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Our Scientific Research - Experiment results Human Spermatogenesis Effects of Pulsed Electromagnetic Radiation Emitted by Video display Terminals on Human Spermatogenesis M.O. North, A.M. Laverdure, J. Surbeck, and J. Tritto Presented at the 20th Annual International Conference of the IEEE (Institute of Electrical and Electronics Engineers) - EMB (Engineering in Medicine and Biology Society) Hong Kong SAR (China), October 29 - November 1, 1998 Published in the Proceedings of the 20th Annual International Conference of the IEEE-EMBS, 1998, pp. 3277-3280. © 1998 IEEE

ABSTRACT We studied a possible action of the radiation emitted by a video display terminal (VDT) on human spermatogenesis. Explants were originated from a 35-year old patient who presented a left varicocele (grade 4). Biopsies were taken off in the two organs and divided in several parts. Except for one untreated control, the other samples were cultivated for 24 hours: one sample (non-exposed) was simply cultivated without any particular treatment and the other two were exposed at 50 cm in front of a computer equipped with an experimental system in order to test its efficacy in ensuring an efficient protection against the radiation (exposed and exposed/protected). On all samples, histopathologic and meiotic studies were done. The results demonstrate a noxious effect of the pulsed electromagnetic radiation (PEMR) on human spermatogenesis. This effect causes a disorganization of the seminiferous tubules, a decrease in spermatide numbers, an increase in apoptotic cells and in pycnotic cells, as well as a typical toxic meiotic impairment (asynapsis), chromosome breakdowns, synaptonemal complex fragmentation, abnormal sex-vesicle). This toxic effect was more pronounced in the varicocele-weakened testis. In the two cases, normal and brittleness testis, presence of the experimental system ensures an efficient protection against the noxious effect of the PEMR. INTRODUCTION It is now well-known that PEMR emitted by cathode ray tubes (Television - TV - or Video Display Terminal - VDT) can exert possible action on living organisms. Among these radiation, the weak Extremely Low Frequency (ELF) were considered for a long time as innocuous to human health. Recently, some experiments related to the possible impact of ELF on living organisms were reported. In chick embryos exposed to well calibrated ELF, a significant increase in the proportion of developmental abnormalities was observed [1, 2]. Considering the worldwide proliferation of TV and VDT units, experiments were conducted to test their impact. In exposed chick embryos, an increase in the percentage of abnormal developments was observed, as well as an increase in the fetal mortality accompanied by a decrease of immunoglobulin G and corticosterone serum levels in the young chicks after porcine thyroglobulin immunization. In white rats exposed during their fetal life, a decrease in body mass was observed as well as some important behavioral changes in adult males and females. In mice fetus injected with a cytosine arabinoside (a teratogen), a high incident of mortality took place after exposure to ELF [5]. In humans, the most important epidemiological studies are related to the risk of miscarriage and birth defects in workers exposed [3, 4, 5]. In the responses addressed to the authors, critics asked for an exact measurement of the VDT weak EFL intensity. Unfortunately, in many biological systems, the actual intensities are at or below the noise limit. Consequently, it was recommended to accept the well standardized biological response to ELF [6]. All these reports are related to whole organisms. In brain tissue cultivated in vitro,pulsed ELF fields brought a frequency-dependent, field-induced enhancement of calcium-ion efflux. In human keraticocyres culativated in vitro, and exposed to ELF, an advanced differentiation at the expense of cell migration and proliferation was demonstrated. These unexhaustive results indicate that ELF emitted by VDT or TV can impact biological material. Recently, it was shown that superimposing spatially coherent electromagnetic noise inhibits field-induced abnormalities in developing chick embryos [7, 8]. So it is possible to test materials designed to protect living organisms. The intrinsic and extrinsic temperature alterations play a major role in testis physiology and male fertility [9] and their implication for genetic alterations were recently demonstrated [10]. Zorgniotti also evoked the role of environmental factors on spermatogenesis and human male fertility. These works and those on the role of biological material led us to study the possible effect of ELF on human male spermatogenesis. MATERIAL AND METHODS We used testicular samples originated from a 35-year old patient who presented a left varicocele (grade 4). Biopsies were