Study of synergistic effects of biochemical and mechanophysical factors on the glucose-mediated inflammatory response

Abstract

ECM proteins exposed to glucose over long periods of time became glycated, and accumulated advanced glycation end-products (AGEs). This glucose-mediated matrix remodeling may change the structure and function of the protein. Previous studies on the mechanism of cellular damage caused by the accumulation of AGES in ECM is proceeding in two directions. First, the activation of the receptor of AGEs (RAGE) signaling pathway by AGEs molecules. Next, the activation of the mechanotransduction signaling pathway via biomechanical factor including increasing stiffness. Previous studies have only studied about the independent effects of RAGE and mechanotransduction pathway. However, in the body, these two signaling pathway was activated simultaneously, and had synergistically affect to induce an inflammatory response. Therefore, we designed an experiment to implement these two mechanisms simultaneously to investigate the mechanism by which ECM induces cellular inflammatory response by simulating a more similar environment in the body. Our hypothesis is that the AGEs accumulated on the ECM memorized glucose-induced stress, so it induces a chronic inflammatory response to cells even in normal blood glucose. To demonstrate this hypothesis, we cultured fibroblasts with collagen and glycated-collagen matrix, and regulated media glucose concentration differently. There was no significant difference in cellular viability between each group, so later results were non-influenced by cell death. In COL, the activation of RAGE and mechanotransduction was increased according to glucose concentration, and the inflammatory response was also induced. On the other hand, in GLY, activation of RAGE was always higher regardless of glucose concentration, whereas mechanotransduction increases according to glucose concentration. As a result, effector molecules regulated through these two signaling pathways will always express high levels in GLY regardless of sugar concentration. So, it was confirmed that in glycated-collagen, the increase in ROS production and the NF-$\kappaB$ activation through nuclear translocation are always highly regulated regardless of glucose concentration. Through this, it was confirmed that, glycated collagen induced chronic inflammation via synergistic effects of the RAGE and mechanotransduction signaling pathway. And it shows that this platform better mimetic the physiologically relevant diabetic tissue microenvironment. In the future this platform may be used for substitute in vitro inflammatory model, and used for test ECM-targeted therapy for chronic disease.