| Objective To observe the protective effects of Fluvastatin on diabetic testis injury and explore the possible mechanism on the reproductive damage in diabetic male rats model.Methods 86 SPF Male SD rats were used in the experiment , 10 rats were taken as normal control group, the others were induced to diabetic model by injected STZ (65mg/kg) in intraperitoneal. The successful model rats were divided into diabetic group (DM), fluvastatin (6,18 mg / kg) group and insulin group according to their blood glucose and body weights. Fluvastatin groups were fed one time per day, the normal control group and diabetic model group were given equal volume of saline, and the insulin group was given protamine zinc insulin 2U·kg once a day in subcutaneously for 8 weeks. Animals were sacrificed after 8 weeks, the specimens of serum and testicular tissue from the rats were collected, the serum testosterone (T) and MDA, SOD, GSH-Px in testicular tissue were detected. The HE staining ,TUNEL staining and immunohistochemistry (Bcl-2 , Bax) indicators were observed under the light microscope, and the ultrastructural changes of testicular tissue were observed by transmission electron microscope.Results1.Effects of fluvastatin on the general situation of diabetic rats: during the experimental period, diabetic rats group showed typical symptoms of the complications of diabetes and higher mortality (41.6%). Compare to the diabetic group, the response of rats to the outside world was more sensitive, and with lesser complications and death rate (36.4% and 27.3%) in each dose of fluvastatin group. In insulin group, the general situation of rats significantly improved, and with lesser death ( 27.3%). 2. Effects of fluvastatin on blood glucose and body weight: after 8 weeks of drug intervention, fasting blood glucose level was significantly higher in the diabetic group than the normal control group (P <0.01); fasting blood glucose level in fluvastatin groups had different extent of decline compared with diabetic group (P <0.01); fasting blood glucose level had no significant difference in insulin group and normal control group (P> 0.05). Compared with normal control group, body weight reduced significantly in diabetic model rats (P <0.01); in each group of fluvastatin and diabetic rats there was no significant difference in body weight (P> 0.05); body weight of insulin group was significantly increased (P <0.01), but still less than the normal group (P <0.01).3. Effects of fluvastatin on diabetic rats testis mass and testicular coefficient: the quality and the coefficient of diabetic model rat testis were significantly lower than the normal control group (P <0.01); testis mass and coefficient in fluvastatin groups were higher than the diabetic model group and 18mg/kg group was significantly different with the diabetic mode group (P <0.01); testis mass was lower in insulin group than normal control group (P <0.01), but there were no significant difference in testis coefficient (P> 0.05).4. Effects of fluvastatin on sperm parameters in diabetic rats: the sperm count and sperm motility in diabetic model group were significantly less than the normal control group (P <0.01); fluvastatin 6mg/kg group had no significant difference with the diabetic mode group (P> 0.05) of sperm count and sperm motility rate, fluvastatin 18mg/kg group was significantly higher than the diabetic model group (P <0.05 or P <0.01)on these, but less than insulin group (P <0.01). Insulin group's sperm count and sperm motility had significant difference with the normal group (P <0.01).5. Effects of fluvastatin on HE staining of diabetic rat's testicular tissue:under the light microscopy, the normal rat's testis seminiferous tubules were full and neatly arranged in spermatogenic cell lumen and the sperm density; the diabetic model group rats'seminiferous tubules were atrophied, germ cells were significant reduced, no sperm in most of the lumens; structure of seminiferous tubules were integrity in fluvastatin groups: mitotic phase of sperm could be seen in 6mg/kg group and some sperms could be seen in 18mg/kg group. The morphologies of spermatogenic cells and seminiferous tubules in insulin group looked like as normal group, more sperm could be seen in the seminiferous tubules.6. Effects of fluvastatin on TUNEL staining of diabetic rats'spermatogenic cells: the amount of apoptotic cells was the most in the diabetic group, and significantly higher than other groups (P <0.01). After the intervention of fluvastatin, the number of apoptotic spermatogenic cells significantly decreased compared with diabetic model group (P <0.01), and it was better in the 18mg / kg group than 6mg/kg group, but still more than insulin group; The number of apoptotic cells in insulin group was similar to the normal group (P> 0.05).7. Effects of fluvastatin on testicular tissue ultrastructure of diabetic rats under the electron microscope: in diabetic rats, Leydig cells were shrunk, cytoplasmic reduced, mitochondrial reduced and cristae dissolved, endoplasmic reticulums were expanded, lipid droplets were increased , Leydig cell nuclear pyknosis , chromatin gathered; Interstitial microvascular endothelial cell proliferated and swelled, basement membrane thickened significantly, showing that a large number of cellulose-like substance deposited. In the fluvastatin group ,organelles increased in Leydig cells ,the structure of mitochondrial cristae became more clear, endoplasmic reticulum increased, lipid droplets, the Phenomenon of Leydig cell nuclear pyknosis and Chromatin gathered better than the diabetic group; The interstitial capillary lumen patency, significantly improved in vascular endothelial cells, vascular basement membrane thickening and fibrinoid deposition was significantly reduced. And the 18mg/kg group was better than 6mg/kg group, close to the normal group.The ultrastructure of interstitial cells and interstitial microvasculars in the Insulin group were close to the normal group.8. Effects of fluvastatin on serum testosterone levels in diabetic rats: testosterone levels in diabetic rats were significantly lower than the normal rats (P <0.01). Testosterone levels were higher in fluvastatin 18mg/kg group than the diabetic model group (P <0.01), but less than insulin group. The testosterone levels of insulin group were lower than the normal group (P <0.05).9. Effects of fluvastatin on testis tissue SOD, GSH-PX activity and MDA content in diabetic rats: compared with normal rats, SOD levels were significantly lower in diabetic rats than normal rats (P <0.01), GSH-PX and MDA levels were significantly higher than normal rats (P <0.01). SOD were significantly higher in the fluvastatin groups than the diabetic group (P <0.01 or P <0.05), GSH-PX level and MDA level were decreased significantly than the diabetic group (P < 0.05 or P <0.01); The fluvastatin groups have no significant differences with the Insulin group in SOD, GSH-PX and MDA level (P> 0.05).10. Effects of fluvastatin on Bax, Bcl-2 positive immunohistochemical expression of spermatogenic cells in diabetic rats: In the normal group, Bcl-2 expression was the most and Bax expression was the least, the Bcl-2 expression was significantly decreased and Bax expression was significantly increased in diabetic group than in the other groups (P <0.01). The Bcl-2 expression in fluvastatin group was significantly increased than in the diabetic group (P <0.01), and Bax expression was significantly decreased (P <0.01)than in the diabetic group, the 18mg / kg group was better than 6mg/kg group. Insulin group was better than the fluvastatin group (P <0.01), but there was significant difference with the normal group (P <0.01).Conclusion1.Fluvastatin can reduces testicular tissue Injury of diabetic rats, it has a protective effect on testicular tissue.2.The Protective mechanism of fluvastatin on the testicular tissue in diabetic rats may related to the reduced oxidative stress and regulation of germ cell Bax, Bcl-2 positive cells... |
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