Dry tobacco leaves: an in vivo and in silico approach to the consequences of occupational exposure.

Exposure of tobacco workers handling dried tobacco leaves has been linked to an increased risk of toxicity and respiratory illness due to the presence of nicotine and other chemicals. This study aimed to evaluate the DNA damage caused by the exposure of tobacco growers during the dry leaf classification process and the relation to cellular mechanisms. A total of 86 individuals participated in the study, divided into a group exposed to dry tobacco (n = 44) and a control group (n = 42). Genotoxicity was evaluated using the alkaline comet assay and lymphocyte micronucleus (MN) assay (CBMN-Cyt), and measurement of telomere length. The levels of oxidative and nitrosative stress were evaluated through the formation of thiobarbituric acid reactive species, and nitric oxide levels, respectively. The inorganic elements were measured in the samples using particle-induced X-ray emission method. The combination of variables was demonstrated through principal component analysis and the interactions were expanded through systems biology. Comet assay, MN, death cells, thiobarbituric acid reactive species, and nitrosative stress showed a significant increase for all exposed groups in relation to the control. Telomere length showed a significant decrease for exposed women and total exposed group in relation to men and control groups, respectively. Bromine (Br) and rubidium (Rb) in the exposed group presented higher levels than control groups. Correlations between nitrate and apoptosis; Br and MN and necrosis; and Rb and telomeres; besides age and DNA damage and death cells were observed. The systems biology analysis demonstrated that tobacco elements can increase the nuclear translocation of NFKB dimers inducing HDAC2 expression, which, associated with BRCA1 protein, can potentially repress transcription of genes that promote DNA repair. Dry tobacco workers exposed to dry leaves and their different agents showed DNA damage by different mechanisms, including redox imbalance.

Fluorosilicic acid and cotinine, separately and in combination, induce genotoxicity and telomeric reduction in human osteoblast cell line MG63.

Skeletal fluorosis is a severe case in which bone deformations and bone tissue weakening occur due to excessive fluorine deposition. Recently, data on smoking have been published that smoke constituents can indirectly influence bone mass and interfere in the metabolism of fluorides in humans. Thus, the present in vitro study aimed to assess the genetic instability in human osteoblast MG63 cells exposed to fluorosilicic acid (FA) and cotinine (COT), separately and in combination, in concentrations found in human plasma. For this, cell cytotoxicity was performed by MTT assay; DNA damage was performed by alkaline comet assay (CA), modified by repair endonucleases (+FPG); micronuclei test (MN) using CBMN-Cyt assay; and telomere length (TL) by qPCR in MG63 cells. No cytotoxicity was observed for all concentrations tested in this study. Alkaline CA results showed a significant increase in DNA damage at all FA concentrations (0.03125-0.300 mg/L), in the two highest concentrations of COT (125 and 250 ng/mL), and the highest concentration of FA+COT (0.300 mg/L+250 ng/mL). Alkaline CA+FPG test was used to detect oxidized nucleobases, which occurred at the two highest concentrations of FA, COT, and FA+COT. Micronuclei test showed an increase in the frequency of MN at all concentrations of FA (0.075-0.300 mg/L) except in the lowest concentration (0.03125 mg/L), in the two highest concentrations of COT (125 and 250 ng/mL), and all concentrations of FA+COT. There was no significant difference in nuclear division index, binucleated cells, nucleoplasmic bridge, and nuclear bud. A TL reduction was observed in cells treated with the highest concentrations of FA alone (0.300 mg/L) and FA+COT (0.300 mg/L+250 ng/mL). Finally, our study showed that FA and COT (mainly alone) at concentrations found in human plasma induced oxidative damage and genetic instability in human osteoblast cells.

Fluorosilicic acid induces DNA damage and oxidative stress in bone marrow mesenchymal stem cells.

Excess fluoride in water can produce changes in tooth enamel mineralization and lead to diseases such as dental or skeletal fluorosis. The present study aimed to assess the genotoxic effects, oxidative stress, and osteoblastic mineralization induced by fluorosilicic acid (FA) in murine bone marrow-derived mesenchymal stem cells (BM-MSCs). BM-MSCs were isolated from the femurs and tibias of rats and cultured under standard conditions. Cells exposure occurred for 3, 7, 14, and 21 days to different concentrations of FA (0.6-9.6 mg/L). Cytotoxicity was observed in 14 and 21 days of exposure for all concentrations of FA (cell proliferation below 60%), and for 3 and 7 days, in which the proliferation was above 80%. Alkaline comet assay results demonstrated significant increased damage at concentrations of 0.3-2.4 mg/L, and the micronucleus test showed increased rates for micronucleus (1.2-2.4 mg/L) and nuclear buds (NBUDs) (0.3-2.4 mg/L) (P < 0.05/Dunnett's test). An alkaline comet assay modified by repair endonuclease (FPG) was used to detect oxidized nucleobases, which occurred at 0.6 mg/L. The oxidative stress was evaluated by lipid peroxidation (TBARS) and antioxidant activity (TAC). Only lipid peroxidation was increased at concentrations of 0.6 mg/L and 1.2 mg/L (P < 0.001/Tukey's test). The osteogenesis process determined the level of extracellular matrix mineralization. The mean concentration of Alizarin red increased significantly in 14 days at the 0.6 mg/L concentration group (P < 0.05/Tukey's test) compared to the control group, and a significant difference between the groups regarding the activity of alkaline phosphatase (ALP) was observed. Unlike other studies, our results indicated that FA in BM-MSCs at concentrations used in drinking water induced genotoxicity, oxidative stress, and acceleration of bone mineralization.