Inhibition of complement C5A rreceptor ameliorates tissue plasminogen activator (T-PA)-mediated increase in blood brain barrier (BBB) permeability.

Background : t- PA can increase BBB permeability and trigger the infiltration of immune cells into the central nervous system. Few plasmin- dependent and - independent pathways participate in this phenomenon, for example, activation of Rho- Kinase 2 and platelet derived growth factor C, respectively. Since plasmin also possesses pro- inflammatory properties, including C3 and C5 convertase activity, we hypothesized that complement activation plays an additional role in this process. Aims : We aimed to evaluate the effect of complement inhibition on t- PA- and plasminogen (plg)- mediated opening of the BBB using an established in vitro model with C5 and C5a- receptor 1 (C5aR1) inhibitors. Methods : An in vitro model of the BBB was assembled on porous membranes of Transwell inserts as a co- culture system of human brain endothelial cells on the luminal surface and human immortalized astrocytes on the abluminal surface. The luminal compartment was stimulated with t- PA and plasminogen in the presence of PMX205 (non- competitive inhibitor of C5aR1), Avacopan (C5aR1 antagonist) or eculizumab (humanised monoclonal inhibitor of human C5). BBB permeability was assessed at both 4hr and 24hr post- stimulation by evaluation of fluorescent albumin passage from the luminal to the abluminal chambers over 1hr. Permeability changes were calculated relative to non- treated controls. Results : PMX205 and Avacopan decreased t- PA mediated increase in BBB permeability by almost 50%, at both the 4 and 24hr time points. Interestingly, our preliminary results indicate that eculizumab was without any significant effect. Conclusions : t- PA- and plasmin- mediated increase in BBB permeability is partly driven by C5a receptor activation. Inhibition of C5 or C5aR1 may have therapeutic implications in traumatic brain injury and stroke thrombolysis, where either endogenous or administered t- PA can compromise the BBB, leading to secondary insults.