Rapid recognition of drug-resistance/sensitivity in leukemic cells by Fourier transform infrared microspectroscopy and unsupervised hierarchical cluster analysis

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Infrared spectroscopy and microscopy in cancer research and diagnosis

Since the middle of 20th century infrared (IR) spectroscopy coupled to microscopy (IR microspectroscopy) has been recognized as a non destructive, label free, highly sensitive and specific analytical method with many potential useful applications in different fields of biomedical research and in particular cancer research and diagnosis. Although many technological improvements have been made to facilitate biomedical applications of this powerful analytical technique, it has not yet properly come into the scientific background of many potential end users. Therefore, to achieve those fundamental objectives an interdisciplinary approach is needed with basic scientists, spectroscopists, biologists and clinicians who must effectively communicate and understand each other's requirements and challenges. In this review we aim at illustrating some principles of Fourier transform (FT) Infrared (IR) vibrational spectroscopy and microscopy (microFT-IR) as a useful method to interrogate molecules in specimen by mid-IR radiation. Penetrating into basics of molecular vibrations might help us to understand whether, when and how complementary information obtained by microFT-IR could become useful in our research and/or diagnostic activities. MicroFT-IR techniques allowing to acquire information about the molecular composition and structure of a sample within a micrometric scale in a matter of seconds will be illustrated as well as some limitations will be discussed. How biochemical, structural, and dynamical information about the systems can be obtained by bench top microFT-IR instrumentation will be also presented together with some methods to treat and interpret IR spectral data and applicative examples. The mid-IR absorbance spectrum is one of the most information-rich and concise way to represent the whole “… omics” of a cell and, as such, fits all the characteristics for the development of a clinically useful biomarker

Prognostic score in liver cirrhosis developed using the Cox's proportional hazard regression model

In order to assess the prognostic value of clinical and laboratory variables in liver cirrhosis, 36 of these variables were statistically analyzed in 151 patients followed up for 8 years. The 'survival time' was taken as the reference variable. In a first step we analyzed by log-rank test and by Cox's proportional hazard regression model the data of 98 patients (study group), obtaining 7 prognostically significant variables (age, leukocytes, calcium, potassium, globulins, cholesterol and previous diagnosis). From the regression coefficients of these variables, a risk score was obtained for each patient. To validate the prognostic value of this score, we computed it, using the same coefficients obtained in the study group, in 53 subsequently examined patients (control group) showing that the prognostic score allows the classification of these patients in 3 risk classes with different observed survival times

Inhibition of erythrocyte glutathione-peroxidase by bromsulphalein.

42 different samples of human erythrocytes were tested for glutathione-peroxidase activity ) in an attempt to study the inhibitory effect of Bromsulphalein (BSP). The mean activity of the enzyme was 11.90 +/- 3.61 U/g Hb, with no significant difference between males and females. BSP was used at different concentrations from 1 to 45 mM and inhibited GSH-Px activity; the inhibition curve showed a sinusoidal pattern. The major effect was obtained at 30 mM BSP when almost 65% of the initial activity was inhibited. The inhibition of GSH-Px by BSP has also been confirmed using partially purified GSH-Px obtained from human erythrocytes, as well as purified bovine GSH-Px. Some difference was noted between males and females: females may be divided into two subgroups, one with a lower and a second with a higher level of GSH-Px. 1 mM BSP increased the activity in the first group, whereas it reduced the activity in the second group. The inhibition by BSP was positively correlated with the basal value of GSH-Px and this effect was particularly evident in females (r = 0.865; p less than 0.001). The possibility that GSH-Px may be inhibited by BSP would be of some importance considering the strategic role of GSH-Px in protecting the cell from oxidative attack

The identification of cystic fibrosis (CF) cells and their pharmacological correction by mid-infrared microspectroscopy and unsupervised data analysis methods

We aimed at demonstrating that non-cystic fibrosis (CF) cells and isogenic, CF bronchial epithelial cell lines can be correctly classified by Fourier transform (FT) mid-infrared (IR) absorbance spectra and unsupervised methods for multivariate data analysis such as principal component analysis (PCA) and hierarchical cluster analysis (HCA). CF cells were exposed \u201cex vivo\u201d to VRT-325, a chemical corrector of defective anion transporter Cystic Fibrosis Transmembrane conductance Regulator (CFTR) caused by the F508del mutation. Unstained epithelial cells from CF patients and a healthy control were deposited on ZnSe window and analyzed with an IR interferometer connected to a FTIR microscope (microFTIR). In order to explain spectral variability reflecting biochemical differences between CF and non-CF cells, we applied PCA to dataset of untreated cells and cells treated for 24 hours with 1x10-5 M VRT-325. The information achieved with IR analysis was cross validated by the findings on CFTR expression and by the results of functional CFTR bioassays carried out in replicate samples. We conclude suggesting the possibility to utilize IR spectra analysis to recognize CF cells and the effect of pharmacological treatment aimed to correct the basic defect of C

Severe impairment of antioxidant system in human hepatoma

Catalase (CAT), glutathione-peroxidase (GSH-Px) activity and reduced glutathione content (GSH) were measured in patients who had hepatocellular carcinoma, and values compared with those of normal liver and liver adjacent to neoplastic tissue. The results showed a remarkable reduction of CAT in tumor and corresponding tumor-free tissue (P less than 0.001 and P less than 0.02, respectively). All neoplastic samples had a significant lower activity of CAT than the corresponding adjacent tumor-free tissue (P less than 0.05). The GSH-Px activity of tumor tissue also was lower than normal (P less than 0.001) but similar to that of adjacent tissue. No correlation was noted between the two enzyme activities. Glutathione content was extremely low in tumor (P less than 0.001) and even in tumor-free tissue (P less than 0.05) when compared with normal liver. In all cases the content of GSH in neoplastic tissue was lower than that of the corresponding tumor-free tissue (P less than 0.05). Whereas in normal liver the activity of GSH-Px was positively correlated with the content of GSH, in the neoplastic tissue such a relationship disappeared. All these findings suggest that the antioxidant system of hepatocellular carcinoma cell is severely impaired

Anti PSMA immunotoxins activity against prostate cancer cell lines.

Immunotherapy with monoclonal antibody (mAb)-vehicled toxins may be a novel treatment option for the management of androgen-independent Prostate Cancer. To investigate the therapeutic potential of anti-prostate specific membrane antigen (PSMA) mAb-ricin A-chain Immunotoxins (ITS) in monolayer and 3D cell cultures of prostate carcinoma cell lines (LNCaP), (J591, PEQ226.5, PM2PO79.1)