Vito Capozzi

Vibrational spectroscopies provide information about the biochemical and structural environment of molecular functional groups inside samples. Over the past few decades, Raman and infrared-absorption-based techniques have been extensively used to investigate biological materials under different pathological conditions. Interesting results have been obtained, so these techniques have been proposed for use in a clinical setting for diagnostic purposes, as complementary tools to conventional cytological and histological techniques. In most cases, the differences between vibrational spectra measured for healthy and diseased samples are small, even if these small differences could contain useful information to be used in the diagnostic field. Therefore, the interpretation of the results requires the use of analysis techniques able to highlight the minimal spectral variations that characterize a dataset of measurements acquired on healthy samples from a dataset of measurements relating to...

Fourier transform infrared micro-spectroscopy (μ-FT-IR) is nowadays considered a valuable tool for investigating the changes occurring in human cells after exposure to ionizing radiation. Recently, considerable attention has been devoted to the use of this optical technique in the study of cells exposed to proton beams, that are being increasingly adopted in cancer therapy. Different experimental configurations are used for proton irradiation and subsequent spectra acquisition. To facilitate the use of μ-FT-IR, it may be useful to investigate new experimental approaches capable of speeding up and simplifying the irradiation and measurements phases. Here, we propose the use of low-e-substrates slides for cell culture, allowing the irradiation and spectra acquisition in transflection mode in a fast and direct way. In recent years, there has been a wide debate about the validity of these supports, but many researchers agree that the artifacts due to the presence of the electromagnetic ...

Raman micro-spectroscopy was used for an in vitro investigation aimed to elucidate the effects of X-rays and polyphenols derived from autochthonous sweet cherries from Campania (Italy) on specific sub-cellular regions of single SH-SY5Y human neuroblastoma cells. Nucleus and cytoplasm regions of single cells were investigated after the exposure to different doses of X-rays (0, 2, 4 Gy). Cells were fixed immediately after irradiation and/or after having been exposed to the cherry polyphenol extracts. Two different concentrations of polyphenol extracts (25 μg/mL and 500 μg/mL) with an exposure time of 48 h to polyphenols were considered. A deconvolution procedure using Lorentzian curves along with the evaluation of ratios between the intensity of selected peaks were used for analyzing the collected spectra in order to highlight the changes due to the different treatments. Biochemical changes occurring in nucleus and cytoplasm regions of single cells as a consequence X-ray irradiation were observed. In particular, the analysis of Raman spectra allowed us to detect modifications in the contribute from proteins, nucleic acids, lipids, and carbohydrates of cells, induced at different extent on the two cell regions. In fact, in the absence of polyphenols, the most relevant effects were observed in the cell nucleus region where modifications in the cell cycle were detected with an increase of DNA/RNA contribution, protein rearrangement and changes in lipids. The presence of the extracts induced changes in the observed variations. Our approach was also able to show the radiomodulating effects induced by the presence of different doses of polyphenols and different exposure times to these substances.

We report vibrational spectra of two types of eumelanin samples: a synthetic one, produced by oxidation of tyrosine with hydrogen peroxide, and a natural one, extracted from Sepia officinalis, with the aim of pointing out the differences between the main vibrational properties of such two pigments. Fourier transform infrared, Raman and surface-enhanced Raman scattering spectra allow the identification of several vibrational modes involving specific monomeric units. The main differences between the spectra of the two types of eumelanin are related to (i) a larger amount of carboxylic acid in the synthetic than in the natural sample; (ii) a shift of spectral position of corresponding peaks; and (iii) a contribution of residual proteins to the signals from the natural sample. Such results suggest that vibrational techniques may be helpful for non-destructively determining the composition of various eumelanin pigments, with potential applications in biomedical, electronic and cultural heritage fields. © 2016 Society of Chemical Industry

Pesticides are chemical compounds widely used in agricultural and industrial activities, as well as domestic ones. A commercial pesticide is usually composed of active ingredients and formulants. Among the active ingredients, Deltamethrin is a pyrethroid chemical widely used for synthesizing pesticides products which are very effective in damaging the central nervous system of pests. In this work, we analyze, by means of Atomic Force Microscopy (AFM), cellular morphological changes induced by exposure to a Deltamethrin-based commercial pesticide. AFM microscopy, in addition to the well-known characterization of the cellular topography, has the ability to monitor interesting biomechanics parameters of the surface as roughness and elastic modulus. In particular, we exposed normal human keratinocytes for 24 hours at different solutions of pesticide, well below the threshold of citotoxicity. The AFM images of exposed cells show alterations of surface cell shape. Moreover exposed cells a...

