Journal: The American journal of pathology

Article Title: The Versican G1 Fragment and Serum-Derived Hyaluronan-Associated Proteins Interact and Form a Complex in Granulation Tissue of Pressure Ulcers.

doi: 10.1016/j.ajpath.2017.10.015

Figure Lengend Snippet: Figure 9 Transient versican G1 fragmenteserum-derived hyaluronan-associated proteinehyaluronan (VG1F-SHAP-HA) complex formation in a mouse model of moist wound healing. A: Representative images from wound tissues from the moist wound-healing model are shown. Edematous granulation tissues were observed at day 3 after wounding; however, the granulation tissues were nonedematous and thin at days 1 and 6. The thickness of the granulation tissues is as follows: day 1, 0.77 0.10 mm; day 3, 2.55 0.17 mm; and day 6, 0.82 0.04 mm. B: Tissue specimens were stained with hematoxylin and eosin. C: Isolated tissues were stained by using the indicated probes. The VG1F-SHAP-HA complex was observed on day 3 in specific lesions, but was absent at days 1 and 6. The color of letters indicating molecules represents the color of the stain. The merged images indicate the VG1F-SHAP-HA complex. D: Immunohis- tochemical analysis of wound tissue taken from day 3. The specimen was stained with the indicated probes, and the merged images show VG1F colocalization, as detected by probes for HA, DPEAAE epitope, and macrophages (CD68). The merged images show macrophage/complex colocalization. Data are expressed as means SD (A). n Z 4 independent experiments (A). Scale bars Z 50 mm (BeD). HC, heavy chain.

Article Snippet: A goat pAb against bikunin (sc-21597) was purchased from Santa Cruz Biotechnology Inc. A mouse monoclonal antibody against the macrophage marker CD68 (STJ-6572) was purchased from St. John’s Laboratory Ltd (London, UK).

Techniques: Derivative Assay, Staining, Isolation

Journal: The American journal of pathology

Article Title: The Versican G1 Fragment and Serum-Derived Hyaluronan-Associated Proteins Interact and Form a Complex in Granulation Tissue of Pressure Ulcers.

doi: 10.1016/j.ajpath.2017.10.015

Figure Lengend Snippet: Figure 11 The versican G1 fragment (VG1F)eserum-derived hyaluronan (HA)eassociated proteineHA complex in human pressure ulcers. A: In human pressure ulcers, edematous wounds (e1 and e2) typically become flat and stabilized (f3 and f4) after appropriate treatments. B: The VG1F-HC complex was detected from the protein extract of wound surfaces shown in A (e1 and e2). The samples are indicated at the top of the figure and correspond to those shown in A. The samples were extracted with 6 mol/L GdnHCl and treated with trypsin and V8 protease (Materials and Methods); separated using SDS-PAGE under nonreducing conditions; and blotted. Incubation with the indicated antibodies revealed the cross-linked heavy chain (HC) and VG1 species (dagger). The bands of cross-linked HC and VG1 species (dagger) disappeared during wound healing. C: Paired samples from edematous or flat wounds were analyzed using Western blotting, and the bands shown in B were quantified. The relative intensities of the bands [VG1-HCs (daggers in B)]/the total polyclonal antibody 6084epositive bands (all VG1) differ significantly (P Z 0.02) between edematous and flat wounds. D: Direct immunofluorescence of wound surface samples obtained from different sites of one individual wound. Samples obtained from the edematous wound (green dashed circle) and flat wound (blue dashed circle) were fixed and incubated with VG1 and HC1 antibodies, as well as biotin-conjugated HAebinding protein, as indicated. A representative result from triplicate experiments with different samples is shown. E: Immunohistochemical analysis of pressure ulcer tissue. The specimen was stained, as indicated. The merged images show VG1F colocalization, as detected by antibodies for DPEAAE epitope, HC1, and macrophage CD68. Data are expressed as means SD (C). n Z 10 (C, each paired sample). Scale bars: 5 mm (D); 100 mm (E).

Article Snippet: A goat pAb against bikunin (sc-21597) was purchased from Santa Cruz Biotechnology Inc. A mouse monoclonal antibody against the macrophage marker CD68 (STJ-6572) was purchased from St. John’s Laboratory Ltd (London, UK).

Techniques: Derivative Assay, SDS Page, Incubation, Western Blot, Immunohistochemical staining, Staining