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Association between arsenic suppression of adipogenesis and induction of CHOP10 via the endoplasmic reticulum stress response.

Hou Y, Xue P, Woods CG, Wang X, Fu J, Yarborough K, Qu W, Zhang Q, Andersen ME, Pi J - Environ. Health Perspect. (2012)

Bottom Line: The effects and associated mechanisms of iAs and its major metabolites on adipogenesis were determined in 3T3-L1 preadipocytes, mouse adipose-derived stromal-vascular fraction cells (ADSVFCs), and human adipose tissue-derived stem cells (ADSCs).In addition, iAs3+, MMA3+, and DMA3+ exhibited a strong inhibitory effect on adipogenesis in primary cultured mouse ADSVFCs and human ADSCs.Arsenic-induced dysfunctional adipogenesis may be associated with a reduced capacity of WAT to store lipids and with insulin resistance.

View Article: PubMed Central - PubMed

Affiliation: Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, North Carolina 27709, USA.

ABSTRACT

Background: There is growing evidence that chronic exposure to inorganic arsenic (iAs) is associated with an increased prevalence of type 2 diabetes (T2D). However, the mechanisms for the diabetogenic effect of iAs are still largely unknown. White adipose tissue (WAT) actively stores and releases energy and maintains lipid and glucose homeostasis.

Objective: We sought to determine the mechanisms of arsenic suppression of adipogenesis.

Methods: The effects and associated mechanisms of iAs and its major metabolites on adipogenesis were determined in 3T3-L1 preadipocytes, mouse adipose-derived stromal-vascular fraction cells (ADSVFCs), and human adipose tissue-derived stem cells (ADSCs).

Results: Exposure of 3T3-L1 preadipocytes to noncytotoxic levels of arsenic, including inorganic arsenite (iAs3+, ≤ 5 μM), inorganic arsenate (≤ 20 μM), trivalent monomethylated arsenic (MMA3+, ≤ 1 μM), and trivalent dimethylated arsenic (DMA3+, ≤ 2 μM) decreased adipogenic hormone-induced adipogenesis in a concentration-dependent manner. In addition, iAs3+, MMA3+, and DMA3+ exhibited a strong inhibitory effect on adipogenesis in primary cultured mouse ADSVFCs and human ADSCs. Time-course studies in 3T3-L1 cells revealed that inhibition of adipogenesis by arsenic occurred in the early stage of terminal adipogenic differentiation and was highly correlated with the induction of C/EBP homologous protein (CHOP10), an endoplasmic reticulum (ER) stress response protein. Induction of CHOP10 by arsenic is associated with reduced DNA-binding activity of CCAAT/enhancer-binding protein β (C/EBPβ), which regulates the transcription of peroxisome proliferator-activated receptor γ and C/EBPα.

Conclusions: Low-level iAs and MMA3+ trigger the ER stress response and up-regulate CHOP10, which inhibits C/EBPβ transcriptional activity, thus suppressing adipogenesis. Arsenic-induced dysfunctional adipogenesis may be associated with a reduced capacity of WAT to store lipids and with insulin resistance.

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iAs3+ activates UPR in 3T3-L1 preadipocytes. Abbreviations: BFA, brefeldin A; TG, thapsigargin; TUN, tunicamycin. (A) Six hours of treatment with iAs3+ concentration-dependently enhanced mRNA expression of Atf4 and Chop10 (n = 3). (B) Time-course of mRNA expression of Atf4 and Chop10 after exposure to 5 μM iAs3+ (n = 3). (C,D) Concentration–response (C) and time course (D) of protein expression of eIF2α, p-eIF2α, ATF4, and CHOP10 in response to iAs3+ treatment. Cells were treated with iAs3+ for 6 hr (C) or 5 μM iAs3+ for the indicated time (D) (n = 3). (E) ER stressors suppress adipogenesis in 3T3-L1 cells. Cells were treated with the stressors at the indicated concentrations during the first 2 days of DMI-induced adipogenesis, followed by ORO staining. *p < 0.05, iAs3+-treated vs. vehicle-treated cells.
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f4: iAs3+ activates UPR in 3T3-L1 preadipocytes. Abbreviations: BFA, brefeldin A; TG, thapsigargin; TUN, tunicamycin. (A) Six hours of treatment with iAs3+ concentration-dependently enhanced mRNA expression of Atf4 and Chop10 (n = 3). (B) Time-course of mRNA expression of Atf4 and Chop10 after exposure to 5 μM iAs3+ (n = 3). (C,D) Concentration–response (C) and time course (D) of protein expression of eIF2α, p-eIF2α, ATF4, and CHOP10 in response to iAs3+ treatment. Cells were treated with iAs3+ for 6 hr (C) or 5 μM iAs3+ for the indicated time (D) (n = 3). (E) ER stressors suppress adipogenesis in 3T3-L1 cells. Cells were treated with the stressors at the indicated concentrations during the first 2 days of DMI-induced adipogenesis, followed by ORO staining. *p < 0.05, iAs3+-treated vs. vehicle-treated cells.

