Time-sequential changes of differentially expressed miRNAs during the process of anterior lumbar interbody fusion using equine bone protein extract, rhBMP-2 and autograft

Abstract

The precise mechanism of bone regeneration in different bone graft substitutes has been well studied in recent researches. However, miRNAs regulation of the bone formation has been always mysterious. We developed the anterior lumbar interbody fusion (ALIF) model in pigs using equine bone protein extract (BPE), recombinant human bone morphogenetic protein-2 (rhBMP-2) on an absorbable collagen sponge (ACS), and autograft as bone graft substitute, respectively. The miRNA and gene expression profiles of different bone graft materials were examined using microarray technology and data analysis, including self-organizing maps, KEGG pathway and Biological process GO analyses. We then jointly analyzed miRNA and mRNA profiles of the bone fusion tissue at different time points respectively. Results showed that miRNAs, including let-7, miR-129, miR-21, miR-133, miR-140, miR-146, miR-184, and miR-224, were involved in the regulation of the immune and inflammation response, which provided suitable inflammatory microenvironment for bone formation. At late stage, several miRNAs directly regulate SMAD4, Estrogen receptor 1 and 5-hydroxytryptamine (serotonin) receptor 2C for bone formation. It can be concluded that miRNAs play important roles in balancing the inflammation and bone formation.

Graphical abstract

Keywords

miRNA / bone protein extract (BPE) / recombinant human bone morphogenetic protein-2 (rhBMP-2) / autograft

References

[1] Hobert O. Gene regulation by transcription factors and microRNAs. Science, 2008, 319(5871): 1785–1786

[2] Lian J B, Stein G S, van Wijnen A J, . MicroRNA control of bone formation and homeostasis. Nature Reviews Endocrinology, 2012, 8(4): 212–227

[3] Palmieri A, Pezzetti F, Brunelli G, . Anorganic bovine bone (Bio-Oss) regulates miRNA of osteoblast-like cells. The International Journal of Periodontics & Restorative Dentistry, 2010, 30(1): 83–87

[4] Palmieri A, Pezzetti F, Avantaggiato A, . Titanium acts on osteoblast translational process. Journal of Oral Implantology, 2008, 34(4): 190–195

[5] Palmieri A, Brunelli G, Guerzoni L, . Comparison between titanium and anatase miRNAs regulation. Nanomedicine, 2007, 3(2): 138–143

[6] Palmieri A, Pezzetti F, Brunelli G, . Anatase nanosurface regulates microRNAs. The Journal of Craniofacial Surgery, 2008, 19(2): 328–333

[7] Palmieri A, Pezzetti F, Brunelli G, . Short-period effects of zirconia and titanium on osteoblast microRNAs. Clinical Implant Dentistry and Related Research, 2008, 10(3): 200–205

[8] Palmieri A, Pezzetti F, Spinelli G, . PerioGlas regulates osteoblast RNA interfering. Journal of Prosthodontics, 2008, 17(7): 522–526

[9] Palmieri A, Pezzetti F, Brunelli G, . Differences in osteoblast miRNA induced by cell binding domain of collagen and silicate-based synthetic bone. Journal of Biomedical Science, 2007, 14(6): 777–782

[10] Palmieri A, Pezzetti F, Brunelli G, . Calcium sulfate acts on the miRNA of MG63E osteoblast-like cells. Journal of Biomedical Materials Research Part B: Applied Biomaterials, 2008, 84B(2): 369–374

[11] Palmieri A, Pezzetti F, Brunelli G, . Zirconium oxide regulates RNA interfering of osteoblast-like cells. Journal of Materials Science: Materials in Medicine, 2008, 19(6): 2471–2476

[12] Palmieri A, Pezzetti F, Brunelli G, . Medpor regulates osteoblast’s microRNAs. Bio-Medical Materials and Engineering, 2008, 18(2): 91–97

[13] Foldager C, Bendtsen M, Zou X, . ISSLS prize winner: positron emission tomography and magnetic resonance imaging for monitoring interbody fusion with equine bone protein extract, recombinant human bone morphogenetic protein-2, and autograft. Spine, 2008, 33(25): 2683–2690

