Effects Of Moderate Intensity Magnetic Field Exposure On Reproductive System Of Male Mice And The Mechanisms

BackgroundWith the development of science and technology,and the wide application of all kinds of electrical and electrical equipment and living electronic products,the electromagnetic radiation of different frequencies and different intensities are distributed to every corner of the earth,which increases the chance of human exposed to the magnetic fields.In the field of biomedicine,the moderate intensity magnetic field refers to a magnetic field with a magnetic induction intensity greater than 1 m T and less than or equal to 1 T.The moderate intensity magnetic field is widely used in the fields of industrial and agricultural production,transportation,medical and health care,and their biological effects have become a hot spot in academic research.The reproductive system is the basis of human reproduction.For male animals,testis is an important reproductive organ for spermatogenesis and maturation.At the same time,the testicular tissue is more sensitive to various stimulating factors,such as temperature,chemical substances,medicine and radiation,under certain conditions all these stimulating factors can affect the spermatogenesis,testicular morphology and function.At present,most studies focus on the biological effects of extremely low frequency magnetic fields and strong static magnetic fields,but the effect of moderate intensity magnetic field on reproductive system and its mechanism are seldom studied.In this paper,the male BALB/c mice were used as the research objects to study the effects of moderate intensity magnetic fields exposure on the reproductive system of male mice,starting from the aspects of sperm quality,testicular morphological structure,serum sex hormone level,testicular antioxidant capacity and spermatogenic cell apoptosis,etc.,and the related mechanism was preliminary discussed.ObjectiveTo explore the effects of moderate intensity magnetic field exposure on the reproductive system of male mice and its mechanism,and to provide a theoretical basis for revising health protection standards.Part I Effects of Moderate Intensity Magnetic Field Exposure on Reproductive System of Male Mice Materials and Methods1.Exposure equipment: The moderate intensity magnetic field generator used a large-capacity pulse capacitor to generate a large pulse current,then a magnetic field was generated through this large pulse current,with the width of 9 ms and a magnetic field intensity peak value of 0 ～ 2 T(adjustable).2.Animal groups and exposure: a total of 270 healthy adult male mice BALB/c mice(body weight: 19.8 ± 2.2 g)were randomized into sham exposure group and 5 exposure groups(10 m T group,50 m T group,100 m T group,400 m T group and 800 m T group),each group has 45 mice,and then the mice in each group were divided into 5 time points,each time point had 9 mice.The moderate intensity magnetic field generator was used to irradiate the mice in the exposure group for 1 h every day and continuously irradiate the mice for 14 days.The treatment for the mice in the sham exposure group was same as that in the exposure group,but the magnetic induction intensity of the magnetic field generator was 0 m T.3.Experimental methods: 1)The changes of the body weight of mice in each group were recorded during 14 days of continuous exposure;2)After 14 days of continuous exposure,sampling was conducted on the time points of 1 d,7 d,14 d,28 d and 60 d after exposure respectively.The bilateral testicles were exfoliated and the weight was recorded to calculate the testicular index;3)Bilateral epididymis were exfoliated,free sperm was dissociated,sperm count,deformity rate and survival rate were observed and counted under light microscope;4)The hematoxylin and eosin(HE)staining was used to observe the morphological structure of the mice testis,and the diameter of seminiferous tubules in the testes of mice were measured by the cross method.Results1.Effects of moderate intensity magnetic field exposure on the body weight and testicular indexes of male miceThe body weight of the mice in the 800 m T group began to decrease after 4 days of exposure,and the body weight of the mice 100 m T group and 400 m T group began to decrease after 7 days of exposure.Compared with the sham group,the high-intensity exposure groups decreased significantly,and the difference was statistically significant(P < 0.05).The body weight of the mice in the six groups was significantly different by statistics at 1 d,7 