Mouse model of cachexia
Six-week-old male BALB/C mice (obtained from the Model Animal Research Center of Nanjing University) were maintained under specific pathogen-free (SPF) conditions and allowed to acclimate for one week. Mice were housed in individually ventilated cages (IVCs) with free access to drinking water and a basal diet under controlled conditions of humidity, light (12 h light/12 h dark cycle), and temperature.
To induce the cachexia mouse model, 100 μl C26 murine adenocarcinoma cells were subcutaneously inoculated into the right flanks of BALB/C mice (5×105 cells/mouse). When the mice lost 20% of their body weight or their tumor volumes reached 1500 mm3, the mice were sacrificed. Tumors, muscles, and other organs, such as the liver and hypothalamus, were rapidly dissected, frozen in liquid nitrogen and stored at -80°C until further analyses or fixed with 10% formaldehyde for immunohistological staining. All experiments involving animals were conducted according to protocols approved by the Institutional Animal Care and Use Committee of Nanjing University, in accordance with the guidelines for animal care and use of Jiangsu Province.