3D Multi-isotope Imaging Mass Spectrometry Reveals Penetration of 18O^{18}O-Trehalose in Mouse Sperm Nucleus

The prevalence of genetically engineered mice in medical research has led to ever increasing storage costs. Trehalose has a significant beneficial effect in preserving the developmental potential of mouse sperm following partial desiccation and storage at temperatures above freezing. Using multi-isotope imaging mass spectrometry, we are able to image and measure trehalose in individual spermatozoa. We provide the first evidence that trehalose penetrates the nucleus of a mammalian cell, permitting tolerance to desiccation. These results have broad implications for long-term storage of mammalian cells

Antioxidant Role for Lipid Droplets in a Stem Cell Niche of Drosophila

SummaryStem cells reside in specialized microenvironments known as niches. During Drosophila development, glial cells provide a niche that sustains the proliferation of neural stem cells (neuroblasts) during starvation. We now find that the glial cell niche also preserves neuroblast proliferation under conditions of hypoxia and oxidative stress. Lipid droplets that form in niche glia during oxidative stress limit the levels of reactive oxygen species (ROS) and inhibit the oxidation of polyunsaturated fatty acids (PUFAs). These droplets protect glia and also neuroblasts from peroxidation chain reactions that can damage many types of macromolecules. The underlying antioxidant mechanism involves diverting PUFAs, including diet-derived linoleic acid, away from membranes to the core of lipid droplets, where they are less vulnerable to peroxidation. This study reveals an antioxidant role for lipid droplets that could be relevant in many different biological contexts

High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry

Background: Secondary-ion mass spectrometry (SIMS) is an important tool for investigating isotopic composition in the chemical and materials sciences, but its use in biology has been limited by technical considerations. Multi-isotope imaging mass spectrometry (MIMS), which combines a new generation of SIMS instrument with sophisticated ion optics, labeling with stable isotopes, and quantitative image-analysis software, was developed to study biological materials. Results: The new instrument allows the production of mass images of high lateral resolution (down to 33 nm), as well as the counting or imaging of several isotopes simultaneously. As MIMS can distinguish between ions of very similar mass, such as ^{12}C^{15}N^{-} and ^{13}C^{14}N^{-}, it enables the precise and reproducible measurement of isotope ratios, and thus of the levels of enrichment in specific isotopic labels, within volumes of less than a cubic micrometer. The sensitivity of MIMS is at least 1,000 times that of ^{14}C autoradiography. The depth resolution can be smaller than 1 nm because only a few atomic layers are needed to create an atomic mass image. We illustrate the use of MIMS to image unlabeled mammalian cultured cells and tissue sections; to analyze fatty-acid transport in adipocyte lipid droplets using ^{13}C-oleic acid; to examine nitrogen fixation in bacteria using ^{15}N gaseous nitrogen; to measure levels of protein renewal in the cochlea and in post-ischemic kidney cells using ^{15}N-leucine; to study DNA and RNA co-distribution and uridine incorporation in the nucleolus using ^{15}N-uridine and ^{81}Br of bromodeoxyuridine or ^{14}C-thymidine; to reveal domains in cultured endothelial cells using the native isotopes ^{12}C, ^{16}O, ^{14}N and ^{31}P; and to track a few ^{15}N-labeled donor spleen cells in the lymph nodes of the host mouse. Conclusion: MIMS makes it possible for the first time to both image and quantify molecules labeled with stable or radioactive isotopes within subcellular compartments

MIMS images from 3 planes and data from a full-thickness study.

<p><b>A</b>) ¹²C¹⁴N quantitative atomic mass images of a mouse sperm head extracted from planes 30, 100 and 200 out of 284 (image field = 15 µm×15 µm; 256×256 pixels; acquisition time per plane = 5.5 mins; total acquisition time = 27 hrs). Green circles: ROIs in sperm head (“N”) and in trehalose film (“T”) used to generate plots of ¹⁸O/¹⁶O values and ²⁸Si counts. <b>B</b>) ¹⁸O/¹⁶O ratios (circles) and ²⁸Si counts (diamonds) extracted from each plane from the ROIs shown in (A). The spikes in the ²⁸Si curves show the point at which the trehalose film (red diamonds) and the sperm head (orange diamonds) have been completely sputtered through. The vertical grey stripe indicates end of sperm head sputtering and the start of silicon exposure.</p