Poly I:C Induced Gene Ablation Negates Potentially Beneficial Effects of A4-Integrin Depletion on Experimental Autoimmune Encephalomyelitis

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2016-04-15

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Abstract

In the experimental autoimmune encephalomyelitis (EAE) murine model of multiple sclerosis, leukocytes transmigrate through the blood brain barrier into the perivascular space and enter the parenchyma of the central nervous system (CNS). This entry is facilitated by the interaction of very late activation antigen-4 (α4β1-integrin) on activated T cells with vascular cell adhesion molecule-1 that is expressed by inflamed endothelium. It has been demonstrated that α4-integrin antagonism in multiple sclerosis patients can be significantly beneficial, but it can also lead to detrimental CNS infections due to impairing of immune surveillance. In this work, we seek to understand cell type specificity of α4-integrin antagonism in order to improve immune competence in the CNS. We generated the conditional knock out mouse strain Mx1.Cre+ α4-integrinfl/fl on the C57BL/6 background that enables α4-integrin gene ablation by the Cre-lox recombination system upon poly I:C treatment. We hypothesize that antagonism of α4-integrin diminishes immunocompetence within the CNS by differentially affecting leukocyte subsets. In order to test this hypothesis, we actively and passively induced EAE on Mx1.Cre+ α4-integrinfl/fl. We observed that disease susceptibility and severity in poly I:C treated Mx1.Cre+ α4-integrinfl/fl mice were similar to that of wild type (WT), but the disease onset was significantly delayed. Additionally, while there was decreased migratory capabilities of CD45+ splenocytes from Mx1.Cre+α4-integrinfl/fl mice, the absolute number and composition of leukocytes in the CNS of EAE mice was similar in WT and poly I:C treated Mx1.Cre+ α4-integrinfl/fl mice. The presence of Evans Blue dye in the parenchyma of poly I:C treated mice suggest a role of poly I:C in this migratory effect. On the other hand, adoptively transferred cells from poly I:C treated Mx1.Cre+ α4-integrinfl/fl mice did not transfer EAE disease into WT. Based on these data, we concluded that Mx1.Cre+ α4-integrinfl/fl strain is limited to study the role of α4-integrin ablation on immunocompetence, however it is successful for the generation of α4-integrin deficient leukocytes. Finally, we determined optimization techniques for CNS leukocyte isolation that can be of benefit for future analyses.

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Subjects

Central Nervous System, Encephalomyelitis, Autoimmune, Experimental, Integrin alpha4, Integrin alpha4beta1, Poly I-C

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