Abstract WP130: Fibroblast-derived Laminin Promotes Blood-brain Barrier Repair After Stroke Not Through Integrinβ1 Receptors

Abstract

Blood-brain barrier (BBB) breakdown has been observed in various neurological disorders, including stroke. More importantly, BBB disruption positively correlates with stroke outcome. Laminin, an extracellular matrix protein, plays an important role in the formation and maintenance of blood brain barrier (BBB) integrity under both homeostatic and pathological conditions. Fibroblasts, mainly located in the meninges and around large blood vessels, also secrete laminin. The function of fibroblast-derived laminin in BBB maintenance, however, remains unknown. Here we generated mutant mice with lamininγ1 deficiency in fibroblast-like cells using the Col1a1-Cre line (γ1FKO). These mutants were born at expected Mendelian ratio and failed to show obvious abnormalities, indicating a dispensable role of fibroblast-derived laminin under homeostatic conditions. Next, we examined BBB integrity in these mutants in a collagenase induced hemorrhagic stroke model. Compared to the controls, the mutant mice exhibited aggravated BBB damage and brain injury, including enhanced BBB permeability, elevated inflammatory cell infiltration, reduced tight junction coverage, reduced PDGFRβ coverage, larger hematoma size, worse neurological dysfunction, and increased neuronal cell death. These results suggest that fibroblast-derived laminin exerts a beneficial role in stroke pathogenesis possibly via repairing damaged BBB. Using γ1FKO:Ai14 and Col1α1-Cre+:Ai14 (control) mice, we found significantly reduced fibroblast density in the peri-hematoma region γ1FKO:Ai14 at day7 after injury. Additionally, laminin γ1 knockout in fibroblast cells by CRISPR Cas9 results in reduced fibroblast migration ability. These results suggest that fibroblast-derived laminin play important roles on fibroblast migration. Classical laminin receptors include integrin-α3β1, -α6β1, -α7β1 and -α6β4. To study the receptor regulating laminin- mediated fibroblasts function, we generated integrin β1 mutant mice using Col1a1-Cre line. In these mutants we did not see difference in terms of stoke outcomes as we saw in γ1FKO mice, indicating that integrin β1 is not the receptor regulating laminin- mediated fibroblasts function.