Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Asiaticoside Combined With Carbon Ion Implantation to Improve the Biocompatibility of Silicone Rubber and to Reduce the Risk of Capsule Contracture

doi: 10.3389/fbioe.2022.810244

Figure Lengend Snippet: Effect of asiaticoside on TGF-β1-induced transformation of fibroblasts into myofibroblasts and collagen deposition. Immunofluorescence staining of human fibroblasts on silicone rubber (SR) and carbon silicone rubber (C-SR) with Col-1A1 and α-SMA (100X).

Article Snippet: The primary antibodies were as follows: Smad2/3 (1:500, Affinity, United States), vimentin (1:500, Proteintech, China), α-SMA (1:500, Proteintech, China), Col-1A1 (1:500, Novus, United States), TGF-β1 (1:500, Affinity, United States), TβRI (1:500, Abcam, United States), TβRII (1:500, Abcam, United States), Smad2/3 (1:500, Affinity, United States), p-Smad2/3 (1:500, Affinity, United States), and β-actin (1:1,000, Abcam, United States).

Techniques: Transformation Assay, Immunofluorescence, Staining

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Asiaticoside Combined With Carbon Ion Implantation to Improve the Biocompatibility of Silicone Rubber and to Reduce the Risk of Capsule Contracture

doi: 10.3389/fbioe.2022.810244

Figure Lengend Snippet: Effect of asiaticoside on transformation of fibroblasts into myofibroblasts, collagen deposition and changes in the TGF-β/Smad signaling pathway in vivo . (A) Representative Western blot analysis of TGF-β1, TβRI, TβRII, Col-1A1, Smad2/3, p-Smad2/3 and α-SMA. One representative blot of three experiments is presented. β-actin was used as loading control. (B–G) Protein signals in A were determined by densitometric analysis of Western blotting using ImageJ software (NIH). Data are reported as the mean ± standard error ( n = 3, ∗ p < 0.05).

Article Snippet: The primary antibodies were as follows: Smad2/3 (1:500, Affinity, United States), vimentin (1:500, Proteintech, China), α-SMA (1:500, Proteintech, China), Col-1A1 (1:500, Novus, United States), TGF-β1 (1:500, Affinity, United States), TβRI (1:500, Abcam, United States), TβRII (1:500, Abcam, United States), Smad2/3 (1:500, Affinity, United States), p-Smad2/3 (1:500, Affinity, United States), and β-actin (1:1,000, Abcam, United States).

Techniques: Transformation Assay, In Vivo, Western Blot, Control, Software

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Asiaticoside Combined With Carbon Ion Implantation to Improve the Biocompatibility of Silicone Rubber and to Reduce the Risk of Capsule Contracture

doi: 10.3389/fbioe.2022.810244

Figure Lengend Snippet: Effect of asiaticoside on the transformation of fibroblasts into myofibroblasts, collagen deposition and fibroblast proliferation in vivo . Immunohistochemical staining of tissue around silicone rubber (SR) and carbon silicone rubber (C-SR) with α-SMA, vimentin, PCNA and COL-1A1 (100X).

Article Snippet: The primary antibodies were as follows: Smad2/3 (1:500, Affinity, United States), vimentin (1:500, Proteintech, China), α-SMA (1:500, Proteintech, China), Col-1A1 (1:500, Novus, United States), TGF-β1 (1:500, Affinity, United States), TβRI (1:500, Abcam, United States), TβRII (1:500, Abcam, United States), Smad2/3 (1:500, Affinity, United States), p-Smad2/3 (1:500, Affinity, United States), and β-actin (1:1,000, Abcam, United States).

Techniques: Transformation Assay, In Vivo, Immunohistochemical staining, Staining

