Effects Of Estrogen On The Expression Of AQPs In Vaginal Tissue Of Female Rats

Objective:In recent years,female sexual dysfunction disease incidence rate is high,one of the common causes is the vagina lubrication decline,but we still unkuown the mechanism exactly. The water channel protein is a group protein family exists in the cell membrane,across a cell membrane can promote small molecules and water. At present,the expression of AQPs in vaginal tissues have been paid more attention and research. The purpose of this study was to investigate the effects of ovariectomy and estrogen replacement on the expression of AQPs in vaginal tissues of rats.Methods:Eighteen female SD rats of 8 weeks were randomly divided into groups A(4-week control),B(4-week ovariectomized),C(4-week ovariectomized and replacement). Group B and group C with 1% sodium pentobarbital intraperitoneal injection of 40 mg,the rats in the Group A only open abdominal wall,then sutured,Group B underwent a bilateral ovariectomy,Group C after surgery weekly subcutaneous injection of estradiol(250μg/kg),group A and group B was injected with vegetable oil. After 4 weeks were measured in rats weight,the level of serum estradiol,vaginal secretion,vaginal weight and the expression of AQPs in vaginal tissues by Western blot and immunohistochemistry. Use 1% pentobarbital sodium 40 mg to anesthetized the animals and made a lower abdomen and median incision. We use sterile needle to remove the bladder contents,in order to avoid the urine caused by the measurement errors. The pelvic nerve identified on the postrolateral aspect of the rectum. A Physiological electrical stimulator was placed around the vaginal/clitoral branch of the pelvic nerve. Electrical stimulation of unilateral pelvic nerve(0.8ms square-wave pulses,60 s duration,and 7V)at 16 Hz frequency was performed in these rats. Using a preweighted cotton-tipped catheter inserted into the distal vaginal cannal, we stimulated the pelvic nerve for 60 seconds and left the cotton-tipped catheter in place for 1 minute. The catheter was removed and weighted. The weight of vaginal fluid was calculated by subtracting the weight of swab from the total weight of swab with absorbed fluid. The full thickness tissue integrity of the vaginal wall was dissected,then ground to a powder and added to the cell lysate,centrifugal supernatant after 15 minutes. Western blot determination of the expression of AQP0,AQP3,AQP5,AQP6,AQP10,AQP11,AQP12 in vaginal tissue of rats. 4% paraformaldehyde in PBS fixed vaginal tissue,embedded in paraffin,sliced,dewaxing to water,dripping antibody,color,drying,dehydration,mount. The detection of AQP0,AQP3,AQP5,AQP6,AQP11 expression and localization in vaginal tissues by immunohistochemistry.Results:Serum estradiol level of group B(19.75±3.66 pg/m L) was significantly decreased as compared with group A(35.96±2.94 pg/m L) and group C(35.58±4.40 pg/m L)(P<0.05). Vaginal secretion was measured by pelvic nerve electrostimulation,group B(2.90±0.48mg)than those in group A(5.94±1.60mg)and group C(6.01±1.76mg)decreased significantly(P<0.05).The vaginal weight of group B(0.18±0.01g)were significantly lower than group A( 0.25±0.02g) and group C(0.25±0.01g).Western blot showed the expression of AQP0,AQP3, AQP5,AQP6,AQP10,AQP11,AQP12 in vaginal tissue of rats in group B than in group A and group C were obviously decreased. The immunohistochemistry results showed AQP0 is mainly expressed in the cytoplasm of the vaginal epithelium,AQP3,AQP5 and AQP6 is mainly expressed in the cytoplasm and plasma membrance of the vaginal epithelium,AQP11 is mainly expressed in the capillaries and venules of the vagina.Conclusion:The reduction of vaginal secretion in group B may be related with the decerase expression of AQP0,AQP3, AQP5,AQP6,AQP10,AQP11,AQP12. Estrogen may down regulate expression of AQP0,AQP3, AQP5,AQP6,AQP10,AQP11,AQP12 to regulate function, and then affect the vaginal lubrication function.