[Cytogenetic, kariopathological and morphological abnormalities of spermatozoa and urothelial cells in human granulocytic anaplasmosis depending on polymorphism of the gene of DNA ligase IV].

AIM: to assess the role of human granulocytic anaplasmosis (HGA) caused by Anaplasma phagocytophilum in the induction of cytogenetic damage of spermatozoa and karyopathological abnormalities of urothelial cells depending on the polymorphism of the gene of enzyme, DNA ligase IV. MATERIAL AND METHODS: A total of 129 male patients with HGA and 84 otherwise healthy donors were examined. The samples of both semen and urothelial cells were obtained from each individuals for microscopic analysis. The diagnosis was confirmed by cytological (microscopic) method, enzyme immunoassay and polymerase chain reaction (PCR). An analysis of the frequencies of damaged spermatozoa and urothelial cells in all participants was carried out. In addition, a molecular cytogenetic study of spermatozoa by fluorescent in situ hybridization (FISH) was carried out using an AneuVysion multicolour for chromosomes 18 and 21 ("Abbott", USA) to determine the frequency of aneuploidy in spermatozoa. The level of DNA fragmentation was studied by SCD (Sperm Chromatin Dispertion Test) using a commercial Halosperm kit ("Halotech DNA", Spain). RESULTS: The cytological analysis revealed the significant increase in the proportion of spermatozoa with cytogenetic abnormalities and urothelial cells with karyopathological damage in the HGA patients. The most significant damage to nuclear structures of cells was determined in the patients with Ile/Ile genotype. The significant effects of HGA in DNA damage and cytogenetic abnormalities in patients were verified by the increased frequency of spermatozoa with DNA fragmentation, monosomy and disomy in 21 and 18 chromosomes, as well as the appearance of urothelial cells with karyopathological abnormalities. In addition, the increased frequencies of pathospermia with pathological abnormalities of head, neck and tail of spermatozoa in HGA patients were found. CONCLUSION: According to our results, the cytological analysis in the patients with HGA demonstrated the significant increase in the frequencies of spermatozoa with head, neck and tail defects and DNA fragmentation, monosomy and trisomy of the 18th and 21st chromosomes, as well as the increase in the frequencies of urothelial cells with karyopathological abnormalities. The genetic polymorphism of the effects of HGA was revealed, and the most significant cytogenetical damage was found in the patients carrying the Ile/Ile genotype of the LIG4 Thr9Ile gene.

[Genetic polymorphism of cytogenetic effects of radon bath therapy in the patients presenting with gonosteoarthritis of different etiology based at the Belokurikha health resort]. / Geneticheskii polimorfizm tsitogeneticheskikh éffektov bal'neoterapii radonovymi vannami kurorta Belokurikha pri gonosteoartritakh razlichnoi étiologii.

