[Effect of Ultrasmall Gold Nanoparticles on the Murine Native Sperm Chromatin].

The effects of ultrasmall (2-3 nm) gold nanoparticles on native epididymal sperm chromatin of CBAxC57BL/6 hybrid mice and 129/IMG mice with a mutation in the DNA-polymerase iota gene were studied. It is shown that for both mouse strains after sperm incubation in a solution containing Au nanoparticles, at 23, 37 and 60 degrees C for 30 min followed by 1 hour treatment in dithiothreitol solution, a decrease in the number of nuclei with fully decondensed chromatin was observed compared with the control. Though, the manifestation of this effect in the population of 129/IMG mice mature sperm, was weaker. Also we have demonstrated that sperm of both strains that were incubated in a sol of Au nanoparticles at 60 degrees C behave differently under the action of dithiothreitol. A considerable part (-80%) of sperm of CBAxC57BL/6 hybrid mice treated with Au nanoparticles showed high resistance to the action of dithiothreitol, whereas in the case of 129/IMG mice only -30% did, and a partial or complete chromatin decondensation takes place in the remaining sperm. In general, using the method of nuclear chromatin decondensation in vitro for the native sperm, the patterns that we have identified in earlier studies on previously demembranized sperm are confirmed.

[Bovine sperm chromatin is not protected from the effects ultrasmall gold nanoparticles].

The response of ejaculated bovine spermatozoa to gold nanoparticles was studied by the standard method of nuclear chromatin decondensation in vitro. After the treatment of semen samples with a hydrosol containing gold nanoparticles with an average diameter of 3.0 nm and a concentration of 1 x 10(15) particles/mL, the ability of sperm nuclei to decondense in the presence of sodium dodecyl sulfate (SDS) and dithiothreitol (DTT) dramatically changed compared to the control. The frequencies of gametes with nondecondensed ("intact"), partially decondensed, and completely decondensed nuclei correlated as 40 : 32 : 28% and 0 : 36 : 64% in the experiment and the control, respectively. Moreover, the appearance of a sufficiently large number of gametes with destructed and almost completely destroyed nuclei was noticed in the spermatozoa treated with gold nanoparticles. This article suggests the putative mechanisms of action of ultrasmall gold nanoparticles on the structural and functional integrity of the deoxyribonucleoprotein (DNP) complex of mature male gametes.

[Effect of gold nanoparticles on mouse spermatogenesis].

The response of the mouse male germ cells exposed to gold nanoparticles (approximately 2.5 nm) was studied. Our investigation demonstrates that treatment with Au nanoparticles for four days does not impair the architecture of the spermatogenic epithelium. Cytogenetic evaluation using micronucleus assay showed that gold nanoparticles can affect the chromosomes of early primary spermatocytes. However, gold nanoparticles did not induce chromosome abnormalities in spermatogonial stem cells. Further, the cauda epididymal sperm was isolated on the 14th day after treatment and was incubated in SDS solution (Na sodium dodecyl) and then in a solution containing DTT (dithiothreitol) to induce nuclear chromatin decondensation. Observations showed that after four days of treatment of spermiogenic (postmeiotic) cells with gold nanoparticles the decondensation process had no differences from the control. On the contrary, in the experiment with the same cells and period of fixation but with a single exposure to gold nanoparticles, the number of mature gametes with totally decondensed nuclei reached 100% as opposed to 44% in the controls.

Regeneration potential of transplanted adult mouse sertoli cells.

The regeneration potential of differentiated Sertoli cells subjected to thermal treatment was studied by the method of cell transplantation. Cells from mice with artificial cryptorchism (1.5 months after fixation of the testes in the body) and after culturing (10 days, 37°C) were transplanted. Transplantation of Sertoli cells from 2-3-month-old and 2-day-old mice served as controls. The cells were transplanted into the testes of recipient mice, from which sex cells and Sertoli cells were removed by busulfan and cadmium salt treatment. Adult mouse Sertoli cells exposed to thermal treatment exhibited much higher regeneration potential than intact cells. Two months after transplantation, mature Sertoli cells subjected to thermal treatment populated the recipient testicular tubules, formed new tubules, and in some cases supported the development of sex cells similarly as immature cells from newborn mice.

[Ageing of the spermatogenesis system].

This review summarizes the data characterizing the effect of ageing on the development of male germ cells and their hereditary structures. We have studied causes of spermatogenesis reduction at late stages of ontogenesis. We have focused on age-specific changes of the structural-functional integrity of stem spermatogonial cells and their microenvironment (niche). We also examined several unique and specific features of the spermatogenic system in senescence-accelerated mutant mice (SAM), with accelerated ageing.

