An increasingly high occurrence of bone metastases in hepatocellular carcinoma (HCC) patients highlights the importance of fundamental research on HCC bone metastasis, which has been limited in its success due to the lack of a model system.
Establishment of animal and cellular models of HCC bone metastasis and discovery of HCC bone metastasis-related genes.
Luciferase-transfected HCC cell lines HCCLM3, MHCC97H, and SMMC-7721 were used to inoculate nude mice intracardially. Formation of bone metastases was examined by bioluminescence imaging, SPECT, and pathology study. Metastatic cells in bone were isolated and subcultured. Differences between bone metastatic cells and their parental cells were studied by in vitro/in vivo assays.
Mouse model of HCC bone metastasis was successfully established. Injected tumour cells formed metastases in the skull, the spine, the hind limbs, and the sternum, causing osteolytic lesions via act of MMP-1 and recruitment of osteoclasts. Four bone metastatic cell lines were extracted from HCCLM3-inoculated mice and were demonstrated to exhibit a much stronger ability to form bone metastases as well as other phenotypes, including enhanced in vitro migration/invasion and colony formation. Moreover, the expression of PTHrP, MMP-1, and CTGF was significantly elevated in bone metastatic cells compared to parental HCC cells.
The nude mouse model and bone metastatic cell lines together provide an effective simulation of HCC bone metastasis. This model system will become powerful tool with which to explore the mechanisms and therapies of HCC bone metastasis. Additionally, PTHrP, MMP-1, and CTGF are candidate genes related to HCC bone metastasis.
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We thank Ph.D. Qing-qing Mo for technical support in bioluminescence imaging and Dr Yan-xia Shen, Yun-feng Han, and Jin-ling Tian for technical support in SPECT scanning. This work is supported by the State Key Project on Infectious Diseases of China (Grant Nos. 2012ZX10002016-004 and 2012ZX10002010-001-004), the Chinese Ministry of Public Health for Key Clinical Projects (No. 439, 2010) to Prof Xiao-ping Chen, the National Nature Science Foundation of China (Nos. 30973498 and 81072001) to Prof Bixiang Zhang, and the National Training Programs of Innovation and Entrepreneurship for Undergraduates (No. 1210487144) to Rui Hou.
Conflict of interest
The authors have declared that there is no conflict of interest.
Electronic supplementary material
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Suppl. Fig. 1. The expression of LUC in PGL4.51-transfected cell lines. (A) BLI images of LUC-labelled cells in double-diluted concentration. (B) The relative luminescence units (RLU) of LUC-labelled cells in different concentration. The results revealed a proportional relationship between RLU and the number of cells, with r2=0.9986, 0.9882, and 0.9916, respectively. Supplementary material 1 (TIFF 3637 kb)
Suppl. Fig. 2. Representative images of mice received intra-tibial injection. (A) BLI images of mice received intra-tibial injection of LM3, 97H, and 7721, respectively. (B) Magnified X-ray image of one tumour bearing mouse. Bone lesion in hind limb is indicated by black arrow. X-ray image of a normal mouse is showed on the right. Supplementary material 2 (TIFF 1851 kb)
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Hou, R., Wang, Y., Liang, H. et al. Animal and cellular models of hepatocellular carcinoma bone metastasis: establishment and characterisation. J Cancer Res Clin Oncol 141, 1931–1943 (2015). https://doi.org/10.1007/s00432-015-1958-6
- Animal model
- Bone metastasis
- Cell line
- Hepatocellular carcinoma