Sexual Dimorphism in Extracellular Matrix Composition and Viscoelasticity of the Healthy and Inflamed Mouse Brain

SIMPLE SUMMARY: In multiple sclerosis (MS), an autoimmune disease of the central nervous system that primarily affects women, gender differences in disease course and in brain softening have been reported. It has been shown that the molecular network found between the cells of the tissue, the extracellular matrix (ECM), influences tissue stiffness. However, it is still unclear if sex influences ECM composition. Therefore, here we investigated how brain ECM and stiffness differ between sexes in the healthy mouse, and in an MS mouse model. We applied multifrequency magnetic resonance elastography and gene expression analysis for associating in vivo brain stiffness with ECM protein content in the brain, such as collagen and laminin. We found that the cortex was softer in males than in females in both healthy and sick mice. Softening was associated with sex differences in expression levels of collagen and laminin. Our findings underscore the importance of considering sex when studying the constitution of brain tissue in health and disease, particularly when investigating the processes underlying gender differences in MS. ABSTRACT: Magnetic resonance elastography (MRE) has revealed sexual dimorphism in brain stiffness in healthy individuals and multiple sclerosis (MS) patients. In an animal model of MS, named experimental autoimmune encephalomyelitis (EAE), we have previously shown that inflammation-induced brain softening was associated with alterations of the extracellular matrix (ECM). However, it remained unclear whether the brain ECM presents sex-specific properties that can be visualized by MRE. Therefore, here we aimed at quantifying sexual dimorphism in brain viscoelasticity in association with ECM changes in healthy and inflamed brains. Multifrequency MRE was applied to the midbrain of healthy and EAE mice of both sexes to quantitatively map regional stiffness. To define differences in brain ECM composition, the gene expression of the key basement membrane components laminin (Lama4, Lama5), collagen (Col4a1, Col1a1), and fibronectin (Fn1) were investigated by RT-qPCR. We showed that the healthy male cortex expressed less Lama4, Lama5, and Col4a1, but more Fn1 (all p < 0.05) than the healthy female cortex, which was associated with 9% softer properties (p = 0.044) in that region. At peak EAE cortical softening was similar in both sexes compared to healthy tissue, with an 8% difference remaining between males and females (p = 0.006). Cortical Lama4, Lama5 and Col4a1 expression increased 2 to 3-fold in EAE in both sexes while Fn1 decreased only in males (all p < 0.05). No significant sex differences in stiffness were detected in other brain regions. In conclusion, sexual dimorphism in the ECM composition of cortical tissue in the mouse brain is reflected by in vivo stiffness measured with MRE and should be considered in future studies by sex-specific reference values.