Considering the increased resistance to antibiotics in the clinic and the ideal antibacterial properties of KR‑12, the effects of KR‑12‑a6, an important analogue of KR‑12, on the osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) were investigated. Osteogenic differentiation‑associated experiments were conducted in hBMSCs, and KR‑12‑a6 was used as an additional stimulating factor during osteogenic induction. Quantitative analysis of alkaline phosphatase (ALP) and alizarin red staining, and reverse transcription‑quantitative PCR analysis of the expression of osteogenesis‑associated genes were performed to determine the effects of KR‑12‑a6 on the osteogenic differentiation of hBMSCs. LDN‑212854 was selected to selectively suppress BMP/SMAD signaling. Western blotting was performed to investigate the underlying mechanisms. The intensity of ALP and alizarin red staining gradually increased with increasing KR‑12‑a6 concentrations. KR‑12‑a6 induced the strongest staining at 40 µg/ml, whereas 60 µg/ml and 80 µg/ml concentrations did not further increase the intensity of staining. The mRNA expression levels of RUNX2 and ALP increased in a dose‑dependent manner as early as 3 days post‑KR‑12‑a6 treatment. The mRNA expression of COL1A1, BSP and BMP2 exhibited significant upregulation from day 7 post‑KR‑12‑a6 treatment. In contrast, the mRNA levels of OSX, OCN and OPN were enhanced dramatically at day 14 following KR‑12‑a6 stimulation. Additionally, KR‑12‑a6 significantly promoted the phosphorylation of Smad1/5. Furthermore, LDN‑212854 suppressed the activation of Smad1/5 and inhibited the upregulation of several osteogenic differentiation‑associated genes in KR‑12‑a6‑treated hBMSCs. KR‑12‑a6 promoted the osteogenic differentiation of hBMSCs via BMP/SMAD signaling.
Keywords: Kr-12-a6; antimicrobial peptide; osteogenic differentiation; human bone marrow mesenchymal stem cells; bone morphogenic protein/SMad signaling.