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The Anti-inflammatory Protein TSG-6 Regulates Chemokine Function by Inhibiting Chemokine/Glycosaminoglycan Interactions.

Dyer DP, Salanga CL, Johns SC, Valdambrini E, Fuster MM, Milner CM, Day AJ, Handel TM - J. Biol. Chem. (2016)

Bottom Line: We also show that the Link_TSG6-binding sites on chemokines overlap with chemokine GAG-binding sites, and that the affinities of Link_TSG6 for these chemokines (KD values 1-85 nm) broadly correlate with chemokine-GAG affinities.Link_TSG6 also inhibits chemokine presentation on endothelial cells not only through a direct interaction with chemokines but also by binding and therefore masking the availability of GAGs.Along with previous work, these findings suggest that TSG-6 functions as a pluripotent regulator of chemokines by modulating chemokine/GAG interactions, which may be a major mechanism by which TSG-6 produces its anti-inflammatory effects in vivo.

View Article: PubMed Central - PubMed

Affiliation: From the Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093-0684, the Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8TA, Scotland, United Kingdom.

No MeSH data available.


Related in: MedlinePlus

Pre-incubation of chemokine with Link_TSG6 inhibits subsequent presentation on collagen. Biotinylated CCL21, CXCL12, or CXCL4 were incubated at different concentrations (10, 50, or 200 nm) on wells pre-coated with collagen, and the amounts bound following washing were detected using labeled streptavidin (A). The biotinylated chemokines CCL21 (50 nm) (B), CXCL12 (50 nm) (C), or CXCL4 (10 nm) (D) were incubated either alone or in combination with different molar ratios of Link_TSG6 (ratios given as Link_TSG6/chemokine) prior to incubation on the collagen-coated surfaces, followed by detection as before. Data are expressed as total binding (relative fluorescence intensity) (A) or as a percentage of maximal binding of chemokine alone (B–D), plotted as mean values (±S.E.) from two independent experiments, each undertaken in duplicate (n = 2). *, p < 0.05; **, p < 0.01 (compared with chemokine-only controls), as determined using repeated measures ANOVA analysis with a Bonferroni post hoc test.
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Figure 9: Pre-incubation of chemokine with Link_TSG6 inhibits subsequent presentation on collagen. Biotinylated CCL21, CXCL12, or CXCL4 were incubated at different concentrations (10, 50, or 200 nm) on wells pre-coated with collagen, and the amounts bound following washing were detected using labeled streptavidin (A). The biotinylated chemokines CCL21 (50 nm) (B), CXCL12 (50 nm) (C), or CXCL4 (10 nm) (D) were incubated either alone or in combination with different molar ratios of Link_TSG6 (ratios given as Link_TSG6/chemokine) prior to incubation on the collagen-coated surfaces, followed by detection as before. Data are expressed as total binding (relative fluorescence intensity) (A) or as a percentage of maximal binding of chemokine alone (B–D), plotted as mean values (±S.E.) from two independent experiments, each undertaken in duplicate (n = 2). *, p < 0.05; **, p < 0.01 (compared with chemokine-only controls), as determined using repeated measures ANOVA analysis with a Bonferroni post hoc test.

Mentions: Chemokines are found abundantly in the ECM (31), and TSG-6 is known to function in the ECM (5, 9, 10). Thus, following our observations that Link_TSG6 can inhibit chemokine presentation on endothelial cells, we investigated whether it inhibits chemokine binding to collagen, an important component of the ECM. In agreement with previous studies describing chemokine/collagen interactions (65, 66), we observed that CCL21, CXCL12, and CXCL4 bind to collagen-coated surfaces (Fig. 9A). Notably, pre-incubation of these chemokines with Link_TSG6 resulted in dose-dependent inhibition of their binding to collagen (Fig. 9, B–D). These findings suggest TSG-6 may function as a general modulator of chemokine presentation on endothelial cell HS and extracellular matrix components, including GAGs and collagen.