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: Effect of BTXA on the viability of mouse L929 fibroblasts. (A) The morphological changes of fibroblasts observed under a microscope. (B) The vimentin identification of fibroblasts with Hoechst 33258 was determined by immunofluorescence assay. (C) BTXA inhibited fibroblast viability, which was detected by cell counting kit-8 assay in a dose- (0, 0.125, 0.25, 0.5, 1 and 2 IU/ml) and time- (12, 24 and 48 h) dependent manner. ** P<0.01 vs. control. BTXA, botulinum toxin type A.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Microscopy, Immunofluorescence, Cell Counting, Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: BTXA suppresses fibroblast viability and promotes apoptosis. (A) BTXA suppressed the high fibroblast viability induced by TGF-β1. (B) BTXA increased the apoptosis of fibroblasts induced by 10 ng/ml of TGF-β1. Dotted line separation represents whether or not fibroblasts were treated with TGF-β1. Data are shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^ P<0.05 and ^^ P<0.01 vs. control with TGF-β1. BTXA, botulinum toxin type A; TGF-β1, transforming growth factor-β1.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: BTXA prevents extracellular matrix (ECM) over-deposition in fibroblasts. BTXA decreased the high expression levels of collagen I, collagen III and α-SMA induced by 10 ng/ml of TGF-β1 in fibroblasts. The mRNA and protein levels of collagen I, collagen III and α-SMA were detected by (A) RT-qPCR and (B) western blot analysis, respectively. (C) α-SMA expression was determined by immunofluorescence. β-actin was used as an internal control. Dotted line separation represents whether or not fibroblast were treated with TGF-β1. Data were shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^ P<0.05 and ^^ P<0.01 vs. control with TGF-β1. BTXA, botulinum toxin type A; TGF-β1, transforming growth factor-β1.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Immunofluorescence, Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: BTXA suppresses the expression levels of MMP-2 and MMP-9. BTXA decreased the high expression levels of MMP-2 and MMP-9 induced by 10 ng/ml of TGF-β1 in fibroblasts. The mRNA and protein levels of MMP-2 and MMP-9 were respectively detected by (A) RT-qPCR and (B) western blot analysis, respectively. β-actin was used as an internal control. Dotted line separation represents whether or not fibroblast were treated with TGF-β1. Data are shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^ P<0.05 and ^^ P<0.01 vs. control with TGF-β1. BTXA, botulinum toxin type A; TGF-β1, transforming growth factor-β1; MMP, matrix metalloproteinase.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: Effect of BTXA on PTEN and caspase-3 expression, as well as PTEN methylation. (A) Effect of treatment of fibroblasts, which were cultured with or without 10 ng/ml of TGF-β1, with various concentrations (0.25, 0.5 and 1 UI/ml) of BTXA on the mRNA expression of PTEN, as determined by RT-qPCR. (B) Effects of BTXA on the protein levels of PTEN, pro-caspase-3 and cleaved-caspase-3, as detected by western blot analysis. (C) PTEN methylation was measured by methylation-specific PCR (MSP). M, methylated; U, unmethylated; MC, methylated control; UC, unmethylated control. β-actin was used as an internal control. Dotted line separation represents whether or not fibroblast were treated with TGF-β1. Data are shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^ P<0.05 and ^^ P<0.01 vs. control with TGF-β1. BTXA, botulinum toxin type A; TGF-β1, transforming growth factor-β1; PTEN, phosphatase and tensin homolog deleted on chromosome ten.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Expressing, Methylation, Cell Culture, Quantitative RT-PCR, Western Blot, Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: BTXA inhibits the activities of DNMTs. (A) The high DNMT activity induced by 10 ng/ml of TGF-β1 was decreased by BTXA. (B and C) The mRNA and protein levels of DNMT1, DNMT3a and DNMT3b were determined by RT-qPCR and western blot analysis, respectively. β-actin was used as an internal control. Dotted line separation represents whether or not fibroblasts were treated with TGF-β1. Data are shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^ P<0.05 and ^^ P<0.01 vs. control with TGF-β1. BTXA, botulinum toxin type A; TGF-β1, transforming growth factor-β1; DNMT, DNA methyltransferase.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Activity Assay, Quantitative RT-PCR, Western Blot, Control

Journal: International Journal of Molecular Medicine

Article Title: Botulinum toxin type A prevents the phenotypic transformation of fibroblasts induced by TGF-β1 via the PTEN/PI3K/Akt signaling pathway

doi: 10.3892/ijmm.2019.4226

Figure Lengend Snippet: BTXA inactivates the phosphorylation of PI3K and Akt. (A) The protein expression of p-PI3K, PI3K, p-Akt and Akt was examined by western blot analysis. (B) BTXA notably suppressed the ratio of p-PI3K/PI3K. (C) BTXA notably suppressed the ratio of p-Akt/Akt. β-actin was used as an internal control. Dotted line separation represents whether or not fibroblasts were treated with TGF-β1. Data are shown as the means ± SD, n=3. * P<0.05 and ** P<0.01 vs. control without TGF-β1; ^^ P<0.01 vs. control with TGF-β1.

Article Snippet: After being blocked in bovine serum albumin (BSA), the fibroblasts were incubated first with anti-α-SMA antibody (A03744, 1:200; BosterBio) overnight at 4°C and then with Cy3-conjugated goat anti-rabbit secondary antibodies (BA1032, 1:500; Beyotime) for 0.5 h. Subsequently, the fibroblasts were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime) and images were captured under a fluorescence microscopy (Nikon).

Techniques: Phospho-proteomics, Expressing, Western Blot, Control

Journal: American Journal of Translational Research

Article Title: RNF128 suppresses the malignancy of colorectal cancer cells via inhibition of Wnt/β-catenin signaling

doi:

Figure Lengend Snippet: Loss of RNF128 increases β-catenin protein level and its target genes in colorectal cancer. (A) The mRNA expressions of cyclin D1, cyclin E1, cyclin A1 and cyclin B1 were quantified in RNF128 knockdown or overexpression cells compared to their respective controls in DLD1. (B) β-catenin, CCND1, and RNF128 expressions were confirmed using Western blotting. β-actin was used as a loading control. (C) Expression of β-catenin and RNF128 in nuclear and cytoplasmic fractions in RNF128-knockdown cells and control cells of DLD1. β-tubulin was used as the cytoplasmic reference protein. LaminB1 was used as the nuclear reference protein. *P<0.05, **P<0.01 and ***P<0.001. No significant difference (P>0.05, ns). Data represented as mean ± SD.

Article Snippet: Supernatants were collected for measuring protein concentration by BCA assay (Thermofisher Scientific, USA). β-tubulin Rabbit mAb, Lamin B1 Rabbit mAb and β-actin Rabbit mAb were purchased from Proteintech (china); β-Catenin Rabbit mAb and CCND1 Rabbit mAb were from Cell Signaling Technology (USA); Ub and RNF128 Rat mAb were from Santa Cruz Biotechnology (USA); goat anti-rabbit and goat anti-rat were from Cell Signaling Technology (USA).

Techniques: Over Expression, Western Blot, Expressing