[Estimation of the frequencies of induced mutations in spermatogenic cells of senescence-accelerated prone mice of the SAMP1 strain].

The results obtained in this work demonstrate the dynamics of cytogenetic changes of spermatogenic cells in senescence-accelerated prone mice, strain SAMP1, after a single exposure to a chemical mutagen, dipin, at a genetically active dose of 30 mg/kg. In the time interval between days 3 and 28 the frequency of induced spermatogonial micronuclei does not significantly exceed the level of spontaneous mutagenesis. The lack of an experimental effect of micronuclei in this time interval is probably a consequence of mitotic delay and (or) of the death of a considerable part of genetically defective cells in the spermatogonial compartment. Different stages of meiosis exhibit different chemical sensibilities: the yield of round spermatids with micronuclei is maximum after treatment of early primary spermatocytes (preleptotene-leptotene stage) with dipin. The high sensibility of preleptotene and leptotene spermatocytes is confirmed by the sperm head shape abnormality assay. Chromosome damage caused by dipin in spermatogonial stem cells is irreversible, as evidenced by a sharp increase in the frequencies of spermatogonial and meiotic micronuclear aberrations within long periods after treatment. Increased genetic instability in the stem compartment does not lead to irreversible degradation of the system of development of male sex cells in senescence-accelerated SAMP1 mice.

[Response of the spermatogenic system to chemical mutagen dipin in SAMP1 senesce-accelerated mice].

Specific features of spermatogenesis were studied in senesce-accelerated mice of the line SAMP-1 after one-time injection of the chemical mutagen dipin. Quantitative and histomorphological changes in the spermatogenic epithelium proved to develop gradually. Cell loss and disorganization of spermatogenesis reached the peak as late as on days 28 and 35 after the injection. Differentiating spermatogonia manifested increased sensitivity to dipin. In prophase I of meiosis, developing spermatocytes proved to be less sensitive to the cytotoxic action of dipin at the pachytene than at the preleptotene-leptotene stages. Spermatogenesis in most seminiferous tubules was restored by day 56 after dipin treatment. At the end of the experiment (day 100), both quantitative parameters and morphological pattern of spermatogenesis did not differ significantly from those in the control. Thus, the cytotoxic action of dipin does not lead to irreversible structural disorganization of the spermatogenic epithelium in SAMP1 mice. Radioautography revealed a large proportion of highly differentiated Sertoli cells with 3H-thymidine-labeled nuclei in experimental animals. In some cases, structures resembling embryonic seminiferous tubules were revealed in the vicinity of rete testis in testis sections of experimental mice. These structures contained the cells morphologically similar to gonocytes and young Sertoli cells.

[Analysis of spermatogenesis in senescence-accelerated mice].

A comparative analysis of age-related dynamics of spermatogenesis has been performed in mutant mouse lines predisposed or resistant to accelerated senescence (SAMP1 and SAMR1 respectively). The results show that quantitative and morphohistological trends in the development of sperm cells and Sertoli cells in both lines are similar in both lines. Their comparison with data obtained in our previous studies (Zakhidov et al., 2001; Gordeeva et al., 2001) shows that sharp quantitative and qualitative changes in the structure of the spermatogenic system have occurred in senescence-accelerated mice of new generations, which confirms the fact of dynamic instability of the germinal lineage. The role of stem spermatogonial cells in restoration of spermatogenesis in animals reaching the critical age is discussed.

[Cytogenetic study of the effect of gossypol on mouse germ cells]. / Tsitogeneticheskoe issledovanie deistviia gossipola na muzhskie polovye kletki myshei.

The clastogenic and mutagenic activities of a new antifertility and antitumor agent gossypol were studied in the mouse male germ cells. Results of the present work indicate that at the doses 125 and 250 mg/kg the drug does not significantly increase frequencies of the micronuclei in the early spermatids and sperm head abnormalities. Hence, genotoxic influence can not be proposed as responsible for the antifertility effect of gossypol.

[DNP complex of the spermatogenic cells of rats with hypovitaminosis A]. / Sostoianie DNP-kompleksa spermatogennykh kletok krys pri gipovitaminoze A.

