frequency Electromagnetic Field (ELF-EMF) stimulus and to a chemical Nitric Oxide treatment. Physically exposed cells showed a decrease of cell proliferation and no change in metabolic activity, cell vitality and apoptotic rate. An increase in the mRNA expression of cardiac and angiogenic differentiation markers, confirmed at the translational level, was also highlighted in exposed cells. Our data, for the first time, provide evidence that physical ELF-EMF stimulus (7 Hz, 2.5 µT), similarly to the chemical treatment, is able to trigger hAMSC cardiac commitment. More importantly, we also observed that only the physical stimulus is able to induce both types of commitments contemporarily (cardiac and angiogenic), suggesting its potential use to obtain a better regenerative response in cell-therapy protocols

Several beneficial effects of the electromagnetic information transfer through aqueous system (EMITTAS) procedure have previously been reported in vitro. The clinical potential of this procedure has also started to be evaluated. Information flow in biological systems can be investigated through chemical and molecular approaches or by a biophysical approach focused on endogenous electrodynamic activities. Electromagnetic signals are endogenously generated at different levels of the biological organization and, likely, play an active role in synchronizing internal cell function or local/ systemic adaptive response. Consequently, each adaptive response can be described by its specific electromagnetic pattern and, therefore, correlates with a unique and specific electromagnetic signature. A biophysical procedure synchronously integrating the EMITTAS procedure has already been applied for the treatment of articular pain, low-back pain, neck pain and mobility, fluctuating asymmetry, early-stage chronic kidney disease, refractory gynecological infections, minor anxiety and depression disorders. This clinical strategy involves a single treatment, since the EMITTAS procedure allows the patient to continue his/ her own personal treatment at home by means of self-administration of the recorded aqueous system. A significant and long-lasting improvement has been reported, showing a potential beneficial use of this biophysical procedure in the management of common illnesses in an efficient, effective and personalized way. Data from recent studies suggest that aqueous systems may play a key role in providing the basis for recording, storing, transferring and retrieving clinically effective quanta of biological information. These features likely enable to trigger local and systemic self-regulation and self-regeneration potential of the organism.

Nanoparticles (NPs) made up of components between 1 nm and 100 nm in size and specifically magnetic iron oxide nanoparticles (IONPs), approved by Food and Drug Administration (FDA), have been extensively studied and have attracted much interest for their intriguing properties employable in a wide range of biomedical applications . NPs are used for diagnosis, prevention and treatment of diseases as much as for tissue engineering and regenerative medicine applications. These implementations demand the cross communication among different disciplines for the success of new therapies in restoring and regenerating the normal function of damaged cells, organs and tissues. The scientific rationale for the present multidisciplinary study is suggested by the need to design innovative and safe strategies to deal with human diseases. We synthetized and characterized ultrafine 3 nm superparamagnetic water-dispersible nanoparticles, prepared by an "arrested precipitation strategy". By a facile and inexpensive one-pot approach, nanoparticles were coated with silica to prevent their degradation/aggregation and to increase their surface functionalization, and contemporarily labelled with fluorescein isothiocyanate (FITC) dye to visualize their intracellular localization. The resulting new sub-5 nm silica-coated magnetic iron oxide fluorescent (sub-5 SIO-Fl) nanoparticles were tested in CaCo-2 cell line, a well characterized model of the intestinal epithelium, commonly used for biopharmaceutical evaluations as well in toxicity studies either as differentiated or undifferentiated cells. We studied sub-5 SIO-Fl nanoparticles cellular uptake and intracellular localization. Furthermore, we investigated if their uptake affected CaCo-2 cell morphology, growth, viability, cell cycle distribution, as well as transcriptional, translational and secretory activities, in a dose-dependent manner. To further shed light on their biocompatibility, the effect of the sub-5 SIO-Fl nanoparticles on CaCo-2 cell differentiation and pro-inflammatory response was analysed. Overall, these results showed the in vitro biocompatibility of the sub-5 SIO-Fl nanoparticles promising their safe employ for diagnostic and therapeutic biomedical applications. Since their magnetic nature, our nanoparticles could be easily in vivo directed toward the desired tissues/organs to shuttle drugs upon the application of an external static magnetic field. They could be used as efficient vehicles for drug/gene delivery for antiblastic therapies, enhancing the efficacy of treatments with reduced systemic toxicity. Moreover, these nanoparticles can maintain the ability to act as antennae in an external alternating magnetic field to convert electromagnetic energy into heat, to synergize the action of the shuttled drugs with hyperthermia.

