Melatonin promotes osteoblastic differentiation through the BMP/ERK/Wnt signaling pathways.
ABSTRACT Although melatonin has a variety of biological actions such as antitumor, antiangiogenic, and antioxidant activities, the osteogenic mechanism of melatonin still remains unclear. Thus, in the present study, the molecular mechanism of melatonin was elucidated in the differentiation of mouse osteoblastic MC3T3-E1 cells. Melatonin enhanced osteoblastic differentiation and mineralization compared to untreated controls in preosteoblastic MC3T3-E1 cells. Also, melatonin increased wound healing and dose-dependently activated osteogenesis markers such as runt-related transcription factor 2 (Runx2), osteocalcin (OCN), bone morphogenic protein (BMP)-2 and -4 in MC3T3-E1 cells. Of note, melatonin activated Wnt 5 α/β, β-catenin and the phosphorylation of c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) in a time-dependent manner while it attenuated phosphorylation of glycogen synthase kinase 3 beta (GSK-3β) in MC3T3-E1 cells. Consistently, confocal microscope observation revealed that BMP inhibitor Noggin blocked melatonin-induced nuclear localization of β-catenin. Furthermore, Western blotting showed that Noggin reversed activation of β-catenin and Wnt5 α/β and suppression of GSK-3β induced by melatonin in MC3T3-E1 cells, which was similarly induced by ERK inhibitor PD98059. Overall, these findings demonstrate that melatonin promotes osteoblastic differentiation and mineralization in MC3T3-E1 cells via the BMP/ERK/Wnt pathways.
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ABSTRACT: Kirenol has been reported to possess anti-oxidant, anti-inflammatory, anti-allergic, anti-adipogenic, and anti-arthritic activities; however, its effect on osteoblast differentiation has not yet been reported. The aim of the present study was to evaluate the effect of kirenol on osteoblast differentiation through activation of the bone morphogenetic protein (BMP) and Wnt/β-catenin signaling pathways in MC3T3-E1 cells. Kirenol markedly promoted alkaline phosphatase (ALP) activity and mineralization. Kirenol not only increased the expression of osteoblast differentiation markers, such as ALP, type I collagen (ColA1), and osteopontin (OPN), but also increased the expression of osteoprotegerin/receptor activator of nuclear factor kappa B ligand (OPG/RANKL) ratio. The effects of kirenol on osteoblast differentiation were accompanied by stimulating the expression of the BMP and Wnt/β-catenin signaling pathways, including BMP2, runt-related transcription factor 2 (Runx2), osterix (Osx), low density lipoprotein receptor related protein 5 (LRP5), disheveled 2 (DVL2), β-catenin, cyclin D1 (CCND1), and phosphorylated glycogen synthase kinase 3β (GSK3β). In addition, kirenol up-regulated the expression of β-catenin, CCND1, ALP, and ColA1 which were down-regulated by siRNA knockdown of β-catenin. Overall, these results demonstrate that kirenol is capable of promoting osteoblast differentiation in MC3T3-E1 cells through activation of the BMP and Wnt/β-catenin signaling pathways, suggesting that it is a potential candidate target for treating or preventing osteoporosis.Fitoterapia 01/2014; · 2.23 Impact Factor
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ABSTRACT: Objective The aim of this study was to evaluate the effect of the topical application of melatonin compared with collagenized porcine bone grafts to accelerate bone formation 2 months after their insertion in tibiae rabbits. Material and Methods Twenty New Zealand rabbits weighing 3,900–4,500 g were used. Twenty collagenized porcine bone (MP3) grafts, twenty melatonin-impregnated bone grafts, and twenty control areas were placed in the proximal metaphyseal area of both rear tibias. Four groups were formed according to the moment in which animal killing was carried out: Group I (15 days), Group II (30 days), Group III (45 days) and Group IV (60 days). Cortical width and cortical length of bone formation was measured. Following implantation, an anteroposterior and lateral radiological study was carried out. Samples were sectioned at 5 μm and stained using hematoxylin-Eeosin, Masson's trichromic, and Gordon-Switt reticulin stains. ResultsAfter 60 days of treatment period, melatonin increased the length of cortical bone formation 99.03 ± 0.61% like control 98.90 ± 3.82% compared with porcine bone 92.73 ± 1.08%. Related to perimeter of cortical bone of the tibiae melatonin new bone was 98.35 ± 1.14% like control 98.0 ± 1.43% more than porcine bone 92.05 ± 1.03%. Histomorphometric values related to porcine bone were connective tissue 49.16 ± 2.4%, graft material (MP3) 23.52 ± 2.3%, and new bone formation 27.32 ± 1.4% compared with test group with melatonin 24.5 ± 1.2%, connective tissue 45.1 ± 1.2%, and new bone formation of 30.4 ± 1.0%. Conclusion Melatonin has proven to regenerate the width and length of cortical bone in tibiae rabbits more quickly than collagenized porcine bone. Melatonin acts as a bone stimulator compared with porcine bone and control sites.Clinical Oral Implants Research 03/2014; · 3.43 Impact Factor
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ABSTRACT: To elucidate the effects of melatonin on cisplatin-induced hepatocellular carcinoma (HepG2) cell death and to identify potential cross-talk pathways. Hepatocellular carcinoma HepG2 cells were treated with melatonin and/or cisplatin for 24 to 48 h. Cell viability and the 50% cytotoxic concentration (CC50) were calculated by MTT assays. The effects and intracellular events induced by the selected concentrations of melatonin (1 mmol/L) and cisplatin (20 μmol/L) were investigated. Cell death and survival detection were primarily evaluated using a fluorescence microscope to assess 4',6 diamideno-2-phenylindol DNA staining and acridine orange lysosome staining and then further analyzed with immunocytochemistry using an anti-LC3 antibody. The potential molecular responses mediated by melatonin against cisplatin after the combined treatment were investigated by reverse transcription-polymerase chains reaction and Western blot analyses of the genes and proteins associated with cell survival and death. A cell cycle analysis was performed using a flow cytometry assay. Melatonin had a concentration-dependent effect on HepG2 cell viability. At 1 mmol/L, melatonin signiﬁcantly increased the cell viability percentage and decreased reactive oxygen species production due to cisplatin. Melatonin reduced cisplatin-induced cell death, decreasing phosphorylated p53 apoptotic protein, cleaved caspase 3 and Bax levels but increasing anti-apoptotic Bcl-2 gene and protein expression. When combined with cisplatin, melatonin induced S phase (DNA synthesis) cell cycle arrest and promoted autophagic events in HepG2 cells. Melatonin also had a concentration-dependent effect on Beclin-1 and its autophagic regulator mammalian target of rapamycin (mTOR) as well as the DNA excision repair cross complementary 1 (ERCC1) protein. The expression levels of these proteins were altered in HepG2 cells during cisplatin or melatonin treatment alone. In the combination treatment, melatonin reversed the effects of cisplatin by suppressing the over-expression of mTOR and ERCC 1 and enhancing the expression levels of Beclin-1 and microtubule-associated protein-light chain3-II, leading to intracellular autophagosome progression. Melatonin attenuated cisplatin-induced cell death in HepG2 cells via a counter-balance between the roles of apoptotic- and autophagy-related proteins.World journal of hepatology. 04/2014; 6(4):230-42.