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Print version ISSN 0034-7094
Rev. Bras. Anestesiol. vol.52 no.1 Campinas Jan./Feb. 2002
Genotoxic effects on professionals exposed to inhalational anesthetics *
Efectos genotóxicos en profesionales expuestos a los anestésicos inhalatorios
Ana Regina Chinelato, M.D.I; Nívea Dulce Tedeschi Conforti Froes, M.D.II
IMestra e Doutoranda em Genética
IIMestra e Doutora em Genética pela UNESP; Pós-doutoramento na Universidade do Texas, TX, USA
BACKGROUND AND OBJECTIVES: This study
compared physicians exposed to anesthetics to non-exposed individuals, aiming
at investigating chromosomal changes, the possible interference of inhalational
anesthetics on cell kinetics and the genotoxic risk associated to occupational
METHODS: Peripheral blood lymphocytes were temporarily cultured to obtain metaphases needed for the analysis of chromosomal aberrations and sister chromatid exchanges.
RESULTS: Cytogenetic analysis has shown an increased frequency in chromosomal aberrations per cell in the exposed group as compared to controls, confirming the clastogenic effect of such compounds. Anesthetic gases, however, had no inducing effects on sister chromatid exchanges. The comparison between proliferation and mitotic indices has also shown no significant differences between exposed and control groups.
CONCLUSIONS: Our data suggest that anesthesiologists may represent a group at risk among exposed people, and that working conditions should be improved.
Key Words: ANESTHETICS, Inhalational; PROFESSIONAL RISK: chronic exposure
JUSTIFICATIVA Y OBJETIVOS: En este estudio
se compararon médicos expuestos a los gases anestésicos con individuos
no expuestos, para la investigación de alteraciones cromosómicas,
verificación de la posible interferencia de los anestésicos inhalatorios
en la cinética celular y evaluación del riesgo genotóxico asociado
a la exposición ocupacional.
MÉTODO: Fueron realizadas culturas temporarias de linfocitos de sangre periférico para la obtención de metafases, necesarias para la análisis de aberraciones cromosómicas y cambios entre cromátidas hermanas.
RESULTADOS: La análisis citogenética mostró un aumento en las frecuencias de aberraciones cromosómicas por célula en el grupo expuesto, cuando comparado al grupo control, denotando el efecto clastogénico de eses compuestos. Con relación al parámetro de cambios entre cromátidas hermanas, los gases anestésicos no demostraron efecto inductor. La comparación entre los índices mitótico y de proliferación celular también mostró que no hay diferencias significativas entre los grupos expuesto y control.
CONCLUSIONES: Los resultados obtenidos sugieren que los anestesiólogos pueden representar un grupo de riesgo entre personas ocupacionalmente expuestas y las condiciones de trabajo deben ser mejoradas.
Human biomonitoring is a useful tool to identify and quantify exposure-related risks. It has been primarily applied to investigate the risk of cancer development by accidental, occupational or therapeutic exposure to substances which may injury genetic material 1. Several studies have shown noxious effects to general welfare as a consequence of chronic exposure in the operating center. There is a proven relative increase in headache, irritability, nausea, itching and fatigue 2,3. Infertility, liver and kidney diseases were also reported. The incidence of cancer seems to be twice as high in professionals working in operating rooms, especially anesthesiologists 4-6.
There are few data in the literature on cytogenetic monitoring and professional exposure to anesthetic gases. In Brazil, there are no studies of this kind. In addition, differences in procedures and working conditions in different countries must be taken in consideration. So, professionals are exposed to a variable amount of anesthetic gases for also variable periods. This study aimed at investigating the genotoxic potential of such compounds through the analysis of chromosomal aberrations and sister chromatid exchanges in peripheral blood lymphocytes of exposed physicians, thus contributing to explain the risk to human health associated to this occupational exposure, and to assure better working conditions for such professionals.
After the Ethics Committee approval, a cytogenetic study was performed with 11 anesthesiologists predominantly exposed to halothane, all of them working for more than 5 years in hospitals of the city of São José do Rio Preto, who were compared to a control group. All individuals were paired by gender and age and supplied information about exposure to physical and chemical agents, alcohol and cigarette consumption, frequency of viral or bacterial diseases and other facts having occurred up to three months before blood collection. Information was obtained through a comprehensive questionnaire and all data were recorded.