d,14 d and 28 d after exposure(P < 0.05).Compared with the sham group,at the time points of 1 d after exposure,the body weight of the mice in 100 m T group decreased significantly,and the difference was statistically significant(P < 0.05);At 1 d,7 d and 14 d after exposure,the body weight of mice in the 400 m T group decreased significantly,and the difference was statistically significant(P < 0.05);At 1 d,7 d,14 d and 28 d after exposure,the body weight of mice in the 800 m T group decreased significantly,and the difference was statistically significant(P < 0.05).It was suggested that in this study,magnetic field exposure with different intensity can affect the body weight increase in male mice.The testicular weight of the mice in the six groups was significantly different by statistics at 1 d,7 d,14 d and 28 d after exposure(P < 0.05).Compared with the sham group,at the time points of 1 d and 7 d after exposure,the testicular weight of the mice in 100 m T group,400 m T group and 800 m T group decreased significantly,and the difference was statistically significant(P < 0.01);At 14 d and 28 d after exposure,the testicular weight of mice in the 800 m T group decreased significantly,and the difference was statistically significant(P < 0.05).The testicular indexes of the mice in the six groups was significantly different by statistics at 1 d,7 d,14 d and 28 d after exposure(P < 0.01).Compared with the sham group,at the time points of 1 d and 7 d after exposure,the testicular indexes of the mice in 100 m T group,400 m T group and 800 m T group decreased significantly,and the difference was statistically significant(P < 0.01);At 14 d after exposure,the testicular indexes of mice in the 800 m T group decreased significantly,and the difference was statistically significant(P < 0.01);At 28 d after exposure,the testicular indexes of mice in the exposure groups decreased significantly,and the difference was statistically significant(P < 0.01).It was suggested that the magnetic field exposure with different intensity under the experimental conditions in this study can affect the testicular index of male mice.2.Effect of moderate intensity magnetic field exposure on sperm quality of male mice2.1 The results of sperm counts showed: The sperm counts of the mice in the six groups was significantly different by statistics at 1 d,7 d and 14 d after exposure(P < 0.01).Compared with the sham group,at 1 d,7 d and 14 d after exposure,the sperm counts of mice in the 400 m T group and 800 m T group decreased significantly,and the difference was statistically significant(P < 0.01),and the greater the magnetic induction intensity,the more obvious the effects.2.2 The results of sperm deformity rate test showed: The sperm deformity rate of the mice in the six groups was significantly different by statistics at 1 d,7 d,14 d and 28 d after exposure(P < 0.05).Compared with the sham group,at 1 d after exposure,the sperm deformity rate of mice in the 100 m T group,400 m T group and 800 m T group increased significantly,and the difference was statistically significant(P < 0.05);At 7 d,14 d and 28 d after exposure,the sperm deformity rate of mice in the 800 m T group increased significantly,and the difference was statistically significant(P < 0.01),and the greater the magnetic induction intensity,the more obvious the effects.2.3 The results of sperm survival rate test showed: The sperm survival rate of the mice in the six groups was significantly different by statistics at 1 d after exposure(P < 0.01).Compared with the sham group,at 1 d after exposure,the sperm deformity rate of mice in the 100 m T group,400 m T group and 800 m T group decreased significantly,and the difference was statistically significant(P < 0.05).The results mentioned above suggested that different intensity magnetic fields can damage the sperm quality of male mice under the experimental conditions in this study,and the greater the exposure intensity,the more obvious the effects.3.Effects of moderate intensity magnetic field exposure on morphological structure of testes in male mice3.1 Changes in testicular morphology: HE staining results showed that mice in the different intensity exposure groups exhibited different trends in pathological changes.Compared with the sham group,at various points in time after exposure in the 10 m T group and 50 m T group,and the seminiferous tubules were normal,the basement membrane was intact,the spermatogenic cells at all levels were arranged in an orderly manner,and the lumen was filled with mature