Bat7001h

For cold exposure experiments shown in Figures 1B and 1D, 14-week-old male C57BL/6N mice (SLC, Japan) were housed individually and exposed to 10°C with ad libitum food and water. The mice were anesthetized with isoflurane at the indicated time points and dissected for further analysis. For cold exposure experiments shown in Figure 7I, 3-h-fasted 11–12-week-old male BAT KO and their littermates (Ctrl) were placed individually in cages and exposed to 4°C with free access to water. The rectal temperature was measured with a thermometer (BAT7001H; Physitemp Instruments Inc.) before exposure to cold and every hour after cold exposure. Infrared thermal images were obtained before and 3 h after cold exposure at 6°C using an infrared thermographic camera (T335; FLIR) (Figure 7J). For the CL316,243 injection experiment (Figure 7K), 11–12-week-old male BAT KO mice and Ctrl mice were anesthetized with isoflurane at concentrations of 1.5%, were placed on a heat block set at 38.5°C. In addition, mice were subcutaneously administered saline or CL316,243 (1 mg/kg; Sigma-Aldrich) after a stable rectal temperature was confirmed; then, rectal temperature was monitored every minute using a thermometer (BAT7001H; Physitemp Instruments Inc.).

For in vitro tumor therapy, HepG2 cells (5 × 10⁴ cells per well) were cultured in 96-well plates for 24 h. Then various concentrations of utReS₂, RSV, and utReS₂@RSV–FA were added into the cell wells and incubated with the cells for an extra 24 h. The cell viability was detected by CCK-8 assay as mentioned above. For in vitro PTT, various concentrations of utReS₂, RSV, and utReS₂@RSV–FA treated cells were irradiated using the 808 nm laser (1 W cm⁻²) for 5 min. The real-time temperature and thermal images (once every 30 seconds) of the cells were first recorded by the thermocouple thermometer (BAT-7001H, Physitemp, USA) and infrared thermal camera (TI25, FLUKE, USA), respectively. These cells were the continuously cultured for 24 h. The cell viabilities were also evaluated by the CCK-8 assay. Simultaneously, these cells were co-stained by calcein-AM/PI (Aladdin, Shanghai, China) for 30 min and then imaged using a confocal laser scanning microscope (calcein-AM E_x = 488 nm, PI E_x = 535 nm).