Use Of Imaging Mass Spectrometry Research Articles

Abstract AP02: EXPOSING THE LOCAL FACTORS THAT CONTRIBUTE TO OVARIAN COLONIZATION OF FALLOPIAN TUBE-DERIVED TUMORS

Abstract High grade serous cancer (HGSC) originates in the fallopian tube and typically invade and colonize the ovary as part of peritoneal metastasis. The purpose of this study was to define the factors that contribute to ovarian specific metastasis of fallopian tube HGSC. While protein factors have been explored that facilitate migration and invasion of fallopian tube cell to the ovary, the small molecule communication between the tissues during metastasis remains largely unexplored. Due to the proximity of ovarian primary metastasis, an experimental design was optimized using imaging mass spectrometry to capture the spatial composition of small molecules uniquely expressed when fallopian tube-derived tumor cells were grown in the microenvironment of the ovary as a model of primary metastasis. The observed mass-to-charge ratios (m/z's) that were induced specifically in co-culture represent small molecules that may contribute to the metastasis of HGSOC selectively to the ovary. Human fallopian tube epithelial HGSOC and tumorigenic murine oviductal epithelial cells, but not normal cell types, repeatedly induced a signal from the ovary at m/z 170. This signal was identified as norepinephrine, which was confirmed to simulate invasion of ovarian cancer cells lacking wildtype p53. Norepinephrine has an established role in ovarian cancer metastasis to secondary sites and the use of beta-blockers in women is associated with increased survival. The use of imaging mass spectrometry also revealed a series of new molecules that are specifically enhanced from the ovary or the fallopian tube during co-culture. These molecules may reveal pathways that contribute to metastasis and biological targets for therapeutic intervention to block ovarian metastasis of fallopian tube-derived HGSOC. The developed mass spectrometry method can be adapted to other mammalian-based model systems for investigation of untargeted metabolomics that facilitate metastasis. Citation Format: Joanna E. Burdette, Katherine Zink, Matthew Dean, and Laura M. Sanchez. EXPOSING THE LOCAL FACTORS THAT CONTRIBUTE TO OVARIAN COLONIZATION OF FALLOPIAN TUBE-DERIVED TUMORS [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr AP02.

Single-Cell Mass Spectrometry Reveals Changes in Lipid and Metabolite Expression in RAW 264.7 Cells upon Lipopolysaccharide Stimulation.

It has been widely recognized that individual cells that exist within a large population of cells, even if they are genetically identical, can have divergent molecular makeups resulting from a variety of factors, including local environmental factors and stochastic processes within each cell. Presently, numerous approaches have been described that permit the resolution of these single-cell expression differences for RNA and protein; however, relatively few techniques exist for the study of lipids and metabolites in this manner. This study presents a methodology for the analysis of metabolite and lipid expression at the level of a single cell through the use of imaging mass spectrometry on a high-performance Fourier transform ion cyclotron resonance mass spectrometer. This report provides a detailed description of the overall experimental approach, including sample preparation as well as the data acquisition and analysis strategy for single cells. Applying this approach to the study of cultured RAW264.7 cells, we demonstrate that this method can be used to study the variation in molecular expression with cell populations and is sensitive to alterations in that expression that occurs upon lipopolysaccharide stimulation. Graphical Abstract.

The Use of Imaging Mass Spectrometry to Study Peptide Toxin Distribution in Australian Sea Anemones

The Use of Imaging Mass Spectrometry to Study Peptide Toxin Distribution in Australian Sea Anemones

Erratum to: Microscopic visualization of testosterone in mouse testis by use of imaging mass spectrometry.

1 Division of Advanced Science and Biotechnology, Graduate School of Engineering of Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan 2 Department of Medical Chemistry, Kansai Medical University, 2-5-1 Shin-machi, Hirakata, Osaka 573-1010, Japan 3 Department of Urology and Andrology, Kansai Medical University, 2-5-1 Shin-machi, Hirakata, Osaka 573-1010, Japan 4 Department of Optical Imaging, Institute for Medical Photonics Research, Preeminent Medical Photonics Education & Research Center, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu, Shizuoka 431-3192, Japan

Microscopic visualization of testosterone in mouse testis by use of imaging mass spectrometry.

