A comprehensive protocol for multiplatform metabolomics analysis in patient-derived skin fibroblasts

Area of Science:

• Biochemistry
• Cell Biology
• Translational Medicine

Background:

• Patient-derived skin fibroblasts are valuable models for studying human diseases.
• Metabolomics profiling reveals metabolic pathway alterations crucial for disease understanding and therapeutic assessment.
• Standardized protocols are essential for reliable metabolomics in fibroblasts.

Purpose of the Study:

• To develop an optimized protocol for measuring amino acids, acylcarnitines, and tricarboxylic acid (TCA) cycle metabolites in human skin fibroblasts.
• To establish a robust workflow for concurrent analysis using gas chromatography-mass spectrometry (GC/MS) and liquid chromatography-mass spectrometry (LC/MS).

Main Methods:

• Evaluated four cell harvesting methods for metabolite recovery using GC/MS and LC/MS.
• Assessed metabolite stability after storage at -80°C for up to one month.
• Focused on amino acids, acylcarnitines, and TCA cycle components.

Main Results:

• Harvesting cells in 80% methanol yielded the best metabolite recovery and preservation.
• Metabolite concentrations remained stable after storage in 80% methanol at -80°C for up to one month.
• Demonstrated suitability for high-throughput multiplatform analysis.

Conclusions:

• A robust workflow for human skin fibroblast metabolomics was developed.
• This method supports high-throughput, multiplatform analysis for comparative studies.
• Enables reliable metabolic assessment in large patient cohorts for disease research.