Wide-Ranging Effects on the Brain Proteome in a Transgenic Mouse Model of Alzheimer’s Disease Following Treatment with a Brain-Targeting Somatostatin Peptide

Abstract

Alzheimer’s disease is the most common neurodegenerative disorder characterized by the pathological aggregation of amyloid-β (Aβ) peptide. A potential therapeutic intervention in Alzheimer’s disease is to enhance Aβ degradation by increasing the activity of Aβ-degrading enzymes, including neprilysin. The somatostatin (SST) peptide has been identified as an activator of neprilysin. Recently, we demonstrated the ability of a brain-penetrating SST peptide (SST-scFv8D3) to increase neprilysin activity and membrane-bound Aβ42 degradation in the hippocampus of mice overexpressing the Aβ-precursor protein with the Swedish mutation (APPswe). Using LC–MS, we further evaluated the anti-Alzheimer’s disease effects of SST-scFv8D3. Following a triple intravenous injection of SST-scFv8D3, the LC–MS analysis of the brain proteome revealed that the majority of downregulated proteins consisted of mitochondrial proteins regulating fatty acid oxidation, which are otherwise upregulated in APPswe mice compared to wild-type mice. Moreover, treatment with SST-scFv8D3 significantly increased hippocampal levels of synaptic proteins regulating cell membrane trafficking and neuronal development. Finally, hippocampal concentrations of growth-regulated α (KC/GRO) chemokine and degradation of neuropeptide-Y were elevated after SST-scFv8D3 treatment. In summary, our results demonstrate a multifaceted effect profile in regulating mitochondrial function and neurogenesis following treatment with SST-scFv8D3, further suggesting the development of Alzheimer’s disease therapies based on SST peptides.

Well-Plate μFASP for Proteomic Analysis of Single Pancreatic Islets

“…The protocol for the well-plate μFASP is adapted from the standard FASP protocol that was previously used to study the effect of a brain-targeting somatostatin peptide on different brain regions. 19 Based on previous findings by Distler et al, reduction and alkylation in the protocol are performed directly on the filter. 3 Further, a second incubation with DTT ensures the removal of IAA.…”

“…For protein purification and tryptic digestion, different amounts of total protein (20, 10, 5, 1, and 0.5 μg) were transferred onto centrifugal filter units (Microcon-30 kDa; Merck, Darmstadt, Germany) and processed as reported previously. 19 Briefly, washing with a buffer containing 8 M urea and 100 mM Tris (pH 8.5) was followed by the reduction with 8 mM DTT (incubation at 56 °C for 15 min). For alkylation, an incubation with 50 mM IAA at room temperature was performed for 20 min.…”

“…The peptides were separated at 40 °C with a gradient run from 3 to 40% (v/v) mobile phase B at a flow rate of 0.3 μL/min over 120 min. Via the reference channel, a lock mass solution composed of [Glu1]-fibrinopeptide B (0.1 μM) and leu-enkephalin (1 μM) was introduced every 60 s. Peptide analysis was performed in positive ionization mode using the ultra-definition MSE (UDMSE) approach. , The reproducibility and stability of the method were controlled with a commercially available HeLa digest (Thermo Scientific, Waltham, MA).…”

“…21 Identified proteins inferred from bottom-up proteomic analysis using PLGS were quantitated with the TOP3 method obtained directly from ISOQuant 1.83 as described elsewhere. 19,20 ISOQuant settings are provided in Supporting Information Table S1.…”

Recent advances in neuropeptide-related omics and gene editing: Spotlight on NPY and somatostatin and their roles in growth and food intake of fish

“…SST has been reported to be associated with Alzheimer’s disease, and identified by LC-MS/MS analysis, somatostatin-related amyloidosis is a novel restricted human amyloid type associated with somatostatin -producing enteropancreatic neuroendocrine tumors ( 36 ). LC-MS analysis using SST peptide (SST-scFv8D3) injected into the hippocampus of mice revealed the elevated degradation of NPY ( 35 ). Using proteomic analysis in renal failure studies, NPY-NPY2R system was found to predict nephrotoxicity and pathogenic effects in the glomerulus ( 91 ).…”