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1.
Ecotoxicol Environ Saf ; 226: 112851, 2021 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-34619480

RESUMO

Long-term excessive intake of fluoride (F) can cause osseous and non-osseous damage. The kidney is the main fluoride excretion organ of the body. This study aimed to explore whether dietary calcium (Ca) supplementation can alleviate kidney damage caused by fluorosis and to further investigate the effects of Ca on the mitigation mechanism of renal cell apoptosis triggered by F. We evaluated the histopathological structure, renal function indicators, and gene and protein expression levels of death receptor-mediated apoptosis pathways in Sprague Dawley (SD) rats treated with sodium fluoride (NaF) and/or calcium carbonate (CaCO3) for 120 days. The results showed that 100 mg/L NaF induced kidney histopathological injury and apoptosis, increased the concentrations of Creatinine (CRE), uric acid (UA), blood urea nitrogen (BUN), potassium (K), phosphorus (P) and F (p < 0.05), and decrease the level of serum magnesium (Mg) (p < 0.05). Moreover, NaF increased the mRNA and protein expression levels of Fas cell surface death receptor (FAS), tumor necrosis factor (TNF), TNF-related apoptosis-inducing ligand (TRAIL), Caspase 8, Caspase 3 and poly ADP-ribose polymerase (PARP) (p < 0.01), which finally activated the death receptor pathway. Inversely, Ca supplementation reversed the decrease of CRE, BUN, UA, F and P levels induced by F, alleviated histopathological damage and apoptosis, and reduced the gene and protein expression levels of death receptor pathway-related markers. In conclusion, 1% Ca alleviates F-induced kidney apoptosis through FAS/FASL, TNFR/TNF, DR5/TRAIL signaling pathways.


Assuntos
Cálcio , Fluoretos , Animais , Apoptose , Cálcio/metabolismo , Cálcio na Dieta , Caspase 8 , Proteína Ligante Fas/genética , Fluoretos/toxicidade , Rim/metabolismo , Ratos , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
2.
Sci Total Environ ; 804: 150184, 2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34517333

RESUMO

As an environmental toxicant, the damage of fluoride to the body has attracted global attention. Because liver is an essential organ for fluoride accumulation and damage. Our previous studies revealed fluoride-induced hepatic injury through interleukin 17A (IL-17A) pathway, but the underlying cellular mechanism remains unclear. Hence, this research explored the mechanism of IL-17A pathway and mitophagy in fluoride-induced liver injury through the use of the mice fluorosis model, IL-17A addition fluorosis cell model, IL-17A gene knockout mice fluorosis model, flow cytometry, immunohistochemistry, fluorescence double staining, ELISA, western blotting, and other techniques. The results showed that fluoride reduced the bodyweight and liver coefficient, increased the bone fluoride content, the aspartate aminotransferase (AST), alanine aminotransferase (ALT), glutamate dehydrogenase (GDH) levels, caspase 8 and caspase 9 activities, and induced liver morphology and ultrastructure damage. Furthermore, the protein expression levels of IL-17A pathway key proteins, IL-17A, IL-17R, and Act1 were increased, but IκB was decreased after fluoride exposure. In addition, fluoride exposure elevated the mitochondrial depolarization percent, the mitochondria damage, the fluorescent spots of mitophagy, and the LC3II/LC3I protein relative expression level. To further verify the role of the IL-17A pathway in fluoride-induced hepatocyte mitochondrial damage and mitophagy disorder, the IL-17A was added and knocked out in cells of animals. The results showed that the addition of IL-17A aggravated fluoride-induced liver morphology and functional damage, activation of the IL-17A pathway, mitochondrial injury, and mitophagy, but the IL-17A knockout mitigated fluoride-induced changes. These results suggested that fluoride exposure induced mitochondrial damage and mitophagy through the IL-17A pathway in hepatocytes.

3.
Toxicol Res (Camb) ; 10(4): 911-927, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34484683

RESUMO

Cholestasis is a severe clinical complication that severely damages the liver. Kidneys are also the most affected extrahepatic organs in cholestasis. The pivotal role of oxidative stress has been mentioned in the pathogenesis of cholestasis-induced organ injury. The activation of the nuclear factor-E2-related factor 2 (Nrf2) pathway is involved in response to oxidative stress. The current study was designed to evaluate the potential role of Nrf2 signaling activation in preventing bile acids-induced toxicity in the liver and kidney. Dimethyl fumarate was used as a robust activator of Nrf2 signaling. Rats underwent bile duct ligation surgery and were treated with dimethyl fumarate (10 and 40 mg/kg). Severe oxidative stress was evident in the liver and kidney of cholestatic animals (P < 0.05). On the other hand, the expression and activity of Nrf2 and downstream genes were time-dependently decreased (P < 0.05). Moreover, significant mitochondrial depolarization, decreased ATP levels, and mitochondrial permeabilization were detected in bile duct-ligated rats (P < 0.05). Histopathological alterations included liver necrosis, fibrosis, inflammation and kidney interstitial inflammation, and cast formation. It was found that dimethyl fumarate significantly decreased hepatic and renal injury in cholestatic animals (P < 0.05). Based on these data, the activation of the cellular antioxidant response could serve as an efficient therapeutic option for managing cholestasis-induced organ injury.

