Proteomic Analysis of Liver in Rats Chronically Exposed to Fluoride

Abstract

Fluoride (F) is a potent anti-cariogenic element, but when ingestion is excessive, systemic toxicity may be observed. This can occur as acute or chronic responses, depending on both the amount of F and the time of exposure. The present study identified the profile of protein expression possibly associated with F-induced chronic hepatotoxicity. Weanling male Wistar rats (three-weeks old) were divided into three groups and treated with drinking water containing 0, 5 or 50 mg/L F for 60 days (n=6/group). At this time point, serum and livers were collected for F analysis, which was done using the ion-sensitive electrode, after hexamethyldisiloxane-facilitated diffusion. Livers were also submitted to histological and proteomic analyses (2D-PAGE followed by LC-MS/MS). Western blotting was done for confirmation of the proteomic data A dose-response was observed in serum F levels. In the livers, F levels were significantly increased in the 50 mg/L F group compared to groups treated with 0 and 5 mg/L F. Liver morphometric analysis did not reveal alterations in the cellular structures and lipid droplets were present in all groups. Proteomic quantitative intensity analysis detected 33, 44, and 29 spots differentially expressed in the comparisons between control vs. 5 mg/L F, control vs. 50 mg/L F, and 5 mg/L vs. 50 mg/L F, respectively. From these, 92 proteins were successfully identified. In addition, 18, 1, and 5 protein spots were shown to be exclusive in control, 5, and 50 mg/L F, respectively. Most of proteins were related to metabolic process and pronounced alterations were seen for the high-F level group. In F-treated rats, changes in the apolipoprotein E (ApoE) and GRP-78 expression may account for the F-induced toxicity in the liver. This can contribute to understanding the molecular mechanisms underlying hepatoxicity induced by F, by indicating key-proteins that should be better addressed in future studies.

Proteomic Analysis of Gastrocnemius Muscle in Rats with Streptozotocin-Induced Diabetes and Chronically Exposed to Fluoride

“…Interestingly, in the absence of F, the expression levels of L-lactate dehydrogenase A chain and L-lactate dehydrogenase C chain were lower in ND rats than in D rats. A previous study reported the absence of L-lactate dehydrogenase A chain in the liver of rats (ND) treated with F [28]. Decreased expression of this enzyme leads to increased aerobic glycolysis.…”

“…Previous studies have treated the animals with F for 6 weeks [4], [28]–[30]; however, a pilot study revealed high levels of mortality prior to completion of the experimental period. Therefore, an experimental time of 22 days was selected.…”

Effects of Fluoride on DNA Damage and Caspase-Mediated Apoptosis in the Liver of Rats

“…The cDNA was reversed transcription (RT) from 1 μg of total RNA with primer oligo(dT)18 by using superscript II RNase (Reverse Transcriptase; Invitrogen Inc., USA). The caspase-3 and −9 cDNA were amplified by polymerase chain reaction (PCR) based on a previously published rat cDNA primer sequence (6)(7)(8). β-Actin was the internal standard.…”

“…Many reports have shown that F can cause progressive degeneration of the structure and functions of skeletal muscles, the brain, and the spine. In addition, it increases the aerobic metabolism and promotes alterations in the metabolism of cellular free radicals in several organs such as the liver, kidneys, and heart [6][7][8][9][10]. Free radicals cause toxicity by attacking membrane phospholipids, leading to membrane injury through lipid peroxidation, depolarization of the mitochondrial membrane, and apoptosis [11].…”

Efeito dose-resposta do fluoreto em parâmetros relacionados à resistência à insulina e na expressão de proteínas hepáticas e musculares em camundongos NOD

“…Além disso, diversos trabalhos sugerem que o F pode interferir em muitas vias metabólicas, inibindo a ação de diversas enzimas e ainda atuando sobre a via glicolítica (DE LA SOTA et al, 1997;WHITFORD, 1996). Estudos realizados com animais experimentais, em laboratório, demonstraram que a ingestão aguda ou crônica de F pode alterar a expressão proteica em tecidos ósseo, hepático, renal, intestinal e dentário, bem como na urina de ratos ou camundongos (LEITE, 2010;KOBAYASHI et al, 2009;KOBAYASHI et al, 2011;KOBAYASHI, 2012;CARVALHO et al, 2013;PEREIRA et al, 2013;CHARONE et al, 2016;MELO et al, 2017;DIONIZIO et al, 2018 (VIÑALS et al, 1993;OTSUKI et al, 2005;ELLIOT et al, 2002;GOH et al, 2003;GUTIÉRREZ-SALINAS et al, 2010).…”