Direct localisation of molecules in tissue sections of growing antler tips using MALDI imaging.

Abstract

The astonishing growth rate of deer antlers offers a valuable model for the discovery of novel factors and regulatory systems controlling rapid tissue growth. Numerous molecules have been identified in growing antlers using a variety of techniques. However, little is known about the spatial distribution of these molecules in situ. A technique that has the potential to help in this regard is direct proteomic analysis of tissue sections by matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS). The present study applied this technique to spatially map molecules in antler tissue sections. Two protonated molecular ions were selected: m/z 6679 and m/z 6200 corresponding to VEGF and thymosin beta-10, respectively. Superimposition of the respective ion images on to histologically stained samples showed distinct spatial distribution across the antler tissue sections which were consistent with the previous reports using in situ hybridization. Two other molecular ions specifically m/z 8100 and m/z 11,800 were also selected, corresponding to reported masses of urocortin precursor and thioredoxin, respectively. As the spatial distribution of these proteins is not specifically known, MALDI-IMS was used as a potential technique to obtain information on their distribution on antler tips. The presence of all these molecules in deer antlers were further confirmed using LC-MS/MS data. The present study also demonstrated that MALDI-IMS could be further used to image antler sections with an extended ion mass range of up to m/z 45,000, thus potentially increasing the ability to discover the distribution of a larger set of molecules that may play an important role in antler growth. We have thus demonstrated that MALDI-IMS is a promising technique for generating molecular maps with high spatial resolution which can aid in evaluating the function of novel molecules during antler growth.