4.
Toxicol Lett ; 349: 12-29, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34089816

RESUMO

The cholestatic liver injury could occur in response to a variety of diseases or xenobiotics. Although cholestasis primarily affects liver function, it has been well-known that other organs such as the kidney could be influenced in cholestatic patients. Severe cholestasis could lead to tissue fibrosis and organ failure. Unfortunately, there is no specific therapeutic option against cholestasis-induced organ injury. Hence, finding the mechanism of organ injury during cholestasis could lead to therapeutic options against this complication. The accumulation of potentially cytotoxic compounds such as hydrophobic bile acids is the most suspected mechanism involved in the pathogenesis of cholestasis-induced organ injury. A plethora of evidence indicates a role for the inflammatory response in the pathogenesis of several human diseases. Here, the role of nuclear factor-kB (NFkB)-mediated inflammatory response is investigated in an animal model of cholestasis. Bile duct ligated (BDL) animals were treated with sulfasalazine (SSLZ, 10 and 100 mg/kg, i.p) as a potent inhibitor of NFkB signaling. The NFkB proteins family activity in the liver and kidney, serum and tissue levels of pro-inflammatory cytokines, tissue biomarkers of oxidative stress, serum markers of organ injury, and the liver and kidney histopathological alterations and fibrotic changes. The oxidative stress-mediated inflammatory-related indices were monitored in the kidney and liver at scheduled time intervals (3, 7, and 14 days after BDL operation). Significant increase in serum and urine markers of organ injury, besides changes in biomarkers of oxidative stress and tissue histopathology, were evident in the liver and kidney of BDL animals. The activity of NFkB proteins (p65, p50, p52, c-Rel, and RelB) was significantly increased in the liver and kidney of cholestatic animals. Serum and tissue levels of pro-inflammatory cytokines (IL-1ß, IL-2, IL-6, IL-7, IL-12, IL-17, IL-18, IL-23, TNF-α, and INF-γ) were also higher than sham-operated animals. Moreover, TGF- ß, α-SMA, and tissue fibrosis (Trichrome stain) were evident in cholestatic animals' liver and kidneys. It was found that SSLZ (10 and 100 mg/kg/day, i.p) alleviated cholestasis-induced hepatic and renal injury. The effect of SSLZ on NFkB signaling and suppression of pro-inflammatory cytokines could play a significant role in its protective role in cholestasis. Based on these data, NFkB signaling could receive special attention to develop therapeutic options to blunt cholestasis-induced organ injury.


Assuntos
Anti-Inflamatórios/farmacologia , Colestase/tratamento farmacológico , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Nefropatias/prevenção & controle , Rim/efeitos dos fármacos , Cirrose Hepática/prevenção & controle , Fígado/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Sulfassalazina/farmacologia , Animais , Colestase/metabolismo , Colestase/patologia , Ducto Colédoco/cirurgia , Modelos Animais de Doenças , Regulação para Baixo , Rim/metabolismo , Rim/patologia , Nefropatias/metabolismo , Nefropatias/patologia , Ligadura , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais
5.
Biol Trace Elem Res ; 2021 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-33629228

RESUMO

Excessive fluoride (F) exposure can lead to liver damage; moreover, recent studies found that the addition of appropriate calcium (Ca) can alleviate the symptom of skeletal fluorosis. However, whether Ca can relieve F-induced liver damage through the mitochondrial apoptosis pathway has not been reported yet. Therefore, we assessed the liver morphology, serum transaminase content, liver oxidative stress-related enzymes, and apoptosis-related gene and protein expression in Sprague Dawley (SD) rats treated with 150 mg/L sodium fluoride (NaF) and different concentrations of calcium carbonate (CaCO3) for 120 days. Our results showed that NaF brought out pathological changes in liver morphology, serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels increased, total antioxidant capacity (T-AOC), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) content decreased, and malondialdehyde (MDA) content increased, suggesting that NaF caused hepatotoxicity and oxidative stress. In addition, the results of quantitative real-time PCR (qRT-PCR) and immunohistochemistry showed that NaF exposure upregulated the expression of Bcl-2-associated x protein (Bax), rho-related coiled-coil kinase 1 (ROCK1), cytochrome C (Cyto-C) mRNA and protein (P < 0.01), and downregulated B cell lymphoma 2 (Bcl-2) protein and mRNA (P < 0.01), indicating that excessive F exposure activated mitochondrial-mediated apoptosis in the liver. However, the addition of 1% CaCO3 to the diet significantly increased the expression of anti-apoptotic gene Bcl-2 (P < 0.01), inhibited the activation of the mitochondrial apoptosis pathway, and reduced mitochondrial damage. In summary, supplementing 1% CaCO3 in the diet can alleviate the NaF-induced liver cell damage through the mitochondrial apoptosis pathway.

