Characterization of lipidic markers of chondrogenic differentiation using mass spectrometry imaging

Abstract

Mesenchymal stem cells (MSC) are an interesting alternative for cell-based therapy of cartilage defects attributable to their capacity to differentiate toward chondrocytes in the process termed chondrogenesis. The metabolism of lipids has recently been associated with the modulation of chondrogenesis and also with the development of pathologies related to cartilage degeneration. Information about the distribution and modulation of lipids during chondrogenesis could provide a panel of putative chondrogenic markers. Thus, the discovery of new lipid chondrogenic markers could be highly valuable for improving MSC-based cartilage therapies. In this work, MS imaging was used to characterize the spatial distribution of lipids in human bone marrow MSCs during the first steps of chondrogenic differentiation. The analysis of MSC micromasses at days 2 and 14 of chondrogenesis by MALDI-MSI led to the identification of 20 different lipid species, including fatty acids, sphingolipids, and phospholipids. Phosphocholine, several sphingomyelins, and phosphatidylcholines were found to increase during the undifferentiated chondrogenic stage. A particularly detected lipid profile was verified by TOF secondary ion MS. Using this technology, a higher intensity of phosphocholine-related ions was observed in the peripheral region of the micromasses collected at day 14.

Optimal bone fracture repair requires 24R,25-dihydroxyvitamin D3 and its effector molecule FAM57B2

“…repair phase seems to be affected by Cyp24a1 or Fam57b deletion, FAM57B2 might link vitamin D 3 metabolism to sphingolipid production to modulate Col2a1 and Acan expression. Increasing evidence highlights the role of sphingolipids in cartilage homeostasis (45,46). Indeed, sphingolipids have been shown to influence Col2a1 expression (47) as well as chondrocyte apoptosis and matrix turnover (48).…”

MALDI mass spectrometric imaging meets “omics”: recent advances in the fruitful marriage

“…146 And beyond this, the image fusion of extracted ion images with H&E stains generated by MALDI MSI and microscopy, respectively, can assist in predicting ion distributions in tissue areas that were not analyzed by MALDI MSI. 165 To go beyond the in situ visualization of metabolic changes, the utilization of additional chemical imaging technique, 166 has been shown as a promising tool, e.g. Within the spatial mapping of lipids, certain classes, such as phospholipids, 156 which can degrade in time, 36 sphingolipids, 157 and ceramides 158,159 have been extensively investigated.…”

Assessment of different sample preparation routes for mass spectrometric monitoring and imaging of lipids in bone cells via ToF-SIMS

“…29 Instead systematic investigations, especially those with a focus on lipids, still rely on other methods like glutaraldehyde fixation 30,31 or gelatin embedding. 9 In 2009, Malm et al compared cryofixation to chemical fixation via glutaraldehyde and freeze-drying (FD) to alcohol drying. 26 In one preparation, osmium tetroxide fixation was performed prior to alcohol drying.…”

“…Especially for cell cultures, this approach has yet not been explored in depth. 9 As ToF-SIMS analysis requires vacuum conditions, cell samples cannot be analyzed directly in the living or native state. 14,15 Sample preparation is thus a crucial factor to maintain sample integrity, and it has to be well adapted to the analytes of interest.…”

“…[1][2][3] On cellular level, lipids act as cell signaling molecules and are involved in the regulation of various cellular functions such as proliferation, apoptosis, inflammation, immunity, and even DNA modulation. 4 Lipids and their metabolism affect pathological processes like atherosclerosis, 5 osteoporosis, 6 cancer, 7 and diabetes 8 and are of potential value as novel biomarkers for such diseases on the one hand and for cellular processes like chondrogenesis, 9 on the other hand. It is thus of strong interest to explore lipidomic pathways in order to understand molecular relationships and especially disease mechanisms.…”