Testosterone is one of the androgens synthesized from cholesterol as a precursor in the Leydig cells of testes. Since the ionization efficiency of testosterone in matrix-assisted laser desorption ionization (MALDI) is quite low, visualization of testosterone by using MALDI-imaging mass spectrometry (MALDI-IMS) has been considered difficult. To overcome this problem, we used two types of on-tissue derivatization techniques, which were achieved by pyridine sulfur trioxide and Girard's T (GT) reagent, to introduce a polar group into testosterone molecule with the aim to increase the sensitivity. Derivatization by use of GT reagent provided excellent results, superior to those obtained with pyridine sulfur trioxide, in terms of ionization efficiency, molecular specificity, and tissue damage. In GT derivatized testis tissues of mice treated with human chorionic gonadotropin (hCG), testosterone was broadly observed both inside and outside the seminiferous tubules by using an iMScope. To evaluate our imaging results, we performed quantification experiments of underivatized testosterone extracted from hCG-treated testes and control testes using LC-MS/MS. We confirmed the 256-fold concentration change between hCG-treated tissues and control tissues. We also confirmed the 228-fold change in detected peak intensities between hCG-treated tissue sections and control tissue sections in imaging results. We consider our tissue preparation methods for IMS provide high sensitivity with high precision. In addition, high-spatial definition IMS was also available, and we confirmed testosterone had mainly accumulated on the surface of the Leydig cells. Graphical abstract Girard's T-testosterone (GT-Ts) provides the fragment ion at m/z 343.24. Clear GT-Ts signal was detected in hCG treated mouse testis not only as spectra but also as a mass image.

MALDI imaging mass spectrometry of Pacific White Shrimp L. vannamei and identification of abdominal muscle proteins.

MALDI imaging mass spectrometry (IMS) has been applied to whole animal tissue sections of Pacific White Shrimp, Litopenaeus vannamei, in an effort to identify and spatially localize proteins in specific organ systems. Frozen shrimp were sectioned along the ventral-dorsal axis and methods were optimized for matrix application. In addition, tissue microextraction and homogenization was conducted followed by top-down LC-MS/MS analysis of intact proteins and searches of shrimp EST databases to identify imaged proteins. IMS images revealed organ system specific protein signals that highlighted the hepatopancreas, heart, nervous system, musculature, and cuticle. Top-down proteomics identification of abdominal muscle proteins revealed the sequence of the most abundant muscle protein that has no sequence homology to known proteins. Additional identifications of abdominal muscle proteins included titin, troponin-I, ubiquitin, as well as intact and multiple truncated forms of flightin; a protein known to function in high frequency contraction of insect wing muscles. The combined use of imaging mass spectrometry and top-down proteomics allowed for identification of novel proteins from the sparsely populated shrimp protein databases.

Imaging Mass Spectrometry Reveals a Decrease of Cardiolipin in the Kidney of NASH Model Mice.

Non-alcoholic steatohepatitis (NASH) can be complicated with chronic kidney disease (CKD). In this study, changes in the distribution of biomolecules in the kidney were studied in NASH model mice with the use of imaging mass spectrometry (IMS). The mass spectra and ion images of IMS showed that the signals of cardiolipin (CL) species were decreased in the kidney cortex of the NASH mice. The decrease of CL might therefore suggest the kidney involvement of NASH.

Alzheimer’s disease is associated with disordered localization of ganglioside GM1 molecular species in the human dentate gyrus

Alzheimer’s disease (AD) is a progressive dementia associated with loss of memory and cognitive dysfunction. In a previous study, we demonstrated a decrease in b-series gangliosides along with a change in ganglioside molecular species in the hippocampal grey matter of patients with AD. The present study demonstrates the use of imaging mass spectrometry for analyzing the spatial arrangement of ganglioside GM1 (GM1) molecular species in the hippocampus. In AD patients, we found a decrease in the ratio of GM1(d20:1/C18:0) to GM1 d18:1/C18:0) in the outer molecular layer (ML) of the dentate gyrus. Because the outer ML is the region of main input into the hippocampus, our findings may have a direct relationship to the mechanism of dysfunction in AD.

