Pressure-actuated cementocyte-on-chip: Maturation-dependent secretory phenotype and compression-induced secretory regulation

Root cementocytes are positioned to sense mechanical stimuli associated with orthodontic loading, yet experimentally accessible models that preserve cementocyte like maturation while allowing controlled compression remain limited. Here, we developed a pressure actuated three dimensional cementocyte on chip platform using IDG-CM6 cementocyte like cells to investigate maturation associated secretory development and compression induced cellular responses. The multilayered microfluidic device combined gravity driven perfusion, a deformable polydimethylsiloxane membrane, and a hydrogel based culture chamber, enabling long term perfused culture followed by superimposed compression. Under unloaded conditions, the chip maintained high cell viability and supported progressive three dimensional cytoskeletal organization, DMP1 associated reporter activity, matrix associated development, and mineralization. Compared with conventional two dimensional culture, on chip culture generated a distinct maturation associated secretory profile characterized by higher FGF23, OPG, OCN, and SOST, lower RANKL, and a progressive increase in the OPG/RANKL ratio. Following 14 days of maturation, direct comparison among unloaded, lower compression, and higher compression three dimensional cultures showed that short term compression did not markedly alter cell viability, gross F actin organization, DMP1 associated GFP reporter activity, or mineralization associated staining, but selectively regulated the secretory profile. OCN, SOST, OPG, and RANKL showed compression magnitude dependent changes, whereas FGF23 remained comparatively stable. Lower compression increased the OPG/RANKL ratio, whereas higher compression reduced the ratio relative to the lower compression condition. These findings establish a controllable three dimensional platform for investigating cementocyte mechanobiology and compression magnitude dependent regulation of resorption associated signaling within the root compartment.