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. 2022 Jun 21;2(7):100239. doi: 10.1016/j.crmeth.2022.100239

Figure 3.

Figure 3

Multi-miR mediated inhibition of tumor suppressor genes in the mouse liver

(A) Diagram depicting the generation of autochthonous liver tumor models using hydrodynamic tail vein injection. The injection delivers transposons expressing the Myc oncogene to the liver, together with GFP-linked Multi-miR shRNAs targeting multiple tumor suppressor genes (Trp53, Pten, Apc). BL6, C57BL/6 mice.

(B) Kaplan-Meier survival curves of mice treated with hydrodynamic tail vein injections of transposon vectors expressing Myc and GFP-linked miR-E shRNAs targeting Trp53, Pten or Apc, or Multi-miR shRNAs targeting Trp53-Pten, Trp53-Apc, or Trp53-Pten-Apc (n = 5). C57BL/6 mice injected with the Myc transposon and Multi-miR shTrp53-shPten-shApc succumbed to liver tumors with a median tumor-free survival of 31 days, while others produced tumors with longer median survival (Myc: no onset of disease, shTrp53;Myc = 65 days, shPten;Myc = 109 days, shApc;Myc = 52 days, shTrp53-shPten;Myc = 59 days, shTrp53-shApc;Myc = 36 days). Statistical significance was calculated using the log rank (Mantel-Cox) test (∗p < 0.05; ∗∗p < 0.01).

(C) Representative images showing typical liver tissue and tumors (top), GFP signals (middle), and hematoxylin and eosin (H&E) staining (bottom). Scale bar, 50 μm.

(D) Quantification of target knockdown in tumor samples. Total RNA was extracted from liver tumors in two independent mice (n = 2) injected with the Myc transposon and the indicated shRNA constructs. Expression levels of Trp53, Pten, and Apc were assessed by qRT-PCR. Each qRT-PCR was done in triplicate. Error bars represent the SD.