H2o2 solution

An H₂O₂‐induced rat cystitis model was created as previously reported for mice (12), with minor modifications. Under 2–3% isoflurane (Pfizer, NY) anesthesia, a polyethylene tube (PE‐50; Clay‐Adams, Parsippany, NJ) was introduced into the bladder transurethrally and then the lower abdomen was pressed gently to withdraw urine. Next, 300 μL of 1.5% H₂O₂ solution (Wako Pure Chemical Industries, Osaka, Japan) suspended in sterile saline was introduced into the bladder through the catheter. The H₂O₂ solution was drained from the bladder after 30 min by pressing the lower abdomen.

GO material was prepared by improved Hummers methods^{6 (link),38 (link)} with natural graphite of Bay carbon graphite from Michigan. The graphite powder (5 g) was inserted in the mixture of 200 mL H₂SO₄ (96%, Wako Chemical Co. Ltd.) and 25 mL H₃PO₄ (85% Wako Chemical Co. Ltd.) followed by addition of 25 g KMnO₄ powder (Wako Chemical Co. Ltd.). The mixing of the chemicals with exothermic reactions was carried out very slowly under cooling with ice-water bath to control the temperature at 311 ± 2 K stirring at 200 rpm for 2 h. Then, the 500 mL of distilled water was added slowly into the mixture followed by addition of 100 mL H₂O₂ solution (10%, Wako Chemical Co. Ltd.). Then, the supernatant by centrifugation was collected and was washed with 1 M HCl (Wako Chemical Co. Ltd.) at 3 times and then washed with distilled water to reach the pH of the washed liquid to 7. Finally, we obtained GO powders by freeze-drying of the GO suspension with liquid nitrogen.

The H₂O₂-induced cystitis model was generated as previously reported (Homan et al., 2013 (link)). Briefly, under 2–3% isoflurane (Pfizer, NY) anesthesia, a polyethylene tube (PE-10; Clay-Adams, Parsippany, NJ) was introduced into the bladder transurethrally and the lower abdomen was pressed gently to withdraw urine. Next, 50 μL of 1.5% H₂O₂ solution (Wako Pure Chemical Industries, Osaka, Japan) in sterile saline was introduced into the bladder through the catheter. The H₂O₂ solution was drained from the bladder after 30 min by pressing the lower abdomen.