6.
Biol Trace Elem Res ; 199(4): 1493-1500, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32710348

RESUMO

To investigate the mechanism of fluoride-induced splenic toxicity, 0, 25, 50, and 100 mg/L sodium fluoride (NaF) were administered in male mice via drinking water for 90 days. After NaF treatment, the histological structure of the spleen, the proportion of helper T 1 cell (Th1) and helper T 2 cell (Th2), and the relative expression levels of cytokines and T-bet and GATA3 were analyzed. The results showed that 50 and 100 mg/L NaF consumption can change the normal structure of mouse spleen and the proportion of Th1/Th2 cells. It also decreased the mRNA expression levels of IL-2, INF-γ, and TGF-ß, but increased the levels of IL-4, IL-6, and IL-10. Importantly, fluoride increased the protein expression of GATA3 but decreased the expression of T-bet. Our findings indicate that superfluous fluoride intake damages the balance of Th1/Th2 cells by changing the levels of T-bet and GATA3 in the spleen, and further changes the expression of Th1/Th2 cell-related cytokines in the spleen microenvironment, eventually leading to spleen injury.


Assuntos
Fluoretos , Células Th2 , Animais , Masculino , Camundongos , Baço , Proteínas com Domínio T/genética , Células Th1
7.
Biol Trace Elem Res ; 199(5): 1919-1928, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-32710350

RESUMO

For this study, we investigate more deeply the effect calcium (Ca) develops on the mechanism underlying fluoride-triggered osteocyte apoptosis. We detected the morphology of osteocytes by HE staining, mitochondrial microstructure by using the transmission electron microscope, and the biochemical indexes related to bone metabolism and the expression of apoptosis-related genes. These results showed that NaF brought out the reduced osteocytes and ruptured mitochondrial outer membrane, with a significantly increased StrACP activity by 10.414 IU/L at the 4th week (P < 0.05), markedly upregulating the mRNA expression of Bax, Cyto-C, Apaf-1, caspase-7, ROCK-1, BMP-2, and BGP (P < 0.01), as well as caspase-6 (P < 0.05), while downregulating Bcl-2 by 61.3% (P < 0.01). Through immunohistochemical analysis, we also found that NaF notably increased the protein expression of ROCK-1 (P < 0.05) and Cyto-C, BMP-2, and BGP (P < 0.01), suggesting that NaF triggered the activation of the mitochondrial apoptotic pathway and Rho/ROCK signaling pathway. Nevertheless, 1% Ca supplementation in diet notably enhanced the mRNA expression of Bcl-2 by 39.3% (P < 0.01), thus blocking the increment of the expression of mitochondrial apoptotic pathway-related genes and ROCK-1. Meanwhile, Ca could attenuate the StrACP activity by 10.741 IU/L at the 4th week (P < 0.05) and protect the integrity of the mitochondrial outer membrane. These findings strongly suggest that 1% Ca abated the mitochondrial apoptosis pathway by increasing the anti-apoptotic gene Bcl-2 expression, and effectively inhibited the hyper-activation of ROCK-1, dually protecting the structural integrity of the mitochondrial outer membrane and maintaining normal cellular metabolic function.


