Figure 2.

miR-101 suppresses human cholangiocarcinoma growth in vivo. A: Construction of human cholangiocarcinoma cell lines with stable overexpression of miR-101. CCLP1 and HuCCT1 cells were stably transduced with miR-101-1 lentivirus (L/miR101) and control lentivirus (L/control), respectively. Left panel: Immunofluorescent microscopy for enhanced green fluorescent protein in cells stably transduced with the miR-101-1 lentivirus (this vector carries the enhanced green fluorescent protein gene under the control of the same cytomegalovirus promoter). Upper row: miR101-1 lentivirus-transduced cells. lower row: Noninfected control cells. Right panel: RT-qPCR analysis for miR-101 in cells infected with miR-101-1 and scramble control lentivirus. The results represent means ± SEM of the miR-101/U6 small nuclear 2 ratio normalized to the scramble control cells. B–F: miR-101–overexpressed or scramble control CCLP1 and HuCCT1 cells (2 × 10⁶) were inoculated subcutaneously into SCID mice (n = 6). Thirty days after inoculation, the mice were sacrificed and the tumors were recovered for analyses. B: Xenograft tumor masses from SCID mice (six mice per group; no tumor development in 1 of 6 mice inoculated with miR-101–overexpressed HuCCT1 cells). C: The volume of xenograft tumors. The data represent means ± SD from six mice. D: The weight of xenograft tumors. The data represent means ± SD from six mice. The bar indicates mean weight of each group. E: The levels of miR-101 in xenograft tumor tissues as determined by RT-qPCR. The data are shown as the means ± SEM from six mice. F: CD31 immunofluorescence staining in xenograft tumor tissues. Left panel: Representative image of CD31 immunofluorescence (CD31 was shown as green and nuclei were counterstained red). Right panel: Normalized capillary numbers in the tumors of each group (the data are expressed as means ± SD from six mice). Scale bars: 100 μm (A and F). ^∗P < 0.05, ^∗∗P < 0.01.