[14] Li H, Zou X, Woo C, . Experimental anterior lumbar interbody fusion with an osteoinductive bovine bone collagen extract. Spine, 2005, 30(8): 890–896

[15] Zou X, Zou L, Foldager C, . Different mechanisms of spinal fusion using equine bone protein extract, rhBMP-2 and autograft during the process of anterior lumbar interbody fusion. Biomaterials, 2009, 30(6): 991–1004

[16] Saeed A I, Sharov V, White J, . TM4: a free, open-source system for microarray data management and analysis. BioTechniques, 2003, 34(2): 374–378

[17] Mountziaris P M, Spicer P P, Kasper F K, . Harnessing and modulating inflammation in strategies for bone regeneration. Tissue Engineering Part B: Reviews, 2011, 17(6): 393–402

[18] Mountziaris P M, Mikos A G. Modulation of the inflammatory response for enhanced bone tissue regeneration. Tissue Engineering Part B: Reviews, 2008, 14(2): 179–186

[19] Kolar P, Schmidt-Bleek K, Schell H, . The early fracture hematoma and its potential role in fracture healing. Tissue Engineering Part B: Reviews, 2010, 16(4): 427–434

[20] Takayanagi H. Osteoimmunology and the effects of the immune system on bone. Nature Reviews Rheumatology, 2009, 5(12): 667–676

[21] Pape H C, Marcucio R, Humphrey C, . Trauma-induced inflammation and fracture healing. Journal of Orthopaedic Trauma, 2010, 24(9): 522–525

[22] Rodig S J, Shahsafaei A, Li B, . The CD45 isoform B220 identifies select subsets of human B cells and B-cell lymphoproliferative disorders. Human Pathology, 2005, 36(1): 51–57

[23] Koarai A, Traves S L, Fenwick P S, . Expression of muscarinic receptors by human macrophages. The European Respiratory Journal, 2012, 39(3): 698–704

[24] Koarai A, Yanagisawa S, Sugiura H, . Cigarette smoke augments the expression and responses of toll-like receptor 3 in human macrophages. Respirology, 2012, 17(6): 1018–1025

[25] Miwa T, Zhou L, Maldonado M A, . Absence of CD59 exacerbates systemic autoimmunity in MRL/lpr mice. Journal of Immunology, 2012, 189(11): 5434–5441

[26] Kalland M E, Oberprieler N G, Vang T, . T cell-signaling network analysis reveals distinct differences between CD28 and CD2 costimulation responses in various subsets and in the MAPK pathway between resting and activated regulatory T cells. Journal of Immunology, 2011, 187(10): 5233–5245

[27] Rouault C, Pellegrinelli V, Schilch R, . Roles of chemokine ligand-2 (CXCL2) and neutrophils in influencing endothelial cell function and inflammation of human adipose tissue. Endocrinolo-gy, 2013, 154(3): 1069–1079

[28] Chou J, Werb Z. MicroRNAs play a big role in regulating ovarian cancer-associated fibroblasts and the tumor microenvironment. Cancer Discovery, 2012, 2(12): 1078–1080

[29] Melo S A, Kalluri R. miR-29b moulds the tumour microenvironment to repress metastasis. Nature Cell Biology, 2013, 15(2): 139–140

[30] Chou J, Lin J H, Brenot A, . GATA3 suppresses metastasis and modulates the tumour microenvironment by regulating microRNA-29b expression. Nature Cell Biology, 2013, 15(2): 201–213

[31] Tural S, Kara N, Alayli G, . Association between osteoporosis and polymorphisms of the bone Gla protein, estrogen receptor 1, collagen 1-A1 and calcitonin receptor genes in Turkish postmenopausal women. Gene, 2013, 515(1): 167–172

[32] Lindahl K, Rubin C J, Brändström H, . Heterozygosity for a coding SNP in COL1A2 confers a lower BMD and an increased stroke risk. Biochemical and Biophysical Research Communications, 2009, 384(4): 501–505

[33] Narisawa S, Yadav M C, Millán J L. In vivo overexpression of tissue-nonspecific alkaline phosphatase increases skeletal minera-lization and affects the phosphorylation status of osteopontin. Journal of Bone and Mineral Research, 2013, 28(7): 1587–1598

