Effects of inhalation anesthetics halothane, sevoflurane, and isoflurane on human cell lines
Introduction
Inhaled anesthetics are probably the most used anesthetics, alone or as a part of a balanced anesthesia. Adverse reactions of inhaled anesthetics, especially mutagenesis and carcinogenesis, were usually in the focus of anesthetic community (Doll and Peto, 1977). The major reason was that inhaled anesthetics were suspected to induce tumors, spontaneous abortions, and congenital anomalies in chronically exposed anesthetic stuff. The first animal and epidemiological studies on effects of chronic exposure to subanesthetic concentration of inhaled anesthetics failed to prove such hypothesis (ASA ad hoc committee, 1974, Mazze et al., 1986, Eger et al., 1978).
The use of inhaled anesthetics in anesthesia for cancer diseases was commonly investigated (Van de Louw et al., 1998, Kvolik et al., 2003). Most observations referring to the use of inhaled anesthetics in cancer surgery and diagnostics were directed to side effects observing, although those could be clinically irrelevant (Fisher et al., 1985). In the clinical setting, the choice of the anesthetic agent may be important in the view of mutagenic potential, impaired metastatic capability, and growth of pre-existing tumor cells. Tumor growth is particularly important in extensive cancer surgery resulting in immune and organ dysfunctions (Melamed et al., 2003).
The major problem concerning in vitro investigations on effects of inhaled anesthetics on tumor growth was a study design. Exposure protocols and the vapor concentration of test chemicals were poorly monitored and different from clinical setting in several points. Some authors have bound anesthetics into drug-micelle complexes with toxic substance DMSO, which allowed concentration adjustment (O'Leary et al., 2000). Others used very high concentrations of anesthetics or prolonged exposure e.g. 3% halothane for 24 h to pronounce toxic effects (Muckter et al., 1998). Hack et al. (1981) used halothane 3 vol.%, enflurane and isoflurane 5 vol.%, and methoxyflurane 2 vol.% in O2: N2O 20: 78% and CO2 2 vol.%.
Investigations in vivo are usually more complex. Numerous factors were different from operative setting: e.g. inhaled anesthetics were delivered in concentrations lower than used in clinical practice over prolonged time, so that spontaneous breathing was allowed (Eger et al., 1978, Hack et al., 1981). In the study of Moudgil and Singal (1997), researching metastatic alterations after exposure to inhaled anesthetics nonexposed tumor cells was inoculated in the anesthetized organism. As experimental conditions were distinctive from those in the anesthesia during surgery, given results were contradictory.
Therefore in this study, clinically relevant doses of halothane, isoflurane, and sevoflurane were used. It was conducted to compare growth effects of single exposure of human cell lines to inhaled anesthetics. Antiproliferative effects on human tumor and normal cells in vitro were measured after 2, 4, and 6 h of exposure to anesthetics. A cell line specific effect and mechanism of death of treated tumor cells were investigated. Our observations pointed that three inhaled anesthetics expressed quantitatively different inhibitory effect against cell lines, and induced apoptosis as a mode of cell death.
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Materials
Human colon carcinoma (Caco-2), human larynx carcinoma (HEp-2), human pancreatic carcinoma cells (MIA PaCa-2), poorly differentiated cells from lymph node metastasis of colon carcinoma (SW-620), and normal fibroblasts (WI-38) were obtained from Institute Rudjer Boskovic, Division of Molecular Medicine, Zagreb, Croatia.
Inhaled anesthetics halothane ('Fluothane', Zeneca Ltd. Macclesfield Cheshire, GB), isoflurane (Forane®), and sevoflurane (Sevorane®) were form Abbott Laboratories, Queensborough,
Cell culture
All cell lines were grown as monolayer in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum, 2 mM glutamine, 100 U penicillin, and 0.1 mg ml− 1 streptomycin. The cultures were equilibrated with humidified 5 vol.% CO2 in air at 37 °C in CO2 incubator (Shell Lab, Sheldon Manufacturing, USA). Cell viability was assessed using the trypan blue dye exclusion method. Cells (2 × 104 cells ml− 1) were plated onto 96-microwell plates and allowed to attach overnight.
