Mouse Meckel's cartilage chondrocytes evoke bone-like matrix and further transform into osteocyte-like cells in culture

Abstract

Background We reported that when Meckel's cartilage was transplanted ectopically, chondrocytes transformed into osteocyte‐like cells accompanying the extracellular calcified matrix. However, we could not determine whether the osteocyte‐like cells were derived from host tissues or from Meckel's cartilage itself. Therefore, we examined whether the Meckel's cartilage chondrocytes, which have a retrogressive ultimate fate, are capable of inducing the observed calcification and further transform into osteocyte‐like cells in culture. Methods Meckelian chondrocytes isolated enzymatically were plated at a low density and grown in α‐MEM containing 10% FBS at 37°C under 5% CO2 in air for up to 4 weeks. Results Chondrocytes were fibroblast‐like cells early in culture, but gradually transformed from polygonal cells into typical chondrocytes showing metachromasia with toluidine blue staining. After an additional week of culture, the chondrocytes transformed from large to small round cells accompanying nodule formations. Small round cells multiple‐layered actively, and showed more intense alkaline phosphatase (ALPase) activity. Immunostaining identified type II collagen in the extracellular matrix at 2 weeks of culture, and type I collagen and osteocalcin were later synthesized by round cells. von Kossa's reaction showed extensive precipitation of calcification throughout the flocculent materials. Ultrastructural analysis showed that the cells surrounded by calcified matrix strongly resembled osteocytes. Conclusions The present study suggested that the Meckel's cartilage chondrocytes can express the osteocyte‐like phenotype in vitro during synthesis of bone‐type marker proteins such as osteocalcin or type I collagen. © 1996 Wiley‐Liss, Inc.

Expression of osteogenic proteins during the intrasplenic transplantation of Meckel's chondrocytes: A histochemical and immunohistochemical study

“…Thick-banded collagen fibrils were easily distinguishable from the fine, type II collagen fibrils existing in the intact cartilage matrix. These fibrils ultrastructurally resembled those reported during the transformation of Meckel's chondrocytes in a cell culture (Ishizeki et al, 1996b(Ishizeki et al, , 1998a and in the femur growth plate of chick embryos (Roach et al, 1995;Roach 1997). They were identified as type I collagen in a bone-composing collagen matrix.…”

“…Although there are some reports on the transformation of chondrocytes in growth plate cartilage of long bones in vivo (Holtrop, 1972;Yoshioka and Yagi, 1988;Galotto et al, 1994;Roach et al, 1995), it is of interest that ectomesenchymal cells originating in the neural crest, such as the chicken chondroid bone (Hall, 1972;Beresford, 1981;Lengelé et al, 1996) and Meckel's cartilage (Ishizeki et al, 1996b(Ishizeki et al, , 1997 as well as mandibular condylar chondrocytes, demonstrate high osteogenic potential in comparison with those of mesoderm-derived cartilage. Chondroid bones appear in some developing membranous bones, such as cranial, lower facial, and mandibular bones (Goret-Nicaise and Dhem, 1982).…”

“…Chondrocytes for transplantation were isolated using 0.1% hyaluronidase (type I-S; Sigma Chemical Co., St. Louis, MO, USA) and 0.15% collagenase (type II; Worthington Biochemical Corp., Freehold, NJ, USA), as described previously (Ishizeki et al, 1996b(Ishizeki et al, , 1998b. Chondrocytes were then collected in a 10 ml centrifuge tube and washed three times with -modified Eagle medium ( -MEM) containing 20% fetal bovine serum.…”

“…Briefly, the specimens surrounded by splenic tissue were fixed with 4% paraformaldehyde, washed thoroughly in PBS, and incubated with 5% silver nitrate for 20 min for von Kossa staining (Ishizeki et al, 1996b).…”

Hypertrophic chondrocytes can become osteoblasts and osteocytes in endochondral bone formation

“…The hypothesis that chondrocytes may become osteoblasts was proposed more than a century ago (32,33); however despite data consistent with such a concept, this has eluded verification in vivo. The plasticity of chondrocytes in culture and their ability to change their phenotype to fibroblastic or osteoblastic states are well documented (34,35). The reversion of HCs to a prehypertrophic-like state in response to endoplasmic reticulum (ER) stress suggests that hypertrophy is not an irreversible state in vivo (25).…”