Hyperglycemia and cancer in human lung carcinoma by means of Raman spectroscopy and imaging.

Raman spectroscopy and Raman imaging were used to identify the biochemical and structural features of human cancer lung cells (CCL-185) and the cancer cells supplemented with glucose and deuterated glucose at normal and hyperglycemia conditions. We found that isotope substitution of glucose by deuterated glucose allows to separate de novo lipid synthesis from exogenous uptake of lipids obtained from the diet. We demonstrated that glucose is largely utilized for de novo lipid synthesis. Our results provide a direct evidence that high level of glucose decreases the metabolism via oxidative phosphorylation in mitochondria in cancer cells and shifts the metabolism to glycolysis via Warburg effect. It suggests that hyperglycemia is a factor that may contribute to a more malignant phenotype of cancer cells by inhibition of oxidative phosphorylation and apoptosis.

Double face of cytochrome c in cancers by Raman imaging.

Cytochrome c (Cyt c) is a key protein that is needed to maintain life (respiration) and cell death (apoptosis). The dual-function of Cyt c comes from its capability to act as mitochondrial redox carrier that transfers electrons between the membrane-embedded complexes III and IV and to serve as a cytoplasmic apoptosis-triggering agent, activating the caspase cascade. However, the precise roles of Cyt c in mitochondria, cytoplasm and extracellular matrix under normal and pathological conditions are not completely understood. To date, no pathway of Cyt c release that results in caspase activation has been compellingly demonstrated in any invertebrate. The significance of mitochondrial dysfunctionality has not been studied in ductal carcinoma to the best of our knowledge. We used Raman spectroscopy and imaging to monitor changes in the redox state of the mitochondrial cytochromes in ex vivo surgically resected specimens of human breast tissues, and in vitro human breast cells of normal cells (MCF 10A), slightly malignant cells (MCF7) and highly aggressive cells (MDA-MB-231). We showed that Raman imaging provides insight into the biology of human breast ductal cancer. Here we show that proper concentration of monounsaturated fatty acids, saturated fatty acids, cardiolipin and Cyt c is critical in the correct breast ductal functioning and constitutes an important parameter to assess breast epithelial cells integrity and homeostasis. We look inside human breast ducts by Raman imaging answering fundamental questions about location and distribution of various biochemical components inside the lumen, epithelial cells of the duct and the extracellular matrix around the cancer duct during cancer development in situ. Our results show that human breast cancers demonstrate a redox imbalance compared to normal tissue. The reduced cytochrome c is upregulated in all stages of cancers development. The results of the paper shed light on a largely non-investigated issues regarding cytochromes and mitochondrial function in electron transfer chain. We found in histopathologically controlled breast cancer duct that Cyt c, cardiolipin, and palmitic acid are the main components inside the lumen of cancerous duct in situ. The presented results show direct evidence that Cyt c is released to the lumen from the epithelial cells in cancerous duct. In contrast the lumen in normal duct is empty and free of Cyt c. Our results demonstrate how Cyt c is likely to function in cancer development. We anticipate our results to be a starting point for more sophisticated in vitro and in vivo animal models. For example, the correlation between concentration of Cyt c and cancer grade could be tested in various types of cancer. Furthermore, Cyt c is a target of anti-cancer drug development and a well-defined and quantitative Raman based assay for oxidative phosphorylation and apoptosis will be relevant for such developments.

The role of pro- and antiangiogenic factors in angiogenesis process by Raman spectroscopy.

Raman spectroscopy and Raman imaging are powerful techniques to monitor biochemical composition around blood vessel. The aim of this study was to understand the role of pro- and antiangiogenic factors in angiogenesis process. Raman imaging and Raman single spectrum measurements allow the diagnosis of cancer biochemical changes in blood vessel based on several biomarkers simultaneously. We have demonstrated that Raman imaging combined with statistical methods are useful to monitoring pro- and antiangiogenic factors responsible for angiogenesis process. In this work Raman markers of proangiogenic and antiangiogenic factors were identified based on their vibrational signatures. Obtained results can help understand how growing tumor create its vascular system.

Raman imaging and statistical methods for analysis various type of human brain tumors and breast cancers.

Spectroscopic methods provide information on the spatial localization of biochemical components based on the analysis of vibrational spectra. Raman spectroscopy and Raman imaging can be used to analyze various types of human brain tumors and breast cancers. The objective of this study is to evaluate the Raman biomarkers to distinguish tumor types by Raman spectroscopy and Raman imaging. We have demonstrated that bands characteristic for carotenoids (1156 cm-1, 1520 cm-1), proteins (1004 cm-1), fatty acids (1444 cm-1, 1655 cm-1) and cytochrome (1585 cm-1) can be used as universal biomarkers to assess aggressiveness of human brain tumors. The sensitivity and specificity obtained from PLS-DA have been over 73%. Only for gliosarcoma WHO IV the specificity is lower and takes equal 50%. The presented results confirm clinical potential of Raman spectroscopy in oncological diagnostics.

Monitoring glycosylation metabolism in brain and breast cancer by Raman imaging.

We have shown that Raman microspectroscopy is a powerful method for visualization of glycocalyx offering cellular interrogation without staining, unprecedented spatial and spectral resolution, and biochemical information. We showed for the first time that Raman imaging can be used to distinguish successfully between glycosylated and nonglycosylated proteins in normal and cancer tissue. Thousands of protein, lipid and glycan species exist in cells and tissues and their metabolism is monitored via numerous pathways, networks and methods. The metabolism can change in response to cellular environment alterations, such as development of a disease. Measuring such alterations and understanding the pathways involved are crucial to fully understand cellular metabolism in cancer development. In this paper Raman markers of glycogen, glycosaminoglycan, chondroitin sulfate, heparan sulfate proteoglycan were identified based on their vibrational signatures. High spatial resolution of Raman imaging combined with chemometrics allows separation of individual species from many chemical components present in each cell. We have found that metabolism of proteins, lipids and glycans is markedly deregulated in breast (adenocarcinoma) and brain (medulloblastoma) tumors. We have identified two glycoforms in the normal breast tissue and the malignant brain tissue in contrast to the breast cancer tissue where only one glycoform has been identified.