Abstract
We have reported that recombinant biglycan (BGN) core protein accelerates bone formation in vivo by enhancing bone morphogenetic protein (BMP)-2 function. The purpose of the present study was to identify the specific domain (“effector”) within the BGN core protein that facilitates BMP-2 osteogenic function. Thus, we generated various recombinant and synthetic peptides corresponding to several domains of BGN, and tested their effects on BMP-2 functions in vitro. The results demonstrated that the leucine-rich repeats 2–3 domain (LRR2-3) of BGN significantly enhanced the BMP-2 induced Smad1/5/9 phosphorylation, osteogenic gene expression, and alkaline phosphatase activity in myogenic C2C12 cells. Furthermore, addition of LRR2-3 to osteoblastic MC3T3-E1 cells accelerated in vitro mineralization without compromising the quality of the mineral and matrix. These data indicate that LRR2-3 is, at least in part, responsible for BGN’s ability to enhance BMP-2 osteogenic function, and it could be useful for bone tissue regeneration.
Highlights
Craniofacial bone defects are a major health threat to our society directly affecting the quality and length of human life
This murine-derived myogenic cell line re-differentiates into osteoblastic cells upon bone morphogenetic protein (BMP)-2 treatment[21,22], as such, it is widely used to evaluate the osteogenic function of BMP-2
When the groups were compared by JMP program (SAS, Cary, NC) using a two-tailed t-test at 0.05 significance level (n = 2/ group), the alkaline phosphatase (ALP) activities of ΔNC-GST + BMP, BGN-GST + BMP and N-terminus to leucine-rich repeat domain 6 (N-LRR6)-GST + BMP were significantly higher than BMP alone while LRR7 to C-terminus (LRR7-C)-GST + BMP was not
Summary
Craniofacial bone defects are a major health threat to our society directly affecting the quality and length of human life. A critical barrier to progress in BMP-based bone regeneration is centered around a lack of understanding the optimal BMP dose to maximize osteogenic function while limiting potential side effects[8]. Biglycan (BGN) is a ~150 kDa molecule composed of a ~45 kDa core protein and two glycosaminoglycan (GAG) chains. It was first identified in bone matrix[9], but it is clear that it is present in non-skeletal tissues modulating various biological processes such as innate immunity, chemotaxis, angiogenesis, and growth factor regulation[10,11,12]. The positive effect of BGN on BMP-2 function is derived from the core protein not the glycosaminoglycan (GAG) component[19], and 3. The analyses showed the bands corresponding to ∆NC (~60 kDa) (lane 1), N-LRR6 (~50 kDa) (lane 2) and LRR7-C (~40 kDa) (lane 3) that were immunopositive with anti-
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