Cerebrospinal fluid biomarkers link toxic astrogliosis and microglial activation to multiple sclerosis severity

Abstract

Background: Once multiple sclerosis (MS) reaches the progressive stage, immunomodulatory treatments have limited efficacy. This suggests that processes other than activation of innate immunity may at least partially underlie disability progression during late stages of MS. Pathology identified these alternative processes as aberrant activation of astrocytes and microglia, and subsequent degeneration of oligodendrocytes and neurons. However, we mostly lack biomarkers that could measure central nervous system (CNS) cell-specific intrathecal processes in living subjects. This prevents differentiating pathogenic processes from an epiphenomenon. Therefore, we sought to develop biomarkers of CNS cell-specific processes and link them to disability progression in MS.Methods: In a blinded manner, we measured over 1000 proteins in the cerebrospinal fluid (CSF) of 431 patients with neuroimmunological diseases and healthy volunteers using modified DNAaptamers (SOMAscan®). We defined CNS cell type-enriched clusters using variable cluster analysis, combined with in vitro modeling. Differences between diagnostic categories were identified in the training cohort (n = 217) and their correlation to disability measures were assessed; results were validated in an independent validation cohort (n = 214).Results: Astrocyte cluster 8 (MMP7, SERPINA3, GZMA and CLIC1) and microglial cluster 2 (DSG2 and TNFRSF25) were reproducibly elevated in MS and had a significant and reproducible correlation with MS severity suggesting their pathogenic role. In vitro studies demonstrated that proteins of astrocyte cluster 8 are noticeably released upon stimulation with proinflammatory stimuli and overlap with the phenotype of recently described neuro-toxic (A1) astrocytes.

Microglia in Multiple Sclerosis: Friend or Foe?

“…Proteomics is one approach that may offer more informative biomarkers of microglial activity in body fluids with the added the ability to assess cell specific processes in living patients. New advances in "cell specific" proteomics have been developed and tested in MS that provide markers of the cell of origin, greatly increasing the utility of these measures (68). Using a predetermined multiplex proteomic scan, CSF of MS patients yielded elevated astrocytic and microglial markers which were correlated with disease severity as measured by two clinical metrics (Age Related MS Severity and MS Disease Severity Scale).…”

“…Using a predetermined multiplex proteomic scan, CSF of MS patients yielded elevated astrocytic and microglial markers which were correlated with disease severity as measured by two clinical metrics (Age Related MS Severity and MS Disease Severity Scale). These approaches await validation on a larger scale but offer an attractive option for disease monitoring and discovery science (68).…”

Emerging Role of Extracellular Vesicles in the Pathophysiology of Multiple Sclerosis

“…In another study, Masvekar and colleague did not find a difference between levels of EVs deriving from cell apoptosis (apoptosomes) isolated in CSF of RRMS (active and non-active) patients and HC, suggesting that apoptotic bodies are not appropriate as disease biomarkers [ 65 ], but further validations are necessary.…”

Drug library screen identifies inhibitors of toxic astrogliosis

“…Identification of CNS cell-specific protein clusters within the MS CSF proteome measured by a DNA-aptamer-based platform (i.e., Somascan®, Somalogic Boulder, CO, USA) detected only two clusters of proteins that reproducibly correlated with the rate of accumulation of disability and CNS tissue damage in the independent validation cohort of MS patients (10). One protein cluster was enriched for microglia-secreted proteins while the second cluster constituted proteins secreted mostly by inflammatory stimuli-activated astrocytes.…”

“…PBMCs were isolated using density gradient centrifugation as described (10). PBMC (1x10 6 cells/ml) were cultured in serum-free X-VIVO (Lonza), and either left untreated (unstimulated) or stimulated with lipopolysaccharide (LPS; Sigma-Aldrich, St. Louis, MO; 100 ng/ml) and CD3/CD28 microbeads (Invitrogen, Carlsbad, CA; at 1:1 beads to cells ratio) to activate simultaneously cells of innate and adaptive immunity.…”