Fgf9 regulated murine mandibular development by promoting endochondral and intramembranous ossification

doi:10.19438/j.cjoms.2021.04.004

Abstract

Abstract: PURPOSE: To investigate the role of fibroblast growth factor 9 (Fgf9) in endochondral and intramembranous ossification during mandibular development by analyzing the mandibular phenotype of Fgf9 knockout mice. METHODS: A Fgf9 gene-knock-out mice model was established for understanding the role of Fgf9 during mandibular development. Micro-CT was applied for analyzing the bone morphology and parameters. In situ hybridization was used for detecting the expression pattern of Fgf9 in mandible. H-E and Safranin O-fast green staining were used for histologic analysis of condyle, Meckel's cartilage, and the region generated by intramembranous ossification. SPSS 25.0 software was performed for statistical analysis. RESULTS: Micro-CT showed defective condyle, coracoid process, and mandibular angle in Fgf9^-/-mice with Tb. N and BMD decreased, and Tb. Sp increased. In situ hybridization indicated that Fgf9 was widely expressed in perichondrium, chondrocyte, osteoblast, endothelial cells, and mesenchyme around the cartilage and trabecular bone. H-E and Safranin O-fast green staining exhibited a malformed condyle cartilage with an inadequate secretion of cartilage matrix, decreased proportion of hypertrophic chondrocyte, and defective and dispersive trabecular bone. Moreover, the trabecular bone in the regions of the anterior part of the Meckel's cartilage and the intramembranous ossification was defective, dispersive, and hypo-mineralized. CONCLUSIONS: During mandibular development, Fgf9 participates in the endochondral ossification of the condyle by promoting maturation and secretion of the chondrocyte. Meanwhile, it is able to facilitate the secretion of osteoblast for pursuit of well-morphed trabecular bone. Furthermore, Fgf9 is involved in intramembranous ossification by regulating the osteoblast during the formation and mineralization of the trabecular bone.

Key words: Fibroblast growth factor 9, Mandibular development, Endochondral ossification, Intramembranous ossification

CLC Number:

R783.5

LI Ruo-mei, WENG Meng-jia, SUN Yi-dan, CHEN Zhen-qi. Fgf9 regulated murine mandibular development by promoting endochondral and intramembranous ossification[J]. China Journal of Oral and Maxillofacial Surgery, 2021, 19(4): 309-314.

