Osteoblast response to fluid induced shear depends on substrate microarchitecture and varies with time.

Osteoblasts are exposed to fluid shear in vivo but the effects are not well understood, particularly how substrate properties or length of exposure modify the response. Short exposure (1 h) to shear reduces the stimulatory effect of micron-scale surface structure on osteoblast differentiation, but the effects of longer term exposures are not known. To test the hypothesis that substrate-dependent responses of osteoblasts to shear depend on the length of exposure to fluid flow, MG63 osteoblasts were grown on tissue culture glass, which has an average roughness (Ra) < 0.2 microm; machined Ti disks (PT, Ra < 0.6 microm); Ti disks with a complex microarchitecture [sand blasted acid etched (SLA), Ra = 4-5 microm); and Ti plasma-sprayed surfaces [Ti via plasma spray (TPS), Ra = 7 microm]. Confluent cultures were exposed to pulsatile flow at shear forces of 0, 1, and 14 dynes/cm(2) for 0, 6, 12, and 24 h. Shear reduced cell number on all surfaces, with greatest effects on TPS. Shear had no effect on alkaline phosphatase on smooth surfaces but increased enzyme activity on SLA and TPS in a time-dependent manner. Its effects on osteocalcin, TGF-beta1, and PGE(2) in the conditioned media were greatest on these surfaces as well. Responses to fluid-induced shear were blocked by the general Cox inhibitor indomethacin and the Cox-2 inhibitor meloxicam, indicating that response to shear is mediated by prostaglandin produced via a Cox-2 dependent mechanism. These results show that the effects of fluid induced shear change with time and are substrate dependent, suggesting that substrate microarchitecture regulates the osteoblast phenotype and effects of shear are determined by the maturation state of the responding population.

Shear force modulates osteoblast response to surface roughness.

Previous studies have shown that osteoblasts are sensitive to surface roughness. When cultured on Ti, MG63 osteoblast-like cells exhibit decreased proliferation and increased differentiation with increasing surface roughness. In vivo, osteoblasts also are subjected to shear force during osseointegration. To examine how shear force modulates osteoblast response to surface roughness, MG63 cells were cultured on glass disks or Ti disks with three different R(a) values and topographies (PT: R(a) = 0.60 microm; SLA: R(a) = 3.97 microm; TPS: R(a) = 5.21 microm) in a continuous flow device, resulting in shear forces of 0, 1, 5, 14, and 30 dynes/cm(2). Confluent cultures were exposed to fluid flow for 1 h. After an additional 23 h, cell number, alkaline-phosphatase-specific activity, and levels of osteocalcin, TGF-beta1, and PGE2 in the conditioned media were determined. Cell numbers on smooth surfaces (glass and PT) were unaffected by shear force. In contrast, shear force caused a dose-dependent reversal of the decrease in cell numbers seen on rough SLA and TPS surfaces. Alkaline-phosphatase-specific activity was unaffected on glass or PT, but shear force caused a biphasic reduction in the roughness-dependent increase on SLA and TPS that was maximal at 14 dynes/cm(2). There was a similar effect seen with TGF-beta1 levels. Osteocalcin was unaffected on smooth surfaces; shear force caused a dose-dependent reduction in the roughness-stimulated increase seen on SLA and TPS. PGE2 production was increased by shear force on all surfaces. There was a twofold increase in PGE2 levels in the media of MG63 cells cultured on glass and PT in response to 14 dynes/cm(2), but on SLA and TPS, 14 dynes/cm(2) shear force caused a 9-10-fold increase. These results show that osteoblastic response to shear force is modulated by surface topography. The shear-force-mediated decrease in osteoblast differentiation seen in cultures on rough surfaces may be due to increased production of PGE2.