BACKGROUND: Borrelia infection is known to develop as a result of the bites of the ixodid ticks while the extremely difficult working conditions and low temperatures contribute to the development of chronic gonosteoarthritis (CGA) associated with ixodid tick-borne borreliosis (ITB) in the residents of the northern part of Siberia. AIM: The objective of the present study was to elucidate the frequency of occurrence of the synovial cells in the knee joints exhibiting the cytogenetic abnormalities with special reference to trisomy of chromosome 7 and the glutathione-S-transferase (GSTM1) gene polymorphism in the residents of the northern part of Siberia presenting with Lyme borreliosis (LB) whose genotype contains some alleles of the GSTM1 gene before and after the treatment of LB-associated CGA with the use of the radon bath therapy. The secondary objective was to compare the respective characteristics and the results of the treatment with those in the patients with post-traumatic gonosteoarthritis (PTGA) and to evaluate the influence of the above abnormalities on the severity of CGA. MATERIAL AND METHODS: The study included 69 patients having gonosteoarthritis associated with ixodid tick-borne borreliosis and residing in the northern areas of Siberia; in addition, 52 other individuals presenting with post-traumatic CGA made up the control group. The synovial cells of the knee joints were obtained from all the participants of the study for their culturing, making preparations, and analyzing chromosomal aberrations. The polymorphism of the GSTM1 gene was investigated with the use of the real-time polymerase chain reaction. RESULTS: The frequency of cytogenetic abnormalities in the synovial cells obtained from the group of patients presenting with LB-associated CGA who were the carriers of the homozygous mutant genotype GSTM1(0/0) was significantly higher than that in the patients having the normal genotype GSTM1(+) before the course of radon bath therapy. This treatment resulted in a significant decrease of the severity of pain syndrome and the improvement of the locomotor function of the affected joints in all groups of the patients involved in the study. The correlation analysis has demonstrated the significant positive correlation between the decrease in the frequency of occurrence of the synovial cells with trisomy of chromosome 7 and the severity of the pain syndrome. CONCLUSIONS: The results of the present study give evidence of the significant association between the genetic polymorphism of the glutathione-S-transferase gene and the frequency of cytogenetic abnormalities in the synovial cells of the knee joints of the patients suffering from CGA associated with LB. The treatment based on radon bath therapy caused a considerable decrease in the frequency of synovial cells with trisomy of chromosome 7 which correlated with the reduction of the intensity of the pain syndrome in these patients.

[Age-related traits of cytogenetic abnormalities in synovial cells of knee-joint in residents of the North of Siberia with arthritis of different etiology depending on the GSTM1 gene polymorphism of glutathion-S-transferase.]

It was studied the frequency of cells with cytogenetic abnormalities in the synovial fluid cells of the knee joint in patients of different age groups suffering from chronic arthritis associated with Lyme borreliosis (CAALB) or post-traumatic arthritis (PTA), depending on the polymorphism of the GSTM1 gene of glutathione-S-transferase. The study included 135 residents of the north of the Tomsk and Tyumen regions, 68 of whom suffered from CAALB, and the rest of the 67 patients who made up the control group were diagnosed with PTA. The results of this study have demonstrated that there are significant age-related differences in the frequency of cytogenetic abnormalities of the synovial fluid cells of the knee joint between young and elderly patients of СAALB. The integrative assessment of clinical and cytogenetic parameters in the group of elderly СAALB patients with mutant GSTM1 (0/0) allele, as compared with the other groups, enable to conclude that there are significant positive correlations between the indices of the severity disruption of articular locomotor function and the frequency of synovial fluid cells with trisomy of chromosome 7.

[Hygienic environmental assessment in the oil-and-gas bearing area on the base of cytogenetical and molecular-genetic methods].

The study have been conducted in settlements located near oilfields of the Nizhnevartovsk area, the Khanty-Mansi autonomous district (Russian Federation). There were examined 802 persons aged of from 18 to 56 years not proximately employed in processes of the oil extraction. Control group was consisted of329 residents of the north of Tomsk Region living in the area without any polluting environment industry. By using such methods of analysis as micronucleus test in human buccal cells, the xenobiotic biotransformation of both GSTM1 and GSTT1 gene polymorphism, as well as the assessment of oil contamination of local drinking water there was executed the hygienic assessment of ecology in the settlements located near oil fields. The elevated rate of cytogenetic disorders was established to be observed most of all in the residents of this region, as well as in persons recently moved to this area. Most significant deviations from the control according to the micronucleus test were detected in individuals with the GSTM1 (0) /GSTT1(0) genotype. In the control group no such consistent pattern was seen.

[Teratozoospermia and karyopathological abnormalities in epithelial urogenital tract cells in male shift workers with varicocele in oil fields of the north of Siberia].

