Determination of Brain Tissue Samples Storage Conditions for Reproducible Intraoperative Lipid Profiling.

Ex-vivo molecular profiling has recently emerged as a promising method for intraoperative tissue identification, especially in neurosurgery. The short-term storage of resected samples at room temperature is proposed to have negligible influence on the lipid molecular profiles. However, a detailed investigation of short-term molecular profile stability is required to implement molecular profiling in a clinic. This study evaluates the effect of storage media, temperature, and washing solution to determine conditions that provide stable and reproducible molecular profiles, with the help of ambient ionization mass spectrometry using rat cerebral cortex as model brain tissue samples. Utilizing normal saline for sample storage and washing media shows a positive effect on the reproducibility of the spectra; however, the refrigeration shows a negligible effect on the spectral similarity. Thus, it was demonstrated that up to hour-long storage in normal saline, even at room temperature, ensures the acquisition of representative molecular profiles using ambient ionization mass spectrometry.

Comparison of Dimensionality Reduction Methods in Mass Spectra of Astrocytoma and Glioblastoma Tissues.

Recently developed methods of ambient ionization allow the collection of mass spectrometric datasets for biological and medical applications at an unprecedented pace. One of the areas that could employ such analysis is neurosurgery. The fast in situ identification of dissected tissues could assist the neurosurgery procedure. In this paper tumor tissues of astrocytoma and glioblastoma are compared. The vast majority of the data representation methods are hard to use, as the number of features is high and the amount of samples is limited. Furthermore, the ratio of features and samples number restricts the use of many machine learning methods. The number of features could be reduced through feature selection algorithms or dimensionality reduction methods. Different algorithms of dimensionality reduction are considered along with the traditional noise thresholding for the mass spectra. From our analysis, the Isomap algorithm appears to be the most effective dimensionality reduction algorithm for negative mode, whereas the positive mode could be processed with a simple noise reduction by a threshold. Also, negative and positive mode correspond to different sample properties: negative mode is responsible for the inner variability and the details of the sample, whereas positive mode describes measurement in general.

Interactive Estimation of Heterogeneity from Mass Spectrometry Imaging.

In this work, we demonstrate a new approach for interactively assessing hyperspectral data spatial structures for heterogeneity using mass spectrometry imaging. This approach is based on the visualization of the cosine distance as the similarity levels between mass spectra of a chosen region and the rest of the image (sample). The applicability of the method is demonstrated on a set of mass spectrometry images of frontal mouse brain slices. Selection of the reference pixel of the mass spectrometric image and a further view of the corresponding cosine distance map helps to prepare supporting vectors for further analysis, select features, and carry out biological interpretation of different tissues in the mass spectrometry context with or without histological annotation. Visual inspection of the similarity maps reveals the spatial distribution of features in tissue samples, which can serve as the molecular histological annotation of a slide.

Assessment of variation of inline cartridge extraction mass spectra.

Recently, mass-spectrometry methods show its utility in tumor boundary location. The effect of differences between research and clinical protocols such as low- and high-resolution measurements and sample storage have to be understood and taken into account to transfer methods from bench to bedside. In this study, we demonstrate a simple way to compare mass spectra obtained by different experimental protocols, assess its quality, and check for the presence of outliers and batch effect in the dataset. We compare the mass spectra of both fresh and frozen-thawed astrocytic brain tumor samples obtained with the inline cartridge extraction prior to electrospray ionization. Our results reveal the importance of both positive and negative ion mode mass spectrometry for getting reliable information about sample diversity. We show that positive mode highlights the difference between protocols of mass spectra measurement, such as fresh and frozen-thawed samples, whereas negative mode better characterizes the histological difference between samples. We also show how the use of similarity spectrum matrix helps to identify the proper choice of the measurement parameters, so data collection would be kept reliable, and analysis would be correct and meaningful.

Inline cartridge extraction for rapid brain tumor tissue identification by molecular profiling.

The development of perspective diagnostic techniques in medicine requires efficient high-throughput biological sample analysis methods. Here, we present an inline cartridge extraction that facilitates the screening rate of mass spectrometry shotgun lipidomic analysis of tissue samples. We illustrate the method by its application to tumor tissue identification in neurosurgery. In perspective, this high-performance method provides new possibilities for the investigation of cancer pathogenesis and metabolic disorders.

Untangling the Metabolic Reprogramming in Brain Cancer: Discovering Key Molecular Players Using Mass Spectrometry.

Cells metabolism alteration is the new hallmark of cancer, as well as an important method for carcinogenesis investigation. It is well known that the malignant cells switch to aerobic glycolysis pathway occurring also in healthy proliferating cells. Recently, it was shown that in malignant cells de novo synthesis of the intracellular fatty acid replaces dietary fatty acids which change the lipid composition of cancer cells noticeably. These alterations in energy metabolism and structural lipid production explain the high proliferation rate of malignant tissues. However, metabolic reprogramming affects not only lipid metabolism but many of the metabolic pathways in the cell. 2-hydroxyglutarate was considered as cancer cell biomarker and its presence is associated with oxidative stress influencing the mitochondria functions. Among the variety of metabolite detection methods, mass spectrometry stands out as the most effective method for simultaneous identification and quantification of the metabolites. As the metabolic reprogramming is tightly connected with epigenetics and signaling modifications, the evaluation of metabolite alterations in cells is a promising approach to investigate the carcinogenesis which is necessary for improving current diagnostic capabilities and therapeutic capabilities. In this paper, we overview recent studies on metabolic alteration and oncometabolites, especially concerning brain cancer and mass spectrometry approaches which are now in use for the investigation of the metabolic pathway.

Feature selection algorithm for spray-from-tissue mass spectrometry.

Detection of the brain tumor margins is one of the most significant problems in neurosurgery. Several mass spectrometry-based approaches have been proposed recently for tumor boundary detection. One of them, spray from tissue does not require sample preparation but needs special algorithms for analysis of its spectra. Here we proposed the feature selection algorithm designed for analysis of spray-from-tissue data.

A novel direct spray-from-tissue ionization method for mass spectrometric analysis of human brain tumors.

Real-time feedback about dissected tissue during the neurosurgical procedure is strongly requested. A novel direct ionization mass spectrometric method for identifying pathological differences in tissues is proposed. The method is based on simultaneous extraction of tissue lipids and electrospray ionization which allows mass spectrometric data to be obtained directly from soft tissues. The advantage of this method is the stable flow of solvent, which leads to stable time-dependent spectra. The tissues included necrotized tissues and tumor tissues in different combinations. Capability for direct analysis of samples of dissected tissues during the neurosurgical procedure is demonstrated. Data validation is conducted by compound identification using precise masses from the MS profile, MS/MS, and isotopic distribution structure analysis. The method can be upgraded and applied for real-time identification of tissues during surgery. This paper describes the technique and its application perspective. For these purposes, other methods were compared with the investigated one and the results were shown to be reproducible. Differences in lipid profiles were observed even in tissues from one patient where distinctions between different samples could be poor. The paper presents a proof of concept for the method to be applied in neurosurgery particularly and in tissue analysis generically. The paper also contains preliminary results proving the possibility of observing differences in mass spectra of different tumors.