Enhancement of the ALP activity of C3H10T1/2 cells by the combination of an oxysterol and apatite

Abstract

Biomimetic apatite coating has been used to load osteogenic biomolecules onto the surface of titanium implants. Apatite on the surface of biomaterials is thought to function as a reservoir of biomolecules as well as enhancing osteoconductivity. In this study, 20alpha-hydroxycholesterol (20alpha-HC), an osteogenic oxysterol, was used to induce differentiation of a mouse embryo fibroblast cell line (C3H10T1/2) by loading the oxysterol on biomimetically coated apatite of titanium discs. We found that the phosphatase (alkaline phosphatase (ALP)) activity of 20alpha-HC was significantly higher with ascorbic acid than alone, suggesting a need for ascorbic acid as a co-factor. When 20alpha-HC was added into the apatite coating solution, the ALP activity of the C3H10T1/2 cells did not increase on the apatite surface, even in the presence of ascorbic acid. However, ALP activity increased dramatically when 20alpha-HC was loaded by volatilization of EtOH from the apatite coat after dipping discs in 20alpha-HC-dissolved EtOH. Interestingly, ascorbic acid was not needed for this increase in ALP activity, suggesting a synergistic effect of 20alpha-HC and apatite. The concentration of calcium ions, a major component of apatite, affected the osteogenic effect of 20alpha-HC, and the increase in ALP activity was attenuated by L-type calcium channel inhibitors, verapamil and nifedipine. These results demonstrate that calcium ions released from apatite are important in the synergistic effect of 20alpha-HC and apatite.

Hair growth-promoting effect of Aconiti Ciliare Tuber extract mediated by the activation of Wnt/β-catenin signaling

“…Cell culture iDPC and immortalized rat vibrissa dermal papilla cells (RvDP) were prepared by Won CH et al and maintained in Dulbecco's Modified Eagle Medium (DMEM) (PAA Laboratories, Austria) containing 10% (vol/vol) fetal bovine serum (FBS) (PAA Laboratories, Austria) and 1% (vol/vol) penicillin/streptomycin (Gibco BRL, NY, USA) in a humidified atmosphere of 95% air/5% CO 2 at 37°C (Son et al, 2010;Won et al, 2010). Human embryonic kidney (HEK-293) cells were maintained in DMEM containing 10% (vol/vol) FBS.…”

Early bone healing onto implant surface treated by fibronectin/oxysterol for cell adhesion/osteogenic differentiation:in vivoexperimental study in dogs

“…This study was based on our two previous studies that demonstrated an increase in cell adhesion on fibronectin and patite-coated titanium surfaces [23] and enhancement of osteoblastic differentiation by oxysterol and apatite-coated titanium surfaces [27]. Since the development of dental implants using titanium, various surface treatments have been studied to make a bioactive titanium surface through the concept of mimicking cellular events in bone formation and remodeling processes [1,20].…”

“…Since the development of dental implants using titanium, various surface treatments have been studied to make a bioactive titanium surface through the concept of mimicking cellular events in bone formation and remodeling processes [1,20]. The two above-mentioned types of implant surfaces were also intended to transform the surface from "biotolerant" to "bioactive" titanium: The fibronectin and apatite coating was intended to promote cellular adhesion onto the surface as a first step in the healing process [23], and the oxysterol or apatite coating was intended to promote differentiation of the adhered cells [27]. However, most dental clinicians still prefer rough surface implants without any biomimetic surface modification, even though various types of biomimetic surface implants have been commercially available.…”

“…Recent studies have introduced a fibronectin-coated dental implant system in the research stage [21,22], and immobilization of fibronectin increased cell attachment [23] and osteoblastic protein expression at the in vitro level [24]. Another strategy in creating biomimetic implant surfaces is the use of growth factors [25,26] or their natural substitutes, such as oxysterol [27], for enhancement of osteogenic cell differentiation. Previous studies have demonstrated that oxysterols regulate differentiation of stem cells into osteogenic cells via the hedgehog pathway, and also prevent adipogenic differentiation in vitro [28,29].…”

“…Even though these two types of biomimetic implant surface developments showed increased cell attachment and differentiation in vitro [23,27], there was a lack of enhancement of osseointegration by using fibronectin or oxysterol in clinically-mimicking an in vivo animal model. Therefore, the present study aimed to evaluate the effects of fibronectin and oxysterol immobilized on machined-surface dental implants for the enhancement of cell attachment and osteogenic differentiation, on peri-implant bone healing in the early healing phase using an experimental model in dogs.…”