Retinol deficiency in rats was shown to cause atypical changes of DNP-complex in spermatogenic cells and a decrease in their fertility. The final stages of spermiogenesis were found to be the most sensitive: ca. 25% of testicular sperms were characterized by abnormal chromatin structure and deficiency in spermiospecific argininerich histones what might be related to a sharp decrease in arginine content in blood plasma. Similar results were obtained with rats kept on retinoic acid. Thus, retinoic acid fails to restore retinol-dependent processes involved in the formation of sperms with normal DNP-complex.

[Quantitative changes in histones during the formation of structures characteristic of the male pronucleus in mammalian spermatozoa in vitro]. / Kolichestvennye izmeneniia gistonov pri obrazovanii v spermatozoidakh mlekopitaiushchikh in vitro struktur, svoistvennykh muzhskomu pronukleusu.

Treatment of bull spermatozoa with DDC--Na/dithiothreitol results in the swelling and decondensation of nuclear chromatin. The structures formed at the final stages of decondensation are morphologically similar to the male pronucleus. Cytophotometric analysis has shown that decondensation of chromatin in the gametes in followed by quantitative changes of basic nuclear proteins. In partly--decondensed sperm nuclei the intensity of histone staining increases as a result of the appearance of extra reactive groups. In fully decondensed nuclei there remain only 54% of histones of the original haploid level. Nucleoproteins revealed in the sperm with fully dispersed chromatin must be histones of the somatic type.

[Effect of gossypol on spermatogenesis and fertility in man and mammals]. / Deistvie gossipola na spermatogenez i fertil'nost' cheloveka i mlekopitauishchikh zhivotnykh.

This review summarizes the main results of experimental and clinical studies of a new antifertile agent, gossypol, a polyphenolic compound isolated from the cotton. This compound disturb the normal course of spermatogenesis and affects, first of all, the locomotory system of mature spermatozoa of the man and some mammal species. The mechanism of gossypol action on the development of germ cells is discussed. Since the action of gossypol is reversible and it does not induce strong side effects, the possibility of its application for the control of birth rate is considered.

[Cytogenetic aberrations in the cells of liver and spermatogenetic epithelium in senescence accelerated SAMP1 and SAMR1 mice]. / Tsitogeneticheskie aberratsii v kletkakh pecheni i spermatogennogo épiteliia u uskorenno stareiushchikh myshei linii SAMP1 i SAMR1.

It was shown that during ontogenesis, the mice prone to (SAMP1) and resistant against accelerates senescence did not differ substantially in the frequency of cytogenetic aberrations in the hepatocytes and spermatogenic cells (spermatozoa and circular spermatids). These data suggest that in the mice of both lines, the processes of appearance, development, and functioning of complex biological systems, such as liver and testis, take place against the background of high genetic instability. The role of genetic instability in senescence is discussed.

[A biological model of accelerated aging. III. Histological characteristics of age-related changes of spermatogenic epithelium in mice SAM (senescence-accelerated mouse)]. / Biologicheskaia model' uskorennogo stareniia. III. Gistologicheskaia kharakteristika vozrastnykh izmenenii spermatogennogo épiteliia u myshei linii SAM (senescence-accelerated mouse).

The data characterizing the age-related morphological changes in the spermatogenic epithelium of SAMP1 (senescence-accelerated prone) and SDAMR1 (senescence-accelerated resistant) mice are presented. In many tubules, "early spermatogenesis" was accompanied by the formation of many morphologically abnormal germ cells on histological sections of the gonads of sexually immature (three-four weeks) mice of both strains. At this stage, destructive processes in the spermatogenic epithelium were more pronounced in SAMR1 mice. In sexually mature (two-three months) SAMP1 and SAMR1 mice, spermatogenesis as a whole proceeded normally. The first signs of regressive changes in the inner structure of most tubules (disintegration, detachment of spermatogenic epithelium from basal membrane) and morphology of germ cells (pycnosis, nuclear and cytoplasmic vacuolization) were found in SAMP1 mice at the age of six-seven months. In the older age groups (9-10 and 12-15 months), all types of spermatogenic cells were represented in both SAMP1 and SAMR1 mice, but most of these cells were atypical. Mitotic figures were recorded in a population of highly differentiated Sertoli cells.

[Biological model of the accelerated aging. I. Dynamics of spontaneous mutation in spermatogenesis in SAM mice (senescence-accelerated mouse)]. / Biologicheskaia model' uskorennogo stareniia. I. Temp spontannogo mutatsionnogo protsessa v spermatogeneze u myshei linii SAM (senescence-accelerated mouse).