THE PURPOSE OF THE RESEARCH: The aim of this work was to study associations between varicocele and the various types of karyopathologic abnormalities in epithelial urogenital tract cells or teratozoospermia in oil field shift workers of the north of Siberia. MATERIALS AND METHODS: We examined 139 male individuals in several oil field of the north of Tomsk and Tyumen regions. The examined individuals were divided in 4 groups: 1) driller-workers with varicocele; 2) driller-workers without varicocele; 3) oil field administrative staff with varicocele, and 4) oil field administrative staff without varicocele. Samples of both the sperm ejaculate and the epithelial urogenital tract cells were obtained from all the individuals for microscopic assay. RESULTS: It was found that the frequencies of both the teratozoospermia and the various karyopathologic abnormalities in epithelial urogenital cells were significantly higher in the workers with varicocele compared to all the other groups. There were correlations between karyopathologic abnormalities in epithelial urogenital cells and teratozoospermia in both the driller-workers with varicocele and the oil field administrative staff with varicocele. However, the frequencies of damaged cells in this workers group were significantly higher than the ones in the administrative staff with varicocele, which can be associated with the genotoxic effects of work conditions at oil fields of the north of Siberia. CONCLUSIONS: From the data it was concluded that the increased frequencies of the abnormalities in the epithelial urogenital cells and the sperm in the driller-workers with varicocele may be associated with both the diseases endogenous factor and the genotoxic effects of work conditions at the oil fields.

[Age features of cytogenetic effects of spring-summer encephalitis among residents of northern western Siberia in connection with polymorphism for genes of glutathione-S-transferase].

The aim of this work was a comparative study of the effects of spring diseases cytogenetic years of tick-borne encephalitis in elderly and young age due to differences in genes of glutathione-S-transferase. Surveyed by routine cytogenetics 120 patients with tick-borne encephalitis residents North of Tomsk region. We have taken in the study persons aged 20-35 years (Group 1) and 65-85 years old (Group 2). Material for study (buccal epithelium) was taken from each subject 3-5 times: 1st-2nd day after hospitalization, in 1 week, 1, 3 and 6 months. Tick-borne encephalitis infection causes a significantly large changes in cytogenetic regimens using buccal epithelium in the elderly than in younger patients. Restoring cytogenetic norms observed in a group of young in 3 months after hospitalization, in the elderly - in 6 months. When comparing cytogenetic effects of encephalitis shows: the young patients tick-borne encephalitis level by routine cytogenetics abnormal cells was significantly higher in carriers of inactive forms of gene GSTM1 (0)/GSTT1 (0) than containing active homozygous variants of these genes. Such patterns have not been noted in a group of elderly patients.

Assessment of the relationship of chronic opisthorchiasis to Epstein-Barr virus infection as well as some cytogenetical and immunological parameters in two comparable Siberian regions.

Previous studies have shown an increase in the frequency of chromosomal abnormalities in the peripheral blood lymphocytes from opisthorchiasis patients. Some evidence suggests that there is an association between chronic opisthorchiasis and certain herpes viruses. To study the relationship of opisthorchiasis to the reactivation of Epstein-Barr virus (EBV) infection as well as the influence of opisthorchis infection on some cytogenetical and immunological parameters, we used the indirect immunofluorescence for measuring some virus specific antibodies, the cytokinesis-block lymphocyte micronucleus assay, and the quantitative immunodiffusion method for measuring immunoglobulin concentrations in serum. A total of 1,580 people were monitored in two comparable Siberian regions: in the Ob River region which is endemically related to opisthorchiasis caused by Opisthorchisfelineus and in the nonendemic control Yenisey River region. There was no significant difference in each of the tested parameters between two uninfected controls from the endemic Ob and nonendemic Yenisey regions. We have found significant difference (p < 0.01) in the frequency of micronucleated cytokinesis-block lymphocytes and the antibody levels against certain EBV antigens between the examined inhabitants of the opisthorchis-infected Ob and uninfected Yenisey regions. Furthermore, there were a good correlation (r = + 0.72) between the increase in titres of antibody to the EBV capsid antigen and the high frequency of micronucleated lymphocytes in the opisthorchis-infected Ob population. Also, levels of both IgG and IgM were increased in opisthorchiasis patients. This study confirms an association between chronic opisthrochiasis and reactivation of EBV which may be implicated in the development of cancer in opisthorchiasis patients.

