Abstract
Summary Statement
The stable inhibition of miR-214 in the aged osteoporotic rats induced by OVX could be achieved by periodic administration of AntagomiR-214 at a dosage of 4 mg/kg and at an interval of 7 days, which will provide a potential bone anabolic strategy for treatment of osteoprosis.
Introduction
MiR-214 has a crucial role in suppressing bone formation and miR-214 inhibition in osteogenic cells may be a potential anabolic strategy for ameliorating osteoporosis (Wang X, et al. 2013). An aged ovariectomised rat has been regarded as a golden model to test bone anabolic agents for reversing established osteoporosis in aged postmenopausal women (Li X, et al. 2009). However, there is still lack of evidence to demonstrate bone anabolic potential of therapeutic inhibition of miR-214 within osteogenic cells in the golden model. So, it should be necessary to establish RNAi-based administration protocol toward stable inhibition of miR-214 at a low level in the golden model. A targeted delivery system specifically facilitating Antagomir-214 approaching osteogenic cells, i.e. (Asp-Ser-Ser)6-liposome (Zhang G, et al 2012), was employed in this study.
Objectives
This study was to investigate optimal dosage and duration for therapeutic inhibition of miR-214 within osteogenic cells in the aged osteoporotic rats induced by ovariectomy.
Materials and Methods
Six-month-old female Sprague-Dawley rats were ovariectomised (OVX) and left untreated for 12 months to establish aged osteoporosis. To determine the optimal dosage for therapeutic inhibition of miR-214, the OVX rats were injected intravenously with the AntagomiR-214 at a dosage of 0.5mg/kg, 1mg/kg, 2mg/kg, 4mg/kg, 6mg/kg and 8mg/kg (n=6 for each dosage group) delivered by (Asp-Ser-Ser)6-liposome, respectively. Thereafter, miR-214 expression level in osteogenic cells from bilateral femur was quantified at day 2 post injection by real-time PCR analysis in combination with laser captured dissection (LCM). To determine the optimal duration of miR-214, the OVX rats were intravenously injected with the AntagomiR-214 (AntagomiR-214 group) or non-sense AntagomiR-214 (NC group) delivered by (Asp-Ser-Ser)6-liposome at the optimal dosage or (Asp-Ser-Ser)6-liposome alone (Vehicle group). Then, the miR-214 level in osteogenic cells from bilateral femur was quantified at 1, 3, 5, 7, 9, 12, 14, 16, 21 day after the single dosing (n=6 for each time-point) by real-time PCR analysis in combination with LCM, respectively. To examine the long-term effect of the AntagomiR-214 after periodic pulsed dosing, the OVX rats were administrated with the AntagomiR-214 at the optimal dosage and duration for 5 repeated injections and then the miR-214 level in osteogenic cells from bilateral femur was quantified by real-time PCR analysis in combination with LCM.
Results
The miR-214 level was efficiently decreased in a dose-dependent manner by the AntogomiR-214 and reached the level lower than 10% of the baseline at a dosage of 4 mg/kg at least in the aged osteoporotic rats. The effective duration for miR-214 at a level lower than 50% of the baseline lasted for 7 days in the osteoporotic rats after the single dosing. The miR-214 level was continuously lowered until 28 days and continuously maintained later at the level lower than 10% of the baseline by the 5 pulsed dosing of the AntagomiR-214 at an interval of 7 days and at a dosage of 4 mg/kg in the osteoporotic rats.
Conclusions
The stable inhibition of miR-214 for bone anabolic strategy in the aged osteoporotic rats induced by OVX could be achieved by periodic administration of AntagomiR-214 at a dosage of 4 mg/kg and at an interval of 7 days.
