Journal of Prevention and Treatment for Stomatological Diseases ›› 2020, Vol. 28 ›› Issue (7): 421-426.doi: 10.12016/j.issn.2096-1456.2020.07.003

• Basic Study • Previous Articles     Next Articles

Effects of casein kinase 2 interacting protein-1 on the osteogenic differentiation ability of human periodontal ligament stem cells

QIN Qing1,SONG Yang2,LIU Jia3,LI Qiang4()   

  1. 1. Department of Dental, The Second Affiliated Hospital of Xi′an Jiaotong University (Xibei Hospital), Xi′an 710003, China
    2. Department of Stomatology, the 986 Hospital of PLA, Xi′an 710054, China
    3. State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shanxi Clinical Research Center for Oral Diseases, Department of Orthodontics, School of Stomatology, The Fourth Military Medical University, Xi′an 710032, China
    4. State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shanxi International Joint Research Center for Oral Diseases, Department of General Dentistry & Emergency, School of Stomatology, The Fourth Military Medical University, Xi′an 710032, China
  • Received:2019-11-04 Revised:2020-02-11 Online:2020-07-20 Published:2020-06-04
  • Contact: Qiang LI E-mail:lqaq726@163.com

Abstract:

Objective To investigate the effects of casein kinase 2 interacting protein-1 (CKIP-1) on the osteogenic differentiation ability of human periodontal ligament stem cells (hPDLSCs). Methods The hPDLSCs were obtained by primary culture with periodontal ligament tissues that were collected from normal humans. Then, a lentiviral vector containing a CKIP-1-specific siRNA sequence was constructed, and the transcriptional level of CKIP-1 in hPDLSCs was downregulated after vector infection. The P4 cells were divided into four groups: the control group, negative control group (infected with a control vector), CKIP-siRNA group (infected by a CKIP-1 siRNA lentivirus) and CKIP-1 group (infected by a CKIP-1 overexpression virus). All of the cells were cultured under osteogenic induction for 21 days. Then, alizarin red staining and quantitative determination were performed to detect the osteogenic differentiation ability of the hPDLSCs. In addition, qPCR was used to detect the transcriptional level of osteogenesis-related regulatory factors, such as Runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), osteocalcin (OCN), and receptor activator of nuclear factor kappa-B ligand (RANKL), and the osteogenesis-related regulatory factors of the bone morphogenetic protein (BMP) signaling pathway. Results There were no differences in the indexes between the negative control group and the control group (P > 0.05). Compared with the negative control group, the CKIP-siRNA group demonstrated more mineralized nodules (P < 0.05), significantly increased calcium salt deposition (P < 0.05), and increased mRNA levels of osteogenesis-related regulatory factors, such as Runx2 , ALP, OCN, and RANKL, and the osteogenesis-related regulatory factors of BMP signaling pathway (P < 0.05). Conclusion Downregulation of CKIP-1 could promote the osteogenic differentiation of hPDLSCs, which is related to the transcription level of osteogenic-related regulatory factors.

Key words: periodontitis, periodontal bone loss, casein kinase 2 interacting protein-1, human periodontal ligament stem cells, osteogenic differentiation, siRNA interference, Runt-related transcription factor 2, alkaline phosphatase, osteocalcin, receptor activator ofnuclear factor kappa-B ligand, bone morphogenetic protein signaling pathway

CLC Number: 

  • R781.4

Table 1

The gene name and primer sequence"

Gene name Primer sequence(5′-3′) bp
CKIP-1 F: 5′-AACCGCTATGTGGTGCTGAA-3′
R: 5′-CAGGGTGAACTTGCTGTGATTT-3′
118
Runx2 F: 5′-CCCGTGGCCTTCAAGGT-3′
R: 5′-CGTTACCCGCCATGACAGTA-3′
92
Coll I F: 5′-CCAGAAGAACTGGTACATCAGCAA-3′
R: 5′-CGCCATACTCGAACTGGAATC-3′
235
ALP F: 5′-AAGTACTGGACAGACCAAGC-3′
R: 5′-AGAGGGCCAAGAAGGGGAACT-3′
239
OCN F: 5′-ATGAGAGCCCTCAGACTCCTC-3′
R: 5′-CGGGCCGTAGAAGCGCCGATA-3′
212
RANKL F: 5′-CAGCTATGATGGAAGGTTCGTG-3′
R: 5′-AACCCGATGGGATGTTGG-3′
188
BMP2 F: 5′-CGGACTGCGGTCTCCTAA-3′
R: 5′-GGAAGCAGCAACGCTAGAAG-3′
217
Smurf1 F: 5′-GAAACCCAATGGCAGAAA-3′
R: 5′-GCAGATGTTGAGGGATGAG-3′
186
Smad4 F: 5′-GACTCACGGGCTTGGATTGA-3′
R: 5′-TCCCCAAAGCAGAAGCTACG-3′
153
β-actin F: 5′-TGGCACCCAGCACAATGAA-3′
R: 5′-CTAAGTCATAGTCCGCCTAGAAGCA-3′
186

