In vitro differentiation of human osteosarcoma cells

人骨肉瘤细胞的体外分化

• 批准号: 62480374
• 负责人: TSUCHIDA Nobuo
• 金额: $ 3.52万
• 依托单位: Faculty of Dentistry, Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1987
• 资助国家: 日本
• 起止时间: 1987 至 1988
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-62480374/
• 关键词: osteosarcoma; alkalinephosphatase; 分化; アルカリフォスファターゼ; オステオネクチン; グラ蛋白質; ハイドロキシアパタイト; 分化誘導; cDNAクローン

Histopathological and electron microscopical observations suggest that the pattern of bone formation in osteosarcoma cells is principally similar to that of normally calcifying tissue. Using osteosarcoma cell line (HOS) established from human osteosarcoma, in the present work we asked questions whether or not (1) HOS cells is induced to differentiate in vitro and (2)KHOS cells, HOS cells transformed by Kirsten murine sarcoma virus (K-MSV) express differentiation markers of osteogenic cells. In addition, we studied on osteonection gene which is expressed in osteogenic cells.(1) The expression of differentiation markers. HOS cells were found to express the bone/liver/kidney-type alkaline phosphatase(B/K/L/ALPase). The expression of ALPase was found to be regulated by growth rate and/or cellular contact. The addition of beta-glycerophosphate and ascorbic acid to the culture medium induced Hos cells to deposition of calcium inside. The electron microscopic observation showed that HOS cells produced hydroxyapatite (HA) crystals, which was identified by Debye-Scherrer ring and micro X-ray analyses. Two initiation sites for calcification were observed as vesicles with HA crystals inside; one type similar to matrix vesicle was 0.3-1 um in diameter and the other 1-3 um. We also observed development of HA crystals along collagen fibers. These results suggest that HOS cells possess an osteogenic potential and thus can be induced to express differentiation markers typical of osteogenic cells.(2) The expression of differentiation markers in KHOS cells. The expression of both ALPase and deposition of calcium was suppressed. KHOS cells contained a single copy of K-MSV proviral integrated DNA from which RNA was transcribed. Therefore, it is highly likely that k-ras oncogene suppressed the expression of differentiation markers.(3) We molecularly cloned osteonection cDNA from human and bovine and determined nucleotide sequences.

组织学和电子显微镜观察表明，骨肉瘤细胞中的骨形成模式基本上与正常钙化组织相似。本研究以人骨肉瘤细胞系（HOS）为研究对象，探讨了（1）HOS细胞在体外是否被诱导分化;（2）KHOS细胞，即Kirsten鼠肉瘤病毒（K-MSV）转化的HOS细胞是否表达成骨细胞的分化标志。此外，我们还对成骨细胞中表达的骨连接基因进行了研究。(1)分化标志物的表达。发现HOS细胞表达骨/肝/肾型碱性磷酸酶（B/K/L/ALP）。发现AL β的表达受生长速率和/或细胞接触的调节。在培养液中加入β-甘油磷酸盐和抗坏血酸可诱导Hos细胞内钙的沉积。电镜观察显示，HOS细胞产生了羟基磷灰石（HA）晶体，并通过Debye-Scherrer环和显微X射线分析鉴定了HA晶体。钙化起始部位为囊泡，囊泡内有HA晶体，一种类似于基质囊泡，直径为0.3 ~ 1 μ m，另一种为1-3 μ m。我们还观察到HA晶体沿着胶原纤维的发展。这些结果表明，HOS细胞具有成骨潜能，因此可以被诱导表达成骨细胞典型的分化标志物。(2)KHOS细胞中分化标志物的表达。抑制了ALP的表达和钙的沉积。KHOS细胞含有单拷贝的K-MSV前病毒整合DNA，从中转录RNA。因此，很可能是k-ras癌基因抑制分化标志物的表达。(3)我们从人和牛中克隆了骨连接素cDNA，并确定了核苷酸序列。

项目成果

片山奈知子: 口腔病学会雑誌. 55. 585-598 (1988)

Katayama，Nachiko：口腔医学会杂志 55. 585-598 (1988)。

Tohru Ikeda: "Differentiation and calcification of human osteosarcoma cell line HOS." The Journal of the Stomatological Society, Japan. 55. 106-118 (1988)

Tohru Ikeda：“人骨肉瘤细胞系 HOS 的分化和钙化。”

Nachiko Katayama: "The suppression of osteogenic differentiation markers in human osteosarcoma cells transformed by Kisrsten murine sarcoma virus." The Journal of the Stomatological Society, Japan.55. 585-598 (1988)

Nachiko Katayama：“Kisrsten 鼠肉瘤病毒转化的人骨肉瘤细胞中成骨分化标记物的抑制。”

池田通: 日腔病学会雑誌. 55. 106-118 (1988)

池田通：日本腔学会杂志 55. 106-118 (1988)。

池田通: 口腔病学会雑誌. 55. 106-118 (1988)

Ikeda, T.：口腔医学会杂志 55. 106-118 (1988)