Biodosimetry results obtained by various cytogenetic methods and electron spin resonance spectrometry among inhabitants of a radionuclide contaminated area around the siberian chemical plant (Tomsk-7).

On April 6, 1993, near the town of Tomsk (Russia) there was an accident at the Siberian Chemical Plant (SCP) which resulted in extensive contamination of an area of 250 km(2) to the north of SCP with long-lived radionuclides such as (239)Pu, (137)Cs and (90)Sr. Cytogenetic methods and electron spin resonance (ESR) spectrometry of tooth enamel were used to estimate the radiation doses received by the population. The ESR signal intensity and the chromosomal aberration frequency in lymphocytes of the tooth donors showed a good correlation. The data showed that 15% of the inhabitants of the Samus settlement received a radiation dose >90 cGy. The exceptions were results of an examination of fishermen, where ESR gave high values (80-210 cGy) but both the chromosome assay and the cytokinesis block micronucleus method gave lower ones (8-52 cGy). A large increase in chromosome damage was observed in people born between 1961 and 1969. It was found that during these years several serious accidents at the Siberian Chemical Plant had occurred causing radiation pollution of the area. The number of cells with chromosome aberrations was significantly less among the people arriving in Samus after 1980. We found good correlations between the level of carotene consumption and a decrease in frequency of both micronuclei in binucleated lymphocytes (r = 0.68, P < 0.01) and chromatid aberrations (r = 0.61, P < 0.01) among the inhabitants. We also examined the inhabitants of Samus for opisthorchis infection, which was present in 30% of the population. The Samus inhabitants affected by Opisthorchis felineus showed significantly increased levels of micronuclei in binucleated lymphocytes and chromatid aberrations as compared with the controls.

Chromosome breakage at sites of oncogenes in a population accidentally exposed to radioactive chemical pollution.

The purpose of the present study was to investigate the level of aberrations at fragile sites of chromosomes in peripheral blood lymphocytes of the population of an area polluted with radionuclides, following an accident at the Siberian Chemical Plant. We carried out the micronucleus test to screen people with radiation-related cytogenetic effects. Of the 1246 inhabitants of the settlement of Samus examined, 148 showed a significantly increased frequency of micronucleated erythrocytes and were selected for chromosome analysis as a radiation-exposed group. Additional analysis was carried out on 40 patients with gastric cancer and atrophic gastritis with stage II-III epithelial dysplasia. Eighty six individuals from a non-polluted area were used as a control group. Chromosomal breaks and exchanges occurred preferentially in chromosomes 3 and 6 among radiation-exposed persons and patients. The regions 3p14-25 and 6p23 were damaged most often. There was a tendency to preferential involvement of q21-25 of chromosome 6 in patients with gastric cancer and atrophic gastritis. Specific damage at certain chromosome sites was observed in the radiation-exposed population as well as in patients with gastric cancer. Most often this damage was located near oncogene loci, which could imply that chromosome damage induced by radiation is likely to be a predisposing factor to the expression of oncogenes and malignant transformation of cells in exposed individuals.

Micronucleated erythrocytes frequency and radiocesium bioconcentration in pikes (Esox lucius) caught in the Tom River near the nuclear facilities of the Siberian Chemical Complex (Tomsk-7).