Figure 1

Identification of hPDLSCs a, b: clone formation assay; c, alizarin red staining; d, cell surface marker CD105 expression"

Figure 2

mRNA expression of CKIP-1 *: vs. shNC group, P < 0.05"

Figure 3

Alizarin red staining and quantitative detection of the osteogenic differentiation ability of hPDLSCs a, control group; b, negative control group; c, CKIP-1 siRNA group; d, CKIP-1 group; e, quantitative determination; *: vs. shNC group, P < 0.05"

Figure 4

mRNA expresssion level of osteogenic regulatory factors *: vs. shNC group, P < 0.05; Runx2: Runt-related transcription factor 2; ALP: alkaline phosphatase; OCN: osteocalcin; RANKL: receptor activator ofnuclear factor kappa-B ligand; Coll I: type I collagen"

Figure 5

mRNA expression level of BMP signal pathway related factors *: vs. shNC group, P < 0.05; BMP2: bone morphogenetic protein 2; Smurf1: Smad ubiquitination regulatory factor 1; Smad4: mothers against decapentaplegic homolog 4"

[1] Guo Z, Chen R, Zhang F , et al. Exendin-4 relieves the inhibitory effects of high glucose on the proliferation and osteoblastic differentiation of periodontal ligament stem cells[J]. Arch Oral Biol, 2018,91:9-16.
doi: 10.1016/j.archoralbio.2018.03.014
[2] 李五一, 谢昊, 刘建国 , 等. 人牙周膜干细胞的分离培养及初步鉴定[J]. 广东牙病防治, 2015,23(3):117-121.
Li WY, Xie H, Liu JG , et al. Isolation culture and identification of human periodontal ligament stem cells[J]. J Prev Treat Stomatol Dis, 2015,23(3):117-121.
[3] Jin L, Lu C, Wu X , et al. Targeting osteoblastic casein kinase-2 interacting protein-1 to enhance Smad-dependent BMP signaling and reverse bone formation reduction in glucocorticoid-induced osteoporosis[J]. Sci Rep, 2017,7:41295.
doi: 10.1038/srep41295
[4] 王也, 林晓萍 . 牙周炎和骨质疏松症共同危险因素及相关机制研究进展[J]. 口腔疾病防治, 2019,27(12):794-798.
Wang Y, Lin XP . Research Progress on common risk factors and related mechanisms of periodontitis and osteoporosis[J]. J Prev Treat Stomatol Dis, 2019,27(12):794-798.
[5] 廖春晖, 李明飞, 叶金梅 , 等. IGF1通过BMP2-Smad1/5信号通路调控犬上颌窦黏膜干细胞成骨分化[J]. 口腔疾病防治, 2020,28(1):16-23.
Liao CH, Li MF, Ye JM , et al. IGF1 regulates osteogenic differentiation of canine maxillary sinus mucosa stem cells through bmp2-smad1/5 signal pathway[J]. J Prev Treat Stomatol Dis, 2020,28(1):16-23.
[6] 李夏宁, 赵红宇, 赵华 . 牙周膜干细胞成脂分化的研究进展[J]. 口腔疾病防治, 2016,24(5):317-320.
Li XN, Zhao HY, Zhao H . The research progress of adipogenic differentiation of periodontal ligament stem cells[J]. J Prev Treat Stomatol Dis, 2016,24(5):317-320.
[7] Wang H, Li J, Zhang X , et al. Priming integrin alpha 5 promotes the osteogenic differentiation of human periodontal ligament stem cells due to cytoskeleton and cell cycle changes[J]. J Proteomics, 2018,179:122-130.
doi: 10.1016/j.jprot.2018.03.008
[8] Han N, Zhang F, Li G , et al. Local application of IGFBP5 protein enhanced periodontal tissue regeneration via increasing the migration, cell proliferation and osteo/dentinogenic differentiation of mesenchymal stem cells in an inflammatory niche[J]. Stem Cell Res Ther, 2017,8(1):210.
doi: 10.1186/s13287-017-0663-6
[9] Diomede F, D′Aurora M, Gugliandolo A , et al. Biofunctionalized scaffold in bone tissue repair[J]. Int J Mol Sci, 2018,19(4):1022.
doi: 10.3390/ijms19041022
[10] Tokuda E, Fujita N, Ohhara T , et al. Casein kinase 2-interacting protein-1, a novel Akt pleckstrin homology domain-interacting protein, down-regulates PI3K/Akt signaling and suppresses tumor growth in vivo[J]. Cancer Res, 2007,67(20):9666-9676.
doi: 10.1158/0008-5472.CAN-07-1050
[11] Xi S, Tie Y, Lu K , et al. N-terminal PH domain and C-terminal auto-inhibitory region of CKIP-1 coordinate to determine its nucleus-plasma membrane shuttling[J]. FEBS Lett, 2010,584(6):1223-1230.
