Chemical Isotope Labeling LC-MS for Monitoring Disease Progression and Treatment in Animal Models: Plasma Metabolomics Study of Osteoarthritis Rat Model

Abstract

We report a chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS) method generally applicable for tracking metabolomic changes from samples collected in an animal model for studying disease development and treatment. A rat model of surgically induced osteoarthritis (OA) was used as an example to illustrate the workflow and technical performance. Experimental duplicate analyses of 234 plasma samples were carried out using dansylation labeling LC-MS targeting the amine/phenol submetabolome. These samples composed of 39 groups (6 rats per group) were collected at multiple time points with sham operation, OA control group, and OA rats with treatment, separately, using glucosamine/Celecoxib and three traditional Chinese medicines (Epimedii folium, Chuanxiong Rhizoma and Bushen-Huoxue). In total, 3893 metabolites could be detected and 2923 of them were consistently detected in more than 50% of the runs. This high-coverage submetabolome dataset could be used to track OA progression and treatment. Many differentiating metabolites were found and 11 metabolites including 2-aminoadipic acid, saccharopine and GABA were selected as potential biomarkers of OA progression and OA treatment. This study illustrates that CIL LC-MS is a very useful technique for monitoring incremental metabolomic changes with high coverage and accuracy for studying disease progression and treatment in animal models.

High tolerance to instrument drifts by differential chemical isotope labeling LC‐MS: A case study of the effect of LC leak in long‐term sample runs on quantitative metabolome analysis

“…The rat model used for metabolomics study of rheumatoid arthritis (RA) has been described previously. 18 The samples collected from the rats consisted of 7 groups, including a group of normal controls (n = 96), RA, and other treated rats. In total, there were 468 samples, which were processed and analyzed in three batches.…”

“…Each sample was labeled using 12 C-dansylation after protein precipitation, and the resultant labeled sample was analyzed by LC-UV to measure the total concentration of labeled metabolites for sample amount normalization. 18,19 Based on the total concentration, an appropriate volume of an individual unlabeled sample was taken to mix with an equal mole amount of other unlabeled samples to generate a pooled sample, which was then labeled by 13 C-dansylation. An equal mole amount of the 12 C-labeled individual sample and the 13 Clabeled pooled sample was mixed for LC-TOF-MS analysis.…”

Hair Metabolomics in Animal Studies and Clinical Settings

“…Well-controlled animal models offer inherent phenotypes for specific diseases or abnormal conditions. Therefore, biological samples from animal models are often used in metabolomics studies [86,87,88]. However, only a few metabolomics studies have been conducted using animal hair.…”

The Metabolome and Osteoarthritis: Possible Contributions to Symptoms and Pathology

“…In rat models of OA, changes to amino acid metabolism are also observed in urine [41]. Again in rats, a longitudinal metabolomic analysis of plasma showed that there are significant and time-dependent changes in products of lysine metabolism after surgical induction of OA [42]. This further suggests that amino acid metabolism may contribute to OA pathology or that select amino acid levels could be used as biomarkers of OA incidence and progression.…”