[34] Kapinas K, Kessler C, Ricks T, . miR-29 modulates Wnt signaling in human osteoblasts through a positive feedback loop. The Journal of Biological Chemistry, 2010, 285(33): 25221–25231

[35] Li Z, Hassan M Q, Jafferji M, . Biological functions of miR-29b contribute to positive regulation of osteoblast differentiation. The Journal of Biological Chemistry, 2009, 284(23): 15676–15684

[36] Kamiya N, Kobayashi T, Mochida Y, . Wnt inhibitors Dkk1 and Sost are downstream targets of BMP signaling through the type IA receptor (BMPRIA) in osteoblasts. Journal of Bone and Mineral Research, 2010, 25(2): 200–210

[37] Kamiya N, Ye L, Kobayashi T, . BMP signaling negatively regulates bone mass through sclerostin by inhibiting the canonical Wnt pathway. Development, 2008, 135(22): 3801–3811

[38] Guo X, Day T F, Jiang X, . Wnt/β-catenin signaling is sufficient and necessary for synovial joint formation. Genes & Development, 2004, 18(19): 2404–2417

[39] Zhang J, He X C, Tong W G, . Bone morphogenetic protein signaling inhibits hair follicle anagen induction by restricting epithelial stem/progenitor cell activation and expansion. Stem Cells, 2006, 24(12): 2826–2839

[40] He X C, Zhang J, Tong W G, . BMP signaling inhibits intestinal stem cell self-renewal through suppression of Wnt–β-catenin signaling. Nature Genetics, 2004, 36(10): 1117–1121

[41] Li J, Huang X, Xu X, . SMAD4-mediated WNT signaling controls the fate of cranial neural crest cells during tooth morphogenesis. Development, 2011, 138(10): 1977–1989

[42] Zhang H, Sucato D J, Welch R D. Recombinant human bone morphogenic protein-2-enhanced anterior spine fusion without bone encroachment into the spinal canal: a histomorphometric study in a thoracoscopically instrumented porcine model. Spine, 2005, 30(5): 512–518

[43] Damien C J, Grob D, Boden S D, . Purified bovine BMP extract and collagen for spine arthrodesis: preclinical safety and efficacy. Spine, 2002, 27(16S): S50–S58

[44] Sandhu H S, Khan S N. Animal models for preclinical assessment of bone morphogenetic proteins in the spine. Spine, 2002, 27(16S): S32–S38

[45] Minamide A, Yoshida M, Kawakami M, . The use of cultured bone marrow cells in type I collagen gel and porous hydroxyapatite for posterolateral lumbar spine fusion. Spine, 2005, 30(10): 1134–1138

[46] Kandziora F, Pflugmacher R, Scholz M, . Comparison of BMP-2 and combined IGF-I/TGF-ss1 application in a sheep cervical spine fusion model. European Spine Journal, 2002, 11(5): 482–493

[47] Burks M V, Nair L. Long-term effects of bone morphogenetic protein-based treatments in humans. Journal of Long-Term Effects of Medical Implants, 2010, 20(4): 277–293

Acknowledgements

The present study was financially supported by National Program on Key Basic Research Project (973 Program; Grant No. 2012CB619100), China Postdoctoral Science Foundation (Grant No. 2013M531876), the National Natural Science Foundation of China (Grant Nos. U0732001, 81000676, 81171682 and 81071512), the Fabrikant Mads Clausens Foundation, Sino-Danish Science and Technology Cooperation Project between Chinese and Danish Governments (AM14:29NNP14), Scientific Research Foundation for the Returned Overseas Chinese Scholars by State Education Ministry (Grant Nos. 2009-136 and 2011-508), Ph.D. Programs Foundation of Ministry of Education of China (Grant No. 20100171110054), and the High-level Talented People Project for Higher Education Institutions of Guangdong Province (Grant No. 80000-3210002). Ossacur AG (Oberstenfeld, Germary) provided free supply of COLLOSS^â E. The authors would like to thank the Orthopaedic Research Laboratory and the Institute of Clinical Medicine at Aarhus University, the staff at the CapitalBio Incorporation of Beijing, China for their assistance during the entire project.

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2014 Higher Education Press and Springer-Verlag Berlin Heidelberg