Journal: The Journal of Biological Chemistry

Article Title: Effects of Ultraviolet-A and Riboflavin on the Interaction of Collagen and Proteoglycans during Corneal Cross-linking *

doi: 10.1074/jbc.M110.169813

Figure Lengend Snippet: SDS-PAGE patterns of cross-linking core proteins. RFUVA caused the monomer band of each PG to virtually disappear. For keratocan, lumican, and mimecan, RFUVA + PG did not cause a higher Mr band to form, instead forming new higher Mr polymers of a wide range of sizes from 100/150 kDa to the base of the gel well (lanes 3, 5, and 7). For decorin, RFUVA caused an entirely new band to form of approximately twice the size of the monomer band (lane 9). Gels were stained with Coomassie Blue.

Article Snippet: Recombinant mouse mimecan and recombinant human decorin proteins were purchased from R&D Systems (Minneapolis, MN).

Techniques: SDS Page, Staining

Journal: The Journal of Biological Chemistry

Article Title: Effects of Ultraviolet-A and Riboflavin on the Interaction of Collagen and Proteoglycans during Corneal Cross-linking *

doi: 10.1074/jbc.M110.169813

Figure Lengend Snippet: Interaction of core proteins keratocan, lumican, and mimecan with type I collagen. Presence of keratocan or lumican prevented RFUVA from causing diminished staining of α chains and β chains of collagen (lanes 5 and 7). In contrast, mimecan did not interfere with collagen cross-linking, thus allowing α chains and β chains to be cross-linked to form high Mr polymers near γ chains in size and toward the top range of the gel, and staining intensity increased sharply at the base of the sample well (lane 9). Gels were stained with Coomassie Blue.

Article Snippet: Recombinant mouse mimecan and recombinant human decorin proteins were purchased from R&D Systems (Minneapolis, MN).

Techniques: Staining

Journal: The Journal of Biological Chemistry

Article Title: Effects of Ultraviolet-A and Riboflavin on the Interaction of Collagen and Proteoglycans during Corneal Cross-linking *

doi: 10.1074/jbc.M110.169813

Figure Lengend Snippet: Western blot of proteoglycan core protein interaction with type I collagen induced by RFUVA. Western blot is shown. A, the presence of keratocan prevented RFUVA from causing collagen type I to undergo cross-linking (lane 4 versus lane 2). The intensities of α and β chains (lane 4) were almost the same as the corresponding bands in lane 1. B, lumican inhibited collagen cross-linking (lane 4 versus lane 2). C, mimecan did not inhibit RFUVA-induced collagen cross-linking (lane 2 versus lane 4), whereas mimecan formed a new band near γ chains and in the top range of the gel (lane 8 versus lane 7). D, decorin did not inhibit RFUVA-induced collagen cross-linking (lane 2 versus lane 4), whereas decorin formed a new band near γ chains (lane 8 versus lane 7). Decorin dimers and oligomers induced by RFUVA were observed (lane 7).

Article Snippet: Recombinant mouse mimecan and recombinant human decorin proteins were purchased from R&D Systems (Minneapolis, MN).

Techniques: Western Blot

Journal: The Journal of Biological Chemistry

Article Title: Effects of Ultraviolet-A and Riboflavin on the Interaction of Collagen and Proteoglycans during Corneal Cross-linking *

doi: 10.1074/jbc.M110.169813

Figure Lengend Snippet: Effect of keratocan or lumican on RFUVA cross-linking of collagen in the simultaneous presence of both mimecan and decorin. Western blot is shown. A, in the simultaneous presence of both mimecan and decorin, collagen cross-linking in response to RFUVA is not inhibited, and both mimecan and decorin each undergo their respective patterns of cross-linking. B, in contrast, in the presence of keratocan, as well as mimecan and decorin, collagen I cross-linking by RFUVA was partly inhibited (lane 3 versus lane 2). In addition, RFUVA-induced cross-linking of both decorin and mimecan in the presence or absence of collagen I are inhibited (lane 4 versus lane 5 and lane 6 versus lane 7). C, similarly, in the presence of lumican, collagen I cross-linking by RFUVA was partly inhibited (lane 3 versus lane 2). In addition, RFUVA-induced cross-linking of both decorin and mimecan in the presence or absence of collagen I is inhibited (lane 4 versus lane 5 and lane 6 versus lane 7).

Article Snippet: Recombinant mouse mimecan and recombinant human decorin proteins were purchased from R&D Systems (Minneapolis, MN).

Techniques: Western Blot

Journal: The Journal of Biological Chemistry

Article Title: Effects of Ultraviolet-A and Riboflavin on the Interaction of Collagen and Proteoglycans during Corneal Cross-linking *

doi: 10.1074/jbc.M110.169813

Figure Lengend Snippet: Patterns of cross-linking collagens and PG core proteins in whole cornea ex vivo. Western blot is shown. A, interaction of keratocan and collagen type I from corneas (with or without RFUVA treatment) treated by CNBr (lanes 4 and 5), N-glycanase (NGase) (lanes 6 and 7), and keratanase II (KSase) (lanes 8 and 9). B, interaction of lumican and collagen type I from corneas (with or without RFUVA treatment) treated by CNBr (lanes 4 and 5), N-glycanase (lanes 6 and 7), and keratanase II (lanes 8 and 9). C, interaction of mimecan and collagen type I from corneas (with or without RFUVA treatment) treated by CNBr (lanes 4 and 5), N-glycanase (lanes 6 and 7), and keratanase II (lanes 8 and 9). D, interaction of decorin and collagen type I from corneas (with or without RFUVA treatment) treated by CNBr (lanes 4 and 5), O-glycanase (OGase) (lanes 6 and 7), and chondroitinase ABC (ABCase) (lanes 8 and 9). All four PG core proteins in the cornea show some molecules that co-migrate in the same region as collagen γ chains (arrowheads).

Article Snippet: Recombinant mouse mimecan and recombinant human decorin proteins were purchased from R&D Systems (Minneapolis, MN).

Techniques: Ex Vivo, Western Blot