Reduction in p53 gene dosage diminishes differentiation capacity of osteoblasts.
Reduction in p53 gene dosage diminishes differentiation capacity of osteoblasts.
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发表时间：
2000-07
影响因子：
2
通讯作者：
N. Chandar;L. Donehower;N. Lanciloti
中科院分区：
文献类型：
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作者：
N. Chandar;L. Donehower;N. Lanciloti
p53功能缺失存在于包括骨肉瘤在内的大量人类肿瘤中，在之前的研究中，我们已经证明了野生型p53与成骨细胞分化之间的密切关系。在本研究中，我们试图通过研究p53基因缺失小鼠的颅骨成骨细胞来了解骨生长和分化与p53基因剂量的关系。结果不同传代时间，p53 -/-型成骨细胞的生长规律均高于p53 +/-型成骨细胞。然而，在p53 +/-细胞的培养过程中，剩余wt等位基因的丢失发生在早期，这些细胞的生长速度和倍增率高于p53 -/-纯合子。将早期成骨细胞置于促分化培养基中，研究成骨细胞体外分化特性。骨钙素（一种分化成骨细胞的标记蛋白）在两种细胞类型中都能合成，但p53 -/-细胞始终表现出更高水平的骨钙素活性，并且在长期培养中形成的骨针状体比p53 +/-细胞多。基础骨钙素启动子活性在两种细胞类型中相似，但将wt p53瞬时转染到细胞中，p53 +/-细胞中的启动子活性比p53 -/-细胞中的启动子活性更大。对新生儿p53 +/-和-/-长骨切片进行组织学检查，未发现明显异常。TUNEL染色显示两种p53基因型的骨生长板和衬细胞均有凋亡细胞，但增生性软骨细胞凋亡仅在p53 +/-骨中可见。总的来说，我们的研究表明，只有在发育过程中至少有一个p53基因的功能拷贝存在时，成骨细胞分化才成为p53依赖的过程。
BACKGROUND Loss of p53 function is seen in a large number of human tumors including osteosarcoma, and in previous studies we have demonstrated a close relationship between wild type p53 and osteoblasts differentiation. In the present study we attempted to understand the relationship of bone growth and differentiation to p53 gene dosage by studying calvarial osteoblasts obtained from p53 deficient mice. RESULTS Growth rules were higher in p53 -/- than p53 +/- osteoblasts when passaged for different lenght of time. However, loss of the remaining wt allele occurred early during culturing of the p53 +/- cells, and these cells became immortalized with growth rates and doubling times reaching values higher than the p53 -/- homozygotes. Early passage osteoblasts were used to study differentiation properties of these osteoblasts in vitro by exposing them to differentiation promoting media. While osteocalcin, a marker protein of differentiated osteoblasts, was synthesized in both cell types, p53 -/- cells consistently showed higher levels of osteocalcin activity, and in long term cultures formed more bone spicules that p53 +/- cells. Basal osteocalcin promoter activity was similar in both cell types, but transient transfection of wt p53 into the cells produced a larger activation of the promoter in p53 +/- cells, than in p53 -/- cells. Sections of long bones from neonatal p53 +/- and -/- were examined histologically, and no gross abnormalities were detected. TUNEL staining of the bones showed apoptotic cells in the growth plate and lining cells of both p53 genotypes, but apoptosis in hypertrophic chondrocytes was seen only in p53 +/- bones. CONCLUSION Overall our studies show that osteoblastic differentiation becomes a p53 dependent process only when at least one functional copy of the p53 gene is present during development.