Giuliano Spremolla

A New Approach to the Study of Human Solid Tumor Cells by Means of FT-IR Microspectroscopy

Cells coming from normal and neoplastic human lung tissue were analyzed by means of FT-IR microspectroscopy. Among the various methods tested to isolate the cells, mechanical treatment alone was found to yield the best results. Monolayers of cells were homogeneously distributed by cytocentrifuge preparation on BaF2 windows, and several spectra were obtained for different circular micro-areas of the order of one hundred microns in diameter. This procedure made it possible to obtain reliable spectra and to reject those containing additional bands due to impurities arising from the isolation treatment. Spectral differences between normal and neoplastic cells reflect an increase in the intensity of the bands corresponding mainly to PO2− symmetrical and asymmetrical vibrations of DNA in pathological samples with respect to normal ones. The value of the ratio of the integrated areas of the bands at 1080 and 1540 cm−1 due to DNA and proteins, respectively, makes it possible to differentiate between normal and abnormal cells, thus suggesting the use of this parameter as an original approach in the recognition of early neoplastic transformation undetectable by means of traditional procedures.

New possibilities of research in chronic lymphatic leukemia by means of fourier transform-infrared spectroscopy-II

Ten samples of lymphocytes coming from patients affected by chronic lymphatic leukemia and ten samples from normal subjects were studied by FT-IR spectroscopy. Spectral differences observed between the two kinds of cells correspond to an increase of the intensities, in the leukemic samples with respect to the normal ones, of the bands corresponding mainly to PO2- symmetrical and asymmetrical stretching vibrations of DNA. The ratios of the integrated areas of the band at 1080 cm-1 mainly involving the symmetrical stretching vibration of the O-P-O linkages of DNA, and of the band at 1540 cm-1, due to the proteic components of the lymphocytes, assume different values for the two kinds of cells. These ratios can constitute an additional marker to diagnose chronic lymphatic leukemia and may be usefully employed to evidence the early phases of the disease.

Analytical infrared spectral differences between human normal and leukaemic cells (CLL) — I

Two series of normal and leukaemic lymphocytes were examined by infrared spectroscopy in order to try to find spectral differences connected with chemical and biological modifications. The bands at 965 and 530 cm-1 present only in the spectra of leukaemic lymphocytes, assume particular significance. The C-H stretching region furnishes useful indications about the different ratios of the methyl groups compared with the methylene ones in the two cases. The infrared bands characteristic of the leukaemic lymphocytes seem to be due to chemical modifications not involving the DNA chain.

Infrared Characterization of Nuclei Isolated from Normal and Leukemic (B-CLL) Lymphocytes: Part III

Nuclei extracted from lymphocytes coming from normal subjects and from patients affected by chronic lymphatic leukemia (B-CLL) were studied by means of infrared spectroscopy. Substantial spectral differences were found above all in the region of the phosphate-group vibrations of DNA. The ratios of the integrated areas of the bands at 1080 and 1540 cm−1, due to the symmetrical stretching vibrations of PO2− groups and to proteic components, respectively, assume increasing values, which are localized in quite separate ranges, for normal lymphocytes, leukemic lymphocytes, normal nuclei, and leukemic nuclei. These values, indicating a different distribution of DNA and of proteic components between normal and leukemic cells and between normal and leukemic nuclei, may assume a remarkable significance in the precocious diagnosis of leukemia (B-CLL) and in following the course of the disease. In the case of leukemic nuclei, the ratio, in weight, of the two components reaches the value of 1:1.

FT-IR analysis of single human normal and leukemic lymphocytes

The infrared spectra of single human normal and leukemic lymphocytes have been obtained by means of the microscope-FT-IR system. Substantial differences between the two kinds of cells have been observed at level of the O-P-O vibrations of the DNA and of the proteic components. This determination opens up the possibility in determining the early stages of leukemia, in following the course of patients under therapy, and in evaluation of the residual neoplastic disease.