doi: 10.1016/j.febslet.2010.02.036
[12] Chen G M, Ding R F, Tan Y D , et al. Role of the CKIP1 gene in proliferation and apoptosis of the human lung cancer cell line H1299[J]. Genet Mol Res, 2015,14(2):4005-4014.
doi: 10.4238/2015.April.27.15
[13] Guo B, Zhang B, Zheng L , et al. Therapeutic RNA interference targeting CKIP-1 with a cross-species sequence to stimulate bone formation[J]. Bone, 2014,59(1):76-88.
doi: 10.1016/j.bone.2013.11.007
[14] Zhang X, Wang Q, Wan Z , et al. CKIP-1 knockout offsets osteoporosis induced by simulated microgravity[J]. Prog Biophys Mol Biol, 2016,122(2):140-148.
doi: 10.1016/j.pbiomolbio.2016.09.007
[15] Jin L, Chao L, Guo B , et al. Increased PLEKHO1 within osteoblasts suppresses Smad-dependent BMP signaling to inhibit bone formation during aging[J]. Aging Cell, 2017,16(2):360-376.
doi: 10.1111/acel.12566
[16] Lu K, Yin X, Weng T , et al. Targeting WW domains linker of HECT-type ubiquitin ligase Smurf1 for activation by CKIP-1[J]. Nat Cell Biol, 2008,10(8):994-1002.
doi: 10.1038/ncb1760
[17] Zhang L, Wu K, Song W , et al. Chitosan/siCKIP-1 biofunctionalized titanium implant for improved osseointegration in the osteoporotic condition[J]. Sci Rep, 2015,5:10860.
doi: 10.1038/srep10860
[18] Liu Q, Guo Y, Wang Y , et al. miR985p promotes osteoblast differentiation in MC3T3E1 cells by targeting CKIP1[J]. Mol Med Rep, 2018,17(3):4797-4802.
[1] HE Mengjiao,LI Lisheng,CHEN Yuling,LUO Kai. Research progress on cell sheet technology and its application in periodontal tissue regeneration [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(7): 458-462.
[2] WU Hongyu,MA Xiaoxin,LU Haixia,FENG Xiping,GU Qin,YE Wei,XIE Yingxin,XIE Danshu,WANG Wenji. Investigation of dental caries and periodontal conditions in maintenance hemodialysis patients [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(5): 313-317.
[3] PENG Haiyan,JIANG Xiaowen,HUANG Huaqing,CHEN Jinyong. The role of the mTORC1 signaling pathway during osteogenic differentiation of mouse bone marrow mesenchymal cells under tension stress [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(4): 219-223.
[4] WANG Zhiheng,ZUO Jie,WANG Mengqi,ZHU Shaojun,LIU Yishan. miR-214 inhibits the osteogenic differentiation of dental follicle cells in vitro [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(3): 146-152.
[5] ZHANG Limu,LIN Xiaoping. Research progress on the mechanism of C-reactive protein mediated periodontitis and systemic diseases [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(3): 184-188.
[6] LIAN Keqian,ZHANG Xin,ZHOU Jieyu,LIAO Yanfen,SI Shanshan. Biocompatibility of bone marrow mesenchymal cells on polyetheretherketone and titanium surfaces in vitro [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(2): 73-78.
[7] LIAO Chunhui,LI Mingfei,YE Jinmei,PENG Wei,CHEN Songling. The regulatory mechanisms of IGF1 in the osteogenic differentiation of canine MSMSCs via BMP2-Smad1/5 signaling pathway [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(1): 16-23.
[8] ZHOU Jiaqi,SHU Linjing,XIONG Yi,ZHANG Yixin,XIANG Lin,WU Yingying. Study on the role of FoxO1 in the regulation of osteoblastic metabolism by 1,25(OH)2D3 in a high glucose environment [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(1): 24-29.
[9] LI Jiatong,ZHOU Xuedong,XU Xin,WANG Yan. Research progress of probiotics in the prevention and treatment of oral infectious diseases [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(9): 598-601.
[10] REN Qingyuan,HE Wulin,WANG Qing,CHU Hongxing,LIN Haiyan. Effect of endoplasmic reticulum stress on the osteogenic differentiation of periodontal ligament cells under continuous static pressure [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(8): 485-489.