Mentions: iAs3+ triggers UPR, which impairs terminal adipogenesis. To investigate whether UPR, especially CHOP10 induction, is involved in the suppression of terminal adipogenesis by iAs3+, the expression of ATF4 and CHOP10 were measured in 3T3-L1 cells. iAs3+ concentration- and time-dependently augmented the expression of ATF4 and CHOP10 at mRNA (Figure 4A,B) and protein (Figure 4C,D) levels. In addition, other ER stress responsive genes, such as growth arrest and DNA-damage-inducible 45a (Gadd45a), X-box binding protein 1 (Xbp1), and spliced Xbp1 (sXbp1) were also significantly induced by iAs3+ [see Supplemental Material, Figure S4 (http://dx.doi.org/10.1289/ehp.1205731)], suggesting that iAs3+ triggers UPR in the cells. Furthermore, the phosphorylation of eukaryotic initiation factor 2α (p-elF2α) was increased in response to iAs3+ treatment, indicating that double-stranded RNA-activated protein kinase (PKR)-like ER kinase (PERK) also plays a role in the response. In 3T3-L1 cells, MMA3+ and DMA3+ also concentration-dependently increased the mRNA expression of many UPR genes, including Atf4, Chop10, Xbp1, and sXbp1 (Figures S5 and S6). In addition, MMA3+ also concentration- and/or time-dependently increased the mRNA expression of Gadd45a (see Supplemental Material, Figure S5) and the protein expression of CHOP10 (see Supplemental Material, Figure S7). These findings suggest that trivalent methylated arsenic may also trigger UPR. In agreement with a previous study (Yu et al. 2009), exposure of 3T3-L1 cells to multiple well-known ER stressors, including BFA, TG, and TUN, at noncytotoxic concentrations resulted in a concentration-dependent suppression of terminal adipogenesis (Figure 4E). Therefore, iAs3+ may activate UPR and induce CHOP10 resulting in suppression of adipogenesis.


Association between arsenic suppression of adipogenesis and induction of CHOP10 via the endoplasmic reticulum stress response.

Hou Y, Xue P, Woods CG, Wang X, Fu J, Yarborough K, Qu W, Zhang Q, Andersen ME, Pi J - Environ. Health Perspect. (2012)

iAs3+ activates UPR in 3T3-L1 preadipocytes. Abbreviations: BFA, brefeldin A; TG, thapsigargin; TUN, tunicamycin. (A) Six hours of treatment with iAs3+ concentration-dependently enhanced mRNA expression of Atf4 and Chop10 (n = 3). (B) Time-course of mRNA expression of Atf4 and Chop10 after exposure to 5 μM iAs3+ (n = 3). (C,D) Concentration–response (C) and time course (D) of protein expression of eIF2α, p-eIF2α, ATF4, and CHOP10 in response to iAs3+ treatment. Cells were treated with iAs3+ for 6 hr (C) or 5 μM iAs3+ for the indicated time (D) (n = 3). (E) ER stressors suppress adipogenesis in 3T3-L1 cells. Cells were treated with the stressors at the indicated concentrations during the first 2 days of DMI-induced adipogenesis, followed by ORO staining. *p < 0.05, iAs3+-treated vs. vehicle-treated cells.
© Copyright Policy - public-domain
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3569692&req=5

