Bone morphogenetic protein&nbsp;2‑enhanced osteogenic differentiation of stem cell spheres by regulation of Runx2 expression

Min,Sae Kyung; Kim,Minji; Park,Jun-Beom

doi:10.3892/etm.2020.9206

Bone morphogenetic protein 2‑enhanced osteogenic differentiation of stem cell spheres by regulation of Runx2 expression

Authors:
- Sae Kyung Min
- Minji Kim
- Jun-Beom Park
View Affiliations

Affiliations: Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul 06591, Republic of Korea, College of Dentistry, Chosun University, Gwangju 61452, Republic of Korea
Published online on: September 11, 2020 https://doi.org/10.3892/etm.2020.9206
Article Number: 79
Copyright: © Min et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Bone morphogenetic protein 2 (BMP‑2) is a growth factor that is used to induce osteogenic differentiation in stem cells. The present study assessed the effects of BMP‑2 on stem cell spheroid morphology, viability and osteogenic differentiation. Stem cell spheres were constructed and treated with BMP‑2 at predetermined concentrations (0‑100 ng/ml) using concave microwells. Cell viability was qualitatively and quantitatively analyzed via microscopy and a water‑soluble tetrazolium salt assay kit, respectively. Alkaline phosphatase activity was assessed and an anthraquinone dye for calcium deposit evaluation was performed to determine osteogenic differentiation. The expressions of (runt‑related transcription factor 2) and collagen 1 were also determined via quantitative PCR. Spherical shapes were formed using concave microwells on day 1, which were maintained up to day 21. On day 1, the relative cell viability of 0, 10 and 100 ng/ml BMP‑2 treated cells was 100.0±1.9, 97.3±4.4 and 101.3±2.6%, respectively. Significantly higher values for alkaline phosphatase activity were determined in the 100 ng/ml treated group when compared with the control group. Additionally, Runx2 mRNA levels were significantly higher in the 100 ng/ml BMP‑2 group compared with the control group, as determined via quantitative PCR. The results of the present study indicated that BMP‑2 enhanced the differentiation of stem cell spheres, which was demonstrated by increased alkaline phosphatase activity and Runx2 expression.

