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Oncogenic transformation of human lung bronchial epithelial cells induced by arsenic involves ROS-dependent activation of STAT3-miR-21-PDCD4 mechanism

View Article: PubMed Central - PubMed

ABSTRACT

Arsenic is a well-documented human carcinogen. The present study explored the role of the onco-miR, miR-21 and its target protein, programmed cell death 4 (PDCD4) in arsenic induced malignant cell transformation and tumorigenesis. Our results showed that treatment of human bronchial epithelial (BEAS-2B) cells with arsenic induces ROS through p47phox, one of the NOX subunits that is the key source of arsenic-induced ROS. Arsenic exposure induced an upregulation of miR-21 expression associated with inhibition of PDCD4, and caused malignant cell transformation and tumorigenesis of BEAS-2B cells. Indispensably, STAT3 transcriptional activation by IL-6 is crucial for the arsenic induced miR-21 increase. Upregulated miR-21 levels and suppressed PDCD4 expression was also observed in xenograft tumors generated with chronic arsenic exposed BEAS-2B cells. Stable shut down of miR-21, p47phox or STAT3 and overexpression of PDCD4 or catalase in BEAS-2B cells markedly inhibited the arsenic induced malignant transformation and tumorigenesis. Similarly, silencing of miR-21 or STAT3 and forced expression of PDCD4 in arsenic transformed cells (AsT) also inhibited cell proliferation and tumorigenesis. Furthermore, arsenic suppressed the downstream protein E-cadherin expression and induced β-catenin/TCF-dependent transcription of uPAR and c-Myc. These results indicate that the ROS-STAT3-miR-21-PDCD4 signaling axis plays an important role in arsenic -induced carcinogenesis.

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Increased miR-21 and suppressed PDCD4 expression by chronic arsenic exposure induces tumorigenesis of BEAS-2B cells in vivo.BEAS-2B cells exposed to indicated concentration of arsenic for 6 months were injected sc into nude mice. After 4 weeks, mice were euthanized using CO2 and tumor was isolated for further examination. (A) Representative images of excised tumors and (B) graphic representation of tumor volume demonstrate increased growth with arsenic treatment. (C) The relative miR-21 level, determined by Taqman real-time PCR, increased with arsenic treatment. (D) Immunohistochemical analysis of PDCD4 protein showed decreased expression with increasing arsenic concentration. (E) Protein levels for PDCD4 and pSTAT3, analyzed by western blot, showed an apparent decrease in PDCD4 and apparent increase in pSTAT3 expressions with arsenic exposure. (F) Increased STAT3 phosphorylation with arsenic exposure was verified by immunohistochemistry. Data presented in the bar graphs are the mean ± SD of three independent experiments. *Indicates a statistically significant difference compared to control with p < 0.05.
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f6: Increased miR-21 and suppressed PDCD4 expression by chronic arsenic exposure induces tumorigenesis of BEAS-2B cells in vivo.BEAS-2B cells exposed to indicated concentration of arsenic for 6 months were injected sc into nude mice. After 4 weeks, mice were euthanized using CO2 and tumor was isolated for further examination. (A) Representative images of excised tumors and (B) graphic representation of tumor volume demonstrate increased growth with arsenic treatment. (C) The relative miR-21 level, determined by Taqman real-time PCR, increased with arsenic treatment. (D) Immunohistochemical analysis of PDCD4 protein showed decreased expression with increasing arsenic concentration. (E) Protein levels for PDCD4 and pSTAT3, analyzed by western blot, showed an apparent decrease in PDCD4 and apparent increase in pSTAT3 expressions with arsenic exposure. (F) Increased STAT3 phosphorylation with arsenic exposure was verified by immunohistochemistry. Data presented in the bar graphs are the mean ± SD of three independent experiments. *Indicates a statistically significant difference compared to control with p < 0.05.

Mentions: Next we investigated whether the increase in miR-21 levels and PDCD4 suppression mediated by chronic arsenic exposure in BEAS-2B cells induces tumors in nude mice. Nude mice were injected sc with BEAS-2B cells that had been exposed to arsenic for 6 months at the indicated concentration. Over a 4-week period post inoculation, we observed visible tumor formation that increased progressively in size for mice injected with arsenic-treated BEAS-2B cells but not in those injected with control cells (Fig. 6A,B). Consistent with our in vitro findings above, we found significantly increased miR-21 levels (Fig. 6C) associated with decreased PDCD4 expression (Fig. 6D,E) and increased phosphorylated STAT3 (Fig. 6F) in xenograft tumors generated with chronic arsenic exposed BEAS-2B cells.


