Journal: Life sciences

Article Title: Collagen matricryptin promotes cardiac function by mediating scar formation.

doi: 10.1016/j.lfs.2023.121598

Figure Lengend Snippet: Fig. 4. p1159 promotes fibroblast migration and this may occur via integrin-α4 (Itgα4). A. LV fibroblasts treated with p1159 (100 nM in serum free media, SFM) presented a significant increase in migration rate, which was similar to the positive control (10 % FBS), demonstrating strong pro-migratory properties. *p < 0.05 p1159 100 nM and 500 nM versus negative control (SFM); n = 6/group. B. p1159 in vivo treatment induces overexpression of Itgα4 post-MI. Left panel: mRNA levels n = 6/group; right panel: representative immunoblot image, TP = total protein stain (loading control), n = 4/group. **p < 0.01. C. The neutralizing antibody against Itga4 inhibited p1159 induced fibroblast migration. Positive control (10 % FBS), negative control (SFM), and p1159 (100 nM and 500 nM) were treated ± Itgα4 neutralizing antibody (Itga4i). The impedance values were normalized before incubation with Itga4i and after wound. *p < 0.05 represents when both concentrations of p1159 statistically differed from its respective groups with Itga4i, and from the negative controls; n = 6/group. D. Cardiac fibroblasts stimulated with p1159 overexpress Itga4 mRNA and this effect is abolished in the presence of Itga4i. Left panel: mRNA levels n = 6/group; right panel: representative immunoblot image, TP = total protein stain (loading control), n = 4/group. *p < 0.05 E. Itgα4 (red) co-localized with p1159 (green, FITC labeled) in human cardiac fibroblasts (HCF) and that effect was inhibited in the presence of Itga4i. Blue = nuclei, yellow = co-localization. Scale bar = 5 μm. All graphs display average ± SEM. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Cell migration was recorded in real time for 48 h. For the experiments testing integrin alpha 4 (Itgα4) as the receptor for p1159, cardiac fibroblasts were cultured in SFM with Itgα4 blocking antibody (#NBP1-26661, Novus Biologicals; Itgα4i) 15 μg/mL for 2 h before wound induction, followed by treatment as listed in Table 1.

Techniques: Migration, Positive Control, Negative Control, In Vivo, Over Expression, Western Blot, Staining, Control, Incubation, Labeling

Journal: Life sciences

Article Title: Collagen matricryptin promotes cardiac function by mediating scar formation.

doi: 10.1016/j.lfs.2023.121598

Figure Lengend Snippet: Fig. 5. p1159-induced migration may occur via Rho GTPase pathways. A. p1159-treatment reduced RhoGDI expression, and this effect was dependent on Itgα4. Gene expression of cardiac fibroblasts treated with p1159 100 nM ± Itgα4 blocking antibody (Ab) and negative control (SFM). *p < 0.05 versus negative control (SFM), #p < 0.05 versus 100 nM p1158/59. B. p1159 induces RhoA activation in fibroblasts. Representative immunofluorescence images of fibroblasts treated with negative control (SFM), positive control (10 % FBS) and SFM ± p1159 100 nM or 500 nM, indicating stimulation of Itgα4 receptor (green) and activation of RhoA (red). Right panels: Representative images of the same groups incubated 2 h prior with blocking antibody against Itgα4 (Itga4 Ab), indicating that both Itgα4 receptor and RhoA signals were strongly decreased by fibroblasts treated with p1159 in the presence of the Ab. Magnification/scale bars: 20× = 50 μm; 40× = 20 μm. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Cell migration was recorded in real time for 48 h. For the experiments testing integrin alpha 4 (Itgα4) as the receptor for p1159, cardiac fibroblasts were cultured in SFM with Itgα4 blocking antibody (#NBP1-26661, Novus Biologicals; Itgα4i) 15 μg/mL for 2 h before wound induction, followed by treatment as listed in Table 1.

Techniques: Migration, Expressing, Gene Expression, Blocking Assay, Negative Control, Activation Assay, Immunofluorescence, Positive Control, Incubation