Melatonin promotes osteoblast differentiation and mineralization of MC3T3‐E1 cells under hypoxic conditions through activation of PKD/p38 pathways

Abstract

Osteoblastic differentiation and bone-forming capacity are known to be suppressed under hypoxic conditions. Melatonin has been shown to influence cell differentiation. A number of in vitro and in vivo studies have suggested that melatonin also has an anabolic effect on bone, by promoting osteoblastic differentiation. However, the precise mechanisms and the signaling pathways involved in this process, particularly under hypoxic conditions, are unknown. This study investigated whether melatonin could promote osteoblastic differentiation and mineralization of preosteoblastic MC3T3-E1 cells under hypoxic conditions. Additionally, we examined the molecular signaling pathways by which melatonin mediates this process. We found that melatonin is capable of promoting differentiation and mineralization of MC3T3-E1 cells cultured under hypoxic conditions. Melatonin upregulated ALP activity and mRNA levels of Alp, Osx, Col1, and Ocn in a time- and concentration-dependent manner. Alizarin red S staining showed that the mineralized matrix in hypoxic MC3T3-E1 cells formed in a manner that was dependent on melatonin concentration. Moreover, melatonin stimulated phosphorylation of p38 Mapk and Prkd1 in these MC3T3-E1 cells. We concluded that melatonin promotes osteoblastic differentiation of MC3T3-E1 cells under hypoxic conditions via the p38 Mapk and Prkd1 signaling pathways.

An insight into the scientific background and future perspectives for the potential uses of melatonin

“…Recently, an in vitro study demonstrates that melatonin promotes differentiation and mineralization of cultured osteoblast cells (MC3T3-E1 cells) under hypoxic conditions [145]. Thus this study concludes that melatonin promotes osteoblastic differentiation of MC3T3-E1 cells under hypoxic conditions via the p38 Mapk and Prkd1 signaling pathways.…”

Behavior of Human Osteoblast Cells Cultured on Titanium Discs in Relation to Surface Roughness and Presence of Melatonin

“…A study by Son, using MC3T3 cells cultured in media containing melatonin at concentrations between 10 and 250 µM. quantified gene expression of ALP , COLIA1 , OCN , and OSX after 1, 3, 7, and 14 days of cell culture showed that gene expression increased in dose and time-dependent manner [46]. However, a study by Nakade using human osteoblast cells and the addition of melatonin to the cell culture media at concentration between 5 and 100 µM, measured different markers of osteoblast differentiation after four days of cell culture.…”

“…Glyceraldehyde 3-phosphate dehydrogenase ( GAPDH ) is one of the most widely used housekeeping genes for managing and normalizing the expression of other genes [7,15,17,20,30,46]. …”

Melatonin promotes osteoblast differentiation by regulating Osterix protein stability and expression

“…One of the mechanisms by which melatonin regulates the bone environment is through its stimulatory effect on osteoblasts 33, 34 . A previous study revealed that melatonin enhances the differentiation of human adult mesenchymal stem cells into mature osteoblasts via MT2 melatonin receptors 35 , and that, during osteoblast differentiation, treatment with melatonin increases the expression levels of osteocalcin (OC), a late, highly specific osteoblast marker of bone formation, followed by enhanced bone mineralization of MC3T3-E1 cells 36 . Although it has been reported that melatonin promotes the expression of Runx2, a master transcription factor during the early stage of osteogenesis 37 , the precise mechanism by which it regulates the differentiation of preosteoblasts into mature osteoblasts remains unknown.…”

“…Moreover, an intraperitoneal administration of melatonin slightly enhanced new cortical bone formation in the femurs of mice 19 . In addition, melatonin promoted mineralization and osteoblastic differentiation by enhancing the expression of Runx2, which is a key transcription factor in the early stage of osteogenic differentiation, via the mitogen-activated protein kinase (MAPK) signaling pathway 20 .…”