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1.
Figure 4

Figure 4. AZX100 treatment reduces TGF-β1-induced cofilin phosphorylation. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

Human keloid fibroblasts were treated as previously indicated.(a) Representative Western blots of cofilin, phospho-cofilin, and β-actin. Cofilin and phospho-cofilin levels were quantified relative to β-actin expression to correct for loading differences and normalized to untreated (U) levels. Data are shown in (b) and are expressed as mean ± standard deviation of the ratio phosphorylated cofilin/cofilin from 3–4 experiments. *P<0.05 compared to the TGF-β1 stimulation.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.
2.
Figure 5

Figure 5. Effect of AZX100 treatment on SMAD3 phosphorylation. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

Human keloid fibroblasts were treated as previously indicated. (a) Representative Western blots of SMAD3, phospho-SMAD3, and β-actin. SMAD3 and phospho-SMAD3 levels were quantified relative to β-actin expression to correct for loading differences and normalized to untreated (U) levels. Data are shown in (b) and are expressed as mean ± standard deviation of the ratio phosphorylated SMAD3/SMAD3 from three experiments. *P<0.05 compared to the TGF-β1 stimulation.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.
3.
Figure 6

Figure 6. AZX100 improved the collagen organization in an in vivo model. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

(a–f) A linear 2-cm incision was made on the upper back of Siberian hamsters and injected with 0.1ml of either saline vehicle (control, a–c) or AZX100 (1mm, d–f). Histology slides of samples taken after 7 (a, d), 14 (b, e), or 21 days (c, f) of healing were stained with trichrome to show collagen fiber organization. (g) Collagen fiber orientation, density, and maturity, as well as the total collagen scores are shown for 7, 14, and 21 days (n=7–8 per group). *P<0.05 for between-group differences at each time point. Bar = 100 µm.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.
4.
Figure 1

Figure 1. AZX100 reduces TGF-β1-induced expression of CTGF and collagen in human keloid fibroblasts. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

Human keloid fibroblasts were serum-starved in DMEM medium containing 0.5% fetal bovine serum for 48 hours before treatment.(a, b) Cells treated with different doses of TGF-β1 for 24 hours. Representative western blots of CTGF and GAPDH are shown in (a). CTGF levels were quantified relative to GAPDH expression to correct for loading differences and normalized to untreated levels. Data are shown in (b), and are expressed as mean ± standard deviation of 3–6 experiments. *P<0.05 compared to the untreated cells. (c–e) TGF-β1-stimulated cells were treated with either AZX100 (50 µm), the protein transduction domain (PTD), or free HSP20 phosphopeptide without the PTD. Representative western blots of CTGF, collagen, and β-actin are shown in (c). CTGF and collagen levels were quantified relative to GAPDH expression to correct for loading differences and normalized to untreated (U) levels. Data are shown in (d) and (e) and are expressed as mean ± standard deviation of 4–6 experiments. *P<0.05 compared to the TGF-β1 stimulation indicates no AZX100 treatment.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.
5.
Figure 3

Figure 3. Effect of TGF-β1 and AZX100 treatment on stress fiber formation and α-SMA expression. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

Human keloid fibroblasts were either untreated (a), treated with AZX100 (50 µm) (b), TGF-β1 at 1.2 ng ml−1 (c), TGF-β1 at 2.5 ngml−1 (d), TGF-β1 (1.2 ng ml−1) + AZX100 (50 µm) (e), or TGF-β1 (2.5 ng ml−1) + AZX100 (50 µM) (f) for 24 hours. The cells were fixed and stained with Alexa 350 phalloidin to visualize the actin cytoskeleton. Bar = 100 µm. Representative western blots of α-SMA, GAPDH, and β-actin in untreated cells, cells treated with AZX100 (50 µm), with TGF-β1 (2.5 ngml−1), or cells treated with TGF-β1 (2.5 ng ml−1) + AZX100 (50 µm). (g) Phase contrast microscopic images of untreated cells and cells treated with AZX100 for 3 days; bar = 200 µm. (h) α-SMA levels were quantified relative to GAPDH expression to correct for loading differences and normalized to untreated levels. (i) Data are expressed as mean ± standard deviation of 4–6 experiments. *P<0.05 AZX100-treated cells compared to untreated and AZX100 + TGF-β1 compared to the TGF-β1 stimulation alone.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.
6.
Figure 2

Figure 2. AZX100 reduces endothelin- and LPA-induced expression of CTGF and collagen in human keloid fibroblasts. From: Cell Permeant Peptide Analogues of the Small Heat Shock Protein, HSP20, Reduce TGF-?1-Induced CTGF Expression in Keloid Fibroblasts.

Cells were serum-starved in DMEM medium containing 0.5% FBS for 48 hours, before treatment. (a–c) Cells were treated with several doses of endothelin (10–50 nm), lysophosphatidic acid (LPA, 10–50 µm), or thrombin (50–150 nm) for 24 hours. Representative Western blots of CTGF, collagen, and GAPDH are shown in (a). CTGF and collagen levels were quantified relative to GAPDH expression to correct for loading differences and normalized to untreated (U) levels. Data are shown in (b) and (c) and are expressed as mean ± standard deviation of 3–4 experiments. (d–f) Cells were stimulated with endothelin (50 nm) and LPA (25 µm) in the presence or absence of AZX100 (50 µm) for 24 hours. Representative western blots of CTGF, collagen, and GAPDH are shown in (d). (e, f) The Western blot bands were quantified by densitometry, and CTGF and collagen expressions were related to GAPDH expression to correct for loading differences. The expression of CTGF and collagen in untreated cells was set to 1 for comparison of different blots. Data expressed as mean ± standard deviation of three experiments. *P<0.05 compared to the TGF-β1 stimulation.

Luciana B. Lopes, et al. J Invest Dermatol. ;129(3):590-598.

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