Nickel nanoparticles induce epithelial-mesenchymal transition in human bronchial epithelial cells via the HIF-1α/HDAC3 pathway

We and others have previously demonstrated that exposure to nickel nanoparticles (Nano-Ni) caused fibrogenic and carcinogenic effects; however, the underlying mechanisms are still not fully understood. This study aimed to investigate the effects of Nano-Ni on epithelial-mesenchymal transition (EMT) in human bronchial epithelial cells (BEAS-2B) and its underlying mechanisms since EMT is involved in both cancer pathogenesis and tissue fibrosis. Our results showed that exposure to Nano-Ni, compared to the control Nano-TiO2, caused a remarkable decrease in the expression of E-cadherin and an increase in the expression of vimentin and α-SMA, indicating an inducible role of Nano-Ni in EMT development in human bronchial epithelial cells. HIF-1α nuclear accumulation, HDAC3 upregulation, and decreased histone acetylation were also observed in the cells exposed to Nano-Ni, but not in those exposed to Nano-TiO2. Pretreatment of the cells with a specific HIF-1α inhibitor, CAY10585, or HIF-1α-specific siRNA transfection prior to Nano-Ni exposure resulted in the restoration of E-cadherin and abolished Nano-Ni-induced upregulation of vimentin and α-SMA, suggesting a crucial role of HIF-1α in Nano-Ni-induced EMT development. CAY10585 pretreatment also attenuated the HDAC3 upregulation and increased histone acetylation. Inhibition of HDAC3 with specific siRNA significantly restrained Nano-Ni-induced reduction in histone acetylation and restored EMT-related protein expression to near control levels. In summary, our findings suggest that exposure to Nano-Ni promotes the development of EMT in human bronchial epithelial cells by decreasing histone acetylation through HIF-1α-mediated HDAC3 upregulation. Our findings may provide information for further understanding of the molecular mechanisms of Nano-Ni-induced fibrosis and carcinogenesis.</p

Mutation of H231 and D232 residues in the HD-GYP domain of <i>Xoc</i> RpfG disrupts the PDE activity and regulatory function.

<p>(A) Purified RpfG_AA-GYP completely lost its PDE activity detected by colorimetric assays. (B–D) RpfG_AA-GYP lost the ability to restore EPS production (B), biofilm formation (C) and virulence to rice (D) of <i>ΔrpfG</i> to the wild-type level. WT: wild-type, EV: empty vector.</p

Bacterial strains and plasmids.

<p>Bacterial strains and plasmids.</p

Virulence assays of the wild-type and <i>rpfG-, hgdA- and hgdC-</i>related single, double and triple mutant strains on rice cv. Jingang 30.

<p>(A) The length of disease lesions was measured at 14 days after pressure inoculation of the wild-type (WT), <i>ΔrpfG</i>, complemented <i>ΔrpfG (rpfG)</i>, <i>ΔhgdA</i>, <i>ΔhgdA/hgdC</i>, <i>ΔhgdA/rpfG</i>, <i>ΔrpfG/hgdC</i>, <i>ΔhgdA/rpfG/hgdC</i> strains, respectively. Ten to 15 leaves were scored for each strain; means ± standard error (SE) are shown. (B) <i>In planta</i> bacterial populations of <i>Xoc</i> RS105, <i>ΔrpfG</i> and <i>ΔrpfG(rpfG)</i> at the specific time points after inoculation. Data are presented as means ± SE.</p

Effects of mutations of genes encoding several putative glycosyltransferases on biofilm formation and EPS production in <i>Xoc</i> wild-type and <i>ΔrpfG</i> mutant backgrounds.

<p>(A) Elevated biofilm formation in <i>Xoc ΔrpfG</i> was reduced to the wild-type level when <i>xagA</i>, <i>xagB</i> and <i>gumD</i>, but not <i>pgaA</i> or <i>pgaC</i> was deleted singly in the <i>ΔrpfG</i> genotype. The full length <i>xagA</i> and <i>gumD</i> genes restored the ability of biofilm formation in <i>ΔrpfG/xagA</i> and <i>ΔrpfG/gumD</i> mutants, respectively. (B) The ability of <i>ΔgumD</i> to form biofilm was greatly attenuated in the wild-type background, but was restored by complementation. Biofilm formation in the <i>ΔpgaA</i>, <i>ΔpgaC, ΔxagA</i> and <i>ΔxagB</i> single mutants was not altered compared to the wild-type strain. (C) EPS production was significantly reduced in the <i>ΔgumD</i> mutant, but not in <i>ΔpgaA</i>, <i>ΔpgaC, ΔxagA</i> and <i>ΔxagB</i> mutants compared to the wild-type. These experiments were repeated at least three times with similar results.</p

The effect of <i>gumD</i> deletion on bacterial virulence in <i>Xoc.</i>

<p><i> Xoc</i> virulence to rice was greatly attenuated when <i>gumD</i> was deleted, but was unaltered in the <i>ΔpgaA</i>, <i>ΔpgaC, ΔxagA</i> and <i>ΔxagB</i> mutant strains. Virulence of the <i>ΔgumD</i> mutant was restored by the introduction of full-length <i>gumD</i> into the <i>ΔgumD(gumD)</i> strain. The length of disease lesions was measured at 20 days after pressure inoculation. Ten to 15 leaves were scored for each strain; means ± SE are shown. The experiments were repeated three times with similar results.</p

The effect of RpfG on the expression of <i>hrp</i>-related genes in <i>Xoc</i>.

<p>(A) Gene expression of <i>hrpG</i>, <i>hrpX</i> and <i>hrpA</i> in wild-type Xoc (RS105), <i>ΔrpfG</i> and complemented strain <i>ΔrpfG(rpfG)</i> was detected by qRT-PCR. 16S RNA was used as an internal control for data analyses. (B) Gene expression of <i>hrpG</i>, <i>hrpX</i> and <i>hrpA</i> in RS105, <i>ΔrpfG</i> and <i>ΔrpfG(rpfG)</i> strains was examined by GUS activities of appropriate promoter-GUS fusions. WT: wild-type.</p