Abstract

OBJECTIVE:  Quantitative real-time PCR (qPCR) is routinely performed for experiments designed to identify the molecular mechanisms involved in the pathogenesis of dental fluorosis. Expression of reference gene(s) is expected to remain unchanged in fluoride-treated cells or in rodents relative to the corresponding untreated controls. The aim of this study was to select optimal reference genes for fluoride experiments performed in vitro and in vivo.

DESIGN:  Five candidate genes were evaluated: B2m, Eef1a1, Gapdh, Hprt and Tbp. For in vitro experiments, LS8 cells derived from mouse enamel organ were treated with 0, 1, 3 and/or 5mM sodium fluoride (NaF) for 6 or 18h followed by RNA isolation. For in vivo experiments, six-week old rats were treated with 0 or 100ppm fluoride as NaF for six weeks at which time RNA was isolated from enamel organs. RNA from cells and enamel organs were reverse-transcribed and stability of gene expression for the candidate reference genes was evaluated by qPCR in treated versus non-treated samples.

RESULTS:  The most stably expressed genes in vitro according to geNorm were B2m and Tbp, and according to Normfinder were Hprt and Gapdh. The most stable genes in vivo were Eef1a1 and Gapdh. Expression of Ddit3, a gene previously shown to be induced by fluoride, was demonstrated to be accurately calculated only when using an optimal reference gene.

CONCLUSIONS:  This study identifies suitable reference genes for relative quantification of gene expression by qPCR after fluoride treatment both in cultured cells and in the rodent enamel organ.

• Related Studies :

  • Possible Association Between Polymorphisms in ESR1, COL1A2, BGLAP, SPARC, VDR, and MMP2 Genes and Dental Fluorosis in a Population from an Endemic Region of West Bengal.

    Dental fluorosis (DF) is the most prevalent form of fluorosis in India affecting millions of people all over the country. As estrogen receptor 1 (ESR1), collagen type 1 alpha 2 (COL1A2), bone ?-carboxyglutamic acid protein (BGLAP), secreted protein acidic and cysteine-rich (SPARC), vitamin D receptor (VDR), and matrix metallopeptidase 2

  • Dental fluorosis and a polymorphism in the COL1A2 gene in Mexican children.

    Highlights Dental fluorosis is a public health problem in the communities evaluated. The rs 412777 polymorphism in the COL1A2 gene was found in Mexican children. An association between the COL1A2 gene polymorphism and dental fluorosis was found. The genetic variant evaluated represents a risk factor to develop dental fluorosis. OBJECTIVE:

  • Multiomics Analysis Revealed the Molecular Mechanism of miRNAs in Fluoride-Induced Hepatic Glucose and Lipid Metabolism Disorders.

    Fluoride-induced liver injury seriously endangers human and animal health and animal food safety, but the underlying mechanism remains unclear. This study aims to explore the mechanism of miRNAs in fluoride-induced hepatic glycolipid metabolism disorders. C57 male mice were used to establish the fluorosis model (22.62 mg/L F–, 12 weeks). The

  • Evaluation of genetic polymorphisms in MMP2, MMP9 and MMP20 in Brazilian children with dental fluorosis.

    Highlights MMP2, MMP9 and MMP20 were expressed in the enamel development of the animalmodels. Polymorphisms in MMP2, MMP9 and MMP2 were not associated with dental fluorosis. Afro-descendants had a higher risk of dental fluorosis than caucasian. Recent studies suggested that genetics contribute to differences in dental fluorosis (DF) susceptibility among individuals

  • Collagenase 1A2 (COL1A2) gene A/C polymorphism in relation to severity of dental fluorosis

    OBJECTIVES:  The aim of this study was to evaluate the putative association between the presence of the COL1A2 gene A/C polymorphism and the severity of dental fluorosis in a sample exposed to high concentrations of fluoride. METHODS:  A cross-sectional study was carried out that included 80 children residing in a community

• Related FAN Content :

  • Dental Fluorosis Is a "Hypo-mineralization" of Enamel

    Teeth with fluorosis have an increase in porosity in the subsurface enamel ("hypomineralization"). The increased porosity of enamel found in fluorosis is a result of a fluoride-induced impairment in the clearance of proteins (amelogenins) from the developing teeth. Despite over 50 years of research, the exact mechanism by which fluoride impairs amelogin

  • Mechanisms by Which Fluoride Causes Dental Fluorosis Remain Unknown

    When it comes to how fluoride impacts human health, no tissue in the body has been studied more than the teeth. Yet, despite over 50 years of research, the mechanism by which fluoride causes dental fluorosis (a hypo-mineralization of the enamel that results in significant staining of the teeth) is not

  • "Mild" Dental Fluorosis: Perceptions & Psychological Impact

    The vast majority of research has found that patients, parents, and the general public alike view mild fluorosis (TF score 3) as a significant blemish of the teeth, one that is likely to embarrass the affected child to a degree that cosmetic treatment would be warranted.

  • Moderate/Severe Dental Fluorosis

    In its "moderate" and severe forms, fluoride causes a marked increase in the porosity of the enamel. After eruption into mouth, the porous enamel of moderate to severe fluorosis readily takes up stain, creating permanent brown and black discolorations of the teeth. In addition to extensive staining, teeth with moderate to severe fluorosis are more prone to attrition and wear - leading to pitting, chipping, and decay.

  • Racial Disparities in Dental Fluorosis

    In 2005, the Centers for Disease Control published the results of a national survey of dental fluorosis conducted between 1999 and 2002. According to the CDC, black children in the United States have significantly higher rates of dental fluorosis than either white or Hispanic children. This was not the first time that black children were found to suffer higher rates of dental fluorosis. At least five other studies -- dating as far back as the 1960s -- have found black children in the United States are disproportionately impacted by dental fluorosis.