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DsbA-L alleviates endoplasmic reticulum stress-induced adiponectin downregulation.

Zhou L, Liu M, Zhang J, Chen H, Dong LQ, Liu F - Diabetes (2010)

Bottom Line: The cellular levels of DsbA-L were altered by RNAi-mediated suppression or adenovirus-mediated overexpression.In addition, inducing ER stress is sufficient to downregulate adiponectin levels in 3T3-L1 adipocytes, which could be protected by treating cells with the autophagy inhibitor 3-methyladenine or by overexpression of DsbA-L.ER stress plays a key role in obesity-induced adiponectin downregulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Texas Health Science Center at San Antonio, San Antonio, Texas, USA.

ABSTRACT

Objective: Obesity impairs adiponectin expression, assembly, and secretion, yet the underlying mechanisms remain elusive. The aims of this study were 1) to determine the molecular mechanisms by which obesity impairs adiponectin multimerization and stability, and 2) to determine the potential role of disulfide-bond-A oxidoreductase-like protein (DsbA-L), a recently identified adiponectin interactive protein that promotes adiponectin multimerization and stability in obesity-induced endoplasmic reticulum (ER) stress and adiponectin downregulation.

Research design and methods: Tauroursodeoxycholic acid (TUDCA), a chemical chaperone that alleviates ER stress, was used to study the mechanism underlying obesity-induced adiponectin downregulation in db/db mice, high-fat diet-induced obese mice, and in ER-stressed 3T3-L1 adipocytes. The cellular levels of DsbA-L were altered by RNAi-mediated suppression or adenovirus-mediated overexpression. The protective role of DsbA-L in obesity- and ER stress-induced adiponectin downregulation was characterized.

Results: Treating db/db mice and diet-induced obese mice with TUDCA increased the cellular and serum levels of adiponectin. In addition, inducing ER stress is sufficient to downregulate adiponectin levels in 3T3-L1 adipocytes, which could be protected by treating cells with the autophagy inhibitor 3-methyladenine or by overexpression of DsbA-L.

Conclusions: ER stress plays a key role in obesity-induced adiponectin downregulation. In addition, DsbA-L facilitates adiponectin folding and assembly and provides a protective effect against ER stress-mediated adiponectin downregulation in obesity.

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Suppression of ER stress improves adiponectin expression in db/db mice. A: The expression levels of adiponectin (Adpn), DsbA-L, and CHOP in white adipose tissue (WAT) and the serum adiponectin levels from saline- or TUDCA-treated lean or db/db mice were determined by Western blot with specific antibodies as indicated. β-actin or immunoglobulin (IgG) was used as a tissue or serum loading control, respectively. The relative protein levels of adiponectin in WAT (B), or in serum (C), and DsbA-L (D) were quantified. Data represent mean ± SEM. **P < 0.01, ***P < 0.001; n = 10 for the saline control mice, and n = 8 per treatment group for the db/db mice. E: Differentiated 3T3–L1 adipocytes were pretreated with or without TUDCA (1 mmol/l) for 1 h followed with TG (0.01 μmol/l) cotreatment for 24 h. The cells were lysed, and the expression levels of adiponectin, DsbA-L, CHOP, TNFα, resistin, and β-tubulin were determined by Western blot analysis with specific antibodies as indicated. Adiponectin levels in conditioned medium were determined with an antiadiponectin antibody. F: The mRNA levels of adiponectin were determined by quantitative reverse transcription PCR. The protein levels of adiponectin as shown in (E) were quantified (G). N = 3, *P < 0.05, **P < 0.01, ***P < 0.001; ns, no statistical difference; ctrl, control.
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Figure 1: Suppression of ER stress improves adiponectin expression in db/db mice. A: The expression levels of adiponectin (Adpn), DsbA-L, and CHOP in white adipose tissue (WAT) and the serum adiponectin levels from saline- or TUDCA-treated lean or db/db mice were determined by Western blot with specific antibodies as indicated. β-actin or immunoglobulin (IgG) was used as a tissue or serum loading control, respectively. The relative protein levels of adiponectin in WAT (B), or in serum (C), and DsbA-L (D) were quantified. Data represent mean ± SEM. **P < 0.01, ***P < 0.001; n = 10 for the saline control mice, and n = 8 per treatment group for the db/db mice. E: Differentiated 3T3–L1 adipocytes were pretreated with or without TUDCA (1 mmol/l) for 1 h followed with TG (0.01 μmol/l) cotreatment for 24 h. The cells were lysed, and the expression levels of adiponectin, DsbA-L, CHOP, TNFα, resistin, and β-tubulin were determined by Western blot analysis with specific antibodies as indicated. Adiponectin levels in conditioned medium were determined with an antiadiponectin antibody. F: The mRNA levels of adiponectin were determined by quantitative reverse transcription PCR. The protein levels of adiponectin as shown in (E) were quantified (G). N = 3, *P < 0.05, **P < 0.01, ***P < 0.001; ns, no statistical difference; ctrl, control.

