Differentiation and proliferation activities of human foetal osteoblast cell line (hFOB 1.19) treated with a polyphenol, tannic acid alone or in combination with pamidronate
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Date
2018-12
Authors
Hashim, Nor Munira
Journal Title
Journal ISSN
Volume Title
Publisher
Pusat Pengajian Sains Perubatan, Universiti Sains Malaysia
Abstract
Bone continuously undergoes bone remodelling and imbalance in bone remodelling leads
to bone pathogenesis, such as osteoporosis. Tannic acid (TA) is a polyphenol with
antioxidant properties which may elevate osteoblast metabolism. Synergism is one of
pharmacological effect that occur when drug combination is used. Hence, the aim of this
study was to evaluate the effect of TA alone and combination of TA and pamidronate on
differentiation, proliferation, mineralisation as well as morphology of human foetal
osteoblast cells (hFOB 1.19). The hFOB 1.19 cells were treated with TA alone and
combination of TA and pamidronate of different combination ratios at different
concentrations (0.1 to 99.0 μg/ml). Half maximal effective concentration (EC50) for TA
alone, pamidronate alone and different combination ratios of TA and pamidronate (25:75,
50:50, 75:25) were measured by using MTT assay. Analysis of synergistic effects was
also performed on the different combination ratios based on Combination Index (CI)
where CI < 1: synergism, CI = 1: additive effect and CI > 1: antagonism. The combination
treatment with the lowest EC50 and CI<1 was then chosen for the next studies. Cell
proliferation assay was conducted to compare the cell viability and morphology of hFOB
1.19 cells was observed by using inverted microscope after treatment from day 1 to day
7. In addition, detection of calcium and phosphate deposits were done by histochemical
staining. Detection of bone specific genes [bone sialoprotein (BSP) and osterix (Osx)]
was conducted by polymerase chain reaction (PCR) test. The EC50 of hFOB 1.19 cellstreated with TA alone (0.56 ± 0.22 μg/ml) and 75:25 ratio of TA and pamidronate (0.48
± 0.03 μg/ml) were more effective compared to pamidronate alone (15.27 ± 0.22 μg/ml)
and other combination ratios treatments [(25:75; 3.80 ± 0.13 μg/ml); (50:50; 2.25 ± 0.11
μg/ml)]. Analysis of synergistic result showed that only 75:25 combination ratio exhibited
synergistic effect while the other two combination ratios manifested antagonistic effects.
All groups showed significant different when compared between day 1 and day 7.
However, all groups except combination group manifested significant different when
comparing between day 3 and 7. When comparing on similar day which was day 7, both
TA alone and pamidronate alone treated groups showed significantly increased in cell
proliferation compared to control (untreated) and combination groups. The morphology
of hFOB 1.19 cells treated with both TA and combination groups had uniform and
elongated shape until the end of study. Treatment with TA alone demonstrated higher
production of calcium and phosphate deposits compared to untreated control,
pamidronate alone and combination groups. All groups manifested the expression of BSP
and Osx until day 7 of treatment periods. In conclusion, TA alone rather than combination
treatment of TA and pamidronate has greater ability in enhancing cell proliferation and
morphology besides improved osteoblast mineralisation.
Description
Keywords
Tannins