Between 1957 and 1993, the Siberian Chemical Complex (Tomsk-7) located in the Tomsk region (Russia) regularly discharged radioactive liquid wastes into the Tom River which resulted in an extensive contamination of large territories with long lived radionuclides such as cesium-137 and strontium-90. In the summers of 1996 and 1997, Research Team of Siberian Medical University conducted biodosimetry and cytogenetic monitoring of pikes (Esox lucius) caught in the Tom River at various distances downstream from the Siberian Chemical Complex (SCC) using the micronucleus test and the gamma spectroscopy. Our findings demonstrated that the difference in frequency of micronucleated erythrocytes between the radiation-exposed fish caught downstream from the SCC and the controls was statistically significant (P < 0.01). Moreover, we found a good correlation between radiocesium concentration and micronucleated erythrocyte frequency in pikes. It was found that both the micronucleated erythrocyte frequency in pike blood and the level of the pike radiocesium concentration depended on the age of pikes. The micronucleated erythrocyte frequency gradually increased from the 1-year-old pikes to the over 20-year-old pikes. On the other hand, the average level of radiocesium concentration gradually increased from the 1-year-old pikes only up to the 10-year-old pikes. There is no correlation between radiocesium concentration and micronucleated erythrocyte frequency in the over 10-year-old pikes.

Estimation of the mutagenic potential of the trematode Opisthorchis felineus in experimentally infected guinea pigs.

We studied the mutagenic potential of the trematode Opisthorchis felineus in experimentally infected guinea pigs by chromosome analysis. The investigation was conducted on 30 treated (experimentally infected) and 20 normal (uninfected) guinea pigs (Cavia porcellus). A sample of living metacercariae of O. felineus was obtained from the muscle of naturally infected crucian carp captured in the Tom River, and 1 ml suspension containing approximately 50 metacercariae in 0.9% NaCl solution was individually given p.o. to guinea pigs that were used as treated specimens, whereas normal guinea pigs from the same stock were given 1 ml 0.9% NaCl solution per individual to serve as controls. The treated and control animals were killed by CO2 asphyxiation and decapitation at 7, 15, 30, 45, and 120 days after contamination. The smears of bone marrow cells for chromosome analysis were prepared by a standard method. Statistical analysis of the data was done using Student's t-test. We found that experimental infection of guinea pigs with O. felineus induced significantly high frequencies of hypoploid cells and, at the 120th day p.i., led to a considerable incidence of chromatid breaks and polyploidy in bone marrow cells.

Frequencies of micronucleated lymphocytes and Epstein-Barr virus contamination in Altai region residents living near the Semipalatinsk atomic testing ground.

We assessed the frequencies of micronucleated lymphocytes in 3,036 individuals living in 16 settlements in the western part of the Altai region. The majority of individuals with significantly high frequencies of micronucleated lymphocytes were detected in settlements adjacent to the Semipalatinsk atomic testing ground (SATG). The most considerable genome instability was found in individuals born in the period of intensive testing at the SATG (1949 to 1962). Moreover, we determined that the residents of the settlements adjacent to the SATG have significantly high levels of antibodies to potentially oncogenic Epstein-Barr virus, in addition to high frequencies of micronucleated lymphocytes. The considerable Epstein-Barr virus contamination among the residents in the radiation-polluted zone around the SATG was supposed to be caused by immunodeficiency disorders in these individuals and was correlated with high frequencies of micronucleated cells.

Micronucleus test of erythrocytes and lymphocytes in the blood of the Altai region residents living near the Semipalatinsk atomic proving ground.

We analysed the erythrocytes and lymphocytes with micronuclei of 3902 individuals living in 16 settlements in the west of the Altai region. It was found that the majority of individuals with high frequencies of micronucleated cells were in the settlements located near to the Semipalatinsk atomic proving ground. In particular, considerable cytogenetic alterations were found in individuals born during the period of intensive testing on the Semipalatinsk proving ground (from 1949 to 1962). The data we have obtained allow us to conclude that radiation damage of cytogenetic structures in the prenatal (predelivery) and early postnatal periods leading to the formation of micronuclei can remain in the human organism for decades and, perhaps, throughout life.

