CC BY
13
Section 11. Medicine
Agzamova Gulnara, applicant for DSc, Department of Faculty and Hospital Therapeutics N1, assistant professor, Tashkent Medical Academy E-mail: [email protected] E-mail: [email protected]
FEATURES OF CYTOGENETIC CHANGES (CHROMOSOMAL ABERRATIONS) IN THE MONONUCLEARS OF PERIPHERAL BLOOD IN PATIENTS WITH CHRONIC LIVER LESIONS AT EXPOSURE TO CHEMICAL SUBSTANCES
Abstract: The pollution of the occupational and surrounding environment with chemical substances appeared to be one of the large problems arising at mankind, due to scientific and technical progress. The anthropogenous pollution gets the increasing significance, both in practical medicine, and in general biological branches. In this article the steps are taken to asses damages of lymphocyte genome at development of pathological states in a liver induced by exposure to the chemical substances.
Keywords: chronic liver damage, chemical substances, chromosome aberrations, peripheral blood lymphocytes.
Introduction The cytogenetic study of the lymphocytes from pe-
The basic sources of involvement of the harmful substanc- ripheral blood of the patients enables to reveal the earliest es into the environment include the enterprises of chemical, signs of development of various pathologies as not specific
metallurgical, oil-producing and other industrial branches. Insufficient use of the methods of waste-free technique, means of gaseous and water waste products clearing from harmful impurities; motor, air, sea and other types of transport working on liquid, firm and gaseous fuels; application in the agriculture of chemical substances of struggle (pesticides) with the plant and weed pests; chemical fertilizers; large power installations, emissive into atmosphere of carbon oxide, sulphur dioxide, nitrogen oxides, heavy resinous products has resulted in distribution of so-called background pathologies expressing in lowering of general body resistance, anti-infectious immunity, when even the insignificant shifts can result in development of serious pathologies, including neoplasias [5, 6].
So now trouble of these factors has made a heavy burden, which presses on all population as a whole. The ecological situation is dangerous to the person, community and state. For the person these are not only diseases connected to threat of life, but also defective breed resulting in accumulation of a genetic burden [1-4]. All above-stated defines necessity of more profound researches of the mechanisms of effects of subthreshold dozes of toxicants, especially at their combined influence. The hemopoietic and reproductive systems appear to be more sensitive to the effects of toxicants [7, 8].
and specific chromosomal disorders in some quantity of these cells in the sporadic defeat, or specific chromosomal disturbances in all cells at inheritance of such disorders [1, 2, 8, 9]. In many diseases there have been already known specific cytogenetic markers. For the patients with hepatitis at influence of chemical substances (pesticides, lead) such markers have not been revealed and consequently the search of them can appear useful for diagnosis of the given pathologies.
The purpose - to estimate damages of lymphocyte genome at development of pathological states in a liver induced on exposure to the chemical substances.
Materials and methods.
For reception of preparations with metaphase plates all studied persons were taken venous blood in volume 2 ml, which then were placed in into vials with medium 199 (6 ml), containing phytohemaglutinin (PHG) and serum of the cattle. Cultivating was performed within 72 hours in thermostat at temperature 37°C. For 2 hours before the cultivating ending into the vials with a mix of cells there was added colchicin to stop mitosis. After hypotonisation and fixation of the cells there were made preparations of metaphase plates with the subsequent staining by Himze-Romanovsky. The analysis of
chromosomes was carried out on microscope Leica, eyepiece 10, objective 100.
For estimation of cytostatic effect of investigated preparations there was defined an index of stimulation PHA (I-PHA), which was calculated by the formula: I-PHA = M: 100, where M - quantity of mitotically divided cells; 100 - total number of cells.
Obj ect of research were 70 patients in the Clinic of Scientific Research Institute of Sanitary, Hygiene and Occupational Diseases of the Ministry of Health of the Republic of Uzbekistan with chronic toxic hepatitis of occupational genesis. Period ofworking of patients was more than 10 years. According to the profession they were linotype operator (direct contact
with lead) and agriculturists - entomologists (contact with pesticides).
I group - Chronic liver damage under the effect of lead (CLD+ LEAD, n = 35) and II group
II group - Chronic liver damage under the effects of pesticides (CLD + PESTICIDES, n = 35),
III group - 50 healthy persons, not contacting with toxic substances.
Results:
At cytogenetic analysis of blood samples of these patients (tabl. 1), the increase of frequency of chromosome aberrations is revealed in comparison with healthy more than in 5, 9 times.
