Madeleine Handsley, Alba Warn

Angiogenesis is the mechanism of neovascularisation by which new blood vessels are formed from pre-existing vasculature. It is an essential feature of the female reproductive cycle, embryonic development and wound repair. Angiogenesis has also been identified as a causal or contributing factor in several pathologies including cancer, where it is a rate-limiting step during tumour progression. The membrane-type matrix metalloproteinases (MT-MMPs) are the membrane-anchored subdivision of the MMPs, a family of proteolytic enzymes whose collective substrate repertoire includes all known protein components of the ECM. MMPs, in particular the gelatinases and MT1-MMP, have been linked to angiogenesis. The vascular phenotype of the MT1-MMP knockout mouse suggests that this protease is a key factor during neovascularisation. Furthermore, observations that in vitro EC-tubulogenesis is inhibited by TIMP-2, -3 and -4 but not TIMP-1 support the involvement of TIMP-1 insensitive MMPs, such as MT-MMP-1, -2, -3, and –5, in this process. Work done in our group has shown that micro- and macro-vascular ECs had a similar tubulogenic response to angiogenic stimulation and TIMP inhibition. In both EC-types TIMP-2 and -4, but not TIMP-1, inhibited tubulogenesis. Further support for the involvement of the MT-MMPs during EC-tubulogenesis was provided by expression profiling data, which revealed that expression of the transmembrane MT-MMPs was specifically increased during EC-tubulogenesis within 3-D extracellular matrices. Angiogenesis Primary endothelial cells embedded in a 3D fibrin matrix forming tubules They are stained with an antibody that recognizes the specific molecular marker CD31, seen in red and counterstained with the nuclear stain DAPI seen in blue. 3D reconstruction of the optical confocal sections, using the Zeiss LMS 510 META confocal microscope, was performed with Volocity software.