TMT-Based Quantitative Proteomics of Cerebrospinal Fluid Reveals Molecular Signatures of Lymphatic Drainage Impairment in Young Rats

The glymphatic system facilitates the clearance of interstitial solutes and waste products from the brain parenchyma, channeling them into the meningeal lymphatic vessels for subsequent drainage into the deep cervical lymph nodes (dcLNs). However, the extent to which this lymphatic outflow via dcLNs is essential for maintaining cerebral homeostasis remains insufficiently understood. To address this, we performed high-resolution proteomic profiling of cerebrospinal fluid (CSF) from 3-month-old rats using Tandem Mass Tag (TMT)–based quantitative mass spectrometry. This approach enabled the sensitive detection of proteomic perturbations in the CSF resulting from impaired lymphatic drainage. A total of six CSF samples from 3-month-old rats (n=6) were analyzed. TMT 10-plex–based quantitative proteomics revealed 4,089 proteins, among which 357 exhibited statistically significant differential expression (p < 0.05). These proteomic alterations indicated early signs of cellular stress and low-grade inflammation. Pathway enrichment analysis further revealed upregulation of molecular cascades associated with neurodegenerative processes, even in this relatively young cohort, following lymphatic disruption.