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Morphological and biochemical characterization of mineralizing primary cultures of avian growth plate chondrocytes: Evidence for cellular processing of Ca2+ and Pi prior to matrix mineralization (1995)

Wu, Licia N. Y. ; Ishikawa, Yoshinori ; Sauer, Glenn R. ; [et al.]

New York, N.Y. : Wiley-Blackwell

Journal of Cellular Biochemistry 57 (1995), S. 218-237

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ISSN: 0730-2312

Keywords: chondrocytes ; cell culture ; mineralization ; calcospherites ; Ca and P mapping ; matrix vesicles ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Chemistry and Pharmacology , Medicine

Notes: Advances in the culture of mineralizing growth plate chondrocytes provided an opportunity to study endochondral calcification under controlled conditions. Here we report that these cultures synthesize large amounts of proteins characteristically associated with mineralization: type II and X collagens, sulfated proteoglycans, alkaline phosphatase, and the bone-related proteins, osteonectin and osteopontin. Certain chondrocytes appeared to accumulate large amounts of Ca2+ and Pi during the mineralization process: laser confocal imaging revealed high levels of intracellular Ca2+ in their periphery and X-ray microanalytical mapping revealed the presence of many Ca2+- and Pi-rich cell surface structures ranging from filamentous processes 0.14 ± 0.02 μm by 0.5-2.0 μm, to spherical globules 0.70 ± 0.27 μm in diameter. Removal of organic matter with alkaline sodium hypochlorite revealed numerous deposits of globular (0.77 ± 0.19 μm) mineral (calcospherites) in the lacunae around these cells. The size and spatial distribution of these mineral deposits closely corresponded to the Ca2+-rich cell surface blebs. The globular mineral progressively transformed into clusters of crystallites. Taken with earlier studies, these findings indicate that cellular uptake of Ca2+ and Pi leads to formation of complexes of amorphous calcium phosphate, membrane lipids, and proteins that are released as cell surface blebs analogous to matrix vesicles. These structures initiate development of crystalline mineral. Thus, the current findings support the concept that the peripheral intracellular accumulation of Ca2+ and Pi is directly involved in endochondral calcification.

Additional Material: 13 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/jcb.240570206

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Induction and characterization of metallothionein in chicken epiphyseal growth plate cartilage chondrocytes (1998)

Sauer, Glenn R. ; Nie, Daotai ; Wu, Licia N.Y. ; [et al.]

New York, N.Y. : Wiley-Blackwell

Journal of Cellular Biochemistry 68 (1998), S. 110-120

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ISSN: 0730-2312

Keywords: cadmium ; zinc ; cell culture ; mineralization ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Chemistry and Pharmacology , Medicine

Notes: Following exposure to cadmium or zinc, chickens were sacrificed and the liver, kidney, and bone epiphyseal growth plates harvested. When cytosolic extracts of the growth plate cartilage were fractionated by gel filtration chromatography, a protein with high metal-binding capacity and low ultraviolet (UV) absorbance eluted in the same position as liver metallothionein (MT) and a MT standard. Cd or Zn treatment resulted in a 25-fold or 5-fold induction in growth plate MT, respectively. In liver the greatest level of MT induction was seen with short-term Cd exposures. In contrast, MT levels in the growth plate increased as the duration of Cd exposure increased. Induction of MT in growth plate chondrocyte cell cultures was observed for media Cd concentrations of ≥0.1 μM and Zn concentrations of ≥100 μM. Basal and inducible levels of MT declined through the culture period and were lowest in the terminally differentiated mineralized late stages of the culture. Alkaline phosphatase activity was also lowest in the late-stage cultures, while total cellular protein increased throughout the culture period. Treatment of chondrocytes with Zn prior to Cd exposure resulted in a protective induction of MT. Pre-treatment of chondrocytes with dexamethasone resulted in suppressed synthesis of MT upon Cd exposure and greater Cd toxicity. Both Cd and Zn resulted in significantly increased levels of MT mRNA in chondrocyte cell cultures. Dexamethasone treatment resulted in an approximate 2- to 3-fold increase in MT mRNA. This is contrary to the finding that MT protein levels were decreased by dexamethasone. The findings suggest that an increased rate of MT degradation in dexamethasone-treated and late-stage chondrocyte cultures may be associated with the terminally differentiated phenotype. J. Cell. Biochem. 68:110-120, 1998. © 1998 Wiley-Liss, Inc.

Additional Material: 8 Ill.

Type of Medium: Electronic Resource

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Inhibition of terminal differentiation and matrix calcification in cultured avian growth plate chondrocytes by Rous sarcoma virus transformation (1998)

Nie, Daotai ; Ishikawa, Yoshinori ; Guo, Yande ; [et al.]

New York, N.Y. : Wiley-Blackwell

Journal of Cellular Biochemistry 69 (1998), S. 453-462

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ISSN: 0730-2312

Keywords: Rous sarcoma virus ; chondrocytes ; matrix calcification ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Chemistry and Pharmacology , Medicine

Notes: Endochondral bone formation involves the progression of epiphyseal growth plate chondrocytes through a sequence of developmental stages which include proliferation, differentiation, hypertrophy, and matrix calcification. To study this highly coordinated process, we infected growth plate chondrocytes with Rous sarcoma virus (RSV) and studied the effects of RSV transformation on cell proliferation, differentiation, matrix synthesis, and mineralization. The RSV-transformed chondrocytes exhibited a distinct bipolar, fibroblast-like morphology, while the mock-infected chondrocytes had a typical polygonal morphology. The RSV-transformed chondrocytes actively synthesized extracellular matrix proteins consisting mainly of type I collagen and fibronectin. RSV-transformed cells produced much less type X collagen than was produced by mock-transformed cells. There also was a significant reduction of proteoglycan levels secreted in both the cell-matrix layer and culture media from RSV-transformed chondrocytes. RSV-transformed chondrocytes expressed two- to- threefold more matrix metalloproteinase, while expressing only one-half to one-third of the alkaline phosphatase activity of mock infected cells. Finally, RSV-transformed chondrocytes failed to calcify the extracellular matrix, while mock-transformed cells deposited high levels of calcium and phosphate into their extracellular matrix. These results collectively indicate that RSV transformation disrupts the preprogrammed differentiation pattern of growth plate chondrocytes and inhibit chondrocyte terminal differentiation and mineralization. They also suggest that the expression of extracellular matrix proteins, type II and type X collagens, and the cartilage proteoglycans are important for chondrocyte terminal differentiation and matrix calcification. J. Cell. Biochem. 69:453-462, 1998. © 1998 Wiley-Liss, Inc.

Additional Material: 8 Ill.

Type of Medium: Electronic Resource

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