Fourier transform infrared (FTIR) micro-spectroscopy has been attracting the interest of many cytologists and histopathologists for several years. This is related to the possibility of FTIR translation in the clinical diagnostic field. In fact, FTIR spectra are able to detect changes in biochemical cellular components occurring when the cells pass to a pathological state. Recently, this interest has increased because it has been shown that FTIR spectra carried out just in the high wavenumber spectral range (2500–4000 cm−1), where information mainly relating to lipids and proteins can be obtained, are able to discriminate cell lines related to different tissues. This possibility allows to perform IR absorption measurements of cellular samples deposited onto microscopy glass slides (widely used in the medical environment) which are transparent to IR radiation only for wavenumber values larger than 2000 cm−1. For these reasons, we show that FTIR spectra in the 2800–3000 cm−1 spectral r...

In recent years, Fourier Transform Infrared (FTIR) micro-spectroscopy has shown promising potential in medical diagnostics at the cellular level. In fact, FTIR spectra can provide information related to DNA, protein, and lipid content and how such a content changes when a pathological state arises. Most of these information is included in the so-called fingerprint region (1000–1800 cm−1), consisting of several spectral peaks related to vibrational modes occurring inside cellular components. Unfortunately, the slides commonly used in cytology (as the glass microscopy slides and coverslips) are opaque to IR radiation in the fingerprint region, whereas they are transparent for wavenumber values larger than 2000 cm−1, where few and broad spectral absorption bands, mainly due to lipids and proteins, are present. Nonetheless, here we show that FTIR spectra performed in the high wavenumber range 2750–3000 cm−1 can be used to discriminate two different types of cells, one from a normal cell...

The toxicological implications of nanoparticles deserve accurate scientific investigation for the protection of human health. Although toxic effects involve specific organs, the events that cause them have their origin from biochemical modifications of some cellular constituents. Therefore, a first analysis to evaluate the effects due to the action of nanoparticles is achieved by investigation of in vitro cells, which allows the identification of the cellular modifications caused by nanoparticles (NPs) even at much lower doses than the lethal ones. This work evaluated the Raman microspectroscopy capability to monitor biochemical changes occurring in human cells as a consequence of exposure to a suspension of gold nanoparticles with a non-cytotoxic concentration. Human keratinocyte cells were used as a model cell line, because they are mainly involved in environmental exposure. A trypan blue assay revealed that the investigated concentration, 650 ng/mL, is non-cytotoxic (about 5% of ...

Single SH-SY5Y human cancer cells Cellular nucleus and membrane Multivariate analysis X-ray effects on DNA, lipids, proteins and carbohydrates a b s t r a c t Raman micro-spectroscopy was performed in vitro on nuclear and membrane regions of single SH-SY5Y human neuroblastoma cells after irradiation by graded X-ray doses (2, 4, 6, 8 Gy). The acquired spectra were analyzed by principal component analysis (PCA) and interval-PCA (i-PCA) methods. Biochemical changes occurring in the different regions of single cells as a consequence of the radiation exposure were observed in cells fixed immediately after the irradiation. The most relevant effects arose from the analysis of the spectra from the cell nucleus region. The observed changes were discussed in terms of the modifications in the cell cycle, resulting in an increase in the DNA-related signal, a protein rearrangement and changes in lipid and carbohydrates profiles within the nucleus. Potential markers of an apoptotic process in cell population irradiated with 6 and 8-Gy X-ray doses could have been singled out. No significant effects were found in spectra from cells fixed 24 h after the irradiation, thus suggesting the occurrence of repairing processes of the X-ray induced damage.

Aims: Until now, studies related to the morphometric and morphological variations produced on peripheral blood leukocytes from healthy donors by exposure to 1.8 GHz electromagnetic radiation (EMR) yielded controversial results. The aim of this research work is to increase the statistics regarding the above mentioned variations. Methodology: By using a reverberation chamber, which provides a controlled EMR intensity, 108 samples of human leukocytes from healthy donors were exposed to EMR of Original Research Article Jirillo et al.; AIR, Article no. AIR.2014.9.001 479 different intensities (12±4 V/m, 22±6 V/m, 42±9 V/m and 78±10 V/m) for times ranging from 5 min to 24 h. Sham exposed blood samples of the same donors were considered as controls. Using a computerized morphometric method, microscopic observations of the area size occupied by each cell were conducted; in each case the dimensional measurements were carried out on three different samples (from different donors). Besides, morphological observations were conducted staining smeared blood samples with May-Grünwald-Giemsa. Results: Exposed and sham exposed leukocytes average size was compared using the Statistical GraphPad Prism 5.0 software. In 18% out of 108 cases examined, no effects dependent on EMR have been revealed. On the contrary, statistically significant variations in area of exposed leukocytes in comparison to non exposed cells were observed in 82% out of 108 cases examined. In 68% out of 108 cases an increase in leukocyte areas was demonstrated along with morphological variations of cells; in 14% out of 108 the cases, a decrease in leukocyte areas was observed. Conclusions: Even though this study needs a functional evaluation of leukocytes exposed to EMR, our results suggest that 1.8 GHz EMR is able to produce increase in the leukocyte areas as well as morphological alterations.