An assessment of frequencies of micronucleated erythrocytes in peripheral blood of pigeons (Columba livia Gm) living in the polluted radioactive area around the Siberian chemical plant.

We have examined approximately 15,000 pigeons living in the radioactive area polluted by releases of the Siberian Chemical Plant (SChP) as a result of an accident in April 1993. We have detected that almost all of the population of pigeons have a high frequency of micronucleated erythrocytes in their peripheral blood. Moreover, we have found that pigeons living in the downwind cities and towns situated closest to the SChP, particularly in the northern districts of the city of Tomsk and in the closed-city of Tomsk-7 (Seversk), have a significantly high frequency of micronucleated erythrocytes. The assessment of the situation in other Siberian cities has shown that frequencies of micronucleated erythrocytes in pigeons living in Tomsk and Seversk are significantly higher than those in pigeons in the control cities. After the radiation accident the highest frequencies of micronucleated erythrocytes was detected after six months and it gradually decreased over three years but there has been no full normalization. However, full normalization in frequencies of micronucleated erythrocytes in pigeons placed in a vivarium occurred within six months. We have found that frequencies of micronucleated erythrocytes are significantly higher in female pigeons than in males.

Micronucleus test of erythrocytes and lymphocytes in the blood of the people living in the radiation pollution zone as a result of the accident at the Siberian chemical plant on April 6, 1993.

We have carried out analysis of the number of blood erythrocytes and lymphocytes with micronuclei in the inhabitants of four settlements located near the place of the accident which happened at the atomic power station of the Siberian chemical plant (Tomsk-7) on April 6, 1993. In all cases, the people examined showed a considerable increase in the number of cells with micronuclei as compared with the control. We observed the same people for 2 years and found a gradual decrease in the number of cells with micronuclei. This study shows that people born between 1963-1970 have a much higher level of cells with micronuclei, which we tend to see as a result of the radiation accident at the Siberian chemical plant in 1963. The data we have obtained allow us to conclude that penetration of radionuclides into the human organism in the prenatal and early postnatal periods can lead to the formation of stable clones of erythroid cells with micronuclei and a higher level of erythrocytes with micronuclei which can remain in the blood for a long time.

The analysis of some indices of immune response, DNA repair, and micronuclei content in cells from tick-borne encephalitis patients.

Patients with tick-borne encephalitis (TBE) had higher counts of red blood cells (RBC) with micronuclei. The majority of patients revealed decreased capacity of blood lymphoid cells for DNA repair except those with a 2-wave pattern of the course of disease; in the latter, the DNA repair was significantly higher than in healthy donors. Patients with TBE revealed lower T-lymphocyte counts due to a decrease in the amount of T-helper cells (the level of T-suppressors was elevated). The intensity of antibody production against TBE virus was significantly enhanced by termination of disease in the majority of patients. The count of natural killer cells was decreased, particularly at the initial stage of disease. At the time of admission to hospital the counts of RBC with micronuclei and of T-helper cells were in reverse proportion. At the terminal stage of disease the same correlation was noted between RBC counts with micronuclei and the antibody level. At the onset of disease a direct correlation was noted between DNA repair and B-lymphocyte and T-helper counts. At the final stage of disease the reverse correlation between the activity of DNA-repair systems and T-suppressor counts was registered. Three months after discharge from hospital, the indices of micronuclear test, natural killer cell activity, and DNA repair returned to normal.

Genome instability as a consequence of DNA-repair and immune system abnormalities.

The cellular level analysis of 720 clinically persons with cytogenetic abnormalities has been performed in this study. Of them 25% showed an increased number of cells with cytogenetic abnormalities. Repeated examination of these patients allowed to "eliminate" sporadic cases of genome instability from the given group. The remaining, 30% of cases, had reliable decrease in excision repair and significant changes in immunoreactivity. At the same time a decrease of activity of DNA-repair synthesis was noticed in donors with a higher number of cells with cytogenetic aberrations.