Hofstetter W, Guenther H L, Stutzer A, Schenk R, Fleisch H, Friis R
Department of Pathophysiology, University of Berne, Switzerland.
J Bone Miner Res. 1991 Jun;6(6):609-22. doi: 10.1002/jbmr.5650060612.
Osteoblastic cells were cloned by culturing rat calvariae cells in agarose in the presence of TGF-beta and EGF. Two bone cell lines were established by immortalizing such an osteoblastic clonal cell population by the introduction of the avian v-mycOK10 gene in the form of a mouse ecotropic retrovirus. Although originating from the same clonal cell population, the two lines exhibited somewhat differing properties. IRC10/30-myc1 expressed alkaline phosphatase (AP), showed PTH- and PGE2-induced cAMP production, synthesized mainly collagen type I and a minor fraction of type III, and produced mRNA for the bone-specific protein osteocalcin. IRC10/30-myc3 did not express AP, showed no PTH responsiveness, and synthesized only about one-third as much collagen as IRC10/30-myc1 (4 versus 12% of total protein synthesis). However, the cell line IRC10/30-myc3 was induced to synthesize cAMP by PGE2 and produced osteocalcin mRNA. When cultured in vivo in diffusion chambers, both lines proved to be osteogenic. Besides bone, both lines also formed cartilage and fibrous tissue. Thus, by immortalizing a clonal cell population of the osteoblastic phenotype, cell lines expressing varying properties can emerge. Furthermore, the expression of alkaline phosphatase and PTH-inducible adenylate cyclase are not prerequisites for a cell to form bone in vivo. Finally, cells expressing the phenotype of differentiated osteoblasts, including osteocalcin synthesis, still have a multipotential differentiation capacity and form bone and cartilage in vivo.
在转化生长因子-β（TGF-β）和表皮生长因子（EGF）存在的情况下，通过在琼脂糖中培养大鼠颅骨细胞，克隆出了成骨细胞。通过以小鼠嗜亲性逆转录病毒的形式导入禽v-mycOK10基因，使这样一个成骨克隆细胞群体永生化，从而建立了两条骨细胞系。尽管这两条细胞系起源于相同的克隆细胞群体，但它们表现出一些不同的特性。IRC10/30-myc1表达碱性磷酸酶（AP），显示甲状旁腺激素（PTH）和前列腺素E2（PGE2）诱导的环磷酸腺苷（cAMP）生成，主要合成I型胶原蛋白和少量III型胶原蛋白，并产生骨特异性蛋白骨钙素的信使核糖核酸（mRNA）。IRC10/30-myc3不表达AP，对PTH无反应，合成的胶原蛋白仅约为IRC10/3个-myc1的三分之一（占总蛋白合成的4%对12%）。然而，细胞系IRC10/30-myc3被PGE2诱导合成cAMP并产生骨钙素mRNA。当在扩散小室中进行体内培养时，两条细胞系都被证明具有成骨能力。除了骨组织外，两条细胞系还形成了软骨和纤维组织。因此，通过使成骨细胞表型的克隆细胞群体永生化，可以产生表达不同特性的细胞系。此外，碱性磷酸酶和PTH诱导的腺苷酸环化酶的表达并非细胞在体内形成骨组织的先决条件。最后，表达分化成骨细胞表型的细胞，包括骨钙素的合成，仍然具有多能分化能力，并在体内形成骨和软骨。
