Icariin enhances differentiation and maturation of rat calvarial osteoblasts in collagen hydrogel three-dimensional culture.
- Author:
Yan-fang XIE
1
,
2
;
Ming-gang WANG
1
;
Ke-ming CHEN
1
;
Wen-gui SHI
1
;
Jian ZHOU
1
;
Yu-hai GAO
1
;
Author Information
- Publication Type:Journal Article
- MeSH: Alkaline Phosphatase; metabolism; Animals; Bone Morphogenetic Protein 2; metabolism; Cell Differentiation; Cell Proliferation; Cells, Cultured; Collagen; chemistry; Core Binding Factor Alpha 1 Subunit; metabolism; Drugs, Chinese Herbal; Flavonoids; pharmacology; Hydrogels; chemistry; Osteoblasts; cytology; drug effects; Rats; Rats, Sprague-Dawley; Skull; cytology; Transcription Factors; metabolism
- From: Journal of Zhejiang University. Medical sciences 2015;44(3):301-307
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate the effects of icariin on the differentiation and maturation of rat calvarial osteoblasts(ROB) in collagen hydrogel three-dimensional culture.
METHODSROB were obtained by enzyme digestion from the segregated neonatal SD rats skull and were embedded in 2 mg/mL rat tail collagen for three-dimensional culture. The growth state of ROB was observed by FDA/PI staining, HE staining and scanning electron microscopy. ROB were treated with icariin at the concentration of 1 × 10⁻⁴, 1 × 10⁻⁵, 1 × 10⁻⁶ and 1 × 10⁻⁷ mol/L respectively. The activity of alkaline phosphatase(ALP) was detected after 3, 6, 9 d of icariin treatment. Three-dimensional cultured ROB were treated with optimal concentration icariin for 12, 24, 36, 48 h and total RNA was extracted and the mRNA expressions of bone morphogenetic protein-2 (BMP-2), Runt-related transcription factor 2 (RUNX-2) and Osterix were detected by real time RT-PCR. The protein expression of BMP-2, RUNX-2 and Osterix were examined by Western-blotting.
RESULTSROB were cultured in collagen hydrogel successfully. FDA/PI staining, HE staining, and scanning electron microscopy showed that ROB adhered with collagen tightly and distributed homogeneously. Icariin at final concentration of 1 × 10⁻⁵, 1 × 10⁻⁶ and 1×10⁻⁷ mol/L all enhanced the activity of ALP of collagen hydrogel three-dimensional cultured ROB, and 1 × 10⁻⁶ mol/L was the optimal concentration. Besides, icariin (1 × 10⁻⁶ mol/L) increased mRNA and protein expression of BMP-2、RUNX-2 and Osterix compared to control group.
CONCLUSIONIcariin can enhance the expression of osteogenic markers of ROB in collagen hydrogel three-dimensional culture significantly.