Proteomic Analysis of Oral Cavity Squamous Cell Carcinoma Specimens Identifies Patient Outcome–Associated Proteins

Global proteomic analysis of oral cavity squamous cell carcinoma was performed to identify changes that reflect patient outcomes. To identify differentially expressed proteins associated with patient outcomes and to explore the use of imaging mass spectrometry as a clinical tool to identify clinically relevant proteins. Two-dimensional separation of digested peptides generated from 43 specimens with high-resolution mass spectrometry identified proteins associated with disease-specific death, distant metastasis, and loco-regional recurrence. RNA expressions had been correlated to protein levels to test transcriptional regulation of clinically relevant proteins. Imaging mass spectrometry explored an alternative platform for assessing clinically relevant proteins that would complement surgical pathologic diagnosis. Seventy-two peptide features were found to be associated with 3 patient outcomes: disease-specific death (9), distant metastasis (16), and loco-regional recurrence (39); 8 of them were associated with multiple outcomes. Functional ontology revealed major changes in cell adhesion and calcium binding. Thirteen RNAs showed strong correlation with their encoded proteins, implying transcriptional control. Reduction of DSP, PKP1, and TRIM29 was associated with significantly shorter time to onset of distant metastasis. Reduction of PKP1 and TRIM29 correlated with poorer disease-specific survival. Additionally, S100A8 and S100A9 reductions were verified for their association with poor prognosis using imaging mass spectrometry, a platform more adaptable for use with surgical pathology. Using global proteomic analysis, we have identified proteins associated with clinical outcomes. The list of clinically relevant proteins observed will provide a means to develop clinical assays for prognosis and optimizing treatment selection.

Development of laser ionization techniques for evaluation of the effect of cancer drugs using imaging mass spectrometry.

Recently, combined therapy using chemotherapy and photodynamic therapy (PDT) has been proposed as a means of improving treatment outcomes. In order to evaluate the efficacy of combined therapy, it is necessary to determine the distribution of the anticancer drug and the photosensitizer. We investigated the use of imaging mass spectrometry (IMS) to simultaneously observe the distributions of an anticancer drug and photosensitizer administered to cancer cells. In particular, we sought to increase the sensitivity of detection of the anticancer drug docetaxel and the photosensitizer protoporphyrin IX (PpIX) by optimizing the ionization-assisting reagents. When we used a matrix consisting of equal weights of a zeolite (NaY5.6) and a conventional organic matrix (6-aza-2-thiothymine) in matrix-assisted laser desorption/ionization, the signal intensity of the sodium-adducted ion of docetaxel (administered at 100 μM) increased about 13-fold. Moreover, we detected docetaxel with the zeolite matrix using the droplet method, and detected PpIX by fluorescence and IMS with α-cyano-4-hydroxycinnamic acid (CHCA) using the spray method.

Organic ion imaging of biological tissue with secondary ion mass spectrometry and matrix-assisted laser desorption/ionization.

Organic secondary ion mass spectrometry (SIMS) and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry can be used to produce molecular images of samples. This is achieved through ionization from a clearly identified point on a flat sample, and performing a raster of the sample by moving the point of ionization over the sample surface. The unique analytical capabilities of mass spectrometry for mapping a variety of biological samples at the tissue level are discussed. SIMS provides information on the spatial distribution of the elements and low molecular mass compounds as well as molecular structures on these compounds, while MALDI yields spatial information about higher molecular mass compounds, including their distributions in tissues at very low levels, as well as information on the molecular structures of these compounds. Application of these methods to analytical problems requires appropriate instrumentation, sample preparation methodology, and a data presentation usually in a three-coordinate plot where x and y are physical dimensions of the sample and z is the signal amplitude. The use of imaging mass spectrometry is illustrated with several biological systems.