Table 1. - Frequency of the chromosome aberrations in the lymphocytes of patients with toxic hepatitis
Groups of patients Number of studied:
metaphases Aberrant metaphases aberrations,%
Group I HLD + LEAD n = 35 700 35 9.4 ± 1,2*,** (66)
Group II HLD+ PESTICIDES, n = 35 1500 60 4.4 ± 0,5* (66)
Group III health n = 50 1000 12 1.6 ± 0,4 (16)
Note: * - p < 0.05, comparison of groups I and II with group ** -p < 0.05, comparison between groups I and II
Among all types of aberrations the chromosome deletions are dominated, chromosome hyper and hypoploidy are on the second place, it is interesting, that there were more often not specific trisomia and monosomia of chromosomes from group A, C, D, E, and D, and as well as DMC and genes, which come to light as the not painted areas of chromosome. They are present on both chromatids as breaks or blanks, the site chromosome is considered fragile, if genes are found more than 2 times in the same point of chromosome. Among revealed by us aberrations there were met deletions more often in chromosomes of group A (1-3 pairs), C (6-12 pairs), D (13-15 pairs), E (16-18 pairs)
Table 2.- Types of chromosome aberrati
and fragments of different size, dicentric chromosomes, fragments with displacement.
At clinical inspection at all patients prior to the beginning there was established background characteristic of karyotype. The instability of genome and presence of high frequency of chromosome aberrations in mononuclears of peripheral blood was noted (tabl. 1, 2).
Cytogenetic changes were presented by polyploid, hyper-aneuploid cells and single cells with not differentiated translocations, as there were found double small chromosomes and deletions (Fig. 1 - normal parameters and Fig. 2, 3, 4 with pathology).
5 and their frequency in the lymphocytes
Types of aberrations,%
Groups of patients deletion hyper-, diploidy and polyploidy translocations DSC Double small chromosomes Genes,%
Group I Lead 3.14 ± 0.65* 2.14 ± 0.54* 0.57 ± 0.28* 1.0 ± 0.37* 2.57 ± 0.59*
Group II and pesticides 1.05 ± 0.24 0.88 ± 0.22* 0.17 ± 0.10 0.7 ± 0.20* 1.52 ± 0.29*
GroupIII n=50, healthy 1.0 ± 0.22 0.1 ± 0.07 0 0 0
Note: * - p < 0.05, comparison I, II with group III
Cytogenetic investigations revealed that each patient has a unique combination of disorders at a level of chromosome (tabl. 3). The individuality of combinations of chromosome aberrations, alongside with some common and specific chro-
Table 3.- Frequency of abei
mosome disorders for different type of diseases at each patient is noted also by other researchers. In table 4 the frequency of chromosome disorders in mononuclears of peripheral blood of the healthy donors (volunteers) is shown.
ions in some chromosomes
№№ chromosomes/group Structural disorders, % Aneuploidy, losses/acquisitions, %
4 34.8 26.7
17 21.7 13.3
5 21.7 13.3
13 21.7 6.7
16 4.4 -
3 - 13.3
6-12 - 13.3
19/20 - 20.0
22/21 - 13.3
1 - 6.7
Table 4.- Frequency of chromosome disorders in the mononuclears of the peripheral blood of healthy donors (volunteers)
№№ Indicators Males. n = 30 Females. n = 20
1. Number of studied metaphases 3000 2000
2. Number of aberrant metaphases 37 20
3. Number of aberrations,% 1.2 ± 0,19 1.0 ± 0,18
4. Number of aberrations per 1 aberrant metaphase 1.0 1.0
Types of aberrations
5. Deletions,% 40.5 ± 0,89 (15) 25.00.96 (5)
6. Large and small fragments,% 56.8 ± 1,10 (21) 60.0 ± 1,09 (12)
7. DSC (double small chromosomes),% 0 0
8. Genes,% 0.8 ± 0,16 -
Note: the absolute values are shown in the round brackets
The analysis of chromosome disorders in lymphocytes of peripheral blood (Fig. 2, 3, 4) allows at a stage of preclinical development of pathological process to determine risk of development of pathological process in each individual: in case of revealing known specific cytogenetic marker to carry out the earliest diagnosis beginning or having already developed disease. On the basis of specific and not specific chromosome disorders this method allows determination of the prognosis of development and outcome of disease.
The possibility of early identification of specific chromosome disorders in the lymphocytes in blood of patients with early clinical signs of disease promotes its appropriate identification that matters for realization of effective treatment.
It has been shown, that on set of specific and not specific chromosome and genome disorders in the somatic cells there is an opportunity with high probability to establish presence in the body of pathological process in its earliest stage, to predict
development of the certain disease, to determine its intensity and stage, character and localization. The opportunity is shown at similar research to trace updating of primary revealed cytogenetic disorders and to determine their interrelation with development of the subsequent clinical signs of disease.
