Fig 2: CCN3 immunostaining in normal and MS brain tissue. CCN3 is present in blood vessels, choroid epithelia and neuron-, and macroglia-like cells of the CNS (black arrows, a–d). Co-labelled sections of MS grey matter with antibodies to the mature neuronal protein HuC/D (green) and anti-CCN3 (red) demonstrates dual-labelling of CCN3 and HuC/D+ neurons (e). Detection of CCN3 transcripts by in situ hybridisation in MS grey matter with a distinct neuronal morphology (f) and respective negative control (f, inset). Area of CCN3 immunoreactivity in MS brain tissue (n = 17 cases) (g). GM, normal-appearing grey matter; GM Lesion, demyelinated GM, GM remy, remyelinated GM; WM, normal-appearing white matter; WM Lesion, demyelinated WM; WM remy, remyelinated WM

Fig 3: Obesity-induced changes in plasma CCN3 levels are evident in MS. Plasma CCN3 levels in lean vs. obese cohorts (a), CCN3 levels (b) and leptin levels (c) in plasma from obese patients before and after 12 weeks of GLP-1 agonist therapy. Corresponding plasma CCN3 levels and BMI in healthy controls and MS (d) and stratified by disease phase and treatment (e). Abbreviations: HC, healthy control; MS, multiple sclerosis; RRMS, relapsing-remitting MS; TN, treatment-naïve; Nat, natalizumab treatment; IFN, interferon-ß treatment; MS-Pr, progressive MS

Fig 4: Neurofilament levels and CCN3 levels correlate in CSF of IIH patients but not MS. Neurofilament light (NFL) levels in CSF of matched IIH and MS donors (a). Relationship between CCN3 levels and NFL levels in CSF of IIH patients (b) and MS patients (c). Abbreviations: CSF, cerebrospinal fluid; IIH, idiopathic intracranial hypertension; MS, multiple sclerosis; NFL, neurofilament light

Fig 5: Plasma CCN3 levels are altered by MS disease phase and treatment. Plasma CCN3 levels are compared in healthy controls and MS (a), controls and MS disease phase (b), RRMS treatment groups (c) and matched samples from RRMS patients during relapse or remittance (d). Detection of plasma-derived CCN3 by western blot (e). Abbreviations: HC, healthy control; MS, multiple sclerosis; RRMS, relapsing-remitting MS; Prog MS, progressive MS; TN, treatment-naïve; Nat, natalizumab treatment; IFN, interferon-ß treatment; Ctl, h295-R-conditioned medium as positive control