Filtration-Based Rapid ECM-Hydrogel Elimination Enables Robust MALDI MSI of Intestinal Organoids

Area of Science:

• Biomedical Engineering
• Metabolomics
• Gastroenterology

Background:

• Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) is crucial for 2D metabolic profiling of intestinal organoids.
• Current methods for extracellular matrix (ECM) hydrogel removal are time-consuming and can damage organoids and metabolites.

Purpose of the Study:

• To develop an efficient method for removing ECM hydrogels from intestinal organoids for improved MALDI MSI analysis.
• To assess the impact of the new method on organoid integrity, metabolic stability, and data quality.

Main Methods:

• Established a filtration-based rapid ECM-hydrogel elimination (FREE) method using a 70 μm filter.
• Compared FREE with conventional methods regarding time, organoid integrity, cell viability, metabolic stability, and data quality.
• Applied the FREE workflow to an LPS-induced inflammatory intestinal organoid model.

Main Results:

• The FREE method reduced sample preparation time from 75 to 25 minutes.
• FREE preserved organoid structure, cell viability, and metabolic stability, while improving spectral quality.
• Achieved a 26.6% increase in metabolite coverage and enhanced signal intensity compared to conventional methods.
• Observed spatial metabolic remodeling and altered lipid distribution (phosphatidylserine and phosphatidylethanolamine) in an inflammatory model, linked to polarity loss.

Conclusions:

• The FREE method is an efficient pretreatment strategy for MALDI MSI of intestinal organoids.
• This approach enhances data quality and enables the study of inflammation-associated metabolic remodeling and its link to cellular processes like apoptosis.