View Figures

View References

1	Liu H, Wei LK, Jian XF, Huang J, Zou H, Zhang SZ and Yuan GH: Isolation, culture and induced differentiation of rabbit mesenchymal stem cells into osteoblasts. Exp Ther Med. 15:3715–3724. 2018.PubMed/NCBI View Article : Google Scholar
2	Fan J, Im CS, Cui ZK, Guo M, Bezouglaia O, Fartash A, Lee JY, Nguyen J, Wu BM, Aghaloo T and Lee M: Delivery of phenamil enhances BMP-2-induced osteogenic differentiation of adipose-derived stem cells and bone formation in calvarial defects. Tissue Eng Part A. 21:2053–2065. 2015.PubMed/NCBI View Article : Google Scholar
3	Lee YH, Lee BW, Jung YC, Yoon BI, Woo HM and Kang BJ: Application of alginate microbeads as a carrier of bone morphogenetic protein-2 for bone regeneration. J Biomed Mater Res B Appl Biomater. 107:286–294. 2019.PubMed/NCBI View Article : Google Scholar
4	Wongwitwichot P and Kaewsrichan J: Osteogenic differentiation of mesenchymal stem cells is impaired by bone morphogenetic protein 7. Adv Med Sci. 62:266–272. 2017.PubMed/NCBI View Article : Google Scholar
5	Qian C, Zhu C, Yu W, Jiang X, Zhang F and Sun J: Bone morphogenetic protein 2 promotes osteogenesis of bone marrow stromal cells in type 2 diabetic rats via the Wnt signaling pathway. Int J Biochem Cell Biol. 80:143–153. 2016.PubMed/NCBI View Article : Google Scholar
6	Qiao Y, Xu Z, Yu Y, Hou S, Geng J, Xiao T, Liang Y, Dong Q, Mei Y, Wang B, et al: Single cell derived spheres of umbilical cord mesenchymal stem cells enhance cell stemness properties, survival ability and therapeutic potential on liver failure. Biomaterials. 227(119573)2020.PubMed/NCBI View Article : Google Scholar
7	Rosenzweig M, Pykett M, Marks DF and Johnson RP: Enhanced maintenance and retroviral transduction of primitive hematopoietic progenitor cells using a novel three-dimensional culture system. Gene Ther. 4:928–936. 1997.PubMed/NCBI View Article : Google Scholar
8	Zhang W, Zhuang A, Gu P, Zhou H and Fan X: A review of the three-dimensional cell culture technique: Approaches, advantages and applications. Curr Stem Cell Res Ther. 11:370–380. 2016.PubMed/NCBI View Article : Google Scholar
9	Brboric A, Vasylovska S, Saarimäki-Vire J, Espes D, Caballero-Corbalan J, Larfors G, Otonkoski T and Lau J: Characterization of neural crest-derived stem cells isolated from human bone marrow for improvement of transplanted islet function. Ups J Med Sci. 124:228–237. 2019.PubMed/NCBI View Article : Google Scholar
10	Kabiri M, Kul B, Lott WB, Futrega K, Ghanavi P, Upton Z and Doran MR: 3D mesenchymal stem/stromal cell osteogenesis and autocrine signalling. Biochem Biophys Res Commun. 419:142–147. 2012.PubMed/NCBI View Article : Google Scholar
11	Naito H, Yoshimura M, Mizuno T, Takasawa S, Tojo T and Taniguchi S: The advantages of three-dimensional culture in a collagen hydrogel for stem cell differentiation. J Biomed Mater Res A. 101:2838–2845. 2013.PubMed/NCBI View Article : Google Scholar
12	Jeong CH, Kim SM, Lim JY, Ryu CH, Jun JA and Jeun SS: Mesenchymal stem cells expressing brain-derived neurotrophic factor enhance endogenous neurogenesis in an ischemic stroke model. Biomed Res Int. 2014(129145)2014.PubMed/NCBI View Article : Google Scholar
13	Lee H and Park JB: Dimethyl sulfoxide leads to decreased osteogenic differentiation of stem cells derived from gingiva via Runx2 and collagen I expression. Eur J Dent. 13:131–136. 2019.PubMed/NCBI View Article : Google Scholar
14	Lee JM, Kim MG, Byun JH, Kim GC, Ro JH, Hwang DS, Choi BB, Park GC and Kim UK: The effect of biomechanical stimulation on osteoblast differentiation of human jaw periosteum-derived stem cells. Maxillofac Plast Reconstr Surg. 39(7)2017.PubMed/NCBI View Article : Google Scholar
15	Yazid MD, Ariffin SHZ, Senafi S, Razak MA and Wahab RMA: Determination of the differentiation capacities of murines' primary mononucleated cells and MC3T3-E1 cells. Cancer Cell Int. 10(42)2010.PubMed/NCBI View Article : Google Scholar
16	Futrega K, Mosaad E, Chambers K, Lott WB, Clements J and Doran MR: Bone marrow-derived stem/stromal cells (BMSC) 3D microtissues cultured in BMP-2 supplemented osteogenic induction medium are prone to adipogenesis. Cell Tissue Res. 374:541–553. 2018.PubMed/NCBI View Article : Google Scholar
17	Hu S, Chen H, Zhou X, Chen G, Hu K, Cheng Y, Wang L and Zhang F: Thermally induced self-agglomeration 3D scaffolds with BMP-2-loaded core-shell fibers for enhanced osteogenic differentiation of rat adipose-derived stem cells. Int J Nanomedicine. 13:4145–4155. 2018.PubMed/NCBI View Article : Google Scholar
