Identification and Relative Quantification of Neuropeptides from the Endocrine Tissues

Abstract

Endocrine tissues like the pituitary, hypothalamus and islets of Langerhans are rich in bioactive peptides. These are used for intercellular signalling and are involved in regulation of almost all physiological processes. Peptidomics is the comprehensive analysis of peptides in tissues, fluids and cells. Peptidomics applied to (neuro-)endocrine tissues aims therefore to identify as many bioactive peptides as possible. Peptidomics of (neuro-)endocrine tissues requires an integrated approach that consists of careful sample handling, peptide separation techniques, mass spectrometry and bioinformatics. Here we describe the methods for isolation and dissection of endocrine tissues, the extraction of bioactive peptides and further sample handling and identification of peptides by mass spectrometry and hyphenated techniques. We also present a straightforward method for the comparison of relative levels of bioactive peptides in these endocrine tissues under varying physiological conditions. The latter helps to elucidate functions of the bioactive peptides.

Platelets of mice heterozygous for neurobeachin, a candidate gene for autism spectrum disorder, display protein changes related to aberrant protein kinase A activity

“…Platelets were isolated as described above with modification of the final wash buffer, which was now replaced by PBS. The procedure for processing the platelet pellets (n = 5 mice/genotype) for mass spectrometry analysis was performed as reported [ 36 ]. DeCyder MS 2.0 (GE Healthcare) is a differential analysis software tool that also allows for easy visualization of liquid chromatography mass spectrometry (LC-MS) runs by creating artificial two-dimensional maps with the m/z values and retention times in the first and second dimension, respectively.…”

“…Time alignment, intensity normalization and statistics were performed using this software. Peptides were identified in additional LC-MS/MS runs of the pooled samples as reported in [ 36 ] using LC quadrupole time-of-flight (Q-TOF) MS.…”

Linking Mass Spectrometric Imaging and Traditional Peptidomics: A Validation in the Obese Mouse Model

“…One half of each pancreas ( n = 6, with n = 3 wt and n = 3 ob / ob samples) was snap-frozen in liquid nitrogen-cooled isopentane and stored prior to crysectioning for subsequent MALDI-MSI analysis of the regions of interest (ROIs), the islets of Langerhans. The second half of each pancreas ( n = 6) was immediately enzymatically digested with collagenase P in order to first specifically isolate the islets of Langerhans, followed by a peptide extraction protocol according to Boonen et al In this way all possible acini-related peptides were excluded from further analysis and interpretation.…”

Crystal structure of Porphyromonas gingivalis dipeptidyl peptidase 4 and structure-activity relationships based on inhibitor profiling

“…The University of Leuven collection of putative neuropeptides and endocrine peptides was based on a combination of peptidomics results and bioinformatics [45,46]. Peptides with a penultimate proline, alanine, glycine or serine were chosen for this study.…”