Ro 31-9790 arrests lymphocytes in the endothelial lining and not in the basement membrane or the lumen of HEVs.

(A) Laminin in the HEV basement membrane was stained and the number of lymphocytes adjacent to or within the basement membrane counted and expressed per unit length of laminin. Although the number of CFSE-labeled cells inside HEVs was visibly increased in Ro 31-9790–treated mice, lymphocyte numbers in the basement membrane were not significantly different (ns). Bar indicates 20 μm. (B) Ultrastructural analysis of HEVs showed the majority of lymphocytes inside HEVs were within the endothelial lining (position 2) and were not attached to the luminal surface of HEVs (position 1). Other types of leukocytes were not seen binding or transmigrating HEVs. Results are percent of lymphocytes in positions 1 and 2 in transmission electron micrographs. A total of 85 to 115 lymphocytes were counted. LNs from perfused mice were fixed and cut and sections stained for transmission electron microscopy using standard techniques. Bar indicates 5 μm.