Assuntos
Cálcio , Intoxicação por Flúor , Animais , Apoptose , Mitocôndrias , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Proteína X Associada a bcl-2
8.
Chemosphere ; 269: 128727, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33213873

RESUMO

Excessive fluoride intake can damage testis by breaking the integrity of sperm DNA and changing the expression profiles of testicular mRNAs and microRNAs. However, the effects of fluoride on the expression of PIWI-interacting RNAs (piRNAs) in mouse testes have not been reported. In this study, we determined the effect of fluoride on PIWI-interacting RNA expression profiling in testis of mice, using deep-sequencing technology. Compared to the control, 50 mg/L sodium fluoride (NaF) exposure led to a reduced testicular organ coefficient, semen quality, and testosterone level, and altered the testicular microstructure. Furthermore, NaF exposure also changed the expression of 28 piRNAs that regulate 182 target genes in mouse testes. In mice given water containing 50 mg/L NaF, the following four pathways were enriched and overexpressed: lysosomal, Jak-STAT, chemokine, and ubiquitin-mediated proteolysis. Among the piRNAs affecting the lysosomal pathway, piR-mmu-1277316, piR-mmu-8060747, and piR-mmu-1566415 levels were increased. We also observed increased levels of the following target gene mRNAs in lysosomal pathwa in the 50 mg/L NaF-treated group: Gga2, Ap4e1, Gla, and Ap1s3. These findings are in line with the results of piRNA-sequencing and suggest that piRNAs in the testis could be potential biomarkers for fluoride reproductive toxicity.


Assuntos
Fluoretos , Testículo , Proteínas Adaptadoras de Transporte Vesicular , Animais , Fluoretos/toxicidade , Humanos , Masculino , Camundongos , RNA Interferente Pequeno/genética , Análise do Sêmen , Espermatozoides
9.
Toxicol In Vitro ; 72: 105074, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33352257

RESUMO

Arsenic (As), a potent toxicant, is known to be a hepatotoxicant. Although As induced liver apoptosis and autophagy, the relationship between apoptosis and autophagy of hepatocytes caused by As remains largely unknown. 3-methyladenine (3-MA) and rapamycin can inhibit and promote autophagy of AML-12 cells, respectively. Hence, in this study, AML-12 cells were treated with different concentrations (0, 2, 4, 6, 8, 10 and 12 µmol/L) of As2O3, and 5 mmol/L 3-MA or 100 nmol/L rapamycin were applied to distinguish the effect of autophagy on apoptosis in AML-12. Results showed that exposure to As induced cell apoptosis and autophagy, which were mediated by the significantly altered expression levels of autophagy markers (mTOR, LC3, PI3K and P62), and apoptosis markers (Bcl-2 and caspase-3). Further analysis indicated that a certain dosage of 3-MA and rapamycin decreased apoptosis and the caspase-3 expression, which suggested that As-induced autophagy regulated AML-12 cells apoptosis through the expressions of PI3K, mTOR, P62 and Bcl-2.

10.
Chemosphere ; 263: 128178, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33297146

RESUMO

The reproductive toxicity of fluoride (F) has been verified by various epidemiological and experimental studies. Our previous work suggested that the interleukin 17A (IL-17A) is involved in the testicular damage induced by excessive F exposure. In this study, we further investigated the role of IL-17A in F-induced testicular injury. Wild type (WT) and IL-17A knockout (IL-17A-/-) mice were exposed to 0, 25, 50, or 100 mg/L sodium fluoride (NaF) for 90 days. We found that exposure to excessive F levels caused testicular damage, decreased semen quality, negatively affected testicular morphology, and increased the inflammatory response. Specifically, excessive F intake increased the expression levels of IL-17A in the testis and increased the protein levels of Act1, NF-κB, IL-17R, C/EBP-α, and TRAF6 in the IL-17A signaling pathway. The increase in IL-17A expression corresponded to increases expression of IL-17R, IL-6, IL-23, IL-1ß, TGF-ß and TNF-α as assessed by RT-PCR and ELISA assays. Remarkably, IL-17A knockout in mice ameliorated the effects of F on testicular damage, semen quality, testicular morphology, and the immune response. Additionally, we found the in vitro exposure of Leydig cells to NaF and recombinant IL-17A led to abnormal apoptosis and a decrease in testosterone secretion. Our findings prove that IL-17A plays a key role in the exacerbation of testicular injuries in F-exposed mice, and that IL-17A deficiency can alleviate F-induced injury by inhibiting the immune response and apoptosis in the testis. These data suggest that targeting IL-17A may be a useful therapeutic strategy for treating F-mediated toxicity in the testis.


