Adult donor female mice (8–16 weeks old) with normal estrus cycles were sacrificed painlessly under the overdose of isoflurane. Uterine horns were excised and dissected into 5 × 2 mm rectangular fragments with all the layers, including myometrium, stroma, and luminal epithelium, and briefly washed with PBS after the removal of connective and fat tissues. For the pregnancy experiments, DUMs were prepared as 10 × 2 mm rectangular fragments. In the ovariectomized mouse DMT model, uterine samples were harvested from the ovariectomized mice that underwent ovariectomy 2 weeks before sacrifice and cut into 3 × 1 mm rectangular fragments.
For decellularization of mouse uteri, the collected uterine fragments were treated by SDS (Wako), as we described previously (3 (link)). Briefly, the tissue fragments from donor mice were immersed in 1% of SDS with PBS solution at room temperature. After SDS treatment, samples were washed with washing buffer containing 0.9% NaCl (WAKO), 0.05 M MgCl2/6 H2O (WAKO), 0.2 mg/ml DNase I (Roche Diagnostics), and 1% Gibco penicillin-streptomycin solution (Thermo Fisher Scientific) for 1 week at 4°C on a shaker set at frequency of 1 Hz with daily buffer exchange.
For decellularization of mouse uteri, the collected uterine fragments were treated by SDS (Wako), as we described previously (3 (link)). Briefly, the tissue fragments from donor mice were immersed in 1% of SDS with PBS solution at room temperature. After SDS treatment, samples were washed with washing buffer containing 0.9% NaCl (WAKO), 0.05 M MgCl2/6 H2O (WAKO), 0.2 mg/ml DNase I (Roche Diagnostics), and 1% Gibco penicillin-streptomycin solution (Thermo Fisher Scientific) for 1 week at 4°C on a shaker set at frequency of 1 Hz with daily buffer exchange.