Mentions: Obesity induces ER stress, and ER stress is correlated with reduced adiponectin levels (15). To determine whether ER stress is involved in downregulating adiponectin expression in obesity, we examined adiponectin levels in db/db mice treated with vehicle or the ER stress–reducing chemical chaperone, TUDCA. Obesity led to a great increase in ER stress in fat tissue of the db/db mice, as demonstrated by increased C/EBP-homologous protein (CHOP) expression (Fig. 1A). Concurrent with increased ER stress, the cellular (Fig. 1B) and circulating (Fig. 1C) levels of adiponectin were significantly reduced in the db/db mice compared with that of the lean mice. Treating the db/db mice with TUDCA greatly reduced obesity-induced CHOP expression, concurrent with an increase in circulating and cellular levels of adiponectin and its interacting protein DsbA-L (Fig. 1A–D). TUDCA treatment had no significant effect on the food intake (data not shown) (16) and body weight of the db/db mice (supplementary Fig. S1A in the appendix available online at http://diabetes.diabetesjournals.org/cgi/content/full/db10-0412/DC1, indicating that the TUDCA-mediated rescuing effect on DsbA-L and adiponectin levels was not a consequence of altered food intake/body weight. The expression levels of DsbA-L and adiponectin were also greatly reduced in WAT of HFD-induced obese mice (10) and this reduction was also rescued by TUDCA treatment (supplemental online Fig. S2). Taken together, these results suggest that ER stress may play a key role in obesity-induced adiponectin downregulation.


DsbA-L alleviates endoplasmic reticulum stress-induced adiponectin downregulation.

Zhou L, Liu M, Zhang J, Chen H, Dong LQ, Liu F - Diabetes (2010)

