Proteomic changes in the extracellular environment of sea bass thymocytes exposed to 17α-ethinylestradiol in vitro

The thymus is an important immune organ providing the necessary microenvironment for the development of a diverse, self-tolerant T cell repertoire, which is selected to allow for the recognition of foreign antigens while avoiding self-reactivity. Thymus function and activity are known to be regulated by sex steroid hormones, such as oestrogen, leading to sexual dimorphisms in immunocompetence between males and females. The oestrogenic modulation of the thymic function provides a potential target for environmental oestrogens, such as 17α-ethynylestradiol (EE2), to interfere with the cross-talk between the endocrine and the immune system. Oestrogen receptors have been identified on thymocytes and the thymic microenvironment, but it is unclear how oestrogens regulate thymic epithelial and T cell communication including paracrine signalling. Much less is known regarding intrathymic signalling in fish. Secretomics allows for the analysis of complex mixtures of immunomodulatory signalling factors secreted by T cells. Thus, in the present study, isolated thymocytes of the European sea bass, Dicentrarchus labrax, were exposed in vitro to 30 nM EE2 for 4 h and the T cell-secretome (i.e., extracellular proteome) was analysed by quantitative label-free mass-spectrometry. Progenesis revealed a total of 111 proteins differentially displayed between EE2-treated and control thymocytes at an α-level of 5% and a 1.3- fold change cut off (n = 5-6). The EE2-treatment significantly decreased the level of 90 proteins. Gene ontology revealed the proteasome to be the most impacted pathway. In contrast, the abundance of 21 proteins was significantly increased, with cathepsins showing the highest level of induction. However, no particular molecular pathway was significantly altered for these upregulated proteins. To the best of our knowledge, this work represents the first study of the secretome of the fish thymus exposed to the environmental oestrogen EE2, highlighting the impact on putative signalling pathways linked to immune surveillance, which may be of crucial importance for fish health and defence against pathogens.