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Androgens are essential for male development and reproductive function. They are transported to their site of action as blood-bome endocrine hormones but can also be produced within tissues to act in intracrine and paracrine fashions. Because of this, circulating concentrations may not accurately reflect the androgenic influence within specific tissue microenvironments. Mass spectrometry imaging permits regional analysis of small molecular species directly from tissue surfaces. However, due to poor ionization and localized ion suppression, steroid hormones are difficult to detect Here, derivatization with Girard T reagent was used to charge-tag testosterone and 5α-dihydrotestosterone allowing direct detection of these steroids in mouse testes, in both basal and maximally stimulated states, and in rat prostate. Limits of detection were ~0.1 pg for testosterone. Exemplary detection of endogenous steroids was achieved by matrix-assisted laser desorption ionization and either Fourier transform ion cyclotron resonance detection (at 150 μm spatial resolution) or quadrupole-time-of-flight detection (at 50 μm spatial resolution). Structural confirmation was achieved by collision induced fragmentation following liquid extraction surface analysis and electrospray ionization. This application broadens the scope for derivatization strategies on tissue surfaces to elucidate local endocrine signaling in health and disease. [ABSTRACT FROM AUTHOR]

Steroid concentrations within tissues are modulated by intracellular enzymes. Such “steroid intracrinology” influences hormone-dependent cancers and obesity and provides targets for pharmacological inhibition. However, no high resolution methods exist to quantify steroids within target tissues. We developed mass spectrometry imaging (MSI), combining matrix assisted laser desorption ionization with on-tissue derivatization with Girard T and Fourier transform ion cyclotron resonance mass spectrometry, to quantify substrate and product (11-dehydrocorticosterone and corticosterone) of the glucocorticoid-amplifying enzyme 11β-HSD1. Regional steroid distribution was imaged at 150–200 μm resolution in rat adrenal gland and mouse brain sections and confirmed with collision induced dissociation/liquid extraction surface analysis. In brains of mice with 11β-HSD1 deficiency or inhibition, MSI quantified changes in subregional corticosterone/11-dehydrocorticosterone ratio, distribution of inhibitor, and accumulation of the alternative 11β-HSD1 substrate, 7-ketocholesterol. MSI data correlated well with LC-MS/MS in whole brain homogenates. MSI with derivatization is a powerful new tool to investigate steroid biology within tissues. [ABSTRACT FROM AUTHOR]

Vitamin D plays a key role in the maintenance of calcium/phosphate homeostasis and elicits biological effects that are relevant to immune function and metabolism. It is predominantly formed through UV exposure in the skin by conversion of 7-dehydrocholsterol (vitamin D3). The clinical biomarker, 25-hydroxyvitamin D (25-(OH)-D), is enzymatically generated in the liver with the active hormone 1,25-dihydroxyvitamin D then formed under classical endocrine control in the kidney. Vitamin D metabolites are measured in biomatrices by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In LC-MS/MS, chemical derivatization (CD) approaches have been employed to achieve the desired limit of quantitation. Recently, matrix-assisted laser desorption/ionization (MALDI) has also been reported as an alternative method. However, these quantitative approaches do not offer any spatial information. Mass spectrometry imaging (MSI) has been proven to be a powerful tool to image the spatial distribution of molecules from the surface of biological tissue sections. On-tissue chemical derivatization (OTCD) enables MSI to image molecules with poor ionization efficiently. In this technical report, several derivatization reagents and OTCD methods were evaluated using different MSI ionization techniques. Here, a method for detection and spatial distribution of vitamin D metabolites in murine kidney tissue sections using an OTCD-MALDI-MSI platform is presented. Moreover, the suitability of using the Bruker ImagePrep for OTCD-based platforms has been demonstrated. Importantly, this method opens the door for expanding the range of other poor ionizable molecules that can be studied by OTCD-MSI by adapting existing CD methods., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)