[11] Gang WANG,Peng SUN,Juan LI,Shiguang HUANG. Expression of the receptor of advanced glycation end products in gingival tissue endothelial cells from type 2 diabetic rats with chronic periodontitis [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(7): 428-434.
[12] DAI Qian,YU Yuanling,HUANG Shiguang. Macrophage IL-33 expression in macrophages of human chronic periapical periodontitis and apical cyst [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(5): 300-303.
[13] XU Xiongcheng,ZHONG Quan,LUO Kai. Application of 3D printing technology in preparation of scaffolds for periodontal tissue regeneration [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(3): 189-193.
[14] ZHU Yanxia,Gegen tana. Research progress of IL-33/ST2 signaling pathway in bone metabolism [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(3): 202-204.
[15] WANG Ye,LIN Xiaoping. Research progress on the common risk factors and related mechanisms of periodontitis and osteoporosis [J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(12): 794-798.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
[1] WU Hongyu,MA Xiaoxin,LU Haixia,FENG Xiping,GU Qin,YE Wei,XIE Yingxin,XIE Danshu,WANG Wenji. Investigation of dental caries and periodontal conditions in maintenance hemodialysis patients[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(5): 313 -317 .
[2] ZHOU Tao,WU Peiyao,YANG Yuqing,CAO Zhiwei,XIE Liang. Research progress on the distribution of primary cilia and related signaling pathways involved in odontogenesis[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(5): 318 -321 .
[3] WANG Shipeng,ZHANG Andong,CHENG Li,CHEN Baoyong,WEN Li,LIU Huawei. Basal cell nevus syndrome: 4 case reports and a literature review[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(1): 41 -45 .
[4] ZOU Xiaolong,CHEN Yuan,WANG Yan,WANG Jiantao. Research progress on animal models of oral mucositis caused by radiotherapy and chemotherapy[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2020, 28(5): 322 -326 .
[5] CUI Ye,HUANG Ziru,WANG Chunlin,LIU Conghua,ZHANG Chao. Effect of three kinds of medium molecular weight proteins on the corrosion resistance of Ni-Ti and stainless steel arch wires[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(2): 83 -89 .
[6] ZHANG Ning,HU Yue,QIAO Chunyan,JI Xin,HAN Ruyu,SUN Lanfang,LI Minghe,HAN Chengmin. Mucoepidermoid carcinoma arising in Warthin’s tumor of the upper lip: a case report and review[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(4): 250 -254 .
[7] LIU Qian,LAN Lufang,YAN Junyi,TIAN Weidong,GUO Shujuan. Research on the surface structure of a dentin matrix with complete demineralization and incomplete demineralization and the osteogenic property promotion of human periodontal ligament cells[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(3): 159 -166 .
[8] LIN Lin,DUAN Ning,WANG Xiang,JIANG Hongliu,WANG Wenmei. Oral verrucous xanthoma in adolescents: a case report and literature review[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(5): 318 -320 .
[9] Shuyu CAI,Xiaoyue LIN,Jin LEI,Song GE. Effects of Porphyromonas gingivalis infection with different fimA genotypes on the secretion of IL-1β, IL-6, and TNF-α by human umbilical vein endothelial cells[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(6): 364 -369 .
[10] LU Weiying,LIU Ping,CHEN Jiawei,LIU Shuying,XU Pingping. Effects of X-ray irradiation on proliferation and RANTES expression of the mouse osteogenic precursor cell line MC3T3-E1[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2019, 27(10): 621 -626 .
This work is licensed under a Creative Commons Attribution 3.0 License.