| This research focused on the mechanism of damage of SD rats’ testis and sperm caused by electromagnetic radiation(EMR) at the frequency of 900 MHz and the protective mechanism of Exocarpium cirti grandis(ECG) in rats’ testis and sperm damage against 900 MHz EMR. Morphological observation, tissue slice, physiological and biochemical tests were used for determining the apoptosis percentage, ROS of testis and sperm of SD rat in different groups. It indicated that 900 MHz EMR would increase the ROS, MDA, LDH, GSH-Px and apoptosis percentage and decrease TAOC, cell numbers, SOD. The results of Western and Southern blot indicated that 900 MHz EMR would increase bax、cytochrome c and caspase-3 expression and decrease bcl-2 expression. Meanwhile, the results of i TRAQ documented that 900 MHz EMR affect some metabolic pathways in SD rats’ testis. 1. Optimization in the extraction process of ECGThe powders of ECG was extracted with ethanol. The optimal extract conditions of the ratio of material to liquid(M/L ratio), concentration of ethanol, extract time and extract temperature were investigated by response surface method. The results indicated that the extraction raio was 5.52% under the conditions that M/L ratio was 1:60, concentration of ethanol was 70%, extract time was 5.5h and extract temperature was 50℃. There was significant impact of concentration of ethanol and M/L ratio in extraction ratio and no significant impact of extract time and temperature in extraction ratio. 2. Research on the damage of SD rats’ sperm against 900 MHz EMR900MHz EMR couldn’t affect the body weight, weight of epididymis, malformation rate of sperm(head and tail malformation). However, 900 MHz EMR could decrease the ratio of epididymis to body, sperm counte, TAOC, bcl-2 expression and increase the ratio of apoptosis and necrosis of sperm, ROS, MDA, bax, cytochrome c and caspase-3 expression. Athough, the results indicated that there was no impaction of 900 MHz EMR on the malformation rate of sperm in SD rats, the results of morphological observation documented that the ratio of sperm with two head, two tails and rotated tails was significantly increased. Meanwhile, the results of electron microscope indicated that the microstructure of sperm in SD rats exposed on 900 MHz electromagnetic field had been changed. All results above indicated that 900 MHz EMR would elevate ROS, which resulted in sperm apoptosis. 3. Research on the damage of SD rats’ testis against 900 MHz EMRThere was no significant impact of 900 MHz EMR on the body weight, weight of testis, ratio of testis to body and diameter of seminiferous tubule. However, 900 MHz EMR could make the seminiferous epithelia cells and spermatogenic cells fell off or necrotized, decrease the sperm counte in seminiferous tubule central cavity.900MHz EMR could increase ROS, MDA, GSH-Px and LDH, and decrease TAOC and SOD value of testis homogenate.5560 prateins were determined and 3888 proteins were quantized in proteomics experiments of SD rats’ testis. There were 378 proteins(179 proteins up-regulated and 199 proteins down-regulated) significant expressing between exposure and control groups. The results of GO analysis indicated that all the significantly expressed proteins accumulated in the pathway of synthesis in Arginine and Proline, the pathway of sterol synthesis, the pathway of PPAR, the pathway of apoptosis and the pathway of PI3K-Akt. The results of RT-QPCR about the genes related with apoptosis were identical with the results of proteomics experiments, indicating that 900 MHz EMR could elevate the apoptosis percentage of testis cells in SD rats distincly. 4. Protective effects of Exocarpium Citri grandis extraction on rat’s spermagainst 900 MHz EMRThere was no significant difference among control, control+ECG and shame exposure, indicating that the operational factors(gavage, irradiation box, et al) and gavage of ECG only could not affect the function of the rat’s reproductive system. There were significant difference between exposure and exposure+ECG groups, indicating that 900 MHz electromagnetism radiation changed the expression of apoptosis-associated genes including bcl-2, bax, cytc and cas3 through increasing the oxidant stress, and consequently resulted in the apoptosis rate of the rat’s sperm cells increasing. However, the Exocarpium Citr Grandis could protect the sperm cell development normally from oxidant stress induced by irradiation. 5. Protective effects of Exocarpium Citri grandis extraction on rat’s testisagainst 900 MHz EMRThere was no significant difference in weight of testis and body weight among the control, control+ECG, shame exposure, exposure and exposure+ECG, indicating that the operational factors(gavage, irradiation box, et al) and gavage of ECG only could not affect the body weight and weight of testis. the seminiferous epithelia cells and spermatogenic cells of SD rats in exposure group had been fell off or necrotized and the sperm counte were decreased in seminiferous tubule central cavity. However, the damage was improved significantly in exposure+ECG group, indicating that ECG could protect the damage of testis caused by 900 MHz EMR.The results of TUNEL indicated that ECG could decrease the apoptosis percentage of spermatogenic cells caused by 900 MHz EMR.There was significant difference in ROS, MDA, TAOC between exposure group and exposure+ECG group, indicating that ECG could protect the oxidative damage caused by 900 MHz EMR.Compared with SD rats in exposure group, the expression of bax、cytochrome c、caspasae3、BAD were decreased and the expression of bcl-2、Hsp90 and Akt were increased in exposure+ECG group. It is indicated that ECG could effectively inhibit the expression of genes which might promote apoptosis and promote the expression of genes which might inhibit apoptosis. |
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