aminoisobutyric acid BAIBA is a natural catabolite of the

β-aminoisobutyric KH CB19 (BAIBA) is a natural catabolite of the branched-chain amino acid thymine. BAIBA was recently found to reduce inflammation in skeletal muscle via the AMP-activated protein kinase (AMPK)-peroxisome proliferator-activated receptor (PPAR)δ signaling pathway. BAIBA reportedly enhances browning of white fat and hepatic β-oxidation via PPARα. BAIBA alleviates hepatic endoplasmic reticular stress, and improves glucose/lipid metabolism in diabetes. Further, BAIBA ameliorates fasting blood glucose levels, insulin tolerance, and glucose tolerance in diabetic mice. We hypothesized that BAIBA may promote proliferation and differentiation of osteoblasts. Therefore, this study investigated whether and how BAIBA promote proliferation and differentiation of osteoblasts.
Methods
Results
Discussion Osteoporosis is a common comorbidity for sarcopenia associated with lowered density and strength of skeleton. BAIBA was initially developed as an anti-obesity agent that acts via increasing fatty acid oxidation (FAO) in the liver and decreasing body fat mass in mice. In the present study, treatment of MC3T3-E1 cells with BAIBA obviously accelerated the proliferation and differentiation of osteoblasts, and the underlying mechanisms were related to the activation of NADPH oxidase/ROS signaling. The homeostasis between bone formation and bone resorption is requisite for maintaining bone growth. Prevention of bone loss or osteoclast activity inhibition may be recommended as therapeutics for osteoporosis. However, as osteoporosis patients generally have already lost bone mass before treatment, promotion of bone formation is a better approach to treat osteoporosis. Osteoblasts are fundamental components in the formation of new bone tissues. Promoting the proliferation and differentiation of osteoblasts is a pivotal event in bone formation. In the present study, we showed that BAIBA dose- and time-dependently promoted the proliferation of MC3T3-E1 cells. qPCR results demonstrated that BAIBA obviously upregulated the mRNA expression of the osteoblast transcription regulators RUNX2 and Osx, as well as the differentiation markers Col1α-1, OC, OPN, and OPG. In addition, we showed that both the activity and mRNA expression of ALP were dose-relatedly increased in MC3T3-E1 cells exposed to BAIBA stimulation. These data indicate that BAIBA dramatically accelerates osteoblasts proliferation and differentiation. ROS are suggested to be crucial in pre-osteoblast proliferation induced by low-level laser therapy and are thought to be key secondary messengers in osteoblast proliferation and differentiation.14, 38 In this study, we showed that treatment of MC3T3-E1 cells with BAIBA led to a significant increase in ROS production as evidenced by DHE fluorescence and increased superoxide anion level, NAD(P)H oxidase activity, and H2O2 content. In addition, the ROS scavenger NAC as well as the NADPH oxidase inhibitor apocynin significantly abolished the proliferation and differentiation of osteoblasts challenged by BAIBA. These results suggested that the ROS signaling pathway is involved in BAIBA-evoked osteoblast proliferation and differentiation. It is worth mentioning that subtype NAD(P)H oxidases such as NOX1, NOX2, and NOX4 are abundantly expressed in osteoblasts. We showed that the protein levels of NOX4 are dramatically upregulated in MC3T3-E1 cells in response to BAIBA, which provides direct evidence that NOX4 may largely contribute to the BAIBA-induced ROS production in MC3T3-E1 cells. In conclusion, the results indicate that BAIBA promotes the proliferation and differentiation of osteoblasts via activation of the ROS signaling pathway, which may help to clarify the mechanism of BAIBA-induced bone formation. It is particularly worth noting that excess ROS cause bone loss. We showed that high dose of BAIBA has the same effect on excessive ROS production as H2O2 in osteoblasts cells, while low dose of BAIBA in association with moderate ROS production plays a protective role in osteoblast proliferation and differentiation. We will evaluate the potential of BAIBA as a bone-forming drug in a future study. The suitable dose of BAIBA in animal studies remains unclear and requires further research.