High Resolution Imaging of Adrenal Glands by DESI-MS

Significance of the Topic

The adrenal glands produce essential steroids that regulate metabolism, cardiovascular function and stress response. Alterations in adrenal lipid composition are linked to diseases such as Cushing’s syndrome, Addison’s disease and congenital adrenal hyperplasia. High-resolution spatial mapping of lipids in these glands can reveal molecular changes underlying pathology and guide biomarker discovery.

Aim and Overview of the Study

This feasibility study evaluates desorption electrospray ionization mass spectrometry (DESI-MS) for imaging mouse adrenal gland sections. Two mass spectrometers—a conventional quadrupole time-of-flight (QToF) and a high-resolution multi-reflecting time-of-flight (MRT) instrument—were compared. Statistical tools were applied to identify lipid markers distinguishing anatomical regions.

Methodology

Healthy wild-type murine adrenal glands were cryo-sectioned at 15 µm thickness. DESI-MS acquisitions were performed in full-scan mode: the QToF at 25 µm pixel size and 2 scans/s, the MRT at 50 µm pixel size and 2 scans/s. A 95:5 methanol:water spray solvent, 5 bar gas flow and 0.7 kV capillary voltage were used. Data processing included:

• High Definition Imaging (HDI) for ROI selection and visualization
• MetaboAnalyst for unsupervised PCA and marker discovery
• MassLynx elemental composition tool and LIPID MAPS for putative ID
• Lipostar MSI for database-driven annotation and image overlays

Main Results and Discussion

Unsupervised PCA separated medulla, cortex and outer capsule regions based on lipid profiles. RGB overlay images showed distinct localization of three marker masses: one in medulla, two in cortex. The MRT instrument, with >200 000 FWHM resolution, resolved a nominal m/z 806.55 signal into three separate peaks, enabling more confident assignment (e.g., PE 36:4, PC 38:6). Conventional QToF provided ~20 000 FWHM and displayed a single broad peak. Targeted MS/MS on m/z 166.086 confirmed phenylalanine in medulla.

Benefits and Practical Applications

• Non-destructive spatial analysis preserves tissue for histology
• Compatibility with conventional and high-resolution MS platforms
• Rapid discovery of regional lipid biomarkers
• Seamless integration into visualization and statistical pipelines
• Targeted MS/MS on same section enhances identification confidence

Future Trends and Applications

Advances may include multimodal imaging combining DESI-MS with microscopy, automated annotation using machine learning, expanded lipid libraries for disease models, translational studies in clinical diagnostics and real-time guided surgery applications.

Conclusion

This study demonstrates that DESI-MS reliably images adrenal gland lipids, differentiating tissue regions and identifying markers. High-resolution MRT instruments significantly enhance molecular specificity. The approach offers a powerful platform for adrenal disease research and biomarker screening.

Instrumentation Used

• DESI-XS source with High Performance sprayer (95:5 methanol:water)
• SYNAPT G2-XS QToF mass spectrometer
• SELECT SERIES MRT multi-reflecting ToF mass spectrometer
• Waters HDI, MassLynx and Lipostar MSI software
• MetaboAnalyst statistical package

References

• US Department of Health Sciences, NIH National Institute of Child Health and Human Development. Adrenal Gland Disorders. https://www.nichd.nih.gov/health/topics/adrenalgland/conditioninfo/types