The sphingosine-1-phosphate/RhoA/Rho associated kinases/myosin light chain pathway in detrusor of female rats is down-regulated in response to ovariectomy.

BACKGROUND

Dysuria is one of the main symptoms of genitourinary syndrome of menopause, which causes serious disruption to the normal life of peri-menopausal women. Studies have shown that it is related to decrease of detrusor contractile function, but the exact mechanism is still poorly understood. Previous results have suggested that the sphingosine-1-phosphate (S1P) pathway can regulate detrusor contraction, and this pathway is affected by estrogen in various tissues. However, how estrogen affects this pathway in the detrusor has not been investigated. In this study, we detected changes of the S1P/RhoA/Rho associated kinases (ROCK)/myosin light chain (MLC) pathway in the detrusor of ovariectomized rats in order to explore the underlying mechanism of dysuria during peri-menopause.

METHODS

Thirty-six female Sprague-Dawley rats were randomly divided into SHAM (sham operation), OVX (ovariectomy), and E groups (ovariectomy + estrogen), with 12 rats in each group. We obtained bladder detrusor tissues from each group and examined the mRNA and protein levels of the major components of the S1P/RhoA/ROCK/MLC pathway using quantitative real-time polymerase chain reaction and Western blotting, respectively. We also quantified the content of S1P in the detrusor using an enzyme linked immunosorbent assay. Finally, we compared results between the groups with one-way analysis of variance.

RESULTS

The components of the S1P pathway and the RhoA/ROCK/MLC pathway of the OVX group were significantly decreased, as compared with SHAM group. The percent decreases of the components in the S1P pathway were as follows: sphingosine kinase 1 (mRNA: 39%, protein: 45%) (both P < 0.05), S1P (21.73 ± 1.09 nmol/g vs. 18.86 ± 0.69 nmol/g) (P < 0.05), and S1P receptor 2/3 (S1PR2/3) (mRNA: 25%, 27%, respectively) (P < 0.05). However, the protein expression levels of S1PR2/3 and the protein and mRNA levels of SphK2 and S1PR1 did not show significant differences between groups (P > 0.05). The percent decreases of the components in the RhoA/ROCK/MLC pathway were as follows: ROCK2 (protein: 41%, mRNA: 36%) (both P < 0.05), p-MYPT1 (protein: 54%) (P < 0.05), and p-MLC20 (protein: 47%) (P < 0.05), but there were no significant differences in the mRNA and protein levels of RhoA, ROCK1, MYPT1, and MLC20 (all P > 0.05). In addition, all of the above-mentioned decreases could be reversed after estrogen supplementation (E group vs. SHAM group) (all P > 0.05).

CONCLUSION

In this study, we confirmed that ovariectomy is closely associated with the down-regulation of the S1P/RhoA/ROCK/MLC pathway in the rat detrusor, which may be one mechanism of dysuria caused by decreased contractile function of the female detrusor during peri-menopause.