Nitric Oxide Production and Smooth Muscle Cell Phenotype of Endothelial Cell-Smooth Muscle Cell Cocultured Model

Abstract

This study investigated nitric oxide (NO) production and phenotype changes of smooth muscle cells (SMC) in a cocultured model (CM) exposed to fluid shear stress. The CM was composed of human umbilical endothelial cells (EC) and SMC, a collagen layer, and a porous membrane. After exposing the CM to shear stress of 1.5 Pa for 24 hours, α-smooth muscle actin (α-SMA) expression of SMC and NO production in culture media were examined. Under static conditions, α-SMA expression in the CM was significantly lower than that of a SMC monocultured model (SMC model). After exposure to shear stress, NO production in the CM increased compared to that in the static CM, and there was no significant difference in α-SMA expression between the CM and SMC model. These results suggest that EC may regulate phenotype changes of cocultured SMC, and NO may be one of the factors which induce dedifferentiation of SMC.

Influence of shear stress on phenotype and MMP production of smooth muscle cells in a co-culture model

“…3). This phenotypic change of SMCs was the same as that observed in previous studies [8,9]. The maximum expression of α-SMA and calponin in SMCs occurred on QM6, as shown in Fig.…”

“…The maximum expression of α-SMA and calponin in SMCs occurred on QM6, as shown in Fig. 2, which is the same result as that obtained with two-dimensionally-cultured SMCs [9]. In this study, the expression of another contractile protein, MHC, was also examined, and the results showed that the maximum expression of MHC in SMCs occurred on QM12, which was later than that of α-SMA and calponin (Fig.…”

“…Therefore, in the present study, we constructed an EC-SMC co-culture model, in which phenotype-controlled SMCs were cultured three-dimensionally in a collagen gel, and ECs were cultured above the SMC layer. Earlier research has mainly focused on the SMC response to an SS of about 1.5-2 Pa, which is similar to the SS in a normal artery [8,9]. In the present study, we explored how high SS affects SMC, and found that high SS could lead to a phenotypic change to a synthetic state and higher MMP-2 and -9 production rates, which could induce blood vessel remodeling.…”

“…Earlier, Sakamoto et al [8,9] reported the construction of a co-culture model with contractile phenotype SMCs cultured on the bottom of a dish as a two-dimensional monolayer with ECs separated from SMCs by a collagen gel. In this study, SMCs were randomly cultured in a collagen gel to allow more intercellular communication between ECs and SMCs, as compared to the previous model.…”

“…SMCs cultured in QM for 15 days (QM15) and ECs on the membrane were combined to complete the co-culture model, and then exposed to different magnitudes of SS (0.2, 2, 6, and 10 Pa) in parallel plate flow chambers for 72 h using a co-culture medium (CM) comprised of 25% PM and 75% QM [8,9] at 37°C in an atmosphere of 95% air/ 5% CO 2 .…”