Temporary lymphocyte cultures were seeded aiming at obtaining metaphases needed to analyze chromosomal aberrations (CA) and sister chromatid exchanges (SCE). Twenty-four hours later, all tubes received 50 µg of 5-bromodeoxyuridine (5-BrdU), according to Korenberg & Freedlender (1974), with modifications according to Ez-Zein et al. 7 in cultures collected both with 50 hours for CA analysis in first division cycle cells and after 70 hours for SCE analysis.
In average, 300 first division metaphases were analyzed per individual to estimate chromosomal aberration frequency. For sister chromatid exchanges, 30 second division metaphases were, in average, analyzed.
Both for chromosomal aberrations and sister chromatid exchanges, best quality metaphases were chosen where chromosomal groups could be safely identified.
Differences in cytogenetic parameters were statistically evaluated by Mann-Whitney test and Chi-square test was used to identify differences in sister chromatid exchange values. Mitotic and cell proliferation indices were analyzed through analysis of variance and Students t test, respectively.
Frequency and structural chromosomal aberrations found in exposed and control groups are shown in figure 1. Chromosomal and chromatidic breakages were computed, as well as structural rearrangements detectable by the above-mentioned technique. Abnormal events were classified according to criteria described in the International System of Human Cytogenetic Naming (ISCN) 8. A breakage was defined as a situation in which the chromosomal segment was separated from the remaining chromosome by a distance longer than the chromatid width. Events were considered chromosomal when affecting both chromatids, and were considered chromatidic when only one chromatid was affected. Chromosomal and chromatidic failures were observed but not computed. Addition was considered a situation when chromosomes had a different than expected size as a result of a breakage and subsequent binding of fragments. Fragments with no centromers and chromosomes resulting from two breakages in different chromosomes followed by reverse binding of fragments, called dicentric were also taken in consideration. Ring chromosomes result from the breakage of telomers and subsequent binding; figures with four chromosomal arms resulting from genetic material rearrangement and called quadriradial were also found.
Mean breakage frequency in the exposed group was 3,52% as compared to 2.14% in the control group. Both groups presented more than one aberration per abnormal cell: 1.4 in anesthesiologists and 1.2 in the control group. Considering the mitotic index, data have shown that anesthetic gases do not seem to interfere in the cell division process.
Sister chromatid exchange frequency is shown in figure 2 for both samples. There were no significant differences on cell proliferation index in both groups.
Changes in human genoma have generated an increasing concern as to the adoption of protecting measures for future generations. This attitude is reinforced by the awareness of an association between genotoxic and carcinogenic properties of chemical radiations and compounds. Furthermore, there is recognition that environmental factors act as etiologic agents in approximately 80% of human cancers. So, tests are becoming more and more urgent to detect such agents, as well to quantify injured DNA in men. This way, well conducted cytogenetic studies may be safely used to indicate excessive exposure to noxious agents as well as to indicate the potential risk for severe health problems in the exposed population 9,10.
Although negative reports on the mutagenicity and carcinogenicity of anesthetic gases, there are still many controversies. Contradictory data were found in the literature, considering the variability of individual responses to different mutagenic and/or carcinogenic agents.
Women normally exposed to the operating room environment are subject to a higher risk for spontaneous abortion during the first trimester of gestation. The incidence is 1.5 to 2 times higher than in non-exposed people. The highest incidence of abortion is seen among anesthesiologists, nurses working with anesthesia and operating room nurses and technicians 2,4,5,11-15. Similar results were found in dentists and assistants using inhalational anesthetics 16,17. However, there are disagreeing data, such as Hemminki et al. 18 who have not detected a significant increase in spontaneous abortion and infant malformation among nurses exposed to anesthetic gases. According to a study of 1970, fetus losses among women occupationally exposed to anesthetics is twice as high as compared to non-exposed women, and premature births are 4.5 times more frequent among anesthesiologists. A change in gender relationship was also noted, from 50/50 to 50/41, favoring women. Anesthesiologists wives, not exposed to the operating room environment, present an abortion rate 3 times higher and a premature birth rate 4 times higher than controls 2,12. However, according to Cohen et al. 19 this increased risk is non-existent. Children of women working in operating rooms show a high incidence of congenital abnormalities 3,5,11, varying from major abnormalities and deformities to less severe defects 2. The risk is twice as high for anesthesiologists as compared to non-exposed pediatricians 5,15. In spite of that, according to Imbriani et al. 14, there is still no understanding as to the frequency of such malformations. Female infertility also seems to be higher in women working in surgical environments 15. Cohen et al. 19 have reported a significant increase in congenital abnormalities in children of non-exposed women married to men working in operating rooms, in line with Knill-Jones et al. 19 who also reported an association between exposed men and total congenital abnormalities of their children. Male exposure was not associated to increased spontaneous abortions. Contrasting with such observations. Cohen et al. 19 have reported a significant spontaneous abortion rate among wives of dentists exposed to anesthetic gases, but birth defects had no significant value. Differences might be the result of different exposure intensity to different anesthetic gases.
