Diet can have a significant impact on normal physiology, with high calorific foods and sedentary lifestyles contributing to the development of obesity. Globally, there is an obesity epidemic with 1 in 3 people overweight (WHO, 2015) and obesity occurring at a younger age. It is also increasingly recognised that being overweight or obese may have a deleterious effects on fertility. The association between obesity and impaired male reproductive function is multifactorial, involving alterations at the level of the hypothalamic-pituitary-gonadal (HPG) axis, as well as direct effects on the testis, including spermatogenesis and crucially somatic cell function (Sermondade et al., 2013). Here, we used a large scale discovery approach to study the effects of diet-induced obesity on the testis, generating a valuable proteomic dataset of potential ‘hits’ which provided insight into the biological pathways and protein networks associated with high fat (HF) diet.
Tissue samples originating from mice fed a HF or chow diet were homogenized and digested with trypsin overnight. Peptides were separated over a 90-minute gradient of 3-40% acetonitrile/0.1% formic acid. MS data were acquired using a Synapt G2-Si operating in IMS-DIA (HDMSE). Samples were acquired in a random fashion and as technical triplicates. The data were processed using Progenesis QI for Proteomics and searched against a Uniprot Mus musculus database, with 1% FDR. Curated results were interrogated using Ingenuity to derive pathways of biological significance.
A total of 4960 proteins were identified across the whole study with 920 those being unique to HF. A number of statistically significant proteins with differential regulation were exhibited and found to be located within potential pathways which are identified in mediating the effects of HF diet on the testis. For example, pathways related to the blood testis barrier were readily identified as statistically significant and disrupted in animals exposed to HF diet.