References

[1] Su N, Jin M, Chen L.Role of FGF/FGFR signaling in skeletal development and homeostasis: learning from mouse models[J]. Bone Res, 2014, 2: 14003.
[2] Naruo K, Seko C, Kuroshima K, et al.Novel secretory heparin-binding factors from human glioma cells (glia-activating factors) involved in glial cell growth. Purification and biological properties[J]. J Biol Chem, 1993, 268(4): 2857-2864.
[3] Harada M, Murakami H, Okawa A, et al.FGF9 monomer-dimer equilibrium regulates extracellular matrix affinity and tissue diffusion[J]. Nat Genet, 2009, 41(3): 289-298.
[4] Chen T M, Hsu CH, Tsai SJ, et al.AUF1 p42 isoform selectively controls both steady-state and PGE2-induced FGF9 mRNA decay[J]. Nucleic Acids Res, 2010, 38(22): 8061-8071.
[5] Weng M, Chen Z, Xiao Q, et al.A review of FGF signaling in palate development[J]. Biomed Pharmacother, 2018, 103: 240-247.
[6] Liu Y, Ma J, Beenken A, et al. Regulation of receptor binding specificity of FGF9 by an autoinhibitory homodimerization [J]. Structure, 2017, 25(9): 1325-1336.e3.
[7] Wang L, Roth T, Abbott M, et al.Osteoblast-derived FGF9 regulates skeletal homeostasis[J]. Bone, 2017, 98: 18-25.
[8] Bird AD, Croft BM, Harada M, et al.Ovotesticular disorders of sex development in FGF9 mouse models of human synostosis syndromes[J]. Hum Mol Genet, 2020, 29(13): 2148-2161.
[9] Sentchordi-Montané L, Diaz-Gonzalez F, Cátedra-Vallés EV, et al.Identification of the third FGF9 variant in a girl with multiple synostosis-comparison of the genotype:phenotype of FGF9 variants in humans and mice[J]. Clin Genet, 2020, 99(2): 309-312.
[10] Rodriguez-Zabala M, Aza-Carmona M, Rivera-Pedroza CI, et al.FGF9 mutation causes craniosynostosis along with multiple synostoses[J]. Hum Mutat, 2017, 38(11): 1471-1476.
[11] Tang L, Wu X, Zhang H, et al.A point mutation in Fgf9 impedes joint interzone formation leading to multiple synostoses syndrome[J]. Hum Mol Genet, 2017, 26(7): 1280-1293.
[12] Ignelzi MA Jr, Wang W, Young AT.Fibroblast growth factors lead to increased Msx2 expression and fusion in calvarial sutures[J]. J Bone Miner Res, 2003, 18(4): 751-759.
[13] Hung IH, Yu K, Lavine KJ, et al.FGF9 regulates early hypertrophic chondrocyte differentiation and skeletal vascularization in the developing stylopod[J]. Dev Biol, 2007, 307(2): 300-313.
[14] Frontini MJ, Nong Z, Gros R, et al.Fibroblast growth factor 9 delivery during angiogenesis produces durable, vasoresponsive microvessels wrapped by smooth muscle cells[J]. Nat Biotechnol, 2011, 29(5): 421-427.
[15] Govindarajan V, Overbeek PA.FGF9 can induce endochondral ossification in cranial mesenchyme[J]. BMC Dev Biol, 2006, 6: 7.
[16] Xu J, Liu H, Lan Y, et al.Hedgehog signaling patterns the oral-aboral axis of the mandibular arch[J]. Elife, 2019, 8: e40315.
[17] Hinton RJ, Jing Y, Jing J, et al.Roles of chondrocytes in endochondral bone formation and fracture repair[J]. J Dent Res, 2017, 96(1): 23-30.
[18] Parada C, Chai Y.Mandible and tongue development[J]. Curr Top Dev Biol, 2015, 115: 31-58.
[19] Motch Perrine SM, Wu M, Stephens NB, et al. Mandibular dysmorphology due to abnormal embryonic osteogenesis in FGFR2-related craniosynostosis mice[J]. Dis Model Mech, 2019, 12(5): dmm038513.
[20] Sugito H, Shibukawa Y, Kinumatsu T, et al.Ihh signaling regulates mandibular symphysis development and growth[J]. J Dent Res, 2011, 90(5): 625-631.
[21] 翁梦佳,章筱悦,陈振琦. 条件性敲除软骨组织中FGF9基因小鼠模型的建立[J]. 中国口腔颌面外科杂志, 2019, 17(5): 412-417.
[22] Colvin JS, Feldman B, Nadeau JH, et al.Genomic organization and embryonic expression of the mouse fibroblast growth factor 9 gene[J]. Dev Dyn, 1999, 216(1): 72-88.
[23] Wagner EF, Karsenty G.Genetic control of skeletal development[J]. Curr Opin Genetics Dev, 2001, 11(5): 527-532.
[24] Mori-Akiyama Y, Akiyama H, Rowitch DH, et al.Sox9 is required for determination of the chondrogenic cell lineage in the cranial neural crest[J]. Proc Natl Acad Sci USA, 2003, 100(16): 9360-9365.
[25] Bhaskar SN, Weinmann JP, Schour I.Role of Meckel's cartilage in the development and growth of the rat mandible[J]. J Dent Res, 1953, 32(3): 398-410.
[26] Yang RT, Zhang C, Liu Y, et al.Autophagy prior to chondrocyte cell death during the degeneration of Meckel's cartilage[J]. Anat Rec, 2012, 295(5): 734-741.
[27] Fakhry A, Ratisoontorn C, Vedhachalam C, et al.Effects of FGF-2/-9 in calvarial bone cell cultures: differentiation stage-dependent mitogenic effect, inverse regulation of BMP-2 and noggin, and enhancement of osteogenic potential[J]. Bone, 2005, 36(2): 254-266.
[28] Behr B, Leucht P, Longaker M T, et al.Fgf-9 is required for angiogenesis and osteogenesis in long bone repair[J]. Proc Natl Acad Sci USA, 2010, 107(26): 11853-11858.
[29] Wallner C, Schira J, Wagner JM, et al.Application of VEGFA and FGF-9 enhances angiogenesis, osteogenesis and bone remodeling in type 2 diabetic long bone regeneration[J]. PLoS One, 2015, 10(3): e0118823.