| Objective: To establish the model of experimental varicocele (varicocele, VC) rat and measure the levels of androgen binding protein, inhibin B in testicular tissue and the levels of testicular testosterone in testicular tissue, in order to understand the damage to secretary function of the Sertoli cells and the influence on secretion of testosterone in testicular tissue and explore the mechanism of infertility resulted by VC.Methods:40 Sprague - Dawley male rats with 8 weeks of age (puberty), weighting 240 ~ 280g, were randomly divided into varicocele group (experimental group) and sham operation group (control group), 20 in each group.The animal model was established based on the Turners method: Anesthetize the rats using 10% hydration chloral hydrate (3.33ml/kg) by intraperitoneal injection. Cut the full abdominal wall by midline incision about 5cm, to expose the abdominal cavity. Confirm the left renal vein, left central vein of suprarenal gland, left spermatic vein. After carefully dissect renal vein, place a metal rod about 0.8 mm in diameter, which is parallelled with left renal vein, between left inferior vena cava and left adrenal vein. Then the rod is ligated together with the left renal vein using 4-0 threads. After take out the metal rod, the renal vein rapidly expands. The wound was closed with 4-0 threads. The rats in control group received only sham operation.The rats are killed 12 weeks after the operation. The diameter of spermatic vein is measured. The criteria of model is that the biggest diameter of spermatic vein is over 1mm, without obvious difference in bilateral kidney weight. Then size of the left testis is observed and recorded when it is pushed into the abdominal cavity, and then it was removed together with epididymis. After carefully cut off the epididymis and surrounding fascia, the testis was washed by cold saline, and bloted by the filter paper. Then the testis would be cut into two parts. For one part, it's white fasia was off, then it would be weighted and added into a glass homogenate containers together with phosphate buffer by 1:2 (weight:volume) ratio, and then grinded, placed in -4℃f reezer for overnight. On the next day the homogenated tissue should be centrifugated at 3,000 R.P.M. for 15min. The supernatant was freezed in -20℃freezer. The other part is fixed in 10% neutral formaldehyde solution for 24 hours, dehydrated, embedded in the paraffin.Androgen-binding protein and inhinbin B in testicular tissue are detected by immunohistochemical staining method. The testosterone concentration in testicular tissue is detected by Access Testosterone (chemiluminescence) method. The testosterone concentration of the sample is detected strictly according to the conventional process of BECKMAN COULTER DXI 800 automatic immune analyzer.The measurement data is represented by x_±s. Apply two independent sample t-test method to compare the mean. Non-parameter test is according to Wilcoxon signed-rank test method. The SPSS 13.0 software is used for statistical analysis. The two-sided test is used, and P<0.05 is considered to be statistically significant.Results:1 The results of androgen-binding protein in testicular tissue : By immunohistochemical method for detection, after 12 weeks, the immunohistochemical score (IHS) of androgen-binding protein (ABP) in left testicular tissue of experimental rats is that Median=6.00,P25=4.00,P50=6.00,P75=8.25,and the IHS of control group is that Median=6.00,P25=4.50,P50=6.00,P75=9.00. The difference is not statistically significant by Wilcoxon signed-rank test, P > 0.05.2 The results of inhibin B in testicular tissue:By immunohistochemical method for detection, after 12 weeks, the immunohistochemical score (IHS) of inhibin B (InhB) in left testicular tissue of experimental rats is that Median=5.00,P25=4.00,P50=5.00,P75=6.00, the expression of which is reduced, compared with Median=7.50,P25=4.50,P50=7.50,P75=9.00 of the control group and the difference is statistically significant by Wilcoxon signed-rank test, P <0.05.3 The results of testosterone in testicular tissue :By Access Testosterone method for detection, after 12 weeks, the testosterone level in left testicular tissue of experimental rats is 94.79±3.31nmol/L, which is obviously reduced, compared with 109.10±6.37nmol/L of control group. And the difference is statistically significant, P <0.05.4 Testicular pathological damage:The pathological changes of testicular tissue of experimental SD rats include the ecclasis of spermatic cells, which would cause serious lumen jams, the basal membrane thickening and fibrosis and hyaline denaturation; mesenchymal edema, leydig cells degeneration (cytoplasmic vacuolar degeneration); mesenchymal small artery disease (wall thickening and lumen narrowing or even blocked), small vein congestion and lymphatic expansion, however, serious pathological changes of only sertoli cells syndrome is not identified.Conclusion:1 After varicocele, the expression of androgen-binding protein in testis was not significantly reduced.2 The decreased production of inhibin B of Sertoli cells caused by varicocele can attenuate its promoting function for leydig cells.3 The decreased level of testosterone in testicular tissue caused by varicocele can affect sperms'development and maturation, which is resulted in male infertility.
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