Exosomal proteome analysis of cerebrospinal fluid detects biosignatures of neuromyelitis optica and multiple sclerosis

Quantitative proteomic analysis of exosomes isolated from cerebrospinal fluid (CSF) of neuromyelitis optica (NMO) patients detected signature proteins differentiating NMO from multiple sclerosis (MS) and idiopathic longitudinally extensive transverse myelitis. Exosomes with good yields were obtained using ultracentrifugation from pooled CSF assisted by chemokine-based clustering strategy, which improved target molecule identification by providing amplified fold change values. 442 significant proteins generated a list of signature molecules of diseases validated primarily by the identification of known markers such as glial fibrillary acidic protein (GFAP) and fibronectin specific to NMO and MS respectively. MetaCore pathway analysis of significant proteins supported the involvement of these proteins in disease progression via neurological pathway. Expression levels of target molecules from orthogonal label-free quantification employing quadrupole-Orbitrap hybrid mass spectrometry were in good agreement with those from Western blotting. Additional investigation of GFAP and fibronectin as representative disease molecules revealed their presence in intact exosomes as detected by flow cytometry. This comprehensive study suggests that the exosomal proteomic analysis of CSF can be applied to the identification and characterization of inflammatory disorders of the central nervous system.