Introduction: The repercussions of long-term exposure to low level mixtures of endocrine disrupting compounds (EDCs) which are present, ubiquitously, in the environment are not well understood. Heat-treated sewage sludge, an EDC-rich product used as fertiliser, has been shown to compromise testicular and ovarian development. However, to date, there has been limited investigation in exposed animals of the adrenal glands, the steroid hormone products of which play key roles in fetal development, parturition and adult metabolic homeostasis.
Methods: An ovine model of EDC exposure was established, comprising pregnant Textel ewes maintained on pastures treated, both before and after conception, with either sewage sludge (T) or inorganic fertiliser (C). Morphological and immunohistochemical analyses were performed on 110 day fetal adrenal glands, while 2-DE coupled with LC-MS/MS were used respectively to analyse and identify proteins. Gene expression was assayed by qPCR using primers predicted from deduced amino acid sequences due to current incomplete sequencing of the ovine genome. Statistical analyses performed using a two-tailed t-test assumed equal variance (CI 95%).
Results: Subcapsular vascularisation was reduced (P<0.05) and separation of cortex and medulla delayed (P<0.01) in T adrenals. 21 protein spots were up-regulated and 50 down-regulated in T versus C groups. 9 of 10 protein spots showing the greatest magnitude of change were down-regulated and corresponding mRNA levels were reduced for all 7 adrenal proteins examined, with 5 of these achieving statistical significance (P<0.05). Identified proteins are involved principally in detoxification and energy metabolism within the adrenal cortex (AKR7A2, AKR7A3, FRIL, IDH1), or adrenal medulla catecholamine metabolism (ANXA7, CMGA, PNMT).
Conclusion: Maternal exposure to EDCs delays adrenal gland development, adversely affecting subcapsular vascularisation, cortical-medulla resolution and transcriptional regulation of adrenal proteins. Further experiments are in progress to evaluate the relative contribution of maternal pre-conception exposure versus gestational exposure to EDCs to fetal adrenal disruption.
Declaration of interest: There is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
Funding: Declaration of Funding: Wellcome Trust (grant number 080388).