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We evaluate a set of protocols for preparation of the secretome from murine Islets of Langerhans for bottom-up proteomic analysis. Of the protocols evaluated, a filter-aided sample preparation based approach using sodium dodecyl sulfate as a detergent to solubilize proteins generated the most protein identifications. A total of 362 protein groups (average of 3.7 peptides/protein) were identified from the secretome using the SDS-FASP protocol; a combination of data from three protocols generated 413 protein group identifications. As expected, the identified proteins included insulin 1 and 2, somatostatin, and glucagon, the four main secreted components from islets. STRING network analysis classified the other proteins as being associated with extracellular exosomes, membrane-bounded vesicles, vesicles, and the extracellular region.

Glucotoxicity is a causative agent of type-2 diabetes, where high glucose levels damage the islets of Langerhans resulting in oxidative damage and endoplasmic reticulum stress. We evaluated the secretomes of healthy CD-1 murine islets. Three experimental conditions were investigated in biological triplicate: a control incubated with 11 mM glucose, 1-day incubation with 25 mM glucose, and 2-day incubation with 25 mM glucose. An SDS-based, filter-aided sample preparation protocol was used to prepare secretomes for analysis. A total of 428 protein groups were identified across the nine samples. Each condition generated between 328-349 protein IDs and intracondition protein overlap was between 66-90% for the biological triplicates. 232 protein groups were identified in all three conditions with 184 quantified at least once in each condition. Significant expression changes were observed for proteins associated with the unfolded protein response, such as proteases, chaperones, and elongation factors, as well as proteins associated with peptide hormone processing and small molecule metabolism.