Brum, F. L. | bioRxiv Science

bioRxiv2023

Spatially resolved distribution of pancreatic hormones proteoforms by MALDI-imaging mass spectrometry

Zinc plays crucial role in the immune system and endocrine processes. Dietary zinc restriction leads to the degeneration of the endocrine pancreas resulting in hormonal imbalance in {beta}-cell. Proteostasis may vary depending on the stage of a pathophysiological process, motivating the development of tools aimed at the direct analysis of biological status. Among proteomics methods, MALDI-ToF-MS can serve as a rapid peptidomics tool from analysis of extracts or by histological imaging. Here we report the optimization of MALDI imaging mass spectrometry analysis of thin histological sections from mouse pancreas, allowing the identification of major islet peptide hormones and major accumulated precursors and/or proteolytic products of peptide hormones. Cross-validation of the identified peptide hormones was performed by LC-ESI-MS from pancreatic islet extracts. Mice fed a zinc-restricted diet had a relatively lower amount of peptide intermediates in comparison with the control group. These data provide evidence for complex modulation of proteostasis by an imbalance of micronutrients, directly accessed by MALDI-MSI.

biochemistry↗

bioRxiv2022

Acquiring thin histological sections of acai(Euterpe oleracea Mart.) seeds for molecular mapping by mass spectrometry imaging

RATIONALEMatrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS) of tissues became popular in the last decade. Consequently, adapting sample preparation methods for different materials turned to be a pivotal step for a successful analysis, due to the requirement of samples slices of 12-20 m thickness. However, preparing thin sections compatible with MALDI-IMS for unusual samples is challenging, as existing histological protocols may not be suitable, thus requiring new methods. Acai (Euterpe oleracea Mart.) seed is an example of a challenging material due to its toughness and resistance to crack. Therefore, our goal was to develop a methodology to obtain thin (<20 m) and entire longitudinal sections of acai seeds for MALDI-IMS analysis. METHODSDifferent strategies were evaluated for obtaining thin cuts of seeds, being the combination of the following steps the most suitable option: (i) softening of seeds by water immersion for 24 h; (ii) longitudinal cut of seeds to obtain half-seeds using a razor blade and a hammer; (iii) half-seeds imbibition in gelatin; (iv) sectioning using a cryostat at -20 {degrees}C to obtain samples with <20 m thickness; and (v) collection of samples in an indium tin oxide coated glass slide covered by a double-face copper tape to avoid sample wrapping and ensure adhesion after unfreezing. (iv) storage of samples in a -80 {degrees}C freezer, if necessary. RESULTSAs a result, this adapted sample preparation method enabled the analysis of acai seeds by MALDI-IMS providing spatial distribution of carbohydrates in the endosperm. CONCLUSIONSThe adaptations developed for sample preparation will help investigate the metabolic and physiological properties of acai seeds in future studies.

plant biology↗