sperms.At 1 d and 7 d after exposure,in the 100 m T group,400 m T group and 800 m T group had gradually abnormal testicular tissue morphology and structure,spermatogenic cells were arranged in a disordered way,the boundary membrane was slightly thickened,the number of spermatozoa in the lumen decreased,and mild cavitation occurred.The morphology and structure of the mice testicular in the 100 m T group,400 m T group and 800 m T group tended to be normal at 28 and 60 days after exposure.3.2 Changes in the diameters of seminiferous tubules: The diameters of seminiferous tubules of the mice in the six groups was significantly different by statistics at 1 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d after exposure,the diameters of seminiferous tubules in the 400 m T group and 800 m T group were significantly shortened and the difference was statistically significant(P < 0.05);At 7 d after exposure,the diameters of seminiferous tubules in the 800 m T group were reduced,and the difference was statistically significant and the difference was statistically significant(P < 0.05).The results mentioned above suggested that different intensity magnetic fields can change the testicular morphology and structure of male mice under the experimental conditions in this study,and the greater the radiation intensity,the more obvious the effect.Part II Mechanisms of Moderate Intensity Magnetic Field Exposure on Reproductive System of Male Mice Materials and Methods1.Exposure device: the same as the first part in this study.2.Animal grouping and exposure: the same as the first part in this study.3.Experimental methods: 1)Detection of apoptosis in seminiferous tubules by TUNEL method;2)Western blotting(WB)was used to detect the expression levels of apoptosis-related proteins(Caspase 3,BAX and BCL;3)Enzyme-linked immunosorbent assays(ELISA)were used to detect the serum testosterone(T),gonadotropin-releasing hormone(Gn RH),luteinizing hormone(LH)and follicle-stimulating hormone(FSH)levels in mice;2)ELISA was used to detect the changes of superoxide dismutase(SOD)activity,glutathione(GSH)and malondialdehyde(MDA)contents.Results1.Effects of moderate intensity magnetic field exposure on testicular cell apoptosis in male mice1.1 Spermatogenic cell apoptosis in testicular tissue: the results of TUNEL staining showed that the proportion of spermatogenic cell apoptosis of the mice in the six groups was significantly different by statistics at 1 d after exposure(P < 0.05).Compared with the sham exposure group,the proportion of spermatogenic cell apoptosis of the mice in the 100 m T group,400 m T group and 800 m T group increased significantly and the difference was statistically significant(P < 0.05).1.2 Expression of apoptosis protein in testis:WB results showed that,compared with the sham group,at 1 d,7 d,14 d,28 d,and 60 d after exposure,there was no significant change in BAX levels in the testicular tissue of mice in different intensity exposure group(P > 0.05);at 1 d and 7 d after exposure,the expression levels of BCL 2 in testicular tissues of mice in 400 m T and 800 m T groups were significantly reduced(P < 0.05);at 7 d after exposure,the expression levels of Caspase 3 in testicular tissues of 50 m T groups Significantly increased(P < 0.05),the expression level of Caspase 3 in the testis tissue of 400 m T group and 800 m T group was significantly reduced(P < 0.05).At 28 days after exposure,the testis tissue of 400 m T group and 800 m T group mice The expression level of Caspase 3 was significantly increased(P < 0.05).At 60 days after exposure,the expression level of Caspase 3 was significantly increased in the testis tissues of 100 m T and 400 m T mice(P < 0.01);at 1 d after exposure in the 800 m T group,the BAX/BCL 2 ratio of testis tissue was significantly increased(P < 0.05).At 7 days after exposure,the BAX/BCL 2 ratio in the 400 m T group and 800 m T group was significantly increased(P < 0.05).The above results suggested that moderate intensity magnetic field can increase testicular cell apoptosis in male mice.2.Effects of moderate intensity magnetic field exposure on serum sex hormones in male mice2.1 Serum T content: The ELISA results showed that the serum T content of mice in the six groups was significantly different by statistics at 1 d and 7 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d after exposure,the serum T content of mice in the 400 m T group and 800 m T group reduced significantly and the difference was statistically significant(P < 0.05);At 7 d after exposure,the