Vasculature-on-a-chip is a powerful tool which may be used to elucidate clinical phenomena in human body including vasculogenesis and pruning of vessel as well as development and treatment of vascular diseases. However, in vitro vascular networks are limited in their uses due to their short availability caused by pruning of vessel and degradation of hydrogel within microfluidic chip. To overcome this limitation, we have added GM6001, broad-spectrum matrix metalloproteinase inhibitor to reduce hydrogel degradation because matrix metallo-proteinase plays a critical role in degrading various components of the extracellular matrix [1, 2]. Microvascular networks in microfluidic chip were created by culturing human umbilical vein endothelial cells (HUVECs) within the fibrin gel and GM6001 was supplemented 5 days after seeding HUVECs. To elucidate effects of matrix metalloproteinase, three different GM6001 concentrations (10μM, 20μM, 40μM) were tested to chips and area of microvascular networks and vessel diameters were quantified to indicate optimal concentration for stable vasculatures. It was confirmed that supplement matrix metalloproteinase helps to establish more stable vascular networks successfully. Our study demonstrates 3D in vitro vascular model could be long-lasting by reducing matrix degradation and promising tool to better investigate long-term clinical phenomena in vessels.