The pathogenic events responsible for the reduction of endothelial progenitor cell (EPC) number and function seen in patients with chronic renal failure (CRF) are poorly understood. Here we investigate the hypothesis that increased concentrations of urea associated with CRF increase ROS production directly in EPCs, causing abnormalities associated with coronary artery disease risk. Human EPCs were isolated from peripheral blood mononuclear cells of healthy donors and cultured in the presence or absence of 20 mmol/L urea. Urea at concentrations seen in CRF induced ROS production in cultured EPCs. Urea-induced ROS reduced the number of endothelial cell colony forming units, uptake and binding of Dil-Ac-LDL and lectin-1, and the ability to differentiate into CD31- and vascular endothelial growth factor receptor 2-positive cells. Moreover, urea-induced ROS generation accelerated the onset of EPC senescence, leading to a senescence-associated secretory phenotype (SASP). Normalization of ...

Microwave based treatment (MWT) of wet wheat kernels induced a striking reduction of gluten, up to <20 ppm as determined by R5-antibodybased ELISA, so that wheat could be labeled as gluten-free. In contrast, analysis of gluten peptides by G12 antibody-based ELISA, mass spectrometry-based proteomics and in vitro assay with T cells of celiac subjects, indicated no difference of antigenicity before and after MWT. SDS-PAGE analysis and Raman spectroscopy demonstrated that MWT simply induced conformational modifications, reducing alcohol solubility of gliadins and altering the access of R5-antibody to the gluten epitopes. Thus, MWT neither destroys gluten nor modifies chemically the toxic epitopes, contradicting the preliminary claims that MWT of wheat kernels detoxifies gluten. This study provides evidence that R5-antibody ELISA alone is not effective to determine gluten in thermally treated wheat products. Gluten epitopes in processed wheat should be monitored using strategies based...

The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption.

In February 2002 a digital OmniPAL receiver was put into operation at the Department of Physics of Bari University (Southern Italy) to record VLF radio signals. The intensity of the VLF signals transmitted by GB (f=16 kHz, United Kingdom), FR (f=20.9 kHz, France), GE (f=23.4 kHz, Germany), IC (f=37.5 kHz, Island) and IT (f=54 kHz, Sicily, Italy) has been monitored with a 5s sampling frequency. In this study we analysed the raw data averaged over 10 min from February 2002 to June 2006. Several decreases of the electric field strength of the radio signals were revealed some days long and generally they occur in not concomitant periods. Such a phenomenology was pointed out mainly looking the ±3 days smoothed trends. The GE signals decrease systematically in winter and summer each year and so, it is well founded to suppose the decreases are related to the transmitter. On the contrary, all the other events are sporadic and the previous justification does not seem realistic. On the time occurrence of these events, we investigated the geomagnetic activity, the meteorological conditions in the receiver area and the regional seismic activity. The main result is that, generally, a pre or post seismic effect seems to give the most convincing justification of the quoted radio decreases.

Starting in 1987, gas contents have been collected with a mean sampling frequency of three days in wells and springs located in the southern area of the Kamchatka peninsula (Russia). We examined the Argon trend at one well and an anomalous increase seems to appear during June--July 1996. On June 21 an earthquake with M = 7.1 occurred at a

A very significant issue today concerns the problem of air pollution caused mainly by human activity. The statistics show that most of the pollutants in the atmosphere is due to emissions caused by anthropogenic factors (e.g. power and industrial plants, traffic and combustion phenomena in general). In this paper we evaluate the implementation of a model using artificial neural networks to forecast short-term rate of air pollution for supporting environmental policy decisions.

Spontaneous photoluminescence (PL) spectra of Cu-doped and undoped e-GaSe have been investigated in the temperature range from 80 to 300 K and at low laser-excitation intensity (P) from 10 to 10 W cm. The main modification of the spectra in doped crystals with respect to those of un-doped samples is the appearance of two bands in the extrinsic part of the PL spectrum and centered at 65S and 678 nm, respectively. The luminescence at energies below the excitonic recombinations (extrinsic bands) is enhanced by doping. Also indirect free-and bound-excitonic lines are also strongly influenced by the impurity concentration; in fact, their emission intensity, which depends linearly on I' in undoped crystals, shows a quadratic dependence in doped samples. The temperature dependence of the PL spectra gives the thermal activation energy of some extrinsic bands, which results equal the ionization energy of the acceptor levels involved in the extrinsic transitions. A simple kinetic model of the radiative recombination is proposed; it accounts for the experimental data of the excitation intensity dependence of the free-and bound-excitonic lines. This model can also explain the different temperature dependence of the PL intensity of these lines: linear for the free-excitonic emissions, exponential for the bound-excitonic recombinations. Some radiative transitions from donor levels located in the energy gap of GaSe are analyzed and a scheme of donor and acceptor states involved in the PL spectra is proposed.