SELECTIVE BINDING OF PERIPHERAL-BLOOD LYMPHOCYTES TO THE WALLS OF CEREBRAL VESSELS IN FROZEN-SECTIONS OF HUMAN BRAIN

In order to identify the factors that control the binding of blood leu cocytes to cerebral blood vessels we have modified and applied the fro zen section assay of Stamper and Woodruff to the study of human brain. Cryostat sections of brain tissue obtained at post mortem were overla id with blood lymphocytes and experimental conditions were defined whi ch permitted optimum binding of the cells to transected blood vessel w alls. The maximal binding of lymphocytes to cerebral vessels occurred when 6 x 10(6) lymphocytes were overlaid onto brain sections for 30 mi n at 7 degrees C with gentle agitation. Only a small proportion (0.01% ) of the added lymphocytes bound to exposed cerebral vessels. However, lymphocytes were far more adherent than monocytes and polymorphonucle ar cells (7-fold and 11-fold respectively: p < 0.001) and activation o f lymphocytes with IL-2 enhanced their binding to blood vessel walls ( mean 130% increase; p < 0.03). Further analysis revealed that CD4-posi tive T lymphocytes were the predominant cell population binding to the blood vessels. Antibody blocking studies showed that lymphocyte bindi ng to cerebral blood vessels was inhibited by pretreating the lymphocy tes with anti-CD11a, anti-CD18 or anti-CD49d (p less than or equal to 0.02) and immunohistochemical studies revealed the presence of the cou nter-receptors ICAM-1 (CD54) and VCAM-1 (CD106) for these adhesion mol ecules in addition to the presence of E-selectin (CD62E) and P-selecti n (CD62P) on the cerebral blood vessels. The establishment of a techni que in situ which measures selective binding of CD4-positive periphera l lymphocytes to sections of cerebral blood vessels will assist in the molecular characterization of factors that control the interaction of leucocytes with the blood-brain barrier in health and disease.