Suppression of ER stress improves adiponectin expression in db/db mice. A: The expression levels of adiponectin (Adpn), DsbA-L, and CHOP in white adipose tissue (WAT) and the serum adiponectin levels from saline- or TUDCA-treated lean or db/db mice were determined by Western blot with specific antibodies as indicated. β-actin or immunoglobulin (IgG) was used as a tissue or serum loading control, respectively. The relative protein levels of adiponectin in WAT (B), or in serum (C), and DsbA-L (D) were quantified. Data represent mean ± SEM. **P < 0.01, ***P < 0.001; n = 10 for the saline control mice, and n = 8 per treatment group for the db/db mice. E: Differentiated 3T3–L1 adipocytes were pretreated with or without TUDCA (1 mmol/l) for 1 h followed with TG (0.01 μmol/l) cotreatment for 24 h. The cells were lysed, and the expression levels of adiponectin, DsbA-L, CHOP, TNFα, resistin, and β-tubulin were determined by Western blot analysis with specific antibodies as indicated. Adiponectin levels in conditioned medium were determined with an antiadiponectin antibody. F: The mRNA levels of adiponectin were determined by quantitative reverse transcription PCR. The protein levels of adiponectin as shown in (E) were quantified (G). N = 3, *P < 0.05, **P < 0.01, ***P < 0.001; ns, no statistical difference; ctrl, control.
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Figure 1: Suppression of ER stress improves adiponectin expression in db/db mice. A: The expression levels of adiponectin (Adpn), DsbA-L, and CHOP in white adipose tissue (WAT) and the serum adiponectin levels from saline- or TUDCA-treated lean or db/db mice were determined by Western blot with specific antibodies as indicated. β-actin or immunoglobulin (IgG) was used as a tissue or serum loading control, respectively. The relative protein levels of adiponectin in WAT (B), or in serum (C), and DsbA-L (D) were quantified. Data represent mean ± SEM. **P < 0.01, ***P < 0.001; n = 10 for the saline control mice, and n = 8 per treatment group for the db/db mice. E: Differentiated 3T3–L1 adipocytes were pretreated with or without TUDCA (1 mmol/l) for 1 h followed with TG (0.01 μmol/l) cotreatment for 24 h. The cells were lysed, and the expression levels of adiponectin, DsbA-L, CHOP, TNFα, resistin, and β-tubulin were determined by Western blot analysis with specific antibodies as indicated. Adiponectin levels in conditioned medium were determined with an antiadiponectin antibody. F: The mRNA levels of adiponectin were determined by quantitative reverse transcription PCR. The protein levels of adiponectin as shown in (E) were quantified (G). N = 3, *P < 0.05, **P < 0.01, ***P < 0.001; ns, no statistical difference; ctrl, control.
Mentions: Obesity induces ER stress, and ER stress is correlated with reduced adiponectin levels (15). To determine whether ER stress is involved in downregulating adiponectin expression in obesity, we examined adiponectin levels in db/db mice treated with vehicle or the ER stress–reducing chemical chaperone, TUDCA. Obesity led to a great increase in ER stress in fat tissue of the db/db mice, as demonstrated by increased C/EBP-homologous protein (CHOP) expression (Fig. 1A). Concurrent with increased ER stress, the cellular (Fig. 1B) and circulating (Fig. 1C) levels of adiponectin were significantly reduced in the db/db mice compared with that of the lean mice. Treating the db/db mice with TUDCA greatly reduced obesity-induced CHOP expression, concurrent with an increase in circulating and cellular levels of adiponectin and its interacting protein DsbA-L (Fig. 1A–D). TUDCA treatment had no significant effect on the food intake (data not shown) (16) and body weight of the db/db mice (supplementary Fig. S1A in the appendix available online at http://diabetes.diabetesjournals.org/cgi/content/full/db10-0412/DC1, indicating that the TUDCA-mediated rescuing effect on DsbA-L and adiponectin levels was not a consequence of altered food intake/body weight. The expression levels of DsbA-L and adiponectin were also greatly reduced in WAT of HFD-induced obese mice (10) and this reduction was also rescued by TUDCA treatment (supplemental online Fig. S2). Taken together, these results suggest that ER stress may play a key role in obesity-induced adiponectin downregulation.

Bottom Line: The cellular levels of DsbA-L were altered by RNAi-mediated suppression or adenovirus-mediated overexpression.In addition, inducing ER stress is sufficient to downregulate adiponectin levels in 3T3-L1 adipocytes, which could be protected by treating cells with the autophagy inhibitor 3-methyladenine or by overexpression of DsbA-L.ER stress plays a key role in obesity-induced adiponectin downregulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Texas Health Science Center at San Antonio, San Antonio, Texas, USA.

ABSTRACT

Objective: Obesity impairs adiponectin expression, assembly, and secretion, yet the underlying mechanisms remain elusive. The aims of this study were 1) to determine the molecular mechanisms by which obesity impairs adiponectin multimerization and stability, and 2) to determine the potential role of disulfide-bond-A oxidoreductase-like protein (DsbA-L), a recently identified adiponectin interactive protein that promotes adiponectin multimerization and stability in obesity-induced endoplasmic reticulum (ER) stress and adiponectin downregulation.

Research design and methods: Tauroursodeoxycholic acid (TUDCA), a chemical chaperone that alleviates ER stress, was used to study the mechanism underlying obesity-induced adiponectin downregulation in db/db mice, high-fat diet-induced obese mice, and in ER-stressed 3T3-L1 adipocytes. The cellular levels of DsbA-L were altered by RNAi-mediated suppression or adenovirus-mediated overexpression. The protective role of DsbA-L in obesity- and ER stress-induced adiponectin downregulation was characterized.

Results: Treating db/db mice and diet-induced obese mice with TUDCA increased the cellular and serum levels of adiponectin. In addition, inducing ER stress is sufficient to downregulate adiponectin levels in 3T3-L1 adipocytes, which could be protected by treating cells with the autophagy inhibitor 3-methyladenine or by overexpression of DsbA-L.

Conclusions: ER stress plays a key role in obesity-induced adiponectin downregulation. In addition, DsbA-L facilitates adiponectin folding and assembly and provides a protective effect against ER stress-mediated adiponectin downregulation in obesity.

Show MeSH
Related in: MedlinePlus