Garcia Ana L H, Matzenbacher Cristina A, Soares Solange, Rohr Paula, da Silva Juliana
Lutheran University of Brazil (ULBRA), Laboratory of Genetic Toxicology, PPGBioSaúde (Postgraduate Program in Cellular and Molecular Biology Applied to Health) and PPGGTA (Postgraduate Program in Genetics and Applied Toxicology), 92425-900, Canoas, RS, Brazil; Laboratory of Genetic Toxicology, La Salle University (UniLaSalle), Canoas, RS, Brazil.
Federal University of Rio Grande do Sul, Department of Genetics, C.P. 15053, 91501-970 Porto Alegre, RS, Brazil.
Mutat Res Genet Toxicol Environ Mutagen. 2022 Apr-May;876-877:503474. doi: 10.1016/j.mrgentox.2022.503474. Epub 2022 Feb 17.
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.
氟骨症是一种严重的病症，因氟过量沉积导致骨骼变形和骨组织弱化。最近，有关于吸烟的数据发表，表明烟雾成分可间接影响骨量并干扰人体氟化物代谢。因此，本体外研究旨在评估人成骨细胞MG63细胞分别及联合暴露于人体血浆中浓度的氟硅酸（FA）和可替宁（COT）时的遗传不稳定性。为此，通过MTT法检测细胞毒性；通过用修复内切酶（+FPG）改良的碱性彗星试验（CA）检测DNA损伤；使用CBMN - Cyt试验进行微核试验（MN）；并通过qPCR检测MG63细胞中的端粒长度（TL）。本研究中测试的所有浓度均未观察到细胞毒性。碱性CA结果显示，在所有FA浓度（0.03125 - 0.300 mg/L）、COT的两个最高浓度（125和250 ng/mL）以及FA + COT的最高浓度（0.300 mg/L + 250 ng/mL）下，DNA损伤均显著增加。碱性CA + FPG试验用于检测氧化核碱基，其在FA、COT和FA + COT的两个最高浓度下出现。微核试验显示，除最低浓度（0.03125 mg/L）外，所有FA浓度（0.075 - 0.300 mg/L）、COT的两个最高浓度（125和2�0 ng/mL）以及所有FA + COT浓度下MN频率均增加。核分裂指数、双核细胞、核质桥和核芽无显著差异。在单独用最高浓度的FA（0.300 mg/L）和FA + COT（0.300 mg/L + 250 ng/mL）处理的细胞中观察到端粒长度缩短。最后，我们的研究表明，人体血浆中浓度的FA和COT（主要是单独作用时）可诱导人成骨细胞的氧化损伤和遗传不稳定性。