That is, the opportunity for use of cytogenic investigation of the lymphocytes of peripheral blood to study of the mechanism of development of diseases, their early diagnosis, prognosis, and prevention has been established.
Therefore, cytogenic investigations of the lymphocytes of peripheral blood - as the model of somatic cells, are of significant importance, as the early definition and diagnosis with clinical methods happens to be inconvenient. Systematization of known, revealing and analysis of new cytogenic criteria, determining higher risk of development of pathologies, and development by their criteria of the principles of diagnosis and prognosis hold promise.
Figure 1. The cell with normal karyotype
Figure 2. Condensation of the chromatin (more frequent in group on exposure to lead)
Figure 3. Polyploid increase in chromosomes, equal to haploid set of the 23 chromosome (liver enlargement and change of the chromosome quantity, are often occurred in group on exposure to the pesticides)
Figurer 4. Hyperaneuploid cell, metaphase
Conclusions:
1. The analysis of chromosome disorders in the lymphocytes of peripheral blood allows at the stage of preclinical development of pathological process to determine risk of progressing of pathological process in every individual: in case of revealing known specific cytogenic marker to carry out the earliest diagnosis beginning or being already devel-
oped disease. On set of specific and not specific chromosome aberrations this method gives an opportunity to determine prognosis the development and outcome of disease.
2. The objective changes of frequency of chromosome aberrations in lymphocytes of peripheral blood in the patients with a chronic liver damage can be used as criterion at early diagnostics chronic toxic hepatitis.
References:
1. Bochkov N. P., Chebotarev A. N., Katocova L. D., Platonova V. I. The database for the analysis of the quantitative characteristics of frequency of the chromosome aberrations in the culture of lymphocytes from peripheral blood of the man // Genetics.-2001.- Vol. 37. No 4.- P. 549-557.
2. Bochkov N. P. Ecological genetics of the man // Med.truda prom. ecol.- 2004.- No. 1.- P. 1-6.
3. Zdolnik T. D. Effect of biological active additives to food on the function of digestive glands under conditions of experimental intoxication with lead and chrome // Hyg.and San.- 2001.- No 2.- P. 46-49.
4. Olikhova S. V. et al. Contents of cadmium, lead and copper in the body of the inhabitants of Tashkent and Tashkent area // Hyg. and San.- 2000.- No 3.- P. 11-12.
5. Forsberg Lars A., Chiara Rasi, Hamid R. Razzaghian, et al. Age-Related Somatic Structural Changes in the Nuclear Genome of Human Blood Cells // Am J Hum Genet.- 2012.- Feb 10.- 90(2).- 217-228. Doi: 10.1016/j.ajhg.2011.12.009
6. Kalfalah Faiza, Sabine Seggewiß, Regina Walter, Julia Tigges, Maria Moreno-Villanueva, Alexander Bürkle, Sebastian Ohse, Hauke Busch, Melanie Boerries, Barbara Hildebrandt, Brigitte Royer-Pokora, Fritz Boege. Structural chromosome abnormalities, increased DNA strand breaks and DNA strand break repair deficiency in dermal fibroblasts from old female human donors // Aging (Albany NY) - 2015.- Feb 7(2).- P. 110-122. Published online - 2015.- Feb 4. Doi: 10.18632/ aging.100723
7. Preiksaitiene Egle, Laima Ambrozaityte, Zivile Maldziene, Ausra Morküniene, Loreta Cimbalistiene, Tautvydas Rancelis, Algirdas Utkus, Vaidutis Kucinskas. Identification of genetic causes of congenital neurodevelopmental disorders using genome wide molecular technologies // Acta Med Litu.- 2016.- 23(2).- P. 73-85. Doi: 10.6001/actamedica.v23i2.3324
8. Sram RadimJ., Blanka Binkova, Olena Beskid, Alena Milcova, Pavel Rossner, Pavel Rossner Jr., Andrea Rossnerova, Ivo Solansky, Jan Topinka. Biomarkers of exposure and effect - interpretation in human risk assessment // Air Qual Atmos Health.-2011.- Dec 4(3-4).- P. 161-167. Published online - 2011. Jan 29. Doi: 10.1007/s11869-011-0133-8.
9. Yong Lee C., Martin R Petersen, Alice J. Sigurdson, Laura A. Sampson, Elizabeth M. Ward. High dietary antioxidant intakes are associated with decreased chromosome translocation frequency in airline pilots // Am J Clin Nutr.- 2009. - No. 90(5).-P. 1402-1410. Published online - 2009.- Sep 30. Doi: 10.3945/ajcn.2009.28207.