Magnetic iron oxide nanoparticles (IONPs), for their intriguing properties, have attracted a great interest as they can be employed in many different biomedical applications. In this multidisciplinary study, we synthetized and characterized ultrafine 3 nm superparamagnetic water-dispersible nanoparticles. By a facile and inexpensive one-pot approach, nanoparticles were coated with a shell of silica and contemporarily functionalized with fluorescein isothiocyanate (FITC) dye. The obtained sub-5 nm silica-coated magnetic iron oxide fluorescent (sub-5 SIO-Fl) nanoparticles were assayed for cellular uptake, biocompatibility and cytotoxicity in a human colon cancer cellular model. By confocal microscopy analysis we demonstrated that nanoparticles as-synthesized are internalized and do not interfere with the CaCo-2 cell cytoskeletal organization nor with their cellular adhesion. We assessed that they do not exhibit cytotoxicity, providing evidence that they do not affect shape, proliferation, cellular viability, cell cycle distribution and progression. We further demonstrated at molecular level that these nanoparticles do not interfere with the expression of key differentiation markers and do not affect pro-inflammatory cytokines response in Caco-2 cells. Overall, these results showed the in vitro biocompatibility of the sub-5 SIO-Fl nanoparticles promising their safe employ for diagnostic and therapeutic biomedical applications.

Heretofore only observed in living systems, we report that weak-field ion cyclotron resonance (ICR) also occurs in inanimate matter. Weak magnetic field (50 nT) hydronium ICR at the field combination (7.84 Hz, 7.5 mu T) markedly changes water structure, as evidenced by finding an altered index of refraction exactly at this combined field. This observation utilizes a novel technique which measures the scattering of a He-Ne laser beam as the sample is exposed to a ramped magnetic field frequency. In addition to the hydronium resonance, we find evidence of ICR coupling to a more massive structure, possibly a tetrahedral combination of three waters and a single hydronium ion. To check our observations, we extended this technique to D2O, successfully predicting the specific ICR charge-to-mass ratio for D3O+ that alters the index of refraction.

In tissue engineering protocols, the survival of transplanted stem cells is a limiting factor that could be overcome using a cell delivery matrix able to support cell proliferation and differentiation. With this aim, we studied the cell-friendly and biocompatible behavior of RKKP glass-ceramic coated Titanium (Ti) surface seeded with human amniotic mesenchymal stromal cells (hAMSCs) from placenta. The sol-gel synthesis procedure was used to prepare the RKKP glass-ceramic material, which was then deposited onto the Ti surface by Pulsed Laser Deposition method. The cell metabolic activity and proliferation rate, the cytoskeletal actin organization, and the cell cycle phase distribution in hAMSCs seeded on the RKKP coated Ti surface revealed no significant differences when compared to the cells grown on the treated plastic Petri dish. The health of of hAMSCs was also analysed studying the mRNA expressions of MSC key genes and the osteogenic commitment capability using qRT-PCR analysis which resulted in being unchanged in both substrates. In this study, the combination of the hAMSCs' properties together with the bioactive characteristics of RKKP glass-ceramics was investigated and the results obtained indicate its possible use as a new and interesting cell delivery system for bone tissue engineering and regenerative medicine applications.

In tissue engineering protocols, the survival of transplanted stem cells is a limiting factor that could be overcome using a cell delivery matrix able to support cell proliferation and differentiation. With this aim, using human amniotic mesenchymal stromal stem cells (hAMSCs) from placenta, the cell-friendly and biocompatible behavior of RKKP glass-ceramic coated Titanium (Ti) surface was studied. The glass-ceramic material was prepared by sol-gel synthesis procedure and then deposited onto the Ti surface by Pulsed Laser Deposition method. Our findings demonstrated that the RKKP coated Ti supports are cell-friendly substrates and the combination of the hAMSCs' properties together with the bioactive characteristics of RKKP glass-ceramics may be used as a new potential tool for bone tissue engineering and regenerative medicine applications

The present work provide evidence that aqueous system could play an additional role in modulating biological functions by generating resonant structure, providing basis for storing and replying an information mediated by electromagnetic signals. External electromagnetic stimuli in resonant conditions with some of the domains of water can induce a dipole moments rearrangement in a way that this structure start to be coherent one to each other. By mean of this procedure, that has been defined as Electro-Magnetic Information Transfer Through Aqueous System (EMITTAS) an external electromagnetic pattern of signals, for instance the one picked up from a biological active molecule, can be stored, translated and transduced by the collective polymorphic water structure to the biological target . Moreover, when this procedure is delivered in contemporaneously with the selected suitable chemical molecule a synergistic effect arise that can even potentiate the molecular action of the source molecule. Remarkably, this innovate method could lead to the possibility to minimize the amount of the dose of a drug to be delivered (up to ten times less in our experiments), consequently their possible side effects as well as their socio-sanitary cost.

