Chondrogenic potential of chick embryonic calvaria: I. Low calcium permits cartilage differentiation

Abstract

Calvaria from day-14 calciumdeficient chick embryos produced by long-term maintenance in shell-less culture, exhibit a cartilage-like phenotype (Jacenko and Tuan 119861 Dev. Biol. 11521L232), which is restored to an osteogenic phenotype upon calcium repletion to the embryo. The expression of cartilage markers in a typically osteogenic tissue under calcium deficiency implies the presence of chondrogenic cells, and questions the conditions associated with calcium deficiency which may cause their divergent pathway of differentiation. In the present study, by explanting normal and shell-less embryonic calvarial pairs in organ culture in vitro and experimentally regulating their calcium supply, the calcium status of the calvaria was modulated as a function of medium calcium. Histological and immunoblotting analyses demonstrated for the first time that calvaria possess cells which can form genuine cartilage. This chondrogenic potential is expressed only in a low-calcium environment, where cartilage forms in both normal and shellless calvarial pairs; their calcium-suplemented counterparts, however, develop as fully osteogenic tissues. Furthermore, chondrogenesis in both normal and shell-less calvaria indicates that the chondrogenic cells must be endogenous constituents of the calvaria, rather than being derived elsewhere in response to systemic calcium deficiency. Finally, the correlation between matrix under-calcification and cartilage expression in the embryonic calvarium suggests that calcium, perhaps in the form of matrix mineral, may modulate cell differentiation during skeletogenesis.

Chondrogenic potential of chick embryonic calvaria: II. Matrix calcium may repress cartilage differentiation

“…Surprisingly, the calcium per dry weight values did not differ significantly; the reason for this phenomenon was unknown. The values of calcium content per bone, however, were within the range of those reported for day-14 and day-17 calvaria developing in vivo, or day-12 calvaria maintained in organ culture for 5 days (see Jacenko and Tuan, 1995, Table 1). This observation suggested that the chondrogenic disposition of the graft could be analyzed as a function of their calcium content, as is demonstrated by the histological analyses below.…”

“…This correlation is clearly indicated by the presence of a cartilage nodule in the superio-orbital region of the calvarium (compare with Fig. 4, Jacenko and Tuan, 1995), as well as by a n increase in the Alcian blue staining at the most temporal region of the bone. Since a major function of the CAM is to transport calcium from the calcareous eggshell into the embryonic circulation (Tuan, 1987), this localized demineralization of the tissue could have resulted from this process.…”

“…The results presented in the preceding report (Jacenko and Tuan, 1995) have demonstrated that chick embryonic calvaria possess genuine chondrogenic potential, and suggested that cartilage is expressed as a function of the calcium status of the tissue. Findings presented in this investigation have confirmed these observations, demonstrating a functional relationship between the mineralization status of the calvarial matrix and the expression of cartilage.…”

“…Additionally, several studies concluded that a critical concentration of intracellular calcium is necessary for normal synthesis of skeletal extracellular matrix and Chojkier, 1986). It is conceivable that in a calcium-(ECM) proteins, among which collagen is more sensi-deprived environment, the appropriate concentration tive to intracellular calcium changes than non-collag-of calcium ions is not accessible to the cells in the cenenous protein (Dietrich and Duffield, 1979; Flaherty tral regions of the calvaria, where cartilage markers ( Jacenko and Tuan, 1986a,b) and cartilage (see preceding report, Jacenko and Tuan, 1995) are detected.…”

“…At day 11-12 of embryonic development, calvaria from N and SL embryos were dissected and cleaned from their periosteum, as described in the preceding report (Jacenko and Tuan, 1995). Each pair of calvaria were then rinsed in physiological saline, and placed onto the chorioallantoic membrane (CAM) of either an SL or N embryo host.…”