Objective To study inhibition of capillary-like formation by Cathepsin B-RNAi-Lentivirus (CTSB-RNAi-LV) EM>in vitro/EM>. Methods The control group and RNAi lentiviral group, and negative control group were tested in this study. Immunohistochemistry method and building capillary-like formation model in vitro were used to study the CTSB-RNAi-LV mouse brain microvascular endothelial cells (bEND.3), and explore affection of Cathepsin B and capillary-like formation. Results BR>Cell immunohistochemistry showed that Cathepsin B positive profiles appeared brow and were mainly located around the nucleus of bEND.3 cells. Of three groups, expression of Cathepsin B was the least in RNAi lentiviral group. Evaluation of bEND.3 the total protein and capillary-like formation showed that RNAi lentiviral group had less total protein content and formation of capillary-like zone than other groups and there was statistically significant (EM>P /EM><0.05).Conclusion CTSB –RNAi-LV can inhibit expression of Cathepsin B in the bEND.3 cell and formation of capillary-like zone in vitro. CTSB-RNAi-LV can inhibited targeted capillary-like formation EM>