The Anti-inflammatory Protein TSG-6 Regulates Chemokine Function by Inhibiting Chemokine/Glycosaminoglycan Interactions.

Dyer DP, Salanga CL, Johns SC, Valdambrini E, Fuster MM, Milner CM, Day AJ, Handel TM - J. Biol. Chem. (2016)

Pre-incubation of chemokine with Link_TSG6 inhibits subsequent presentation on collagen. Biotinylated CCL21, CXCL12, or CXCL4 were incubated at different concentrations (10, 50, or 200 nm) on wells pre-coated with collagen, and the amounts bound following washing were detected using labeled streptavidin (A). The biotinylated chemokines CCL21 (50 nm) (B), CXCL12 (50 nm) (C), or CXCL4 (10 nm) (D) were incubated either alone or in combination with different molar ratios of Link_TSG6 (ratios given as Link_TSG6/chemokine) prior to incubation on the collagen-coated surfaces, followed by detection as before. Data are expressed as total binding (relative fluorescence intensity) (A) or as a percentage of maximal binding of chemokine alone (B–D), plotted as mean values (±S.E.) from two independent experiments, each undertaken in duplicate (n = 2). *, p < 0.05; **, p < 0.01 (compared with chemokine-only controls), as determined using repeated measures ANOVA analysis with a Bonferroni post hoc test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4933465&req=5

Figure 9: Pre-incubation of chemokine with Link_TSG6 inhibits subsequent presentation on collagen. Biotinylated CCL21, CXCL12, or CXCL4 were incubated at different concentrations (10, 50, or 200 nm) on wells pre-coated with collagen, and the amounts bound following washing were detected using labeled streptavidin (A). The biotinylated chemokines CCL21 (50 nm) (B), CXCL12 (50 nm) (C), or CXCL4 (10 nm) (D) were incubated either alone or in combination with different molar ratios of Link_TSG6 (ratios given as Link_TSG6/chemokine) prior to incubation on the collagen-coated surfaces, followed by detection as before. Data are expressed as total binding (relative fluorescence intensity) (A) or as a percentage of maximal binding of chemokine alone (B–D), plotted as mean values (±S.E.) from two independent experiments, each undertaken in duplicate (n = 2). *, p < 0.05; **, p < 0.01 (compared with chemokine-only controls), as determined using repeated measures ANOVA analysis with a Bonferroni post hoc test.
Mentions: Chemokines are found abundantly in the ECM (31), and TSG-6 is known to function in the ECM (5, 9, 10). Thus, following our observations that Link_TSG6 can inhibit chemokine presentation on endothelial cells, we investigated whether it inhibits chemokine binding to collagen, an important component of the ECM. In agreement with previous studies describing chemokine/collagen interactions (65, 66), we observed that CCL21, CXCL12, and CXCL4 bind to collagen-coated surfaces (Fig. 9A). Notably, pre-incubation of these chemokines with Link_TSG6 resulted in dose-dependent inhibition of their binding to collagen (Fig. 9, B–D). These findings suggest TSG-6 may function as a general modulator of chemokine presentation on endothelial cell HS and extracellular matrix components, including GAGs and collagen.

Bottom Line: We also show that the Link_TSG6-binding sites on chemokines overlap with chemokine GAG-binding sites, and that the affinities of Link_TSG6 for these chemokines (KD values 1-85 nm) broadly correlate with chemokine-GAG affinities.Link_TSG6 also inhibits chemokine presentation on endothelial cells not only through a direct interaction with chemokines but also by binding and therefore masking the availability of GAGs.Along with previous work, these findings suggest that TSG-6 functions as a pluripotent regulator of chemokines by modulating chemokine/GAG interactions, which may be a major mechanism by which TSG-6 produces its anti-inflammatory effects in vivo.

View Article: PubMed Central - PubMed

Affiliation: From the Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093-0684, the Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8TA, Scotland, United Kingdom.

No MeSH data available.


Related in: MedlinePlus