Assuntos
Fluoretos , Interleucina-17 , Animais , Apoptose , Humanos , Imunidade , Interleucina-17/genética , Masculino , Camundongos , Análise do Sêmen
11.
Sci Total Environ ; 742: 140533, 2020 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-32721723

RESUMO

Increasing investigations suggest that fluoride (F) exposure was associated with gastrointestinal diseases, but related literatures were still largely insufficient and the underlying mechanisms have not been fully elucidated. Moreover, previous study in our lab reported F toxicity has the reversible tendency, but it still needs to be further explored. To address this issue, we established a 90 days F exposure and 15 days & 30 days self-recovery mice model, including control and three F groups (25, 50 and 100 mg/L sodium fluoride (NaF)) in each period. The results revealed that after 90 days F exposure, histological structure and ultrastructure of small intestine were markedly disrupted; the value of villus height to crypt depth, and expressions of tight junctions related mRNA and proteins were significantly decreased; intestinal permeability, pro-inflammatory cytokines and pyroptosis related mRNA and proteins were notably increased in duodenum, jejunum and ileum. However, intriguingly, after 30 days recovery period, indices in F groups almost all have recovered towards normalcy. Collectively, this study demonstrated that F exposure could impair the structure and epithelial barrier function of small intestine, leading to the intestinal inflammation, and pyroptosis may contribute to this damage; Furthermore, F toxicity on small intestine is reversible, and could be restored when off the F exposure environment for a certain period of time. Additionally, among the three regions of small intestine, duodenum seems more vulnerable to F exposure than jejunum and ileum.


Assuntos
Fluoretos , Piroptose , Animais , Inflamação , Intestino Delgado , Jejuno , Camundongos
12.
Chemosphere ; 256: 127105, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32450357

RESUMO

Previous studies have shown that waterborne fluoride exposure has adverse effects on the reproductive system of zebrafish. However, the underlying toxic mechanisms were still not clear. In the present study, female zebrafish were exposed to different concentrations of 0.787 (Control), 18.599, 36.832 mg/L of fluoride for 30 d and 60 d, and the effects of different doses of fluoride on ovary development, reproductive hormones, oogenesis, ROS content, antioxidant levels, and the expression of apoptosis-related genes and proteins in the ovaries of female zebrafish were analyzed. The results showed that ovarian weight and GSI were significantly decreased, FSH, LH and VTG levels were significantly reduced, the transcriptional profiles of oogenesis-related genes (tgfß1, bmp15, gdf9, mprα, mprß, ptg2ß) were remarkably altered, ROS levels was notably increased, the SOD, CAT, GPx activities and GSH content as well as their mRNA expressions were significantly decreased, MDA content was remarkably increased, the expressions of apoptosis-related genes and proteins (caspase3, caspase8, caspase9, Fas-L, Cytochrome C, Bax and Bcl-2) were significantly changed, the ratio of Bax/Bcl-2 protein levels were notably increased. Taken together, this study demonstrated that fluoride exposure significantly affected ovarian development, decreased the reproductive hormones, affected oogenesis, induced oxidative stress, caused apoptosis through both extrinsic and intrinsic pathways in ovary of zebrafish. Indicating that oogenesis, oxidative stress, and apoptosis were responsible for the impairment of ovarian development.


Assuntos
Fluoretos/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Feminino , Oogênese/efeitos dos fármacos , Ovário/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Diferenciação Sexual , Peixe-Zebra/metabolismo
13.
Chemosphere ; 246: 125772, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31901658

RESUMO

Excessive fluoride (F) ingestion via drinking water interfered with spermatogenesis and lowered sperm quality of human and animals. However, it is still unclear why the effects of fluoride on sperm quality focus on mostly sperm motility rather than sperm count. The objective of this study is to investigate the potential relationship between alteration in the structure and function of sperm flagellum and fluoride exposure in the environment. 40 male mice were allocated to four groups which were treated with 0, 25, 50, 100 mg/L NaF deionized water, respectively, for 8 weeks continuously. The testicular morphology, ultra-structure of fibrous sheath and axoneme of sperm flagellum, and eleven key function genes Akap3, Akap4, Dnah1, Eno4, Cfap43, Cfap44, Hydin, Spef2, Spag6, Spag16, and Cfap69 were examined by histology, transmission electron microscopy, and real-time PCR methods respectively. The results displayed that fluoride damaged the typical "9 + 2″ microtubule structure including fibrous sheathes and axoneme of sperm flagellum in testes of mice. Furthermore, the mRNA and protein expression levels of AKAP3 and AKAP4 related to fibrous sheathes formation, and CFAP43, CFAP44 and HYDIN in axoneme were down-regulated by fluoride exposure. Taken together, we revealed that fluoride altered the structures of the fibrous sheathes and axonemal in sperm flagellum via down-regulating the mRNA and protein expression levels of AKAP3, AKAP4, CFAP43, CFAP44, and HYDIN, which may be one of the reasons that fluoride lowered sperm quality and male reproductive function.


