ID: 1008 Quercus infectoria semi-purified fractions promoted BMP-2, Runx2 and osteopontin expression in human fetal osteoblastic cell line

Quercus infectoria (QI) is traditional medicines in Asia which contain many active phytochemical compounds that have the potential to stimulate bone formation. However, the precise mechanism of the therapeutic effect of QI on osteoblast has not been elucidated. The present study was carried out to extract the active compound of QI through a series of purification in order to obtain semi-purified fractions of QI followed by examining its phytochemical profile through liquid chromatography mass spectrometry (LC-MS) as well as to delineate the molecular mechanism of QI semi-purified fraction-enhanced bone formation by investigating the protein expression of bone morphogenic protein-2 (BMP-2), Runx2 and osteopontin (OPN) in the hFOB 1.19 human fetal osteoblastic cell line. The semi-purified fractions of QI were produced in series of column chromatography and thin layer chromatography (TLC) technique. The obtained fractions were then tested in a series of bio-guided assay through MTT assay. Three most potent fractions with lowest EC50 value (Fraction A-10.85 µg/ml, Fraction B-12.00 µg/ml and Fraction C-11.60 µg/ml) were selected to be treated with hFOB1.19 cell followed by quantification of BMP-2, Runx2 and OPN protein expression at day 1, 3 and 7. Compound profiling through LC-MS analysis showed that the chosen fractions consist of main active component known as gallic acid that has been showed previously to promote bone formation. Results show that QI semi-purified fractions increased the activity of BMP-2, Runx2 and OPN at day 1 and day 3. Meanwhile as the expression of BMP-2 and Runx2 continue to increase at day 7, the expression of OPN shows a down regulation at day 7. Furthermore, the study showed that combination of Fraction A, B and C wit osteoporotic drug (pamidronate) further increase the expression of BMP-2 and Runx2 contrary to OPN expression. These finding demonstrated that semi-purified fraction of QI enhanced bone formation through expression of investigated bone-related marker.

Download Full-text

The Role of Semipurified Fractions Isolated from Quercus infectoria on Bone Metabolism by Using hFOB 1.19 Human Fetal Osteoblast Cell Model

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2018/5319528 ◽

2018 ◽

Vol 2018 ◽

pp. 1-13 ◽

Cited By ~ 1

Author(s):

Amira Raudhah Abdullah ◽

Hermizi Hapidin ◽

Hasmah Abdullah

Keyword(s):

Gene Expression ◽

Bone Formation ◽

Column Chromatography ◽

Cell Model ◽

Osteoblast Cell ◽

Isolation And Purification ◽

Traditional Medicines ◽

Phytochemical Compounds ◽

Layer Chromatography ◽

Quercus Infectoria

Background. Quercus infectoria (QI) is a plant used in traditional medicines in Asia. The plant was reported to contain various active phytochemical compounds that have potential to stimulate bone formation. However, the precise mechanism of the stimulation effect of QI on osteoblast has not been elucidated. The present study was carried out to isolate QI semipurified fractions from aqueous QI extract and to delineate the molecular mechanism of QI semipurified fraction that enhanced bone formation by using hFOB1.19 human fetal osteoblast cell model. Methods. Isolation of QI semipurified fractions was established by means of column chromatography and thin layer chromatography. Established QI semipurified fractions were identified using Liquid Chromatography-Mass Spectrometry (LC-MS). Cells were treated with derived QI semipurified fractions and investigated for mineralization deposition and protein expression level of BMP-2, Runx2, and OPN by ELISA followed gene expression analysis of BMP-2 and Runx2 by RT-PCR. Results. Column chromatography isolation and purification yield Fractions A, B, and C. LC-MS analysis reveals the presence of polyphenols in each fraction. Results show that QI semipurified fractions increased the activity and upregulated the gene expression of BMP-2 and Runx2 at day 1, day 3, and day 7. OPN activity increased in cells treated with QI semipurified fractions at day 1 and day 3. Meanwhile, at day 7, expression of OPN decreased in activity. Furthermore, the study showed that combination of Fractions A, B, and C with osteoporotic drug (pamidronate) further increased the activity and upregulated the gene expression of BMP-2 and Runx2. Conclusions. These findings demonstrated that polyphenols from semipurified fractions of QI enhanced bone formation through expression of the investigated bone-related marker that is its potential role when combined with readily available osteoporotic drug.

Download Full-text