Downhill running increases bone mass accumulation and bone tissue morphology through TGF- β /Smad pathway in mice

Abstract Background: Compelling evidence has demonstrated that impact exercise can help to accumulate more bone mass. However, the cellular mechanism is less clear. The current study aimed to examine the effects of downhill running on osteoblast formation and the ability of mineralization in mice. The expression of transforming growth factor-b (TGF-b)/Smad pathway molecules was also determined. Methods: Twenty-eight 4-week-old C57BL/6 female mice were randomly assigned to either downhill running group (DG) or control group (CG). DG mice received a daily forty-minute downhill training on a decline treadmill (9 o decline) for 8 weeks. Eight weeks later, all mice were killed. Bone marrow stromal cells were collected and cultured for assessment of osteoblastic differentiation and the capacity of osteoblastic mineralization. We determined the protein (Western Blot) and mRNA (RT-PCR) expression of TGF-b, p-Smad2/3/4, and Runx2 in bone tissue. The bone mineral density (BMD) and histomorphological changes were also examined. Results: Eight-week training significantly increased bone mineral density (BMD) and enhanced the expression of alkaline phosphatase positive (ALP + ) osteoblasts in tibia epiphysis. Downhill running also promoted bone volume fraction (BV/TV), trabecular number (Tb.N) and trabecular thickness (Tb.Th), while it decreased the trabecular separation (Tb.Sp). Additionally, the numbers of alkaline phosphatase positive colony forming units-fibroblastic (ALP ＋ CFU-f) cells and the area of mineralized nodule formation were significantly higher in training group compared with the control group. Furthermore, the mRNA expression of TGF-b, Smad2, Smad3, Smad4 and Runt-related transcription factor 2 (Runx2) were significantly elevated by downhill training, and the protein expression of p-Smad2, p-Smad3, p-Smad4 and Runx2 also increased in exercise group. Conclusion: The findings suggested that downhill running enhanced bone accrual in mice mainly by promoting osteoblastic differentiation and the ability of mineralization. The beneficial changes partly be regulated by TGF-b/Smad signaling pathway.