Assuntos
Fluoretos/toxicidade , Cauda do Espermatozoide/ultraestrutura , Testículo/metabolismo , Proteínas de Ancoragem à Quinase A , Animais , Dineínas , Poluentes Ambientais , Fluoretos/metabolismo , Humanos , Infertilidade Masculina , Masculino , Camundongos , Proteínas dos Microtúbulos , Fenótipo , RNA Mensageiro/metabolismo , Motilidade Espermática , Cauda do Espermatozoide/efeitos dos fármacos , Espermatogênese , Espermatozoides/metabolismo
14.
Biol Trace Elem Res ; 193(1): 195-203, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30887282

RESUMO

The gap junction protein plays an important role in the bone formation and alteration of these proteins leading to cause bone development. Aim to determine the effects of different concentration of fluoride on gap-junctional intercellular communication (GJIC) related genes and proteins in the rats' osteoblast cells. We treated the osteoblast cells with various concentrations (0, 0.01, 0.1, 0.5, and 1.0 mM) NaF for 24 and 72 h. We used the scrape loading and dye transfer technique to research the intracellular connectivity. Moreover, the mRNA expression levels of connexin 43 (Cx43), connexin45 (Cx45), collagen I, and osteocalcin (OCN) were analyzed by qRT-PCR, the protein expression levels of connexin43 (Cx43) were analyzed by western blotting and immunofluorescence. Our results suggested that the osteoblast proliferations were decreased in the 0.5 and 1 mM NaF groups, after 24 and 72 treatments. The scrape loading and dye transfer experiment showed that the GJIC were increased in the 0.01 mM NaF group and decreased in the 0.5 and 1 mM NaF groups. In addition, the mRNA expressions of Cx43, Cx45, and OCN, and the protein expressions of Cx43 were increased in the 0.01 mM NaF group and decreased in the 0.5 and 1 mM NaF groups. In summary, these results suggest that the low concentration NaF is good for the GJIC, but the high concentration NaF damages the GJIC.


Assuntos
Comunicação Celular/efeitos dos fármacos , Fluoretos/farmacologia , Junções Comunicantes/metabolismo , Osteoblastos/metabolismo , Animais , Células Cultivadas , Conexina 43/biossíntese , Conexinas/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Osteocalcina/biossíntese , Ratos
15.
Food Funct ; 11(1): 1155-1164, 2020 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-31872845

RESUMO

Bone is the main target of fluorosis, and it has been perfectly elaborated that a moderate dosage of calcium (Ca) can alleviate bone fluorosis. However, whether Ca can alleviate fluorosis through the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT) signaling pathway has not yet been reported. Hence, we evaluated the histopathological structure, the imbalance of the biochemical index of bone metabolism, and the expression levels of PI3K/AKT apoptosis signaling pathway-related genes in rats treated with sodium fluoride (NaF, F) and/or calcium carbonate (CaCO3) for 120 days. Our results suggest that 100 mg L-1 NaF induced histopathological injury as alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (StrACP) activity increased, with a decrease in the serum Ca levels (p < 0.05). Moreover, the results of qRT-PCR and western blotting showed that F increased the expression levels of transglutaminase 2 (TGM2), focal adhesion kinase (FAK), PI3K, AKT, forkhead box O1 (Foxo1), Bcl-2 interacting mediator of cell death (BIM), Bcl2-associated x protein (Bax) and Caspase 3 (p < 0.05, p < 0.01). It also decreased the expression of AnnexinA5 (Anxa5), 3'-phosphoinositide-dependent kinase 1 (PDK1) and B-cell lymphoma-2 (Bcl-2) (p < 0.05, p < 0.01), which finally activated the PI3K/AKT pathway. On the other hand, CaCO3 supplementation reversed the histopathological injury along with the levels of ALP, StrACP and serum Ca, alleviating the gene expression levels of PI3K/AKT pathway-related markers. Altogether, we can conclude that CaCO3 supplementation mitigated F-induced bone damage via the PI3K/AKT signaling pathway.


Assuntos
Osso e Ossos/efeitos dos fármacos , Cálcio/metabolismo , Fluoretos/efeitos adversos , Transdução de Sinais , Animais , Apoptose , Osso e Ossos/patologia , Intoxicação por Flúor/terapia , Masculino , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-Dawley
16.
J Agric Food Chem ; 67(37): 10285-10295, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31443611

