Artikel
Touchless optical assessment of the IDH1 mutation status of experimental gliomas
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Autoren
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Wenmin Yao
- Universitätsklinikum Carl Gustav Carus, Dresden, Deutschland
| Veröffentlicht: | 18. Juni 2018 |
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Gliederung
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Objective: Mutation of the isocitrate dehydrogenase -1 (IDH1) gene has profound implications for diagnosis, treatment and prognosis of glioma patients. Current detection methods of the mutation status of IDH1 cannot applied intra-operatively. Raman spectroscopy is a rapid, simple, and real time diagnostic method that detects the biochemical composition of cells and tissues.Our aim is to use Raman spectroscopy as an online tool to identify the mutation status of IDH1 gene in vivo using the chorion allontoic membrane model of glioma growth.
Methods: We used five different glioma cell lines that underwent genetic modification to provide both IDH1 wild-type (wt) and mutant (mut) variants: U87 EGFP, U87 IDH-1wt, U87 IDH-1mut. HT12346 IDH-1wt/mut, HT12347 IDH-1wt/mut, SVGP12wt/mut, HT7060 IDH-1wt/mut. Cells were implanted on embryonic day 10 on the CAM (4x106 cells/egg, n=10 per cell line). Tumor growth of U87-EGFP cell lines was monitored by fluorescence microscopy. Tumors were allowed to grow for 8 days until ), in ovo /in vivo Raman spectroscopy was performed. Raman spectra were analyzed in Matlab v7.2. Thereafter, tumors were removed from the egg and fixed in formalin. 10 µm cryosections were analyzed using H&E staining and immunohistochemistry for Ki67, IDH1 and a-SMA.
Results: Our results demonstrate that all of the cell lines can be used for tumor induction in the CAM model and form glioma within 8 days. The tumors exhibited angiogenic effects, high proliferation rates and the morphological characteristics of glioma. Significant differences in the Raman spectra between the IDH1 mutation and the IDH1 wide type were found, particularly in the spectra ranges of 760, 826,1003(C-C ring breathing of Phenylalanine), 1337 (Ch2 and Ch3 groups in lipids/ protein), 1660cm-1,which primarily contain signals related to protein, DNA, and lipids. The peaks at 826,1337 cm-1 were significantly decreased in wt-IDH1 (p<0.05). Bsaed on these two peaks, the classification of Raman spectra according to IDH1 genotype with a correct rate of 89% by using discriminant analysis with Mahalanobis distances.
Conclusion: Raman spectroscopy constitutes a simple, rapid and safe procedure for determination of the IDH1 mutation in vivo in the CAM model reflecting the intraoperative setting.