Osteoblast differentiation is an important process needed to maintain the continuous supply of mature osteoblast cells for growth, repair and remodelling of bones. The regulation of this process has also an important and significant impact on clinical strategies and future applications of cell therapy. We studied the effect of Electro Magnetic Field (EMF) tuned at calcium-ion cyclotron frequency of 50 Hz exposure on bone differentiation of human mesenchymal stem cells (hMSC) alone or in synergy with dexamethasone, their canonical chemical differentiation agent. Five days of continuous exposure to calcium-ion cyclotron resonance (Ca2+-ICR) affect hMSC proliferation, morphology and cytoskeletal actin reorganization. We also observed an increase of osteoblast differentiation markers' expression such as Runx2, Alkaline Phosphatase, (ALP), Osteocalcin (OC), and Osteopontin (OPN) together with the Osteoprotegerin (OPG) mRNA modulation. Moreover, in these cells, the increase of the protein expression of Osteopontin and Alkaline Phosphatase was also demonstrated. These results demonstrate bone commitment of hMSCs through a non-invasive and biocompatible differentiating physical agent treatment and highlights its possible applications in new biophysical regenerative medicine protocols.

Over the last decade, nanotechnology has become more relevant in medicine. Among magnetic nanomaterials the future of iron oxide nanoparticles (IONPs) for clinical applications relies on their biocompatibility in moderate doses, their ability to be produced in a wide range of sizes and shapes with biofunctionalization potential. Additionally, they show great promise to serve as a cell tracking system in cell-based therapies, and to generate local temperature increases in the magnetic thermotherapy of solid tumours. Thus, the study and development of novel magnetic nanoparticles for biomedical applications is one of the key topics in the field of nanotechnology. Extremely small-sized Fe3O4 superparamagnetic nanoparticles were prepared by coprecipitation, thinly coated with silica and conjugated with FITC, as molecular model specimen. Nanoparticles were characterized by dinamic light scattering (DSL), transmission electron microscopy (TEM) and X-ray diffraction analysis (XRD) and surface functional groups and composition were analysed by infrared spectroscopy (FTIR). The aim of this study was to determine the biocompatibility of the magnetic nanoparticles carriers with biofunctional coating (FITC-conjugated) on colon carcinoma CaCo-2 cell line as human cellular model. Phase contrast, immunofluorescence and confocal microscopy analyses were performed to study nanoparticles internalization and up-take. By transmission electron microscopy technique was investigated the effect of their internalization on ultrastructural features and intracellular compartments. Cellular growth and viability resulted unaffected following nanoparticles up-take and lack of toxicity was confirmed at transcriptional and translational level. Finally, even when used at high concentration, the cytotoxicity effect of the nanoparticles was not significant compared with control experiments, demonstrating their high potential in the applications of nanomedicines for a diagnostic and therapeutic tool.

Over the last decade, nanotechnology has become more relevant in medicine. Among magnetic nanomaterials the future of iron oxide nanoparticles (IONPs) for clinical applications relies on their biocompatibility in moderate doses as well as their ability to be produced in a wide range of sizes and shapes with biofunctionalization potential. Additionally, they show great promise to serve as a cell tracking system in cell-based therapies, and to generate local temperature increases in the magnetic thermotherapy of solid tumours. Thus, the study and development of novel magnetic nanoparticles for biomedical applications is one of the key topics in the field of nanotechnology. Extremely small-sized Fe3O4 superparamagnetic nanoparticles were prepared by coprecipitation, thinly coated with silica and conjugated with FITC, as molecular model specimen. Nanoparticles were characterized by dynamic light scattering (DSL), transmission electron microscopy (TEM) and X-ray diffraction analysis (XRD) and surface functional groups and composition were analysed by infrared spectroscopy (FTIR). We assessed the biocompatibility of the magnetic nanoparticles carriers with biofunctional coating (FITC-conjugated) using a colon carcinoma cell line (CaCo-2) as human cellular model. Phase contrast, fluorescence and confocal microscopy analyses were performed to study nanoparticles up-take and internalization