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. 2017 Apr 3;108(3):512–519. doi: 10.1111/cas.13149

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© 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

This is an open access article under the terms of the Creative Commons Attribution‐NonCommercial‐NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.

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Establishment of human VASH2 mutant. (a) MLTC‐1 cells, in which VASH2 is not endogenously expressed, were stably transfected with WT‐hVASH2 or Mut‐hVASH2 cDNA. Stable transfection of these genes was confirmed by RT‐PCR analysis. (b) Cell proliferation was determined as described in the “Materials and Methods”. We repeated this experiment three times and confirmed the reproducibility. Representative data are presented as the mean and SD (n = 4 wells). *P < 0.05 (vs mock). (c) Transfectants were inoculated into Vash2 ^lacZ/lacZ mice, and tumor growth was monitored. Data are presented as the mean and SD (n = 9). *P < 0.05 (vs mock), NS (vs mock). (d) Tumor vessels were examined immunohistochemically with anti‐CD31 Ab (green), and the vascular luminal area was quantified. Data are presented as the mean and SD (n = 4). *P < 0.05 (vs mock). HRG, hygromycin‐resistance gene; NS, not significant.