Fig 1: Skin biopsies from MDE patient (b, d, f, and h) and healthy donor (a, c, e, and g). M1 macrophages (iNOS positive) are present in MDE areas together with iNOS negative macrophages (b). No reaction is noticed in the middermis of control patients (a). A weak reaction for MMP-2 (c) and TIMP-2 (e) is detected on cell membranes in the middermis of a control subject. A positive reaction for MMP-2 (d) and TIMP-2 (f) is evident on the cell surface of middermal fibroblasts and inflammatory cells. A diffuse reaction for MMP-14 is localized in large areas of the middermis from MDE patient (h). A mild reaction for MMP-14 is present also on cell membranes in the middermis from a control subject (g). (a-b) Original magnification ×100; (c-d) original magnification ×200; (e-f) original magnification ×100; original magnification ×40.
Fig 2: Purified human fibrinogen inhibits gelatinolytic activity of recombinant human MMP-2. (a) Lineweaver-Burk plot of the proteolytic processing of DQ-gelatin by MMP-2 in the absence or presence of 3 mg/mL FBG; [MMP-2] = 0.001 mg/mL (13.9 nM) and [DQ-gelatin] = 0.02, 0.04, 0.06, 0.08 or 0.1 mg/mL. (b) Bar graphs showing the effect of intact fibrinogen (3 mg/mL) on the activity of MMP-2 (0.001 mg/mL); MMP-2 alone and human serum albumin (3 mg/mL) plus MMP-2 were used as a control. *P < 0.05 vs MMP-2 determined by student’s t-test. ns, not significant. Data are shown as mean of triplicates. (c) Left: SDS-PAGE confirming complete degradation of fibrinogen (6 mg/mL) when incubated with plasmin (0.001 mg/mL) for 12 hours at 37 °C. Right: Bar graphs showing that and plasmin-degraded fibrinogen fragments (3 mg/mL) vs intact fibrinogen (3 mg/mL) have no effect on the activity of MMP-2 (0.001 mg/mL). *P < 0.05 vs MMP-2 determined by student’s t-test. ns, not significant. (d) Bar graph showing restoration of MMP-2 activity when FBG is selectively removed from solution by an anti-FBG antibody. [MMP-2] = 0.001 mg/mL (13.9 nM); [DQ-gelatin] = 0.05 mg/mL; [FBG] = 2 mg/mL (5.88 µM); [IgG] and [anti-FBG] = 5.88 µM. *P < 0.05; determined by two-tailed student’s t-test (n = 4). (e) Plot showing the effect of increasing fibrinogen concentrations (0.0 to 10 mg/mL) on MMP-2 (0.001 mg/mL) activity. Note that high circulating FBG concentrations (as those found in RA patients (Fig. 2a)) effectively inhibit MMP-2 activity by more than 50%. Data are presented as mean ± standard error of mean. FBG, fibrinogen; RFU, relative fluorescence unit.
Fig 3: Fibrinogen and Marimastat bind at a common region of the catalytic domain of MMP-2. (a) Molecular docking of MMP-2 to FBG. Labelled residues of MMP-2 (green) that form hydrogen bonds (blue dotted lines) with FBG residues (magenta) are presented. (b) Molecular docking of MMP-2 to Marimastat. Labelled residues of MMP-2 (green) that form hydrogen bonds (blue dotted lines) with Marimastat (red) are presented.
Fig 4: Degradation of fibrillary collagens by active MMP-2. (A–C) Expression mRNA levels of collagen type I, III and V. n = 5 for Sham, n = 3 for Day 3 and 7, n = 4 for Day 14, and n = 5 for Day 28. (D) Representative image of MMP-2 gelatin acrylamide gel zymography. (E and F) Quantitative analysis of the relative levels of MMP-2 activity in liver lysates. n = 2 for Sham, n = 3 for Day 3, n = 4 for Day 7 and 14, and n = 5 for Day 28. Data are presented as means ± SD. One-way ANOVA followed by post-hoc Tukey’s test was used. §p < 0.05 vs. BH untreated of each time point, §§p < 0.01 vs. BH untreated of each time point, §§§p < 0.001 vs. BH untreated of each time point, #p < 0.05 vs. BH penumbra of each time point, ##p < 0.01 vs. BH penumbra of each time point, ###p < 0.001 vs. BH penumbra of each time, *p < 0.05 vs. sham, **p < 0.01 vs. sham and ***p < 0.001 vs. sham.
Fig 5: The 10 kDa VTN fragment originates from the reduction of the disulfide bond between the V65 and V10 subunits. (A) C-VTN non-reducing western blotting analysis of 1:10 dilutions of a NASH serum sample in the presence or absence of MMP-2 and -9. Total protein staining by Ponceau S on the nitrocellulose membrane is shown. (B) C-VTN western blotting analysis of 1:10 dilutions of a NASH serum sample in the presence or absence of a sample-reducing agent (SRA). Total protein staining by Ponceau S on the nitrocellulose membrane is shown. (C) Non-reducing western blotting analysis of 1:10 dilutions of a NASH serum sample probed with antibodies specific for the vitronectin C-terminal (C-VTN) and N-terminal (N-VTN) ends. These images are representative of experiments carried out in triplicate.
Supplier Page from Abcam for Recombinant human MMP2 protein (Active)