Avenanthramides 2c, 2f and 2p regulate osteoblast gene expression and survival in vitro
Authors: Pellegrini Gretel G , Morales Cynthya C , Johnson Jodee , Wallace Taylor C , Bellido Teresita 
Published: November 2023

Increasing evidence suggests that long term consumption of some whole grain cereals, promotes good health. Oat is a commonly consumed whole‐grain cereal and contains different types of phytochemicals with antioxidant properties, such as tocotrienols, phenolic acids, flavonoids, sterols, and phytic acid. Oats are gaining increasing scientific and public interest for their antioxidant‐associated health benefits. Avenanthramides, a group of alkaloids uniquely found in oats that contain a phenolic group, have been reported to exhibit strong antioxidant activity both in vitro and in vivo in different cell types. To date, there are no studies addressing the effect of avenanthramides in bone in vivo or in bone cells in vitro, on bone homeostasis or the function of the cells of bone. Here, we investigated in vitro the ability of avenanthramides 2c, 2f and 2p to regulate the function of osteoblasts by determining their effects on gene expression and on basal and induced apoptosis in osteoblastic cells. In order to determine in vitro the ability of avenanthramides to regulate the function of osteoblasts, a 24‐hour dose response for osteoblast mRNA gene expression was conducted. OB‐6 osteoblastic cells were cultured in αMEM + 1% PSG + 10% FBS media for one day. Medium was changed and cells were cultured overnight. The next day, medium was removed and cells were cultured for 24‐h with fresh αMEM + PSG + 2% FBS (500 μl/well) with control (DMSO) and 0.01, 0.1, 1, 10 and 100 μM of avenantramide 2c, 2f and 2p. RNA was extracted and gene expression was determined by quantitative PCR and quantified by the delta CT method. Although the expression of osteoblasts markers osteocalcin, runx2, collagen 1, osterix and alkaline phosphatase was not affected by avenanthramides 2c, 2f and 2p, and RANKL was not detected, there was an increase in OPG (osteoprotegerin) gene expression with the lowest doses (0.01 and 0.1 μM) of avenanthramides 2f and 2c. To examine the effect of avenanthramide 2f on basal and induced apoptosis in OB‐6 cells, cells were culture for 20‐h in αMEM + PSG + 10% FBS. Medium was changed to αMEM + PSG + 2% FBS+ control (DMSO) or avenanthramide 2f at 1, 10 and 100 μM (300 μl/well) and cultured for 1 h. The pro‐apoptotic agent etoposide (50 μM) was added to the corresponding wells for 6 h. Cell viability was determined by trypan blue uptake. The presence of avenanthramide 2f did not affect basal levels of apoptosis of osteoblastic OB‐6 cells. However, avenanthramide 2f prevented etoposide‐induced apoptosis in a dose dependent manner. These results demonstrate that avenanthramides may directly affect gene expression and survival of osteoblasts upregulating the expression of the anti‐osteoclastogenic cytokine OPG and preventing osteoblast apoptosis. Thus, the beneficial effects of avenanthramides, antioxidant compounds found in oats, might be related to regulation of bone cell function and life span.Support or Funding InformationThis study was supported by the National Osteoporosis Foundation through unrestricted educational grants from the Dairy Management Inc. and the Egg Nutrition Center and PepsiCo, Inc., Quaker Oats Center of Excellence, PepsiCo R & D, Inc, Barrington, IL, USA