Emodin suppresses lung metastasis and CXCR4 expression in orthotopic mice model.

Spontaneous metastatic model of human liver cancer cells orthotopically implanted in nude mice. Cubes of HCCLM3_Luc tumor were implanted orthotopically into the liver of female Balb/c nude mice. Mice were euthanized when the detectable liver tumor signal was >1×10¹¹ p/s. A, At the time of necropsy, bioluminescent imaging was employed to monitor signals from lung metastasis. The lung biopsy was imaged to detect metastasis in the lungs. The color bar depicts the photon flux (p/s) emitted. # = mouse ID; D = day at necropsy. Lung and liver signals are in p/s unit. B, Comparison of the lung metastasis signals detected at time of necropsy for the untreated and emodin-treated groups. C, Tumor tissue extracts were prepared as described in Methods. 30 µg of protein was taken and analyzed by Western blot analysis with antibodies against CXCR4. The same blots were stripped and reprobed with β-actin antibody to show equal protein loading. D, Immunohistochemical analysis of representative section of tumor tissues stained with anti-CXCR4 antibody demonstrate strong cytoplasmic staining for CXCR4 in the control section and weak cytoplasmic staining in the emodin treated section E, Emodin decreases CXCR4 levels in tumor tissues. Tumor tissue homogenate was prepared as described in Methods. 150 µg of the tissue supernatant was taken for ELISA assay. Assay was performed according to manufacturer’s instructions using ELISA kit to determine the levels of CXCR4 in the control and treated group. Data is represented as Mean ± SE. * indicates p value <0.05.