mouse anti human smad4  (Boster Bio)

Journal: Mediators of Inflammation

Article Title: The TGF- β 1 Signaling Pathway as an Attractive Target in the Fibrosis Pathogenesis of Sjögren's Syndrome

doi: 10.1155/2018/1965935

Figure Lengend Snippet: (a) Immunohistochemical localization of TGF- β 1, pSMAD2/3, and SMAD4 in healthy (A, C, E) and pSS (B, D, F) SG tissues. The analysis revealed TGF- β 1 expression in the cytoplasm of most interstitial infiltrating mononuclear cells, fibroblasts, and acinar and ductal cells of all the SG specimens analyzed. A strong positivity for TGF- β 1 was detected in acinar and ductal cells of pSS sections (B), and the intensity of labeling was strongly reduced in healthy salivary tissue (A); high levels of pSMAD2/3 were detected in pSS specimens, and expression analysis showed that the number of positive cells for pSMAD2/3 was higher in pSS samples (D) when compared with normal tissue (C). A prominent immunostaining for SMAD4 was observed in acinar and ductal cells of pSS biopsies (F) compared with healthy subjects (E). Brown staining shows positive immunoreaction; blue staining shows nuclei. (A, B, C, D, E, F) original magnification, ×20; bar = 20 μ m. All images were scanned and analyzed with Aperio ImageScope instrument. (b) panels G, H, and I represent immunohistochemistry signal quantification of TGF- β 1, pSMAD2/3, and SMAD4 positivity, respectively, performed by the Aperio ImageScope Software on healthy and pSS SG sections. Absorbance measurements performed by Aperio confirmed the microscopy observation and showed that staining for TGF- β 1, pSMAD2/3, and SMAD4 were significantly increased in pSS glands than in the control glands ( ∗∗ p < 0.01).

Article Snippet: The following antibodies were used for the study: mouse anti-human TGF- β 1 monoclonal antibody (mAb) (1 : 50, Santa Cruz Biotechnology, Santa Cruz, CA, USA), goat anti-human Smad2/3 polyclonal Ab (pAb) (1 : 100, R&D Systems, Minneapolis, MN, USA), goat anti-human p-SMAD2/3 (Ser 423/425) pAb (1 : 100, Santa Cruz Biotechnology), mouse anti-human SMAD4 mAb (1 : 50, Santa Cruz Biotechnology), rabbit anti-human Snail mAb (1 : 50, Cell Signaling Technology, Danvers, MA), mouse anti-human β -actin mAb clone AC-15 (1 : 100, Sigma-Aldrich, St. Louis, MO), mouse anti-human E-cadherin mAb (1 : 100, Dako, Santa Clara, CA, USA), mouse anti-human vimentin mAb (1 : 100, Thermo Fisher Scientific, Waltham, MA, USA), and rabbit anti-human collagen type I pAb (1 : 500, Thermo Fisher Scientific, Waltham, MA, USA).

Techniques: Immunohistochemical staining, Expressing, Labeling, Immunostaining, Staining, Immunohistochemistry, Software, Microscopy

Journal: Mediators of Inflammation

Article Title: The TGF- β 1 Signaling Pathway as an Attractive Target in the Fibrosis Pathogenesis of Sjögren's Syndrome

doi: 10.1155/2018/1965935

Figure Lengend Snippet: Schematic illustration of a working model for the TGF- β 1/SMAD/Snail pathway in modulation of EMT-dependent fibrosis in SGEC. TGF- β 1 stimulates epithelial cells by binding and activating transmembrane kinase receptors leading to phosphorylation and activation of Smad2/3. Once activated, pSmad2/3 form heterocomplexes with Smad4, which collectively translocate to the nucleus to mediate signaling events linked to EMT activation. The activation of the transcription factor Snail and induction of EMT markers have, as consequence, the upregulation of the mesenchymal marker Vimentin and the downregulation of the epithelial marker E-cadherin. During the activated EMT program, SGEC exhibit dramatic morphological changes and the gain of mesenchymal properties including increased migratory capacity and contractility. Finally, these mesenchymal cells become myofibroblasts which are responsible for progressive SG fibrosis.

Article Snippet: The following antibodies were used for the study: mouse anti-human TGF- β 1 monoclonal antibody (mAb) (1 : 50, Santa Cruz Biotechnology, Santa Cruz, CA, USA), goat anti-human Smad2/3 polyclonal Ab (pAb) (1 : 100, R&D Systems, Minneapolis, MN, USA), goat anti-human p-SMAD2/3 (Ser 423/425) pAb (1 : 100, Santa Cruz Biotechnology), mouse anti-human SMAD4 mAb (1 : 50, Santa Cruz Biotechnology), rabbit anti-human Snail mAb (1 : 50, Cell Signaling Technology, Danvers, MA), mouse anti-human β -actin mAb clone AC-15 (1 : 100, Sigma-Aldrich, St. Louis, MO), mouse anti-human E-cadherin mAb (1 : 100, Dako, Santa Clara, CA, USA), mouse anti-human vimentin mAb (1 : 100, Thermo Fisher Scientific, Waltham, MA, USA), and rabbit anti-human collagen type I pAb (1 : 500, Thermo Fisher Scientific, Waltham, MA, USA).

Techniques: Binding Assay, Activation Assay, Marker

Journal: Experimental and Therapeutic Medicine

Article Title: Identification of an activation-related protein in B cells in the ABO incompatible condition

doi: 10.3892/etm.2019.8234

Figure Lengend Snippet: Analysis of the interaction protein of HP. (A) Bioinformatics analysis of protein interactions with HP (3240). The numbers in the figure are the Gene IDs obtained from the National Center for Biotechnology Information (NCBI) database (https://www.ncbi.nlm.nih.gov/gene/?term=). The indegree is indicated by the size of the nodes. The edge betweenness and shared interaction are indicated with the width and length of the lines, respectively. The direction of interaction is indicated with an arrow. (B) The protein-protein interaction network of HP and Smad3 (4088). (C) Interaction analysis of HP and Smad3. PVDF membranes were blotted with mouse anti-human HP (dilution 1:1,000), followed by incubation with cell protein extract of the HK2+PBMC group, rabbit anti-human Smad3 (dilution 1:200) and goat anti-rabbit IgG-HRP (dilution 1:5,000). (D) Interaction analysis of HP and Smad4. PVDF membranes were blotted with rabbit anti-human HP (dilution 1:200), followed by incubation with cell protein extract of the HK2+PBMC group, mouse anti-human Smad4 (dilution 1:200) and goat anti-mouse IgG-HRP (dilution 1:5,000). (E) Interaction analysis of Smad3 and Smad4. PVDF membranes were blotted with rabbit anti-human Smad3 (dilution 1:200), followed by incubation with cell protein extract of the HK2+PBMC group, mouse anti-human Smad4 (dilution 1:200) and goat anti-mouse IgG-HRP (dilution 1:5,000). The experiments were repeated at least 3 times. PBMC, peripheral blood mononuclear cell; HP, haptoglobin.

Article Snippet: After 1 μl mouse anti-human HP (dilution 1:1,000; Abcam; cat. no. ab13429), rabbit anti-human HP (dilution 1:200), or mouse anti-human Smad4 (dilution 1:200; BosterBio) was applied to PVDF membranes, the membranes were blocked with blocking buffer at room temperature for 1 h. Following 3 washes with TBS for 5 min each, the cell protein extraction of the HK2+PBMC group (dilution 1:50) was added to the membranes and incubated at room temperature for 1 h and then rinsed 3 times with TBS for 5 min each.

Techniques: Incubation