Infrared parameters in the characterization of the neoplastic cell transformation

Abstract In this paper we report the results on the characterization of neoplastic cells by means of FT-IR spectroscopy. Spectral differences have been found in examining normal and leukemic lymphocytes. Through analyzing also nuclei extracted from normal and leukemic lymphocytes an increase of DNA with respect to proteic components was observed in the order: normal lymphocytes, leukemic lymphocytes, normal nuclei, and leukemic nuclei. Spectral differences were observed between lipids extracted from normal and leukemic cells. Recently the infrared spectra of single normal and leukemic cells have been obtained. The later opens new possibilities for revealing the early phase of the disease, in following the trends of leukemia in patients who are under treatment with drugs, and in evaluating the residual neoplastic disease. This research has extended to the study of human tumor lung cells. Spectral differences were also observed analyzing normal and neoplastic cells. Spectroscopic parameters distinguishing normal from tumoral cells have been suggested.

Micro FT-IR Characterization Of Human Lung Tumor Cells

FT-IR spectroscopy has opened up a new approach to the analytical study of cell transformation. Investigations carried out in normal and leukemic lymphocytes have evidenced an increase in DNA with respect to proteic components in neoplastic cells.(1) The evaluation of the ratio of the integrated areas(A) of the bands at 1080 cm-1 (mainly DNA) and at 1540 cm-1 (proteic components) has allowed us to establish a parameter which indicates, for values above 1.5, the neoplastic nature of cells. Recently, this approach has been applied to the study of human lung tumor cells. Several monocellular suspension procedures of the tissue fragment (mechanical and/or chemical) were tested to obtain reproducible and reliable spectra able to differentiate clearly between normal and patological cells. Chemical treatment (EDTA, Pepsin, Collagenase, etc.) produced additional bands in the spectra of the cells causing distortion of the profiles of some absorptions, and as a result, mechanical treatment was preferred. The normal and neoplastic cells homogeneously distributed by cytospin preparation on BaF2 windows were examined by means of FT-IR microscopy. An examination of several microareas of each sample yielded reproducible spectra, with values of the A 1080 cm-1 / A 1540 cm-1 parameter within a very narrow range for each sample, even if certain differences still remained among the different cases, in good agreement with the results obtained for leukemic cells.(1) The value of this parameter was found to be lower for cells isolated from the normal area of lung, than in the case of those corresponding to the tumoral area, meaning that an increase occurs in DNA with respect to the proteic components. These insights, which provide a basis to obtain indications at the molecular level, can open up new possibilities in clinical practice, in order to obtain diagnosis confirmation, to detect early stages of disease and to offer additional indications in cases of dubious interpretation.

Analysis Of Pulmonary Tissue Of Natural Mummy Of XIII Century (Saint Zita, Lucca-Tuscany) By Means Of FT-IR Microspectroscopy.

During a recognition carried out by one of us on the S.Zitas body, preserved for seven hundred years in the S.Fredianos Basilica (Lucca, Tuscany-Italy) it has been possible to verify the good condition of many orgains and systems. In this comunication we report preliminary results of an analysis of the pulmonary tissue which to the histologic examination showed a typical aspect. It was observed the presence of a massive anthracosis. In order to obtain indication, at molecular level, FT-IR microspectroscopy measurements were carried out. The infrared spectra relative to different microareas allowed us to evidence regions of tissue decomposed, and others in which the infrared spectra showed absorptions characteristic of proteic components at 1650 cm-1 and 1540 cm-1 ascribed to the Amide I and Amide II vibrations respectively. In the 1350-950 cm-1 range bands due to nucleic acids were detected. Also in the CH stretching region bands due to methyl and methylene groups were observed, likely due to the presence of paraffinic segments of phospholipids of cellular membranes. Previous FT-IR studies carried out by us on biological systems such as normal and leukemic lymphocytes (1), on lymphoblast (2), and recently on cells isolated from neoplastic pulmonary tissues (3), represent a basis which allows us to perform a sufficiently complete assignment on the spectra obtained on this finding. These spectra are surprisingly very similar to those obtained by us on human lung tissue from surgical specimens. As far as we know, FT-IR microspectroscopy, which was first employed by our group in the study of human neoplasia at level of single cell (4) has been applied in the analysis of ancient pulmonary tissue, certainly not submitted to any conservative treatment. This kind of approach can open up new possibilities in obtaining indication, at molecular level, on findings of paleophanthology of different origin and age.