Targeted Blood Brain Barrier Opening With Focused Ultrasound Induces Focal Macrophage/Microglial Activation in Experimental Autoimmune Encephalomyelitis

Abstract

Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis (MS). EAE reflects important histopathological hallmarks, dissemination, and diversity of the disease, but has only moderate reproducibility of clinical and histopathological features. Focal lesions are less frequently observed in EAE than in MS, and can neither be constrained to specific locations nor timed to occur at a pre-specified moment. This renders difficult any experimental assessment of the pathogenesis of lesion evolution, including its inflammatory, degenerative (demyelination and axonal degeneration), and reparatory (remyelination, axonal sprouting, gliosis) component processes. We sought to develop a controlled model of inflammatory, focal brain lesions in EAE using focused ultrasound (FUS). We hypothesized that FUS induced focal blood brain barrier disruption (BBBD) will increase the likelihood of transmigration of effector cells and subsequent lesion occurrence at the sonicated location. Lesion development was monitored with conventional magnetic resonance imaging (MRI) as well as with magnetic resonance elastography (MRE) and further analyzed by histopathological means. EAE was induced in 12 6–8 weeks old female C57BL/6 mice using myelin oligodendrocyte glycoprotein (MOG) peptide. FUS-induced BBBD was performed 6, 7, and 9 days after immunization in subgroups of four animals and in an additional control group. MRI and MRE were performed on a 7T horizontal bore small animal MRI scanner. Imaging was conducted longitudinally 2 and 3 weeks after disease induction and 1 week after sonication in control animals, respectively. The scan protocol comprised contrast-enhanced T1-weighted and T2-weighted sequences as well as MRE with a vibration frequency of 1 kHz. Animals were sacrificed for histopathology after the last imaging time point. The overall clinical course of EAE was mild. A total of seven EAE animals presented with focal T2w hyperintense signal alterations in the sonicated hemisphere. These were most frequent in the group of animals sonicated 9 days after immunization. Histopathology revealed foci of activated microglia/macrophages in the sonicated right hemisphere of seven EAE animals. Larger cellular infiltrates or apparent demyelination were not seen. Control animals showed no abnormalities on MRI and did not have clusters of activated microglia/macrophages at the sites targeted with FUS. None of the animals had hemorrhages or gross tissue damage as potential side effects of FUS. EAE-animals tended to have lower values of viscoelasticity and elasticity in the sonicated compared to the contralateral parenchyma. This trend was significant when comparing the right sonicated to the left normal hemisphere and specifically the right sonicated compared to the left normal cortex in animals that underwent FUS-BBBD 9 days after immunization (right vs. left hemisphere: mean viscoelasticity 6.1 vs. 7.2 kPa; p = 0.003 and mean elasticity 4.9 vs. 5.7 kPa, p = 0.024; right vs. left cortex: mean viscoelasticity 5.8 vs. 7.5 kPa; p = 0.004 and mean elasticity 5 vs. 6.5 kPa; p = 0.008). A direct comparison of the biomechanical properties of focal T2w hyperintensities with normal appearing brain tissue did not yield significant results. Control animals showed no differences in viscoelasticity between sonicated and contralateral brain parenchyma. We here provide first evidence for a controlled lesion induction model in EAE using FUS-induced BBBD. The observed lesions in EAE are consistent with foci of activated microglia that may be interpreted as targeted initial inflammatory activity and which have been described as pre-active lesions in MS. Such foci can be identified and monitored with MRI. Moreover, the increased inflammatory activity in the sonicated brain parenchyma seems to have an effect on overall tissue matrix structure as reflected by changes of biomechanical parameters.

Acute Effects of Focused Ultrasound-Induced Blood-Brain Barrier Opening on Anti-Pyroglu3 Abeta Antibody Delivery and Immune Responses

“…In line with the previous literature showing an acute increase in proinflammatory cytokines and activated glia following sonication [45,46], we probed for the general microglia/macrophage marker Iba1 and the phagocytic marker CD68 in the brain and observed an overall increase in their immunoreactivity from 4 to 72 h in the mice that received combination treatment (mAb + bilateral FUS) compared to the mice that received 07/2a antibody alone. Such a slight increase in glial or phagocytic activity in this cohort could be probably explained by a low degree of BBB leakiness (cannot confirm) after sonication as estimated with lower levels of anti-IgG2a immunostaining seen in these sections compared to the IgG2a immunoreactivity seen in the unilateral sonicated cohort.…”

Pulsed Low-Intensity Focused Ultrasound (LIFU) Activation of Ovarian Follicles

“…2a-2b). These effects have been observed to have the capacity to stimulate an immunological response, specifically the activation of macrophages [21], [22], [23]. Consequently, alterations in the macrophage population inside ovarian tissue were identified after LIFU stimulation, and we observed increased numbers of macrophages (Fig.…”

Novel Developments to Enable Treatment of CNS Diseases with Targeted Drug Delivery

“…These patterns are also found in traumatic brain injury. Recently Schregel et al [82] delivered an Optison ® microbubble dose in combination with the same peak negative pressure used in the experiment by Kovacs et al [83] to intentionally induce focal lesions resulting in experimental autoimmune encephalomyelitis (EAE) as a disease model of multiple sclerosis in mice.…”