FTIR microspectroscopy of malignant fibroblasts transformed by mouse sarcoma virus

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Abstract

Fourier transform infrared microspectroscopy (FTIR-MSP), which is based on the characteristic molecular vibrational spectra of cells, was used to investigate spectral differences between normal primary rabbit bone marrow (BM) cells and bone marrow cells transformed (BMT) by murine sarcoma virus (MuSV). Primary cells, rather than cell lines, were used for this research because primary cells are similar to normal tissue cells in most of their characteristics. Our results showed dramatic changes in absorbance between the control cells and MuSV124-transformed cells. Various biological markers, such as the phosphate level and the RNA/DNA obtained, based on the analysis of the FTIR-MSP spectra, also displayed significant differences between the control and transformed cells. Preliminary results suggested that the cluster analysis performed on the FTIR-MSP spectra yielded 100% accuracy in classifying both types of cells.

Introduction

Progress in deciphering the molecular mechanisms of carcinogenesis has been enhanced substantially by the study of the action of viruses as one of the causatives of cancer [1]. Viruses consist of either DNA or RNA as nucleic acid and have been well studied in the last 20 years. Among them, retroviruses containing RNA are known to be responsible for various types of malignancies in humans [2], [3]. Cellular transformation by retroviruses involves the insertion and activation of viral proto-oncogenes [4]. More than 70 different proto-oncogenes are documented in the literature [5]. Cervical cancer is the second type of malignancy causing mortality in women in the World. The human papiloma viruses (HPV) have been incriminated in the transformation of epithelia in cervical cancer [6]. Powerful molecular techniques are available today to detect the virus in the cells obtained from patients [7]. However, these methods are expensive and time-consuming in nature.

Fourier transform infrared (FTIR) spectroscopy has been widely applied in biology and medicine. FTIR has expanded our knowledge of the structure, conformation and dynamics of various molecular components of the cell [8]. With the introduction of microscopy in modern FTIR instrumentation, FTIR analysis of cells and tissues has become a reality. Naumann's group successfully achieved the classification of different classes of bacteria using FTIR spectroscopy [9]. In recent years, there has been tremendous interest in applying FTIR as a tool for the diagnosis of cancer. Successful diagnoses of lung [10], breast [11], cervical [12] and prostate [13] cancers have been reported. In addition to the in vitro studies on cells [14], our group has contributed to the accurate diagnosis of colon cancer using FTIR microspectroscopy (FTIR-MSP) [15].

There is enough evidence in the literature that viral infections, especially in cervical cancer, could be detected using FTIR-MSP [16]. This is the first controlled study in which the malignant transformation of fibroblasts in ex vivo was performed by retroviral infection and detected by FTIR-MSP. Our initial results indicated that retroviral infection could be successfully detected in fibroblasts by means of FTIR-MSP.

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Cells and viruses

Primary rabbit cells obtained from the bone marrow (BM) of 1.5-kg rabbits were grown at 37 °C in Roswell Park Memorial Institute (RPMI) medium, supplemented with 10% newborn calf serum (NBCS) and antibiotics: penicillin, streptomycin and neumycin. All the chemicals used were obtained from Biological Industries (Bet ha-Emeq, Israel). Clone 124 of TB cells (mouse fibroblast cells), chronically releasing Moloney MuSV-124, was used to prepare the appropriate virus stock. TB cells were grown in a

Cell characteristics

In the present study, primary cells (1–2 passages/culture) and malignant cells transformed by retroviruses (MuSV) were used.

Primary rabbit BM fibroblasts, grown in plastic dishes in RPMI medium with 10% NBCS, appear as flat cells under an inverted light microscope (Fig. 1a). These cells are completely unable to grow in soft agar. When BM cells were infected by MuSV, transformed cells, with a highly refractive shape, were detected growing randomly in a criss-cross fashion (Fig. 1b). These BMT

Discussion

Our FTIR-MSP results on normal and MuSV-infected fibroblasts, isolated from the bone marrow of rabbits, have suggested directions for the applications of advanced optical technology in the detection of malignancies caused by retroviruses. Significant differences in absorbance were observed between BM and BMT. Similar changes were observed in cases where the same virus infected the NIH 3T3 cell line [23]. In addition, in our earlier report on FTIR-MSP studies on H-ras-transfected fibroblast

Acknowledgements

We gratefully acknowledge the Harry Stern Applied Research Grant Program and the Israel Science Foundation (ISF grant number: 788/01) for their financial support.

References (29)

  • J. Versalovic et al.

    Molecular detection and genotyping of pathogens: more accurate and rapid answers

    Trends Microbiol.

    (2002)
  • H.H. Mantsch et al.

    Infrared spectroscopy of biomolecules

    (1996)
  • D. Naumann et al.

    Microbiological characterizations by FT-IR spectroscopy

    Nature

    (1991)
  • T. Gao et al.

    Human breast carcinomal tissues display distinctive FTIR spectra: implication for the histological characterization of carcinomas

    Anal. Cell. Pathol.

    (1999)
  • Cited by (0)

    1

    Present address: Advanced Biomedical Science and Technology Group, Life Sciences Division, Oak Ridge National Laboratory, TN 37831-6101, USA.

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