Proteomic analysis of cerebrospinal fluid exosomes derived from cerebral palsy children
Zhang Houjun, Deng Bowen, Jiang Shengyuan, Zhao Yi, Ren Jingpei, Xu Lin, Mu Xiaohong
Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing 100105, China
Abstract: BACKGROUND: Cerebral palsy is a wide range of nerve injury diseases. Its etiology is still unclear to a great extent, and the molecular indicators used for classification or monitoring of the disease have not been reported.
OBJECTIVE: To isolate and culture cerebrospinal fluid derived exosomes from children with cerebral palsy, and analyze the proteomic expression profile characteristics of exosomes by label-free quantitative mass spectrometry proteomics, so as to provide molecular biological basis for typing, diagnosis and prognosis evaluation of children with cerebral palsy.
METHODS: Among cerebral palsy patients undergoing selective posterior rhizotomy at Dongzhimen Hospital of Beijing University of Chinese Medicine, there were four patients with spastic cerebral palsy and three patients with mixed cerebral palsy. 3 mL cerebrospinal fluid was extracted from each patient. The exosomes in the cerebrospinal fluid of children with cerebral palsy were separated by ultracentrifugation, and characterized by nanoparticle tracking analysis, transmission electron microscope and protein mass spectrometry.
RESULTS AND CONCLUSION: (1) Transmission electron microscopy showed that there were secretory vesicles in cerebrospinal fluid of children with cerebral palsy. Western blot assay detected the expression of secrete marker proteins Syntenin-1 and Flotillin-1. (2) Nanoparticle tracking analysis showed that the size and concentration of vesicles were consistent with the heterogeneity of exosomes. The cerebrospinal fluid exosomes of spastic cerebral palsy and mixed cerebral palsy were analyzed by protein mass spectrometry. A total of 551 proteins were identified. (3) Bioinformatics analysis showed that the identified cerebrospinal fluid exosome proteins were related to the molecular signal mechanism of exosome function and neural development. These proteins were mainly concentrated in brain regions such as frontal cortex and motor cortex, which were closely related to the pathology of cerebral palsy. It is concluded that there are exosomes in cerebrospinal fluid of children with cerebral palsy, and exosome proteins may be related to the molecular pathological mechanism of cerebral palsy.
Key words: cerebral palsy, cerebrospinal fluid, exosome, proteomic analysis