serum T content of mice in the 100 m T group,400 m T group and 800 m T group reduced significantly and the difference was statistically significant(P < 0.05).2.2 Serum Gn RH content: The ELISA results showed that the serum Gn RH content of mice in the six groups was significantly different by statistics at 1 d and 7 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d and 7 d after exposure,the serum Gn RH content of mice in the 800 m T group increased significantly and the difference was statistically significant(P < 0.05);At 7 d after exposure,the serum Gn RH content of mice in the 400 m T group and 800 m T group increased significantly and the difference was statistically significant(P < 0.05).2.3 Serum LH content: The ELISA results showed that the serum LH content of mice in the six groups was significantly different by statistics at 7 d and 14 d after exposure(P < 0.05).Compared with the sham exposure group,at 7 d after exposure,the serum LH content of mice in the 50 m T group and 100 m T group increased significantly and the difference was statistically significant(P < 0.05),and 800 m T group reduced significantly and the difference was statistically significant(P < 0.01).At 14 d after exposure,the serum LH content of mice in the 50 m T group and 100 m T group increased significantly and the difference was statistically significant(P < 0.05).2.4 Serum FSH content: The ELISA results showed that the serum FSH content of mice in the six groups was significantly different by statistics at 1 d and 14 d after exposure(P < 0.01).Compared with the sham exposure group,at 1 d and 14 d after exposure,the serum FSH content of mice in the 800 m T group reduced significantly and the difference was statistically significant(P < 0.05).The above results suggested that moderate intensity magnetic field can affect the serum sex hormones in male mice.3.Effects of moderate intensity magnetic field exposure on the antioxidative capacity of testes in male mice3.1 SOD activity: The ELISA results showed that the SOD activity in the testicular tissue of mice in the six groups was significantly different by statistics at 1 d,7 d,14 d and 28 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d after exposure,the SOD activity in the testicular tissue of mice in the 400 m T group and 800 m T group reduced significantly and the difference was statistically significant(P < 0.05);At 7 d,14 d and 28 d after exposure,the SOD activity in the testicular tissue of mice in the 800 m T group reduced significantly and the difference was statistically significant(P < 0.05).3.2 MDA content: The ELISA results showed that the MDA content in the testicular tissue of mice in the six groups was significantly different by statistics at 1 d,7 d and 14 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d after exposure,the MDA content in the testicular tissue of mice in the 100 m T group,400 m T group and 800 m T group increased significantly and the difference was statistically significant(P < 0.05);At 7 d and 14 d after exposure,the MDA content in the testicular tissue of mice in the 800 m T group increased significantly and the difference was statistically significant(P < 0.05).3.3 GSH content: The ELISA results showed that the GSH content in the testicular tissue of mice in the six groups was significantly different by statistics at 1 d and 7 d after exposure(P < 0.05).Compared with the sham exposure group,at 1 d after exposure,the GSH content in the testicular tissue of mice in the 400 m T group and 800 m T group reduced significantly and the difference was statistically significant(P < 0.05);At 7 d after exposure,the GSH content in the testicular tissue of mice in the 800 m T group reduced significantly and the difference was statistically significant(P < 0.05).The above results suggested that the moderate intensity magnetic field can affect the antioxidative capacity of testes in male mice.ConclusionsModerate intensity magnetic field exposure under this experimental condition in this study:1.It can affect the weight gain of male mice and cause the change of testicular index;2.It can reduce the sperm quality of male mice,and the greater the intensity of exposure,the more obvious the effect;3.It can cause changes in morphological structure of testes in male mice,and the greater the intensity of exposure,the more obvious the effect;4.It can induce testicular cell apoptosis;5.These effects may be related to the changes of sex hormone levels,testicular antioxidant capacity,and increased testicular cell apoptosis.