Cytotoxicity of chlorhexidine digluconate to murine macrophages and its effect on hydrogen peroxide and nitric oxide induction

Abstract

Chlorhexidine, even at low concentrations, is toxic for a variety of eukaryotic cells; however, its effects on host immune cells are not well known. We evaluated in vitro chlorhexidine-induced cytotoxicity and its effects on reactive oxygen/nitrogen intermediate induction by murine peritoneal macrophages. Thioglycollate-induced cells were obtained from Swiss mice by peritoneal lavage with 5 ml of 10 mM phosphate-buffered saline, washed twice and resuspended (10 6 cells/ ml) in appropriate medium for each test. Cell preparations contained more than 95% macrophages. The cytotoxicity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay and the presence of hydrogen peroxide (H 2 O 2 ) and nitric oxide (NO) by the horseradish peroxidase-dependent oxidation of phenol red and Griess reaction, respectively. The midpoint cytotoxicity values for 1-and 24-h exposures were 61.12 ± 2.46 and 21.22 ± 2.44 µg/ml, respectively. Chlorhexidine did not induce synthesis or liberation of reactive oxygen/nitrogen intermediates. When macrophages were treated with various sub-toxic doses for 1 h (1, 5, 10, and 20 µg/ml) and 24 h (0.5, 1, and 5 µg/ml) and stimulated with 200 nM phorbol myristate acetate (PMA) solution, the H 2 O 2 production was not altered; however, the NO production induced by 10 µg/ml lipopolysaccharide (LPS) solution varied from 14.47 ± 1.46 to 22.35 ± 1.94 µmol/ l and 13.50 ± 1.42 to 20.44 ± 1.40 µmol/l (N = 5). The results showed that chlorhexidine has no immunostimulating activity and sub-toxic concentrations did not affect the response of macrophages to the soluble stimulus PMA but can interfere with the receptor-dependent stimulus LPS. Correspondence

Effect of Fish or Soybean Oil-Rich Diets on Bradykinin, Kallikrein, Nitric Oxide, Leptin, Corticosterone and Macrophages in Carrageenan Stimulated Rats

“…These results are similar to other [33] in neutrophils from rats fed a soybean-rich diet. H 2 O 2 is an important citotoxic agent [34] and its lower release could contribute to the anti-inflammatory effect of the hyperlipidic diets, as observed previously [12,27] and reproduced in the present study. Some reports have shown decreased activity of superoxide-dismutase (SOD), the enzyme catalyzing H 2 O 2 formation, in plasma and in different tissues of animals treated with nY3 or nY6 PUFA-rich diets [35,36].…”

Essential Oils from Ugandan Medicinal Plants: In Vitro Cytotoxicity and Effects on IL‐1β‐Induced Proinflammatory Mediators by Human Gingival Fibroblasts

“…However, in vitro studies have shown chlorhexidine to be highly cytotoxic even in low concentration to many cell types, including gingival fibroblasts [39], macrophages [40], and osteoblastic and endothelial cells [41]. Other studies have also shown that topical application of chlorhexidine may result in its penetration through the epithelial barrier, thus triggering tissue damage [42].…”

The role of DNA damage and caspase activation in cytotoxicity and genotoxicity of macrophages induced by bisphenol‐A‐glycidyldimethacrylate

“…BisGMA with its hydrophobic property needs organic solvents to facilitate absorbtion in a hydrophilic environment for biocompatibility studies. Macrophages have been used for immunocytotoxicity testing because they permit the measurement of cytotoxic responses directly in cell culture and because of their ability to maintain immunological functions in the presence of many chemical agents (Bonacorsi et al. 2004); thus, RAW264.7 cells were selected in this study.…”