RESUMO

Fluoride (F) is capable of promoting abnormal proliferation and differentiation in primary cultured mouse osteoblasts (OB cells), although the underlying mechanism responsible remains rare. This study aimed to explore the roles of wingless and INT-1 (Wnt) signaling pathways and screen appropriate doses of calcium (Ca2+) to alleviate the sodium fluoride (NaF)-induced OB cell toxicity. For this, we evaluated the effect of dickkopf-related protein 1 (DKK1) and Ca2+ on mRNA levels of wingless/integrated 3a (Wnt3a), low-density lipoprotein receptor-related protein 5 (LRP5), dishevelled 1 (Dv1), glycogen synthase kinase 3ß (GSK3ß), ß-catenin, lymphoid enhancer binding factor 1 (LEF1), and cellular myelocytomatosis oncogene (cMYC), as well as Ccnd1 (Cyclin D1) in OB cells challenged with 10-6 mol/L NaF for 24 h. The demonstrated data showed that F significantly increased the OB cell proliferation rate. Ectogenic 0.5 mg/L DKK1 significantly inhibited the proliferation of OB cells induced by F. The mRNA expression levels of Wnt3a, LRP5, Dv1, LEF1, ß-catenin, cMYC, and Ccnd1 were significantly increased in the F group, while significantly decreased in the 10-6 mol/L NaF + 0.5 mg/L DKK1 (FY) group. The mRNA expression levels of Wnt3a, LRP5, ß-catenin, and cMYC were significantly decreased in the 10-6 mol/L NaF + 2 mmol/L CaCl2 (F+CaII) group. The protein expression levels of Wnt3a, Cyclin D1, cMYC, and ß-catenin were significantly increased in the F group, whereas they were decreased in the F+CaII group. However, the mRNA and protein expression levels of GSK3ß were significantly decreased in the F group while significantly increased in the F+CaII group. In summary, F activated the canonical Wnt/ß-catenin pathway and changed the related gene expression and ß-catenin protein location in OB cells, promoting cell proliferation. Ca2+ supplementation (2 mmol/L) reversed the expression levels of genes and proteins related to the canonical Wnt/ß-catenin pathway.


Assuntos
Cálcio/metabolismo , Fluoretos/efeitos adversos , Osteoblastos/efeitos dos fármacos , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Suplementos Nutricionais/análise , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , Camundongos , Osteoblastos/classificação , Osteoblastos/metabolismo , Proteínas Wnt/genética , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/genética
17.
J Agric Food Chem ; 67(39): 10832-10843, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31464433

RESUMO

Excessive fluoride mainly causes skeletal lesions. Recently, it has been reported that an appropriate level of calcium can alleviate fluorosis. However, the appropriate concentration and mechanism of calcium addition is unclear. Hence, we evaluated the histopathology and ultrastructure, DNA fragmentation, hormonal imbalances, biomechanical levels, and expression of apoptosis-related genes after treating the rats with 150 mg/L NaF and different concentrations of CaCO3. Our results suggested that NaF induced the histopathological and ultrastructural injury, with a concomitant increase in the DNA fragmentation (P < 0.05) and serum OC (17.5 ± 0.89 pmoL/L) at 120 days. In addition, the qRT-PCR and western blotting results indicated that NaF exposure upregulated the mRNA and protein expression of Bax, Calpain, Caspase 12, Caspase 9, Caspase 7, Caspase 3, CAD, PARP, and AIF while downregulated Bcl-2 (P < 0.01) and decreased the bone ultimate load by 27.1%, the ultimate stress by 10.1%, and the ultimate deformity by 23.3% at 120 days. However, 1% CaCO3 supplementation decreased the serum OC (14.7 ± 0.65 pmoL/L), bone F content (P < 0.01), and fracture and breakage of collagen fibers and changed the expression of endoplasmic reticulum pathway-related genes and proteins at 120 days. Further, 1% CaCO3 supplementation increased the bone ultimate load by 20.9%, the ultimate stress by 4.89%, and the ultimate deformity by 21.6%. In summary, we conclude that 1% CaCO3 supplementation alleviated fluoride-induced bone damage by inhibiting endoplasmic reticulum stress and mitochondrial dysfunction.


Assuntos
Osso e Ossos/efeitos dos fármacos , Cálcio/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Fluoretos/toxicidade , Mitocôndrias/efeitos dos fármacos , Animais , Osso e Ossos/metabolismo , Caspases/genética , Caspases/metabolismo , Fragmentação do DNA/efeitos dos fármacos , Masculino , Mitocôndrias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Ratos Sprague-Dawley
18.
Chemosphere ; 226: 201-209, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30927672

