Adipose Mesenchymal Stem Cells-Derived Microvesicles Ameliorate Cyclophosphamide-Induced Cystitis via Inhibiting Pyroptosis and Ferroptosis in Rats

Aims: Cyclophosphamide (CYP) may through its toxic metabolite, acrolein, induce hemorrhagic cystitis and bladder hyperactivity. Previous studies demonstrated intra-iliac arterial administration of adipose derived mesenchymal stem cells (ADSC)-derived microvesicles with less immune response and adverse effects than ADSC itself may confer anti-oxidative stress and anti-inflammatory potential to improve bladder dysfunction. We explored whether ADSC-derived microvesicles may prevent CYP-induced bladder cystitis and overactivity. Methods: Female Wistar rats were divided into control (Con), CYP (Cy), CYP+microvesicles (CyM), and microvesicles treated control (CoM) groups. Con rats were intraperitoneally treated with saline, while the Cy rats were induced by intraperitoneally administered CYP (100 mg/kg body weight). We injected ADSC-derived microvesicles at the dosage of 15 μg/ml via intra-iliac artery to the rats with or without CYP treatment. We measured the responses of transcystometrogram, pathology, expression of muscarinic receptors (M3) and purinergic receptors (P2X7), pyroptosis related Caspase 1 and IL-1β, xCT/Gpx4 related ferroptosis by western blot in CYP-treated bladders. Wire myography of the urinary bladder was determined. Results: ADSC-derived microvesicles effectively decreased micturition frequency (overactivity), inflammation and fibrosis in CyM rats versus Cy rats. ADSC-derived microvesicles efficiently downregulated P2X7 and M3 receptor expression, Caspase 1/IL-1β mediated pyroptosis, xCT/Gpx4 regulated ferroptosis and restored Bcl-2/HO-1 mediated antioxidant defense mechanisms in CYP-induced cystitis. The pathologic results also displayed the effective reduction of bladder immune cell infiltration (inflammation) and fibrosis, and the preservation of the integrity in the urothelium by the treatment of ADSC-derived microvesicles. Conclusion: ADSC-derived microvesicles can ameliorate CYP-induced bladder overactivity, inflammation, fibrosis, ferroptosis and pyroptosis.