Cell viability assay
The
Growth inhibition
Cytotoxic activity was measured on exponentially growing cells. In order to determine antiproliferative potency of single exposure to anesthetic gas mixture, cell proliferation was performed after 72 h. Thus, a time for normal cell cycle progression was allowed. All human cell lines showed growth alterations as a consequence of single anesthetic exposure to the inhaled anesthetic gas mixture. Halothane causes the most pronounced growth inhibition in all cancer cell lines.
Results of MTT test are
Discussion
This study investigated proliferative effects of inhalation anesthetics halothane, isoflurane, and sevoflurane on human normal and tumor cells growth, and some mechanisms of such actions. Quantitative differences among effects of various anesthetics could be considered due to changes in dynamics of tumor cells growth during and after anesthesia. A tumor cell proliferation after exposure to clinically relevant concentrations of inhaled anesthetics appears important in the view of altered
Acknowledgements
A support of this study by the Ministry of Science (Project No 0127 111) is gratefully acknowledged.
References (45)
- et al.
Rapid colorimetric assay for cell viability: application to the quantitation of cytotoxic and growth inhibitory lymphokines
Journal of Immunological Methods
(1984) - et al.
Immunohistochemical demonstration of the expression of CYP2E1 in human breast tumour and non-tumour tissues
Cancer Letters
(2003) - et al.
Differential effects of isoflurane and i.v. anaesthetic agents on metabolism of alveolar type II cells
British Journal of Anaesthesia
(1999) - et al.
A novel apparatus for the exposure of cultured cells to volatile agents
Journal of Pharmacological and Toxicological Methods
(1998) - et al.
Antiproliferative actions of inhalational anaesthetics: comparisons to valproat teratogen
International Journal of Developmental Neuroscience
(2000) - et al.
Remodeling for demolition: changes in mitochondrial ultrastructure during apoptosis
Molecular Cell
(2002) - et al.
Apoptotic nuclear morphological changes without DNA fragmentation
Current Biology
(1999) - et al.
Use of sevoflurane for surgery of phaeochromocytoma
Annales Francaises d'Anesthesie et de Reanimation
(1998) - et al.
Formation of trifluoroacetylated protein antigens in cultured rat hepatocytes exposed to halothane in vitro
Biochemical Pharmacology
(1994) - et al.
Apoptosis in rectal carcinoma
Cancer
(2001)
Occupational disease among operating room personnel: a national study
Anaesthesiology
The DNA of annexin V-binding apoptotic cells is highly fragmented
Cancer Research
Attenuation of the tumor-promoting effect of surgery by spinal blockade in rats
Anesthesiology
Timing within the oestrous cycle modulates adrenergic suppression of NK activity and resistance to metastasis: possible clinical implications
British Journal of Cancer
Role of alcohol dehydrogenase 3 and cytochrome P-4502E1 genotypes in susceptibility to cancers of the upper aerodigestive tract
International Journal of Cancer
Cytotoxicity of millimolar concentrations of ethanol on HepG2 human tumor cell line compared to normal rat hepatocytes in vitro
Journal of Cancer Research and Clinical Oncology
Isoflurane pretreatment inhibits cytokine-induced cell death in cultured rat smooth muscle cells and human endothelial cells
Anesthesiology
Mortality among doctors in different occupations
British Medical Journal
A test of the carcinogenicity of enflurane, isoflurane, halothane, methoxyflurane, and nitrous oxide in mice
Anesthesia and Analgesia
Comparison of enflurane, halothane, and isoflurane for diagnostic and therapeutic procedures in children with malignancies
Anesthesiology
Midazolam metabolism by modified Caco-2 monolayers: effects of extracellular protein binding
The Journal of Pharmacology and Experimental Therapeutics
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