18	Dilogo IH, Phedy P, Kholinne E, Djaja YP, Fiolin J, Kusnadi Y and Yulisa ND: Autologous mesenchymal stem cell implantation, hydroxyapatite, bone morphogenetic protein-2, and internal fixation for treating critical-sized defects: A translational study. Int Orthop. 43:1509–1519. 2019.PubMed/NCBI View Article : Google Scholar
19	Zhang Y, Chen H, Zhang T, Zan Y, Ni T, Cao Y, Wang J, Liu M and Pei R: Injectable hydrogels from enzyme-catalyzed crosslinking as BMSCs-laden scaffold for bone repair and regeneration. Mater Sci Eng C Mater Biol Appl. 96:841–849. 2019.PubMed/NCBI View Article : Google Scholar
20	Paidikondala M, Kadekar S and Varghese OP: Innovative strategy for 3D transfection of primary human stem cells with BMP-2 expressing plasmid DNA: A clinically translatable strategy for ex vivo gene therapy. Int J Mol Sci. 20(56)2018.PubMed/NCBI View Article : Google Scholar
21	Kato S, Kawabata N, Suzuki N, Ohmura M and Takagi M: Bone morphogenetic protein-2 induces the differentiation of a mesenchymal progenitor cell line, ROB-C26, into mature osteoblasts and adipocytes. Life Sci. 84:302–310. 2009.PubMed/NCBI View Article : Google Scholar
22	Kim SY, Kim YK, Kim KS, Lee KB and Lee MH: Enhancement of bone formation on LBL-coated Mg alloy depending on the different concentration of BMP-2. Colloids Surf B Biointerfaces. 173:437–446. 2019.PubMed/NCBI View Article : Google Scholar
23	Park JB: Combination of simvastatin and bone morphogenetic protein-2 enhances the differentiation of osteoblasts by regulating the expression of phospho-Smad1/5/8. Exp Ther Med. 4:303–306. 2012.PubMed/NCBI View Article : Google Scholar
24	Lysdahl H, Baatrup A, Foldager CB and Bünger C: Preconditioning human mesenchymal stem cells with a low concentration of BMP2 stimulates proliferation and osteogenic differentiation in vitro. Biores Open Access. 3:278–285. 2014.PubMed/NCBI View Article : Google Scholar
25	Yarygin NV, Parshikov MV, Prosvirin AA, Gur'ev VV, Govorov MV, Bosykh VG, Akatov VS and Chekanov AV: Effect of morphogenetic protein BMP-2 on X-ray density of bone defect in the experiment. Bull Exp Biol Med. 168:574–577. 2020.PubMed/NCBI View Article : Google Scholar
26	Lee MH, Kwon TG, Park HS, Wozney JM and Ryoo HM: BMP-2-induced Osterix expression is mediated by Dlx5 but is independent of Runx2. Biochem Biophys Res Commun. 309:689–694. 2003.PubMed/NCBI View Article : Google Scholar
27	Rickard DJ, Sullivan TA, Shenker BJ, Leboy PS and Kazhdan I: Induction of rapid osteoblast differentiation in rat bone marrow stromal cell cultures by dexamethasone and BMP-2. Dev Biol. 161:218–228. 1994.PubMed/NCBI View Article : Google Scholar
28	Lee H, Son J, Yi G, Koo H and Park JB: Cellular viability and osteogenic differentiation potential of human gingiva-derived stem cells in 2D culture following treatment with anionic, cationic, and neutral liposomes containing doxorubicin. Exp Ther Med. 16:4457–4462. 2018.PubMed/NCBI View Article : Google Scholar
29	Yasui Y, Ando W, Shimomura K, Koizumi K, Ryota C, Hamamoto S, Kobayashi M, Yoshikawa H and Nakamura N: Scaffold-free, stem cell-based cartilage repair. J Clin Orthop Trauma. 7:157–163. 2016.PubMed/NCBI View Article : Google Scholar
30	Ong CS, Yesantharao P, Huang CY, Mattson G, Boktor J, Fukunishi T, Zhang H and Hibino N: 3D bioprinting using stem cells. Pediatr Res. 83:223–231. 2018.PubMed/NCBI View Article : Google Scholar
31	Takeoka Y, Matsumoto K, Taniguchi D, Tsuchiya T, Machino R, Moriyama M, Oyama S, Tetsuo T, Taura Y, Takagi K, et al: Regeneration of esophagus using a scaffold-free biomimetic structure created with bio-three-dimensional printing. PLoS One. 14(e0211339)2019.PubMed/NCBI View Article : Google Scholar
32	Kang W, Liang Q, Du L, Shang L, Wang T and Ge S: Sequential application of bFGF and BMP-2 facilitates osteogenic differentiation of human periodontal ligament stem cells. J Periodontal Res. 54:424–434. 2019.PubMed/NCBI View Article : Google Scholar
33	Cruz ACC, Cardozo FTGS, Magini RS and Simões CMO: Retinoic acid increases the effect of bone morphogenetic protein type 2 on osteogenic differentiation of human adipose-derived stem cells. J Appl Oral Sci. 27(e20180317)2019.PubMed/NCBI View Article : Google Scholar
34	Zhang W, Zhang X, Li J, Zheng J, Hu X, Xu M, Mao X and Ling J: Foxc2 and BMP2 induce osteogenic/odontogenic differentiation and mineralization of human stem cells from apical papilla. Stem Cells Int. 2018(2363917)2018.PubMed/NCBI View Article : Google Scholar
35	Han K, Ko Y, Park YG and Park JB: Associations between the periodontal disease in women before menopause and menstrual cycle irregularity: The 2010-2012 Korea national health and nutrition examination survey. Medicine (Baltimore). 95(e2791)2016.PubMed/NCBI View Article : Google Scholar