taken off in the testes and divided in four fragments. From each testis (N-normal, V-varicocele weakened), a part of the sample (A) was immediately fixed in Bouin's solution as a control, and the other parts (B, C, D) were submitted to a touch imprint. Samples B, C, D were immersed in a standard culture medium (Ham F10) and maintained for 24 hours into thermostated incubators. Temperature was continuously measured by an AMR Type 2290-3 and a Testoterm Type 451 registering apparatus. Sample B was cultivated without any particular treatment. In the same experimental conditions, samples C and D were exposed at 50 cm in front of a computer (Philips, 14 inches, design type 7CM 5279/30T, serial numbers HD 0097 062 18 910 and HD 0097 062 18 900), but one of the two screens was equipped with an experimental system consisting of two mini-spheres filled with a rare earth oxides solution (EMF-Bioshield®, from A_Nox® Technology), in order to test its efficacy in granting an effective protection against radiation. At the end of the experiment, the explants were cut in two pieces. The first pieces were fixed as before, dehydrated, embedded in paraffin and 5µm-sectionned. The sections were mounted in a 0.2% gelatin (sigma)/0.05% chrome-alun and dried. After deparaffinization in xylene and rehydration, the slides were colored by hematoxylin-eosin or treated for apoptosis study according to the instructions of the supplier (Boehringer Mannheim). The slides with touch imprint were fixed into 70º alcohol and colored with eosine-hematoxylin. With the second pieces, air-dried meiotic preparations were done according to standard procedures [[11]. Meiotic abnormalities entered in the Class IIC of the Meiotic Abnormalities Classification (North and al., 1977). RESULTS NORMAL TESTIS Compared with the control piece (A), biopsy B showed a good survival rate after 24 hours of culture with all kinds of cells present with normal distribution rates and normal meiotic intercourse. The rate of of apoptosis remained extremely low in both (<5%). In sample C, some degenerating signals appeared. The meiotic abnormalities were typical impairment secondary to unfavorable physiological background: autosomal synaptic failure (asynapsis, breakdown, high level of sutosome-sex vesicle association), degeneration (high level of background silver deposition, fragmentation of synaptonemal complex, XY excredence). The rate of meiotic abnormalities represented 20% of a;; meiotic cells. Histopathologic degenerating signals consisted in appearance of slight seminiferous tubules modification (thickening of walls), low pycnotic cell rate, low spermatid number decreasing. The rate of apoptosis became significantly higher (20%) correlated to histopathological degenerating aspects and to meiotic abnormalities. For the D sample, meiotic, histopathological and apoptosis studies, the results were not significantly different from these of the A and B samples. VARICOCELE-WEAKENED TESTIS Samples A and B showed the same effect of varicocele on spermatogenesis: high rate of meiotic abnormalities (50% in this case), low rate of spermatid cell with rare spermatozoa, seminiferous tubule desorganization, high rate of apoptosis (50% of the cells). In sample C, the degradation of spermatogenesis was amplified with 80% of meiotic abnormalities, a high rate of seminiferous tubule desorganization, a high number of pycnotic cells, a rare spermatiods and no spermatozomo, and 90% of apoptotic cells. In sample D, we found again the same aspects as in samples A and B. Click on picture to zoom in Click on picture to zoom in Click on picture to zoom in Click on picture to zoom in DISCUSSION These results, after eliminating a possible effect of culture (comparison of samples A and B), demonstrate the noxious effect of pulsed electromagnetic radiation on human testes (sample C). This effect consists in meiotic disturbance, degenerating aspects of the cells (from apoptosis to pycnosis and cell death) and seminiferous tubules disorganizaton. Our study demonstrated also that a testis submitted to an unfavorable micro-environment (a slight increase in temperature secondary to a varicocele in our study) was more sensitive to an external agression. In both cases, normal and weakened testes, the presence of the experimental system provided an effective protection against the noxious effects of the pulsed electromagnetic radiation. This study has important and epidemiological consequences. It is well known that in men, failure in meiotic pairing and in spermiogenesis wre accompanied by a diminished production of spermatozoa which had a reduced viability. Then these men presented spontaneous reproductive failure and required the assistance of costly artificial reproductive technologies. ACKNOWLEDGEMENTS We thank the Centro Internazionale di Andrologia, Centro Diagnostico, Roma, Italy, for its technical assistance.