MicroRNA-28-5p Regulates Liver Cancer Stem Cell Expansion via IGF-1 Pathway

Figure 3

miR-28-5p overexpression suppresses liver CSC expansion. (a) Hepatoma cells were infected with the miR-28-5p mimic virus and control virus. The miR-28-5p overexpression effect was determined by RT-PCR assay (). (b) The mRNA expression of liver CSC markers in the miR-28-5p mimic and control HCC cells was checked by RT-PCR assay (). (c) The protein expression of liver CSC markers in the miR-28-5p mimic and control HCC cells was checked by western blot assay. GAPDH acted as a loading control. (d) The mRNA expression of stemness-associated genes in the miR-28-5p mimic and control HCC cells was checked by RT-PCR assay (). (e) The protein expression of stemness-associated genes in the miR-28-5p mimic and control HCC cells was checked by western blot assay. GAPDH acted as a loading control. (f) The EpCAM-positive cells in the miR-28-5p mimic and control HCC cells was determined by flow cytometry (). (g) The self-renewal ability of the miR-28-5p mimic and control HCC cells was compared by spheroid formation assay (). (h) In vitro limiting dilution assay of the miR-28-5p mimic and control HCC cells. The results are shown as a natural logarithm of the proportion of CSCs (). (i) The tumorigenicity of liver CSCs in the Huh7 miR-28-5p mimic and its control cells was compared by in vivo limiting dilution assay. Tumors were observed over 2 months; for each group. (j) The protein expression of liver CSC markers and stemness-associated genes in the above xenograft tumors was checked by western blot assay. GAPDH acted as a loading control. Data are represented as ; ; two-tailed Student’s -test.

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