Zhang Zhengguo, Li Kuanxin, Zhang Xiaomei, Fang Zhong, Xiong Wei, Chen Qi, Chen Wenjian, Li Feng
Department of Orthopaedics, Huazhong University of Science and Technology, Wuhan, 430030, China.
Department of Orthopaedics, First Affiliated Hospital, School of Medicine, Shihezi University, Shihezi, 832008, China.
J Huazhong Univ Sci Technolog Med Sci. 2012 Jun;32(3):364-369. doi: 10.1007/s11596-012-0063-1. Epub 2012 Jun 9.
The formation of osteolytic bone lesions is a key process for osteolytic cancer to metastasize to the bone and is under the control of a set of transcription factors. Recently, the inhibitor of differentiation 1 (Id1) has been linked with angiogenesis, tumorigenesis, metastasis and bone formation. However, the function of Id1 during the process of bone destruction caused by cancer in vivo has not yet been elucidated. We, therefore, examined whether and how Id1 affects the ability of cancer to form osteolytic lesion in vivo. The study used a lentiviral vector overexpressing short hairpin RNA (shRNA) targeting Id1 gene. PC3 cells, a prostate cancer cell line, were transduced with Id1 shRNA or negative control (NC) shRNA before implantation in BALB/c mice. Cells were implanted in a tibial injection model. Tumor formation in bone was monitored by X-ray. The relationship between parathyroid hormone-related protein (PTHrP), an osteolytic factor, and Id1 was analyzed by using immunohistochemistry in tissue sections from osteolytic lesion of the BALB/c mice. Our results showed that Id1 shRNA delivery to PC3 cells by lentivirus caused efficient and stable Id1 gene silencing. In the intratibial model, PC3 cells produced primarily osteolytic lesions in the bone. Eleven of 14 mice in Id1 shRNA group but only 4 of 14 mice in the NC shRNA group developed osteolytic lesions with cortical destruction at 4th week. Mice treated with Id1 shRNA had larger tumor volume in the bone and larger cortical destruction. The expression of PTHrP protein in PC3 cells was not affected by Id1 knockdown in vivo. These results indicate that Id1 may down-regulate the ability of PC3 cells to form osteolytic lesions in vivo and the signal pathway needs to be further investigated.
溶骨性骨病变的形成是溶骨性癌症转移至骨的关键过程，且受一组转录因子的调控。最近，分化抑制因子1（Id1）已与血管生成、肿瘤发生、转移及骨形成相关联。然而，Id1在体内癌症所致骨破坏过程中的功能尚未阐明。因此，我们研究了Id1是否以及如何影响癌症在体内形成溶骨性病变的能力。该研究使用了一种过表达靶向Id1基因的短发夹RNA（shRNA）的慢病毒载体。在将前列腺癌细胞系PC3细胞植入BALB/c小鼠之前，用Id1 shRNA或阴性对照（NC）shRNA进行转导。将细胞植入胫骨注射模型中。通过X射线监测骨中的肿瘤形成。使用免疫组织化学方法对来自BALB/c小鼠溶骨性病变组织切片中的溶骨因子甲状旁腺激素相关蛋白（PTHrP）与Id1之间的关系进行分析。我们的结果表明，通过慢病毒将Id1 shRNA递送至PC3细胞可导致高效且稳定的Id1基因沉默。在胫骨内模型中，PC3细胞在骨中主要产生溶骨性病变。Id1 shRNA组14只小鼠中有11只在第4周出现了伴有皮质破坏的溶骨性病变，而NC shRNA组14只小鼠中只有4只出现此类病变。用Id1 shRNA处理过的小鼠在骨中的肿瘤体积更大，皮质破坏也更大。体内Id1基因敲低并未影响PC3细胞中PTHrP蛋白的表达。这些结果表明，Id1可能在体内下调PC3细胞形成溶骨性病变的能力，其信号通路有待进一步研究。
