MK2 Signaling Differentially Regulates Chemokines During Aggregatibacter Actinomycetemcomitans Challenge

Objectives: Aggregatibacter actinomycetemcomitans is a bacteria often isolated from patients diagnosed with aggressive periodontal disease. During periodontal disease pathogenesis, chemokine and chemokine receptor signaling regulates inflammatory cell recruitment. We previously showed that A. actinomycetemcomitans activates intracellular MK2 signaling in macrophages, which are derived from the hematopoietic cells. Non-hematopoietic cells such as fibroblasts also secrete chemokines. Thus, the goal of this study was to determine the role of MK2 in chemokine and chemokine receptor signaling in hematopoietic and non-hematopoietic cells during A. actinomycetemcomitans pathogenesis.
Methods: We utilized a murine calvarial model and bone marrow transplantation combined with an air pouch in Mk2^+/+ and Mk2^-/- mice. Mice were challenged with A. actinomycetemcomitans or PBS vehicle by subcutaneous injection. Chemokines and chemokine receptors were assessed by analysis of RNA, protein, and flow cytometry.
Results: In calvarial tissue, RT-qPCR and multiplex analysis revealed that chemokines, Ccl3 and Ccl4 RNA and CCL3 protein, were significantly upregulated in Mk2^+/+ mice compared to Mk2^-/- mice treated with A. actinomycetemcomitans (P≤0.05). Nanostring RNA analysis showed that Ccr5, which encodes a receptor that binds CCL3 and CCL4, was also significantly upregulated in Mk2^+/+ mice compared to Mk2^-/- mice challenged with A. actinomycetemcomitans (P≤0.05). MK2 did not regulate circulating CD11b⁺CCR5⁺ and CD11b⁺CCR1⁺ monocytes during bacterial challenge. ELISA detected that MK2 induced CCL4 in the non-hematopoietic cells, but not CCL3 in the A. actinomycetemcomitans air pouch challenge. Lastly, MK2 deficiency in the hematopoietic compartment significantly enhanced Ly6C⁺CCR1⁺ monocytes compared to the control group (P≤0.05).
Conclusions: MK2 deficiency in the hematopoietic compartment increased Ly6C⁺CCR1⁺ despite decreases in CCL3 and CCL4, which may be a result of a compensatory mechanism. These novel findings suggest that MK2 differentially regulates CCL3 and CCL4 RNA and protein in the hematopoietic and non-hematopoietic compartments during A. actinomycetemcomitans pathogenesis.