We studied the dynamics of age-related cytogenetic changes in the developing male germ cells of mice prone to accelerated senescence (strain SAMP1) and mice resistant to accelerated senescence (strain SAMR1) by counting the spermatogonial and meiotic micronuclei and testing the defects of the spermatozoon head shape. In these animals, the accumulation of germinative mutations has a nonlinear pattern. During the entire ontogenesis of SAMP1 mice, the frequency of circular spermatids with micronuclei corresponded to the level observed upon induced mutagenesis.

[Biological model of accelerated senescence. II. Age-related changes in the number of developing male germ cells and Sertoli cells in gonads of senescence-accelerated mice]. / Biologicheskaia model' uskorennogo stareniia. II. Vozrastzavisimye izmeneniia chisla razvivaiushchikhsia muzhskikh polovykh kletok i kletok Sertoli v gonadakh myshei linii SAM (senescence-accelerated mouse).

It was shown that in immature three- to four-week-old mice prone to accelerated senescence (SAMP1 strain), the number of spermatogonia, pachytene spermatocytes, and circular spermatids exceeded that in mice resistant to accelerated senescence (SAMR1 strain) by more than two times. Differences were found in the pattern of age-related changes in the number of meiotic and postmeiotic cells in the sexually mature SAMP1 and SAMR1 mice. In the gonads of SAMP1 and SAMR1 mice, the number of Sertoli cells was unstable.

[The chromosomal sensitivity of hematopoietic cells in Xenopus laevis larvae to the action of the chemical mutagen fotrin]. / Chuvstvitel'nost' khromosom krovetvornykh kletok lichinok Xenopus laevis k deistviiu khimicheskogo mutagena fotrina.

Using the method of counting micronuclei it was shown that, in peripheral blood of larva of frog exposed to high doses of alkylating agent fotrin, the abundance of erythrocytes with signs of chromosomal aberrations significantly exceeds the level of spontaneous mutations on the 16th, 24th and 30th days of fixation but not on the 7th day following the experiment onset. In addition, with all terms of fixation under examination, an increased in comparison with norm number of binuclear erythrocytes was observed in the population of erythroid cells. Our observations combined with the data obtained by other authors confirm that amphibian hemopoietic cells can be used as a reliable test system for the detection of cyto- and genotoxic compounds as well as for the genetic monitoring of water environment.

[The modifying cytogenetic effects of gossypol and its derivatives]. / Modifitsiruiushchie tsitogeneticheskie éffekty gossipola i ego proizvodnykh.

The method of micronuclei account allowed to demonstrate protective effect of gossypol and its derivatives megacin and batriden of chromosomes of rat male germ cells from mutagenic effect of nitrosomethylurea (NMU) and, conversely, enhancement of clastogenic effect of another alkylating agent, fotrin. Strong antimutagenic effect of gossypol from NMU was also observed in external granular layer of young rat cerebellum. Possible mechanisms of antimutagenic action of this unique polyphenol of cotton plant are discussed.

[The effect of chemical mutagens on the chromatin properties of mouse spermatogenic cells]. / Vliianie khimicheskikh mutagenov na svoistva khromatina spermatogennykh kletok myshei.

The main components of chromatin (DNA and protamine-like proteins) were studied in the Dipin- and NMU-treated mouse spermatogenic cells using quantitative cytochemistry. Marked variation of DNA content and deficiency of the basic nuclear proteins have been shown in epididymal spermatozoa developed from cells, which were at the moment of treatment either at the early prophase I or at intermediate stages of spermiogenesis. These stages are, respectively, one of genetically most sensitive stages and a stage of active synthesis of spermiospecific proteins, which play an important role in condensation and stabilization of gamete chromatin. The combined use of DNAse I and Feulgen reaction allowed us to obtain new data about cytochemical properties of chromatin in the other types of mouse germ cells after treatment with chemical mutagens.

[The effect of chemical mutagens on mammalian spermatogenesis. A cytogenetic analysis]. / Vliianie khimicheskikh mutagenov na spermatogenez mlekopitaiushchikh. Tsitogeneticheskii analiz.

It was shown using the micronucleus test and estimating the defects of sperm heads that premeiotic and meiotic mammalian cells are genetically very sensitive to Dipin and nitrosomethylurea. In rats, unlike mice, the stem and differentiating spermatogonia with serious chromosomal defects are not eliminated and pass through a "sieve" of mitotic and meiotic divisions reaching the stage of round spermatids. Our observations suggest long-term preservation Dipin-induced, rather than nitrosomethylurea-induced, mutations in the stem cells of both mice and rats.