Several analysis were performed in an attempt to estimate cancer frequency among workers exposed to anesthetics. In one of them, the American Society of Anesthesiologists (ASA) performed a retrospective study with 49.595 individuals working in operating centers in all North-American states. An increase of 1.3 to 2 times in cancer frequency among ASA and American Association of Nurse Anesthetists female members was observed as compared to non-exposed control groups; however, no increase in cancer frequency was observed among studied males 1. In epidemiological surveys several types of risks were identified: abortions, premature births and stillborns, congenital defects, cancer, liver diseases, fatigue, decreased welfare and performance 2. Several studies suggest that chronic exposure to anesthetics in operating rooms represents a risk for the health of occupationally exposed people 14. However, due to epidemiological analysis interpretation problems, many investigators decided for different tests to obtain information on the carcinogenic potential of specific anesthetic drugs 1.
In our study, the fact of having found a higher chromosomal aberration frequency, especially chromosomal breakage, in individuals exposed to anesthetic gases shows the presence of errors during DNA replication or deficiencies in repair mechanisms. Chemical compounds, when acting on the DNA, may be divided in two classes: S-independent compounds, or those causing aberrations in all cell cycle phases; and those depending on DNA synthesis to manifest their effects (S-dependent). S-independent compounds (for instance, radiomimetic drugs) induce, as the ionizing radiation, chromosomal breakages in G0 and G1, chromosomal and chromatidic aberrations in S and chromatidic aberrations in G2. S-dependent compounds (such as alkylating drugs) produce, as UV light, their effects in phase S or when the cells go through a synthesis phase between exposure and presence of effects 20,21. Anesthetic gases, then, seem to act similarly to radiomimetic drugs.
Another relevant fact in this study was the observation of rare events, such as additions, dicentric and acentric, which are classified as unstable events which may lead to cell death 22 Such rare events may lead to deficiencies or complex rearrangements. Deficiencies lead to the loss of multiple genes, so, after cell division, daughter cells may not survive. Recent evidences show that dead cells would stimulate the division of new cells, which could allow for the appearance of dormant cancer cells 9. Another possibility are rearrangements which would not necessarily cause cell death; however, due to gene juxtaposition, chromosomal rearrangements may lead to the abnormal expression of translocated genes and the abnormality will be transmitted to daughter cells. This type of aberration also seems to be related to cancer development through proto-oncogene activation 22-24.
For what has been said, together with results from other studies, one may state that a lot is still to be done to determine the actual risk to which anesthesiologists are exposed. Another important factor to be taken in consideration is that most contradictions come from the fact that most studies were performed in operating rooms with air renewal systems, not present in many hospitals, including those where individuals here sampled work. During prolonged anesthesia, physicians are constantly breathing an atmosphere overloaded with alcohol, ether, anesthetic gases and other products. Radiological procedures, stress and long hours exposed to cleaning products used in the surgical area are also factors which may affect anesthesiologists. In addition, several compounds, including alcohol, therapeutic drugs, environmental and agricultural pollutants may act as liver enzyme inducers, which may cause severe problems.
Taking into account the multifactorial nature of health professionals exposure, measures must be taken to minimize occupational exposure to chemical agents with known or possible genotoxic potential. This way it will be possible to effectively prevent noxious effects to the health of such professionals. Better equipped operating rooms, with adequate ventilation and exhaustion systems, as well as their maintenance should be demanded.
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Submitted for publication January 18, 2001 *
Received from Laboratório de Epidemiologia Molecular da Universidade Estadual
Paulista (UNESP) de São José do Rio Preto, SP
Accepted for publication October 03, 2001
Submitted for publication January 18, 2001
* Received from Laboratório de Epidemiologia Molecular da Universidade Estadual Paulista (UNESP) de São José do Rio Preto, SP