Identification of the most effective isomer of vitamin E as antiosteoporotic and anabolic agent using a static three-dimensional human osteoblast-osteoclast co-culture system

Abstract

Osteoporosis is a metabolic bone disorder affecting more than 200 million elderly worldwide. Many animal and cellular studies have been conducted in order to find a new antiosteoporotic agent to combat this disease. To date, naturally available vitamin E from plant oils such as palm and annatto oils have been shown to possess antiosteoporotic properties and bone anabolic properties. Therefore, this study was aimed to determine the antiosteoporotic effects of individual tocotrienols (alpha, beta, gamma, delta) and alpha-tocopherol isomers using a static three-dimensional human osteoblast-osteoclast co-culture system on bovine bone scaffold. Eight analyses were done namely designing bone scaffold from bovine trabecular bone, optimization of co culture time- point, optimization of osteoblast-osteoclast co-culture ratio, bone mechanical strength, bone histomorphometry, micro-CT imaging system, scanning electron microscopy and dual-energy X-ray absorptiometry. On day three, which was the optimized co- culture time point, human osteoclasts derived from peripheral blood mononuclear cells were added to the culture of human fetal osteoblasts cell line (hFOB 1.19) at the ratio of one osteoclast to two osteoblasts, as pre-determined based from the TRAP staining result. The co-culture was then seeded onto demineralized osteoporotic bovine bone scaffold and treated with individual tocotrienol isomers (alpha, beta, gamma, delta) and alpha-tocopherol at the concentration of 100 nM for 21 days under standard culture conditions (37°C, 95% humidity and 5% CO2 concentration). Delta- and gamma-tocotrienols gave the greatest improvement on bone microstructure as compared to other isomers; as seen from bone histomorphometry (bone surface, bone parameter, bone volume fraction, bone trabecular thickness, bone trabecular number, bone trabecular separation), micro-CT imaging (total bone volume and total bone porosity), scanning electron microscopy (highest cells adhesion and proliferation) and dual-energy X-ray absorptiometry (bone mineral content and bone mineral density). However, for bone biomechanical strength, the delta isomer showed the highest total bone strength and was significantly higher as compared to gamma and other isomers. In conclusion, delta- and gamma-tocotrienols were the most effective isomers of vitamin E for improving the demineralized bone. Thus, they are potentially the best tocotrienol isomers to be developed as antiosteoporotic agents.