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Fig. 2 | Breast Cancer Research

Fig. 2

From: Upregulation of Nox4 induces a pro-survival Nrf2 response in cancer-associated fibroblasts that promotes tumorigenesis and metastasis, in part via Birc5 induction

Fig. 2

Targeting Nox4 inhibited tumor growth and metastasis in animal models. (A) Primary tumor growth induced by murine mammary carcinoma cell line, 4T1 when implanted orthotopically into Balb/c mice. Mice were injected with 2.5 × 104 4T1 cells into 4th mammary gland. When tumors were palpable on day 6, animals were given daily doses of control vehicle or GKT137831 (60 mg/kg body weight) by oral gavage. * represents p < 0.0001 versus control on day 30. (B) After 30 days post-tumor cell implantation, mice were euthanized and lungs were harvested, fixed and embedded for metastasis analysis. Lung micro-metastasis was scored based on H&E stained sections. N = 9 per group. (C) Primary tumor growth of E0771 murine mammary carcinoma cell line in syngeneic C57BL/6 mice, both wild type (WT) and Nox4 knock-out models (Nox4−/−). Mice were orthotopically injected with 2 × 105 E0771 cells. One group of animals were given daily doses of GKT137831 (60 mg/kg body weight) by oral gavage starting on day 7. * represents p < 0.0005 versus control or wild type group. Mice were euthanized on day 20 and macro-metastasis (including lungs, lymph nodes, and peritoneal metastatic nodules) was scored as shown in (D). N = 6–7 per group. Representatives of two independent animal studies were shown here. (E) Representative immunofluorescent image (merged with the corresponding bright-field image) of an E0771 induced primary tumor section showing αSMA-positive cells in the stroma region as compared to the tumor area. Random fields of tumor stroma area as indicated by the boxed area are analyzed and quantified for αSMA positivity. (F) Representative immunofluorescent images of E0771 tumors post-GKT treatment and Nox4 deletion. (G) The number of positive αSMA cells in stroma were quantified for 6–10 fields of view. (H) Representative immunofluorescent labeling of α SMA of 4T1 tumors post-GKT treatment. (I) The number of positive αSMA cells in stroma was quantified for 6–10 fields of view. (J) Representative immunofluorescent labeling of αSMA of 4T1-induced metastatic nodules in lungs. (K) Fluorescent intensities of αSMA staining in metastatic sites were quantified by ImageJ. (G, I, K) * represents P < 0.05 vs WT. Error bars are standard deviation of N = 3

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