Effects of resveratrol supplementation on bone growth in young rats and microarchitecture and remodeling in ageing rats

Abstract

Osteoporosis is a highly prevalent skeletal disorder in the elderly that causes serious bone fractures. Peak bone mass achieved at adolescence has been shown to predict bone mass and osteoporosis related risk fracture later in life. Resveratrol, a natural polyphenol compound, may have the potential to promote bone formation and reduce bone resorption. However, it is unclear whether it can aid bone growth and bone mass accumulation during rapid growth and modulate bone metabolism during ageing. Using rat models, the current study investigated the potential effects of resveratrol supplementation during the rapid postnatal growth period and in late adulthood (early ageing) on bone microarchitecture and metabolism. In the growth trial, 4-week-old male hooded Wistar rats on a normal chow diet were given resveratrol (2.5 mg/kg/day) or vehicle control for 5 weeks. In the ageing trial, 6-month-old male hooded Wistar rats were treated with resveratrol (20 mg/kg/day) or vehicle for 3 months. Treatment effects in the tibia were examined by μ-computer tomography (μ-CT) analysis, bone histomorphometric measurements and reverse transcription-polymerase chain reaction (RT-PCR) gene expression analysis. Resveratrol treatment did not affect trabecular bone volume and bone remodeling indices in the youth animal model. Resveratrol supplementation in the early ageing rats tended to decrease trabecular bone volume, Sirt1 gene expression and increased expression of adipogenesis-related genes in bone, all of which were statistically insignificant. However, it decreased osteocalcin expression (p = 0.03). Furthermore, serum levels of bone resorption marker C-terminal telopeptides type I collagen (CTX-1) were significantly elevated in the resveratrol supplementation group (p = 0.02) with no changes observed in serum levels of bone formation marker alkaline phosphatase (ALP). These results in rat models suggest that resveratrol supplementation does not significantly affect bone volume during the rapid growth phase but may potentially have negative effects on male skeleton during early ageing.

Figure 1

The effects of resveratrol supplementation on growth plate, primary spongiosa heights, and bone volume in the tibia of young rats. 3D reconstruction images showing trabecular bone in (A) a control rat and (B) a resveratrol treated rat; (C) The effects of resveratrol supplementation on trabecular bone volume/tissue volume (BV/TV) from μ-CT analysis in young animals; (D) Histological analysis showing H & E stained section showing the growth plate and primary spongiosa of a control rat (bar = 500 μm); (E) Measurements of growth plate thickness; (F) Measurements of primary spongiosa heights.

Figure 2

Effects of resveratrol supplementation on the trabecular bone structure and levels of mRNA expression of osteogenesis-related factors in the metaphysis of the tibial bone of ageing rats. 3D reconstruction images showing trabecular bone in (A) a control rat and (B) a resveratrol treated rat; (C) Effects on trabecular BV/TV from μ-CT analysis; (D) Effects on cortical bone volume from μ-CT analysis; (E) RT-PCR gene expression analyses for osterix (p = 0.34); (F) RT-PCR gene expression analyses for osteocalcin (p = 0.02). * p < 0.05 compared to control group, n = 7.

Figure 3

Effects of resveratrol supplementation on osteoclast, osteoblast, and adipocyte densities within the secondary spongiosa in early ageing rats. Histological analyses demonstrating (A) TRAP stained section of a control tibial bone showing stained osteoclasts (cells stained red containing three + nuclei as indicated by arrows) (bar = 100 μm); (B) Osteoclast density at the secondary spongiosa (n = 7); (C) H & E-stained section showing osteoblasts in secondary spongiosa as indicated by arrows (bar = 125μm); (D) Osteoblast density within the secondary spongiosa (n = 7); (E) H & E-stained section showing adipocytes in lower secondary spongiosa as indicated by arrows (bar = 100 μm); (F) Adipocyte density within the lower secondary spongiosa (n = 7).

Figure 4

Effects of resveratrol supplementation on mRNA expression levels of adipogenesis-related factors and Sirtuin 1 (Sirt1) in early ageing rats. RT-PCR gene expression analyses at tibial metaphysis for (A) C/EBPα (p = 0.06); (B) FABP4 (p = 0.34) and (C) Sirt1 (p = 0.36) (n = 7). C/EBPα, CCAAT/enhancer-binding protein alpha; FABP4, fatty acid-binding protein 4.

Figure 5

Effects of resveratrol supplementation on serum levels of bone resorption marker collagen type I cross-linked C-telopeptide (CTX-1) and bone formation marker alkaline 12hosphatase (ALP) in the ageing rats. Serum levels of (A) CTX-1 (p = 0.02); (B) ALP (p = 0.36).