f4: iAs3+ activates UPR in 3T3-L1 preadipocytes. Abbreviations: BFA, brefeldin A; TG, thapsigargin; TUN, tunicamycin. (A) Six hours of treatment with iAs3+ concentration-dependently enhanced mRNA expression of Atf4 and Chop10 (n = 3). (B) Time-course of mRNA expression of Atf4 and Chop10 after exposure to 5 μM iAs3+ (n = 3). (C,D) Concentration–response (C) and time course (D) of protein expression of eIF2α, p-eIF2α, ATF4, and CHOP10 in response to iAs3+ treatment. Cells were treated with iAs3+ for 6 hr (C) or 5 μM iAs3+ for the indicated time (D) (n = 3). (E) ER stressors suppress adipogenesis in 3T3-L1 cells. Cells were treated with the stressors at the indicated concentrations during the first 2 days of DMI-induced adipogenesis, followed by ORO staining. *p < 0.05, iAs3+-treated vs. vehicle-treated cells.
Mentions: iAs3+ triggers UPR, which impairs terminal adipogenesis. To investigate whether UPR, especially CHOP10 induction, is involved in the suppression of terminal adipogenesis by iAs3+, the expression of ATF4 and CHOP10 were measured in 3T3-L1 cells. iAs3+ concentration- and time-dependently augmented the expression of ATF4 and CHOP10 at mRNA (Figure 4A,B) and protein (Figure 4C,D) levels. In addition, other ER stress responsive genes, such as growth arrest and DNA-damage-inducible 45a (Gadd45a), X-box binding protein 1 (Xbp1), and spliced Xbp1 (sXbp1) were also significantly induced by iAs3+ [see Supplemental Material, Figure S4 (http://dx.doi.org/10.1289/ehp.1205731)], suggesting that iAs3+ triggers UPR in the cells. Furthermore, the phosphorylation of eukaryotic initiation factor 2α (p-elF2α) was increased in response to iAs3+ treatment, indicating that double-stranded RNA-activated protein kinase (PKR)-like ER kinase (PERK) also plays a role in the response. In 3T3-L1 cells, MMA3+ and DMA3+ also concentration-dependently increased the mRNA expression of many UPR genes, including Atf4, Chop10, Xbp1, and sXbp1 (Figures S5 and S6). In addition, MMA3+ also concentration- and/or time-dependently increased the mRNA expression of Gadd45a (see Supplemental Material, Figure S5) and the protein expression of CHOP10 (see Supplemental Material, Figure S7). These findings suggest that trivalent methylated arsenic may also trigger UPR. In agreement with a previous study (Yu et al. 2009), exposure of 3T3-L1 cells to multiple well-known ER stressors, including BFA, TG, and TUN, at noncytotoxic concentrations resulted in a concentration-dependent suppression of terminal adipogenesis (Figure 4E). Therefore, iAs3+ may activate UPR and induce CHOP10 resulting in suppression of adipogenesis.

Bottom Line: The effects and associated mechanisms of iAs and its major metabolites on adipogenesis were determined in 3T3-L1 preadipocytes, mouse adipose-derived stromal-vascular fraction cells (ADSVFCs), and human adipose tissue-derived stem cells (ADSCs).In addition, iAs3+, MMA3+, and DMA3+ exhibited a strong inhibitory effect on adipogenesis in primary cultured mouse ADSVFCs and human ADSCs.Arsenic-induced dysfunctional adipogenesis may be associated with a reduced capacity of WAT to store lipids and with insulin resistance.

View Article: PubMed Central - PubMed

Affiliation: Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, North Carolina 27709, USA.

ABSTRACT

Background: There is growing evidence that chronic exposure to inorganic arsenic (iAs) is associated with an increased prevalence of type 2 diabetes (T2D). However, the mechanisms for the diabetogenic effect of iAs are still largely unknown. White adipose tissue (WAT) actively stores and releases energy and maintains lipid and glucose homeostasis.

Objective: We sought to determine the mechanisms of arsenic suppression of adipogenesis.

Methods: The effects and associated mechanisms of iAs and its major metabolites on adipogenesis were determined in 3T3-L1 preadipocytes, mouse adipose-derived stromal-vascular fraction cells (ADSVFCs), and human adipose tissue-derived stem cells (ADSCs).

Results: Exposure of 3T3-L1 preadipocytes to noncytotoxic levels of arsenic, including inorganic arsenite (iAs3+, ≤ 5 μM), inorganic arsenate (≤ 20 μM), trivalent monomethylated arsenic (MMA3+, ≤ 1 μM), and trivalent dimethylated arsenic (DMA3+, ≤ 2 μM) decreased adipogenic hormone-induced adipogenesis in a concentration-dependent manner. In addition, iAs3+, MMA3+, and DMA3+ exhibited a strong inhibitory effect on adipogenesis in primary cultured mouse ADSVFCs and human ADSCs. Time-course studies in 3T3-L1 cells revealed that inhibition of adipogenesis by arsenic occurred in the early stage of terminal adipogenic differentiation and was highly correlated with the induction of C/EBP homologous protein (CHOP10), an endoplasmic reticulum (ER) stress response protein. Induction of CHOP10 by arsenic is associated with reduced DNA-binding activity of CCAAT/enhancer-binding protein β (C/EBPβ), which regulates the transcription of peroxisome proliferator-activated receptor γ and C/EBPα.

Conclusions: Low-level iAs and MMA3+ trigger the ER stress response and up-regulate CHOP10, which inhibits C/EBPβ transcriptional activity, thus suppressing adipogenesis. Arsenic-induced dysfunctional adipogenesis may be associated with a reduced capacity of WAT to store lipids and with insulin resistance.

Show MeSH
Related in: MedlinePlus