[Function of bladder smooth muscle autophagy in cyclophosphamide-induced cystitis].

OBJECTIVE

To explore the existence of bladder smooth muscle autophagy and its function in cyclophosphamide (CYP) -induced cystitis.

METHODS

A total of 30 Sprague-Dawley rats were randomized into control (n = 6) and experimental groups (4 h, 48 h, 72 h, 96 h, n = 24) . The experimental group received a single intraperitoneal injection of 150 mg/kg CYP in 0.6 ml saline while the control group had the same volume of saline injection. The changes of microtubule-associated protein light chain 3 (LC3), interleukin-6 (IL-6), IL-1β and tumor necrosis factor-alpha (TNF-α) proteins in bladder muscular layer were measured with Western blot. Co-location of LC3 and alpha-small muscle action (α-SMA) was detected with double-labeled immunofluorescence. Autophagic vacuoles were observed with electron microscopy. The changes in bladder inflammation were measured by hematoxylin and eosin staining.

RESULTS

Autophagy was confirmed in detrusor myocytes with electron microscope, LC3 Western blot and double-labeled immunofluorescence. As compared with the control group, the expression level of LC3 in experimental groups decreased (0.08 ± 0.01 vs 0.13 ± 0.01, P < 0.05) at 4 h and increased 1.19, 0.75 and 0.20 fold (0.29 ± 0.04 vs 0.13 ± 0.01, P < 0.05; 0.23 ± 0.03 vs 0.13 ± 0.01, P < 0.05;0.16 ± 0.03 vs 0.13 ± 0.01, P > 0.05) at 48, 72 and 96 h respectively.Electron microscopy and double-labeled immunofluorescent also supported the above change of LC3 by Western blot.In addition, as compared with the control groups, IL-6, IL-1β and TNF-α and bladder histological scores of bladder muscular layer significantly increased in experimental groups (all P < 0.05) .

CONCLUSIONS

Autophagy exists in bladder smooth muscle. In cyclophosphamide-induced cystitis, bladder smooth muscle autophagy activation may not be sufficient to protect bladder muscle layer inflammation in rats.