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Detrusor myocyte autophagy protects the bladder function via inhibiting the inflammation in cyclophosphamide-induced cystitis in rats.

Zhao J, Song Q, Wang L, Dong X, Yang X, Bai X, Song B, Damaser M, Li L - PLoS ONE (2015)

Bottom Line: The expressions of microtubule-associated protein 1 light chain 3 (LC3), p-p70s6k (the phosphorylated form of ribosomal protein S6), SOD2 (superoxide dismutase 2) in the bladder muscular layer were measured using western blot.Inflammation and oxidative stress were significantly decreased and the bladder histology and micturition function were significantly improved with rapamycin (RAPA, autophagy agonist) pre-treatment.The autophagy agonist RAPA significantly decreased the inflammation and protected the bladder function, which might be considered as a potential treatment for interstitial cystitis.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Second Affiliated Hospital, Third Military Medical University, Chongqing, China.

ABSTRACT
Autophagy, a highly conserved homeostatic cellular process that removes and recycles damaged proteins and organelles in response to cellular stress, is believed to play a crucial role in the immune response and inflammation. The role of autophagy in bladder cystitis, however, has not well been clarified. Here we investigate the role of detrusor myocytes autophagy (DMA) in cyclophosphamide-induced cystitis animal model. 164 female Sprague-Dawley rats were randomized into three experimental groups and compared to three control groups, respectively. The expressions of microtubule-associated protein 1 light chain 3 (LC3), p-p70s6k (the phosphorylated form of ribosomal protein S6), SOD2 (superoxide dismutase 2) in the bladder muscular layer were measured using western blot. The co-location of LC3, alpha-smooth muscle actin (α-SMA), and autophagic vacuoles were investigated with double-labeled immunofluorescence and transmission electron microscopy (TEM). The expression of lL-1β, IL-6, IL-8, malondialdehyde (MDA), and glutathione (GSH) in the detrusor layer were analyzed using ELISA. The bladder inflammation and the number of mast cells in the muscular layer were analyzed by histology. The bladder function was evaluated using cystometry. In cyclophosphamide-induced cystitis, autophagy was detected in detrusor myocytes by increased LC3, p-p70s6k expression, and autophagosomes. However, the presence of enhanced inflammation and oxidative stress in the cyclophosphamide-treated group suggest autophagy of detrusor myocytes may not be sufficiently activated. Inflammation and oxidative stress were significantly decreased and the bladder histology and micturition function were significantly improved with rapamycin (RAPA, autophagy agonist) pre-treatment. In contrast, inflammation and oxidative stress were dramatically increased and the bladder histology and function were negatively affected with chloroquine (CQ, autophagy blocker) pre-treated. These findings preferentially provide evidence of the association between DMA and cyclophosphamide-induced cystitis in rats. The autophagy agonist RAPA significantly decreased the inflammation and protected the bladder function, which might be considered as a potential treatment for interstitial cystitis.

No MeSH data available.


Related in: MedlinePlus

Pathology and inflammation scores in CYP-treated rats using histological evaluation.(A) Representative bladder showing pathologic changes in rat specimens. Staining with hematoxylin and eosin; arrows demonstrate inflammatory cells, (scale bars, 50μm). Inset images show mucosal change. (B) The statistical chart demonstrates the inflammation grading in control, CYP, CYP+RAPA and CYP+CQ treatment, n = 7. Data are expressed as the mean ± SD, a indicates a significant difference (P ˂0.01) from the control value at each time point. b indicates a significant difference (P <0.01) from the CYP group value at each time point.
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pone.0122597.g006: Pathology and inflammation scores in CYP-treated rats using histological evaluation.(A) Representative bladder showing pathologic changes in rat specimens. Staining with hematoxylin and eosin; arrows demonstrate inflammatory cells, (scale bars, 50μm). Inset images show mucosal change. (B) The statistical chart demonstrates the inflammation grading in control, CYP, CYP+RAPA and CYP+CQ treatment, n = 7. Data are expressed as the mean ± SD, a indicates a significant difference (P ˂0.01) from the control value at each time point. b indicates a significant difference (P <0.01) from the CYP group value at each time point.

Mentions: Compared with the control group (Figs 6A and 7A), bladder tissue form CYP-treated rats indicated mssive ulcers, obvious edema and hemorrhage, and increased inflammatory cell infiltration (particularly mast cell) in the submucosal and muscular layer. This trend is more obvious in the CYP+CQ group. However, in the CYP+RAPA group, bladder tissue indicated less mssive ulcers, edema and hemorrhage, and inflammatory cell infiltration (particularly mast cell) in the submucosal and muscular layer. Additionally, the quantitative assessment of histological score and mast cell count (Figs 6B and 7B) demonstrated that the inflammatory response was more severe in the CYP+CQ group, but more varied in the CYP+RAPA group compared to the CYP and control groups.


