Effect of zoledronate on protein interaction between Ca2+/calmodulin-dependent protein kinaseⅡ and calmodulin and expression of downstream genes during osteoclast differentiation
10.3760/cma.j.issn.1002-0098.2017.02.014
- VernacularTitle: 唑来膦酸对破骨细胞分化中Ca2+/钙调蛋白依赖性蛋白激酶Ⅱ与钙调蛋白结合及下游基因表达的影响
- Author:
Hui WANG
1
;
Wei DONG
1
;
Mengchun QI
1
;
Hong SUN
2
;
Xiaojie FENG
1
;
Liming WEN
1
Author Information
1. Department of Oral and Maxillofacial Surgery, College of Stomatology, North China University of Science and Technology, Tangshan Hebei 063000, China
2. Department of Basic Medical Sciences, Department of Pathology, North China University of Science and Technology, Tangshan Hebei 063000, China
- Publication Type:Journal Article
- Keywords:
Calcium-calmodulin-dependent protein kinase type 2;
Calmodulin;
Zolderonate;
Immunoprecipitation
- From:
Chinese Journal of Stomatology
2017;52(2):120-125
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effect of zoledronate on protein interaction between Ca2+/calmodulin-dependent protein kinaseⅡ(CaMKⅡ) and calmodulin and protein expression of nuclear factor of activation of T cells-1 (NFATc1) and tartrate resistant acid phosphatase (TRAP) during osteoclast differentiation.
Methods:Mouse RAW264.7 cells were divided into group A and B and were cultured. Group A was induced with 50 mg/L receptor activator of NF-κB ligand (RANKL) for osteoclastogenesis, and group B was treated with 1×10-6 zoledronate for two days from day 2. Co-immunoprcipitation (Co-IP) and reverse Co-IP were used to detect the protein-binding between CaMKⅡ and calmodulin. Western-blotting and immunofluorescent cytochemistry were also used to detect the protein level of NFATc1 and TRAP in both groups. Osteoclast formation was also analyzed.
Results:In group B, the number of osteoclasts, number and size of dentin resorption lacunaes were 11.3±1.5, 8.7±2.1 and (5 034.4±775.4) μm2 respevtively, which were significantly lower than those (37.7±5.7, 23.0±4.0 and [15 042.7±1 906.0] μm2) in group A (P<0.01). Co-IP and reverse Co-IP examination indicated that protein-binding between CaMKⅡ and calmodulin significantly decreased by 59.8% and 50.9% in group B compared with group A (P<0.01). The protein level of calmodulin and CaMKⅡ in total cellular proteins also significantly decreased by 52.1% and 51.5% in group B compared with group A (P<0.01). NFATc1 and TRAP protein decreased by 52.4% and 38.9% in group B than in group A (P<0.01), respectively.
Conclusions:Zoledronate could significantly inhibit protein-binding between CaMKⅡ and calmodulin and down-regulate protein level of NFATc1 and TRAP.
