Supplemental Information
An Interactive Cluster Heat Map to Visualize and Explore
Multidimensional Metabolomic Data
Paul H. Benton†*, Julijana Ivanisevic†*, Duane Rinehart†, Adrian Epstein§, Michael E.
Kurczy†, Michael D. Boska+, Howard E. Gendelman§ and Gary Siuzdak†*
†
Scripps Center for Metabolomics, The Scripps Research Institute, 10550 North Torrey Pines
Road, La Jolla, CA 92037, United States
§
Departments of Pharmacology and Experimental Neuroscience and +Radiology, University
of Nebraska Medical Center, Omaha, NE 68198-5880, United States
AUTHOR INFORMATION
*these authors contributed equally
Corresponding Author
E-mail: siuzdak@scripps.edu, phone : (858) 784-9415, fax : (858) 784-9496
ABBREVIATED TITLE
An Interactive Heat Map for Metabolomic Data Visualization
MATERIALS AND METHODS
Animals and Tissue preparation. NOD scid IL2 receptor gamma chain knockout,
NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ, (NSG) mice (The Jackson Laboratories, Bar Harbor,
Maine, USA; stock number 005557) were obtained from an established breeding colony and
housed under pathogen-free conditions in accordance with ethical guidelines for care of
laboratory animals at the National Institutes of Health and the University of Nebraska
Medical Center (Gorantla et al. 2012).
Mice were anesthetized with 1–2 % isoflurane in oxygen then aligned in waterjacketed holder for microwave irradiation in a Muromachi Microwave Fixation System
(10 kW model). Irradiation time was 800 ms at 4.9 kW (Epstein et al. 2013). Single voxel
localized spectra were acquired at the midbrain using point resolved spectroscopy (PRESS).
Spectra were acquired with a repetition time of 4 seconds, echo time of 50 ms, 128 averages,
using volume coil transmit and birdcage coil receive on a 7 Tesla/16 cm Bruker Pharmascan
(Karlsure, Germany) MRI/MRS system. Single-scan, localized, unsuppressed water signals
were acquired as a reference for metabolite normalization.
Sub-regional dissection followed anatomical boundaries to separate hemi-brains into
cerebellum, hindbrain (pons and medulla), cerebral cortex, hippocampus, striatum, and
midbrain (midbrain and thalamus). Cortex was further divided into frontal, dorsal
(somatosensory through visual areas) and ventral (pirriform through perirhinal areas)
sections. Following dissection all tissues were flash frozen in liquid nitrogen and stored at -80
Celcius.
Chemical materials. Ammonium acetate (NH4Ac) and ammonium hydroxide
(NH4OH) were purchased from Sigma Aldrich. Acetonitrile was purchased from Fisher
Scientific (Morris Plains, NJ, USA), and water was purchased from J.T. Baker (Phillipsburg,
NJ, USA).
Supplementary Figure 1. Interactive XCMS Online screen shot displaying the results for the
untargeted multigroup comparison of anatomical regions of mouse brain. The top panels provide an
overlay of total ion chromatograms before and after (Aligned TICs) the retention time correction
including the retention time deviation curves across all runs. The lower panels present the cloud plot,
multidimensional scaling (MDS) and principal component analysis (PCA) output. The interactive
tools are listed on the left as clickable panels that enable the detailed exploration of data through the
interactive visualization of cloud plot, heat map or PCA.
REFERENCES
Epstein, A. A., Narayanasamy, P., Dash, P. K., High, R., Bathena, S. P., Gorantla, S., et al.
(2013). Combinatorial assessments of brain tissue metabolomics and histopathology
in rodent models of human immunodeficiency virus infection. Journal of
neuroimmune pharmacology : the official journal of the Society on NeuroImmune
Pharmacology, 8(5), 1224-1238.
Gorantla, S., Poluektova, L., & Gendelman, H. E. (2012). Rodent models for HIV-associated
neurocognitive disorders. Trends Neurosci, 35(3), 197-208.