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成年小鼠骨细胞的分离鉴定及其成纤维细胞生长因子受体的表达

, PP. 718-722

Keywords: 骨细胞,细胞分离,成纤维细胞生长因子受体

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Abstract:

目的建立稳定的成年小鼠骨细胞分离和鉴定方法,检测其标志性基因和成纤维细胞生长因子受体(fibroblastgrowthfactorreceptor,FGFR)表达。方法将3月龄C57小鼠的胫骨和股骨分离后剪除干骺端,冲去骨髓,切割成约1mm碎片,用Ⅰ型胶原酶和EDTA溶液交替消化以获取骨细胞。分别通过形态观察、ALP染色、OC染色和E11/gp38染色鉴定骨细胞。qRT-PCR检测骨细胞标志性基因表达,并与骨细胞系IDG-SW3和MLO-Y4比较。检测FGFRs在骨细胞的表达。结果分离出的骨细胞呈多树突状或星状,并借突触彼此连接。ALP染色可见成骨细胞染色阳性,骨细胞染色阴性。OC染色成骨细胞和骨细胞均阳性,成纤维细胞阴性,而E11/gp38染色仅见骨细胞染色阳性。实验所得细胞表达骨细胞特异基因DMP1、SOST、FGF23,表明骨细胞成功分离,且此原代骨细胞与IDG-SW3和MLO-Y4细胞系表达有差异。FGFR1~3在骨细胞中均有表达。结论成功分离成年小鼠原代骨细胞,发现FGFR1~3在原代骨细胞中均有表达。

References

[1]  Karsenty G, Oury F. The central regulation of bone mass, the first link between bone remodeling and energy metabolism[J]. J Clin Endocrinol Metab, 2010, 95(11): 4795-4801. [2]Komori T. Functions of the osteocyte network in the regulation of bone mass[J]. Cell Tissue Res, 2013, 352(2): 191-198. [3]Teti A, Zallone A. Do osteocytes contribute to bone mineral homeostasis? Osteocytic osteolysis revisited[J]. Bone, 2009, 44(1): 11-16. [4]Bonewald L F. The amazing osteocyte[J]. J Bone Miner Res, 2011, 26(2): 229-238. [5]Cherian P P, Siller-Jackson A J, Gu S, et al. Mechanical strain opens connexin 43 hemichannels in osteocytes: a novel mechanism for the release of prostaglandin[J]. Mol Biol Cell, 2005, 16(7): 3100-3106. [6]van-der-Plas A, Nijweide P J. Isolation and purification of osteocytes[J]. J Bone Miner Res, 1992, 7(4): 389-396. [7]Xiao Z, Zhang S, Mahlios J, et al. Cilia-like structures and polycystin-1 in osteoblasts/osteocytes and associated abnormalities in skeletogenesis and Runx2 expression[J]. J Biol Chem, 2006, 281(41): 30884-30895. [8]李晓刚, 苏楠, 唐玉彬, 等. FGF2对破骨细胞体外分化以及骨吸收功能的直接调控作用[J]. 第三军医大学学报, 2014, 36(7): 636-639. [9]朱莹, 戚华兵, 王权, 等. 构建FGFR3基因沉默慢病毒载体及其对ATDC5细胞增殖和分化的影响[J]. 第三军医大学学报, 2013, 35(11): 1043-1047. [10]Gu G, Nars M, Hentunen T A, et al. Isolated primary osteocytes express functional gap junctions ?in vitro?[J]. Cell Tissue Res, 2006, 323(2): 263-271. [11]van-der-Plas A, Nijweide P J. JBMR anniversary classic. Isolation and purification of osteocytes. A van der Plas A, PJ Nijweide. Originally published in Volume 7, Number 4, pp 389-96 (1992)[J]. J Bone Miner Res, 2005, 20(4): 706-714. [12]Stern A R, Stern M M, Van-Dyke M E, et al. Isolation and culture of primary osteocytes from the long bones of skeletally mature and aged mice[J]. Biotechniques, 2012, 52(6): 361-373. [13]Kalajzic I, Matthews B G, Torreggiani E, et al. ?In vitro? and ?in vivo? approaches to study osteocyte biology[J]. Bone, 2013, 54(2): 296-306. [14]Uchihashi K, Aoki S, Matsunobu A, et al. Osteoblast migration into type I collagen gel and differentiation to osteocyte-like cells within a self-produced mineralized matrix: a novel system for analyzing differentiation from osteoblast to osteocyte[J]. Bone, 2013, 52(1): 102-110. [15]Pirraco R P, Cerqueira M T, Reis R L, et al. Fibroblasts regulate osteoblasts through gap junctional communication[J]. Cytotherapy, 2012, 14(10): 1276-1287. [16]Zhang K, Barragan-Adjemian C, Ye L, et al. E11/gp38 selective expression in osteocytes: regulation by mechanical strain and role in dendrite elongation[J]. Mol Cell Biol, 2006, 26(12): 4539-4552. [17]Du X, Xie Y, Xian C J, et al. Role of FGFs/FGFRs in skeletal development and bone regeneration[J]. J Cell Physiol, 2012, 227(12): 3731-3743.
[2]  王亚萍,佟明华,雷香丽,等.一种改进的大鼠胎肝干细胞分离纯化方法[J].第三军医大学学报,2013,35(04):366. [2]罗茜,王斌,雷增杰,等.肝星状细胞具有向肝细胞样细胞分化的干细胞潜能[J].第三军医大学学报,2013,35(07):604.  Luo Xi,Wang Bin,Lei Zengjie,et al.Hepatic stellate cells might possess potential capabilities of stem cells to differentiate into hepatocyte-like cells in vitro[J].J Third Mil Med Univ,2013,35(08 ):604. [3]周强,李起鸿,许建中,等.兔原代肋软骨细胞的三步酶消化法高效分离及体外培养观察[J].第三军医大学学报,2005,27(16):1667. [4]刘美,李少林,陈宓,等.人结肠癌细胞株CW-2干细胞富集方法的比较[J].第三军医大学学报,2010,32(09):912.  Liu Mei,Li Shaolin,Chen Mi,et al.Comparison on enriching methods for colon cancer stem cells from human colon cancer cell line CW-2[J].J Third Mil Med Univ,2010,32(08 ):912. [5]董华英,吴诚义,陈元文.乳腺癌新辅助化疗后的癌干细胞分离培养[J].第三军医大学学报,2010,32(20):2172.  Dong Huaying,Wu Chengyi,Chen Yuanwen.Isolation and culture of breast cancer stem cells derived from breast cancer after neoadjuvant chemotherapy[J].J Third Mil Med Univ,2010,32(08 ):2172.

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