Subcutaneous injection of CTHs induces leukocyte recruitment and local production of CXCL10.

(A) Balb/c mice (CTHs (n = 7) and PBS (n = 7)) were challenged with 100 μl of a 10 μg/ml CTHs solution and the number of leukocytes that infiltrated the air pouch were measured. (B) Identification of cellular subsets recruited into the mice air pouch was done as described in Material and Methods. Monocytes (CD11b+Ly6C+CD64+), neutrophils (CD11b+Ly6G+), eosinophils (CD11b-CD64-Ly6CintSSC-Ahigh) and NK cells (CD11b-Ly6G-NK1.1+) were identified after gating on PI- CD3- B220- events. (C) Mice received an injection with 100 μl of a 10 μg/ml CTH solution and the air pouch supernatants from C57BL/6 wild-type (n = 12), TLR4 KO (n = 7), and MyD88 KO (n = 7) mice were used to measure the local levels of CXCL10. (D) Mice received an injection with 100 μl of a 10 μg/ml CTH solution and the number of leukocytes that infiltrated the air pouch, are depicted for wild-type (n = 7), TLR4 KO (n = 7), and MyD88 KO (n = 7) and mice. (E) Mice received an injection with 100 μl of a 100 μM IRR20 solution and the air pouch supernatants from wild-type (n = 6) and TLR4 knockout (n = 6) mice were used to measure the concentration of CXCL10. (F) Mice received an injection with 100 μl of a 100 μM IRR20 solution and the number of leukocytes that infiltrated the air pouch, was determined in wild-type (n = 6) and TLR4 knockout (n = 6) mice.