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pNaKtide inhibits Na/K-ATPase reactive oxygen species amplification and attenuates adipogenesis

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Science Advances  16 Oct 2015:
Vol. 1, no. 9, e1500781
DOI: 10.1126/sciadv.1500781
  • Fig. 1 Effect of increasing pNaKtide concentrations on adipogenesis in mouse preadipocytes.

    Adipogenesis was measured as the relative absorbance of Oil Red O at day 7 after inducing adipogenesis as described in Materials and Methods. (A) Representative images. (B) Quantitative data expressed as means ± SE, n = 7; *P < 0.05 versus control.

  • Fig. 2 pNaKtide increased adiponectin levels and decreased adipogenic markers in 3T3L1 adipocytes.

    (A) Adiponectin levels were determined in conditioned media obtained from 3T3L1 cells after treatment with pNaKtide for 7 days. Cells were treated with varying concentrations of pNaKtide, and 0.7 μM pNaKtide was determined to be the optimal concentration for increasing adiponectin levels. Results are means ± SE, n = 4; *P < 0.05 versus control (CTR). (B to D) Expression of (B) FAS, (C) MEST, and (D) PPARγ was determined by Western blot analysis in 3T3L1 cells after treatment with pNaKtide (0.7 μM) for 7 days. Quantitative densitometric evaluation of protein ratios was done. Data are expressed as means ± SE, n = 6; *P < 0.05 versus control.

  • Fig. 3 Effect of increasing doses of pNaKtide on body weight in mice fed a high-fat diet.

    Treatment with pNaKtide at doses of 1 and 5 mg/kg did not significantly reduce body weight in C57Bl6 mice fed a high-fat diet. However, administration of pNaKtide at 25 mg/kg every 8 days in mice fed a high-fat diet for 8 weeks significantly reduced body weight as compared to high-fat diet–fed animals. There were no significant changes in food intake among the groups. Results are means ± SE, n = 7 to 14 per group; *P < 0.05 versus control, #P < 0.05 versus high-fat diet.

  • Fig. 4 Effect of increasing doses of pNaKtide on visceral and subcutaneous fat content in mice fed a high-fat diet.

    (A and B) C57Bl6 mice fed a high-fat diet (HF) for 8 weeks were injected with pNaKtide at doses of 1, 5, and 25 mg/kg every 8 days. Administration of pNaKtide at 25 mg/kg in mice fed a high-fat diet significantly reduced visceral (A) and subcutaneous (B) fat content as compared to high-fat diet–fed animals. Results are means ± SE, n = 7 to 14 per group; *P < 0.05 versus control, #P < 0.05 versus high-fat diet.

  • Fig. 5 Effect of increasing doses of pNaKtide on adipogenic markers and adiponectin expression in visceral adipose tissue in mice fed a high-fat diet.

    (A and B) Western blot and densitometric analysis of adipogenic markers in visceral adipose tissue, FAS (A) and MEST (B). Results are means ± SE, n = 7 per group; *P < 0.05 versus control, #P < 0.05 versus high-fat diet. Data are shown as mean band density normalized to β-actin. (C) Western blot and densitometric analysis of adiponectin expression. Results are means ± SE, n = 7 per group; *P < 0.05 versus control, #P < 0.05 versus high-fat diet. Data are shown as mean band density normalized to β-actin.

  • Fig. 6 Effect of increasing doses of pNaKtide on metabolic profile in mice fed a high-fat diet.

    (A) HOMA-IR. (B) Glucose tolerance test. (C) Thiobarbituric acid–reactive substance (TBARS), a marker of oxidative injury, was measured in visceral adipose tissue. (D) Plasma adiponectin levels. Results are means ± SE, n = 7 to 14 per group; *P < 0.05 versus control, #P < 0.05 versus high-fat diet. (E to G) Effect of pNaKtide on protein carbonylation and phosphorylation of c-Src and ERK1/2 in fat tissue in mice fed a high-fat diet. (E) Protein carbonylation. (F) Phosphorylation of c-Src. (G) Phosphorylation of ERK1/2. Results are means ± SE, n = 4 to 7 per group; **P < 0.01 versus control, #P < 0.01 versus high-fat diet.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/1/9/e1500781/DC1

    Fig. S1. pNaKtide distribution in 3T3L1 cells and adipose tissue.

    Fig. S2. pNaKtide decreased large lipid droplets and oxidative stress and increased small lipid droplets in 3T3L1 adipocytes.

    Fig. S3. pNaKtide decreased lipid accumulation and adipogenic markers in 3T3L1 adipocytes exposed to fructose.

    Fig. S4. pNaKtide decreased lipid accumulation and adipogenic markers in 3T3L1 adipocytes exposed to glucose oxidase.

    Fig. S5. pNaKtide decreased carbonylation and phosphorylation of Src and ERK in 3T3L1 adipocytes exposed to glucose oxidase.

  • Supplementary Materials

    This PDF file includes:

    • Fig. S1. pNaKtide distribution in 3T3L1 cells and adipose tissue.
    • Fig. S2. pNaKtide decreased large lipid droplets and oxidative stress and increased small lipid droplets in 3T3L1 adipocytes.
    • Fig. S3. pNaKtide decreased lipid accumulation and adipogenic markers in 3T3L1 adipocytes exposed to fructose.
    • Fig. S4. pNaKtide decreased lipid accumulation and adipogenic markers in 3T3L1 adipocytes exposed to glucose oxidase.
    • Fig. S5. pNaKtide decreased carbonylation and phosphorylation of Src and ERK in 3T3L1 adipocytes exposed to glucose oxidase.

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