Abstract: Advanced Glycation End Products (AGEs) in bone matrix are the products of non-enzymatic glycation of glucose and collagen, which are closely related to the mechanism of weightless bone loss. However, how AGEs accumulate in bone matrix remains unclear. The type L microvessels with slower blood flow velocity were increased under microgravity, which may be related to the accumulation of AGEs in bone. To study the effects of flow velocity on the transport of intravascular glucose molecules into bone matrix and the formation of AGEs in bone matrix, a bilayer channel microfluidic chip experimental system was constructed to simulate the blood vessel-bone matrix interface in vitro. A self-developed double-injection-pump continuous directional liquid supply system was applied to the chip, and the biocompatibility, stability and interorganizational interactivity of the system were verified by experiments. The results show that the fluid stress distribution in the main region of the channel in fabricated chip is uniform, and the flow is laminar. The endothelial cells in microfluidic chips could grow normally after loading high sugar medium of 2.88 mL·min^–1 and 0.38 mL·min^–1 for 72 h, respectively. The diffusion rate of glucose molecules from to the lower collagen channel through the microporous membrane was higher under low flow rate loading than under high flow rate loading, and more AGEs generated in collagen. The experimental system constructed in this paper has excellent biocompatibility, long-term operational stability and interorganizational interactivity, which lays a technical foundation for further in-depth research on biophysical mechanisms related to AGEs accumulation in bone matrix, and has the potential to be applied to space life science research.