RESUMO

The mechanism of GSTO1, as a high-risk factor for neurological damage, in sodium fluoride (NaF)-induced learning and memory impairment remained still unclear. Hence, in this study, we used the siRNA-GSTO1 HT22 model to explore the effect of NaF and siRNA-GSTO1 on the viability, and proliferation rate of HT22 cells, as well as the mRNA and protein expression levels of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB), neural cell adhesion molecule (NCAM), stem cell factor (SCF) and brain-derived neurotrophic factor (BDNF). The results of MTT showed that 10-3, 10-4, and 10-5 moL/L sodium fluoride (NaF) exposure could significantly promote the proliferation of HT22 cells at 24 h, 36 h, and 48 h, respectively. In addition, our results showed that exposure to 10-3, 10-4, and 10-5 moL/l NaF increased GSTO1 mRNA and protein expression, but decreased CREB and BDNF expression levels in a dose and time-dependent manner. The mRNA and protein expressions of GSTO1, CREB and BDNF were significantly decreased in the siRNA-GSTO1 and NaF + siRNA-GSTO1 group (P < 0.05). We have shown that various NaF doses affected the learning and memory ability by down-regulation the expressions of CREB, BDNF, NCAM and SCF. In summary, we concluded that GSTO1 plays a mediator role in NaF-induced neurological damage.


Assuntos
Fator Neurotrófico Derivado do Encéfalo , Proteínas de Transporte/fisiologia , Glutationa Transferase/fisiologia , Hipocampo/efeitos dos fármacos , Moléculas de Adesão de Célula Nervosa , Fluoreto de Sódio/efeitos adversos , Animais , Fator Neurotrófico Derivado do Encéfalo/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Linhagem Celular , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/efeitos dos fármacos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regulação para Baixo/efeitos dos fármacos , Glutationa Transferase/efeitos dos fármacos , Glutationa Transferase/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Deficiências da Aprendizagem/induzido quimicamente , Transtornos da Memória/induzido quimicamente , Camundongos , Moléculas de Adesão de Célula Nervosa/efeitos dos fármacos , Moléculas de Adesão de Célula Nervosa/metabolismo , Fator de Células-Tronco/efeitos dos fármacos , Fator de Células-Tronco/metabolismo
19.
Arch Toxicol ; 92(11): 3277-3289, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30225638

RESUMO

Previous studies have reported that excessive fluoride exposure induced liver damage. However, the underlying mechanism of fluoride-induced hepatic toxicity is still unclear. Hence, this study was aimed to evaluate the fluoride-induced apoptosis, autophagy, and IL-17 signaling pathway-related genes to explore the possible mechanisms of NaF-induced liver injury in mice. For this, 48 male mice were allotted randomly to four groups, treated with deionized water, 25, 50, 100 mg/L NaF for 150 days continuously. Our results suggested that treatment with NaF decreased the PAS staining-positive area, with a concomitant increase in liver score, and serum ALT and AST levels which indicated that NaF induced the liver injury. In addition, the qRT-PCR, immunohistochemistry, and western blotting results indicated that NaF exposure activated IL-17 signaling, apoptosis, and autophagy pathways. In summary, these results suggested that NaF induced apoptosis and autophagy in liver by activating the IL-17 signaling pathway, eventually leading to impaired liver function.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Interleucina-17/fisiologia , Transdução de Sinais/efeitos dos fármacos , Fluoreto de Sódio/toxicidade , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Hepatócitos/patologia , Hepatócitos/ultraestrutura , Masculino , Camundongos , Células Th17/efeitos dos fármacos , Células Th17/fisiologia
20.
Food Chem Toxicol ; 116(Pt B): 189-195, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29642045

RESUMO

In reviewing the literature, the cellular mechanism of fluoride F-induced osteoblast OB cells apoptosis is diverse and perplexing, but detailed regulatory pathway, targets and role of extracellular Ca2+ remains still unclear. Hence, in the present study, we investigated the effects of F (9 mg/L F ion) and different Ca2+ (0.5, 1, 2, 4, 8 mmol/L) levels treatment on the proliferation rate of osteoblast cells, intracellular free Ca2+ ([Ca2+]i) and endoplasmic reticulum (ER) stress apoptosis pathway related gene levels of rabbit OB cells. Our results demonstrated that F exposure had a pronounced negative effect on osteoblast survival, further different Ca2+ levels treatment suggested that low concentration of Ca2+ (0.5-1 mmol/L) relieved the damaged effect, on the contrary, high concentration of Ca2+ (2-8 mmol/L) enhanced the effect. In addition, F significantly increased [Ca2+]i levels and the expression of ER stress-induced cell apoptosis pathway related genes. Treatment with 0.5-1 mmol/L Ca2+ markedly reversed the F-induced harmful effects, but high dose Ca2+ (2-8 mmol/L) enhanced these effects. In summary, 0.5-1 mmol/L Ca2+ can alleviate F-induced OB cells injure through ER stress apoptosis pathway, which provided a dose basis for the future study on the treatment of skeletal fluorosis with Ca2+.


Assuntos
Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Fluoretos/metabolismo , Osteoblastos/efeitos dos fármacos , Animais , Coelhos
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