Detrusor myocyte autophagy protects the bladder function via inhibiting the inflammation in cyclophosphamide-induced cystitis in rats.

Zhao J, Song Q, Wang L, Dong X, Yang X, Bai X, Song B, Damaser M, Li L - PLoS ONE (2015)

Pathology and inflammation scores in CYP-treated rats using histological evaluation.(A) Representative bladder showing pathologic changes in rat specimens. Staining with hematoxylin and eosin; arrows demonstrate inflammatory cells, (scale bars, 50μm). Inset images show mucosal change. (B) The statistical chart demonstrates the inflammation grading in control, CYP, CYP+RAPA and CYP+CQ treatment, n = 7. Data are expressed as the mean ± SD, a indicates a significant difference (P ˂0.01) from the control value at each time point. b indicates a significant difference (P <0.01) from the CYP group value at each time point.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4382282&req=5

pone.0122597.g006: Pathology and inflammation scores in CYP-treated rats using histological evaluation.(A) Representative bladder showing pathologic changes in rat specimens. Staining with hematoxylin and eosin; arrows demonstrate inflammatory cells, (scale bars, 50μm). Inset images show mucosal change. (B) The statistical chart demonstrates the inflammation grading in control, CYP, CYP+RAPA and CYP+CQ treatment, n = 7. Data are expressed as the mean ± SD, a indicates a significant difference (P ˂0.01) from the control value at each time point. b indicates a significant difference (P <0.01) from the CYP group value at each time point.
Mentions: Compared with the control group (Figs 6A and 7A), bladder tissue form CYP-treated rats indicated mssive ulcers, obvious edema and hemorrhage, and increased inflammatory cell infiltration (particularly mast cell) in the submucosal and muscular layer. This trend is more obvious in the CYP+CQ group. However, in the CYP+RAPA group, bladder tissue indicated less mssive ulcers, edema and hemorrhage, and inflammatory cell infiltration (particularly mast cell) in the submucosal and muscular layer. Additionally, the quantitative assessment of histological score and mast cell count (Figs 6B and 7B) demonstrated that the inflammatory response was more severe in the CYP+CQ group, but more varied in the CYP+RAPA group compared to the CYP and control groups.

Bottom Line: The expressions of microtubule-associated protein 1 light chain 3 (LC3), p-p70s6k (the phosphorylated form of ribosomal protein S6), SOD2 (superoxide dismutase 2) in the bladder muscular layer were measured using western blot.Inflammation and oxidative stress were significantly decreased and the bladder histology and micturition function were significantly improved with rapamycin (RAPA, autophagy agonist) pre-treatment.The autophagy agonist RAPA significantly decreased the inflammation and protected the bladder function, which might be considered as a potential treatment for interstitial cystitis.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Second Affiliated Hospital, Third Military Medical University, Chongqing, China.

ABSTRACT
Autophagy, a highly conserved homeostatic cellular process that removes and recycles damaged proteins and organelles in response to cellular stress, is believed to play a crucial role in the immune response and inflammation. The role of autophagy in bladder cystitis, however, has not well been clarified. Here we investigate the role of detrusor myocytes autophagy (DMA) in cyclophosphamide-induced cystitis animal model. 164 female Sprague-Dawley rats were randomized into three experimental groups and compared to three control groups, respectively. The expressions of microtubule-associated protein 1 light chain 3 (LC3), p-p70s6k (the phosphorylated form of ribosomal protein S6), SOD2 (superoxide dismutase 2) in the bladder muscular layer were measured using western blot. The co-location of LC3, alpha-smooth muscle actin (α-SMA), and autophagic vacuoles were investigated with double-labeled immunofluorescence and transmission electron microscopy (TEM). The expression of lL-1β, IL-6, IL-8, malondialdehyde (MDA), and glutathione (GSH) in the detrusor layer were analyzed using ELISA. The bladder inflammation and the number of mast cells in the muscular layer were analyzed by histology. The bladder function was evaluated using cystometry. In cyclophosphamide-induced cystitis, autophagy was detected in detrusor myocytes by increased LC3, p-p70s6k expression, and autophagosomes. However, the presence of enhanced inflammation and oxidative stress in the cyclophosphamide-treated group suggest autophagy of detrusor myocytes may not be sufficiently activated. Inflammation and oxidative stress were significantly decreased and the bladder histology and micturition function were significantly improved with rapamycin (RAPA, autophagy agonist) pre-treatment. In contrast, inflammation and oxidative stress were dramatically increased and the bladder histology and function were negatively affected with chloroquine (CQ, autophagy blocker) pre-treated. These findings preferentially provide evidence of the association between DMA and cyclophosphamide-induced cystitis in rats. The autophagy agonist RAPA significantly decreased the inflammation and protected the bladder function, which might be considered as a potential treatment for interstitial cystitis.

No MeSH data available.


Related in: MedlinePlus