Ferric nitrate enneahydrate was obtained from Wako, and nitrilotriacetic acid disodium salt was obtained from TCI (Tokyo, Japan); they were dissolved in deionized water to make 300 and 600 mM solutions, respectively. The solution of Ferric nitrate enneahydrate and nitrilotriacetic acid disodium was mixed immediately before use to make Fe‐NTA solution, with a ratio of 1:2 (v/v); the pH was adjusted to 7.4 with sodium carbonate (Wako). Finally, the Fe‐NTA solution was diluted 10 times with deionized water. All the chemicals used were of analytical grade.
From 4 weeks of age, 58 male wild‐type A/J mice were maintained under a standard diet at the laboratory animal facility of Nagoya University Graduate School of Medicine. All the mice were injected i.p. with Fe‐NTA, with a dose of 5 mg iron/kg five times a week for the first 2 weeks, and injected with a dose of 7 mg iron/kg five times a week during the next 10 weeks. The mice that were found dying were killed. The fresh kidney tissues were either fixed in 10% PBS for H&E staining/immunohistochemistry or frozen at −80°C for other analyses.
From 4 weeks of age, 58 male wild‐type A/J mice were maintained under a standard diet at the laboratory animal facility of Nagoya University Graduate School of Medicine. All the mice were injected i.p. with Fe‐NTA, with a dose of 5 mg iron/kg five times a week for the first 2 weeks, and injected with a dose of 7 mg iron/kg five times a week during the next 10 weeks. The mice that were found dying were killed. The fresh kidney tissues were either fixed in 10% PBS for H&E staining/immunohistochemistry or frozen at −80°C for other analyses.