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. 2022 May 30;45(6):388–402. doi: 10.14348/molcells.2022.2232

Fig. 6. Subcutaneous tumors in nude mice.

Fig. 6

(A) Growth curves of tumors in vivo. The formula for calculating the tumor volume was as follows: V = 0.5 × a × b × b, where a and b represent the long and short diameters of the tumor. The number of animals in one group was 12 (n = 12). *P < 0.05, **P < 0.01, vs model group; #P < 0.05, vs model + Lv-KLF4 group. Data are expressed as the means ± SD. (B) Immunohistochemical staining of Ki67 protein in tumors cells from each group. For statistics, 5 visual fields of each sample were randomly selected for counting and calculating the average value, the Ki67 positive rate is expressed as the percentage of positive cells in all. *P < 0.05,**P < 0.01, vs model group; ##P < 0.01, vs model + Lv-KLF4 group (t-test). Data are expressed as the means ± SD. (C) Immunofluorescence staining of CD34 in tumors from each group. Five visual fields of each sample were randomly selected for counting, microvessel density was expressed as the number of microvessels in a unit field of view. **P < 0.01, vs model group; ##P < 0.01, vs model + Lv-KLF4 group (t-test). Data are expressed as the means ± SD. (D) RT-qPCR was used to detect the relative levels of IMAT1, hsa-miR22-3p, KLF4, and Snai1 in tumors from each group. β-actin and U6 were used as reference genes, respectively. *P < 0.05, **P < 0.01, vs model group; ##P < 0.01, vs model + Lv-KLF4 group (t-test). Data are expressed as the mean ± SD. (E) Changes in the of relevant protein expressions (KLF4, Snai1, α-SMA, and vimentin) in tumors from each group were analyzed by western blotting. The x-coordinate represents the experimental groups, and the y-coordinate represents the relative protein level. β-actin was used as the reference protein. The results are shown as the mean ± SD. *P < 0.05, **P < 0.01, vs model group; #P < 0.05, ##P < 0.01, vs model + Lv-KLF4 group (t-test). Data are expressed as the means ± SD. (F) Survival analysis of the tumor-bearing mice. Survival events of mice were recorded once a week until the 13th week following recombinant virus injection.