Oncogenic transformation of human lung bronchial epithelial cells induced by arsenic involves ROS-dependent activation of STAT3-miR-21-PDCD4 mechanism
Increased miR-21 and suppressed PDCD4 expression by chronic arsenic exposure induces tumorigenesis of BEAS-2B cells in vivo.BEAS-2B cells exposed to indicated concentration of arsenic for 6 months were injected sc into nude mice. After 4 weeks, mice were euthanized using CO2 and tumor was isolated for further examination. (A) Representative images of excised tumors and (B) graphic representation of tumor volume demonstrate increased growth with arsenic treatment. (C) The relative miR-21 level, determined by Taqman real-time PCR, increased with arsenic treatment. (D) Immunohistochemical analysis of PDCD4 protein showed decreased expression with increasing arsenic concentration. (E) Protein levels for PDCD4 and pSTAT3, analyzed by western blot, showed an apparent decrease in PDCD4 and apparent increase in pSTAT3 expressions with arsenic exposure. (F) Increased STAT3 phosphorylation with arsenic exposure was verified by immunohistochemistry. Data presented in the bar graphs are the mean ± SD of three independent experiments. *Indicates a statistically significant difference compared to control with p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5120334&req=5

f6: Increased miR-21 and suppressed PDCD4 expression by chronic arsenic exposure induces tumorigenesis of BEAS-2B cells in vivo.BEAS-2B cells exposed to indicated concentration of arsenic for 6 months were injected sc into nude mice. After 4 weeks, mice were euthanized using CO2 and tumor was isolated for further examination. (A) Representative images of excised tumors and (B) graphic representation of tumor volume demonstrate increased growth with arsenic treatment. (C) The relative miR-21 level, determined by Taqman real-time PCR, increased with arsenic treatment. (D) Immunohistochemical analysis of PDCD4 protein showed decreased expression with increasing arsenic concentration. (E) Protein levels for PDCD4 and pSTAT3, analyzed by western blot, showed an apparent decrease in PDCD4 and apparent increase in pSTAT3 expressions with arsenic exposure. (F) Increased STAT3 phosphorylation with arsenic exposure was verified by immunohistochemistry. Data presented in the bar graphs are the mean ± SD of three independent experiments. *Indicates a statistically significant difference compared to control with p < 0.05.
Mentions: Next we investigated whether the increase in miR-21 levels and PDCD4 suppression mediated by chronic arsenic exposure in BEAS-2B cells induces tumors in nude mice. Nude mice were injected sc with BEAS-2B cells that had been exposed to arsenic for 6 months at the indicated concentration. Over a 4-week period post inoculation, we observed visible tumor formation that increased progressively in size for mice injected with arsenic-treated BEAS-2B cells but not in those injected with control cells (Fig. 6A,B). Consistent with our in vitro findings above, we found significantly increased miR-21 levels (Fig. 6C) associated with decreased PDCD4 expression (Fig. 6D,E) and increased phosphorylated STAT3 (Fig. 6F) in xenograft tumors generated with chronic arsenic exposed BEAS-2B cells.

View Article: PubMed Central - PubMed

ABSTRACT

Arsenic is a well-documented human carcinogen. The present study explored the role of the onco-miR, miR-21 and its target protein, programmed cell death 4 (PDCD4) in arsenic induced malignant cell transformation and tumorigenesis. Our results showed that treatment of human bronchial epithelial (BEAS-2B) cells with arsenic induces ROS through p47phox, one of the NOX subunits that is the key source of arsenic-induced ROS. Arsenic exposure induced an upregulation of miR-21 expression associated with inhibition of PDCD4, and caused malignant cell transformation and tumorigenesis of BEAS-2B cells. Indispensably, STAT3 transcriptional activation by IL-6 is crucial for the arsenic induced miR-21 increase. Upregulated miR-21 levels and suppressed PDCD4 expression was also observed in xenograft tumors generated with chronic arsenic exposed BEAS-2B cells. Stable shut down of miR-21, p47phox or STAT3 and overexpression of PDCD4 or catalase in BEAS-2B cells markedly inhibited the arsenic induced malignant transformation and tumorigenesis. Similarly, silencing of miR-21 or STAT3 and forced expression of PDCD4 in arsenic transformed cells (AsT) also inhibited cell proliferation and tumorigenesis. Furthermore, arsenic suppressed the downstream protein E-cadherin expression and induced &beta;-catenin/TCF-dependent transcription of uPAR and c-Myc. These results indicate that the ROS-STAT3-miR-21-PDCD4 signaling axis plays an important role in arsenic -induced carcinogenesis.

No MeSH data available.


Related in: MedlinePlus