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Gottron's papules exhibit dermal accumulation of CD44 variant 7 (CD44v7) and its binding partner osteopontin: a unique molecular signature.

Kim JS, Bashir MM, Werth VP - J. Invest. Dermatol. (2012)

Bottom Line: Mechanically stretching cultured fibroblasts for 6 hours induced CD44v7 mRNA and protein, whereas IFN-γ treatment induced OPN mRNA and protein.OPN alone did not induce CD44v7, but stretching dermal fibroblasts in the presence of OPN increased human acute monocytic leukemia cell line (THP-1) monocyte binding, which is blunted by anti-CD44v7 blocking antibody.C4S, CD44v7, and OPN are three molecules uniquely present in Gottron's papules that contribute to inflammation individually and in association with one another.

View Article: PubMed Central - PubMed

Affiliation: New York University School of Medicine, New York, New York, USA.

ABSTRACT
The accumulated mucin in non-Gottron's dermatomyositis (DM) lesions is primarily chondroitin-4-sulfate (C4S), which is immunomodulatory in vitro. Gottron's papules are a particularly resistant manifestation of DM that often persist after other lesions have resolved with therapy. We examined non-Gottron's DM lesions and Gottron's papule skin biopsies for C4S, CD44 variant 7 (CD44v7), a chondroitin sulfate-binding isoform causally implicated in autoimmunity, and osteopontin (OPN), a CD44v7 ligand implicated in chronic inflammation. Gottron's papule dermis contained more C4S and CD44v7 than non-Gottron's lesions. Normal skin showed less CD44v7 over joints relative to Gottron's lesions. All DM dermis had increased OPN compared with healthy skin. Mechanically stretching cultured fibroblasts for 6 hours induced CD44v7 mRNA and protein, whereas IFN-γ treatment induced OPN mRNA and protein. OPN alone did not induce CD44v7, but stretching dermal fibroblasts in the presence of OPN increased human acute monocytic leukemia cell line (THP-1) monocyte binding, which is blunted by anti-CD44v7 blocking antibody. C4S, CD44v7, and OPN are three molecules uniquely present in Gottron's papules that contribute to inflammation individually and in association with one another. We propose that stretch-induced CD44v7 over joints, in concert with dysregulated OPN levels in the skin of DM patients, increases local inflammatory cell recruitment and contributes to the pathogenesis and resistance of Gottron's papules.

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Mechanical stretching induces CD44v7 in cultured human dermal fibroblastsConfluent cultured human dermal fibroblasts grown on flexible membranes were subjected to tonic mechanical stretching. All cells were plated simultaneously and stretched for the final 6 or 12 h of incubation, after which RNA was harvested simultaneously. a. CD44v7 mRNA levels were assayed by qRT-PCR, normalized to PPIA mRNA levels, and then expressed relative to unstretched controls. Every data point from a representative experiment is displayed; wide and short horizontal bars represent mean and SEM, respectively. P=0.0017 by ANOVA; (**, P<0.01; n.s., not significant); b. CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours were assayed by western blot and expressed relative to unstretched controls; c. Mean CD44v7 protein detected in one representative experiment; horizontal bar represents SEM. P=0.0321 by ANOVA (*, p<0.05).
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Figure 4: Mechanical stretching induces CD44v7 in cultured human dermal fibroblastsConfluent cultured human dermal fibroblasts grown on flexible membranes were subjected to tonic mechanical stretching. All cells were plated simultaneously and stretched for the final 6 or 12 h of incubation, after which RNA was harvested simultaneously. a. CD44v7 mRNA levels were assayed by qRT-PCR, normalized to PPIA mRNA levels, and then expressed relative to unstretched controls. Every data point from a representative experiment is displayed; wide and short horizontal bars represent mean and SEM, respectively. P=0.0017 by ANOVA; (**, P<0.01; n.s., not significant); b. CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours were assayed by western blot and expressed relative to unstretched controls; c. Mean CD44v7 protein detected in one representative experiment; horizontal bar represents SEM. P=0.0321 by ANOVA (*, p<0.05).

Mentions: Because mechanical stretching has been reported to induce expression of CD44v7 and v8 in porcine ocular trabecular meshwork cells (Keller et al., 2007), we hypothesized that the increased CD44v7 seen in the IP skin of DM patients and healthy volunteers is induced by chronic stretching of the skin over joints. Thus, we stretched primary dermal fibroblasts on cell culture membrane inserts and then determined CD44v7 mRNA expression by quantitative real-time PCR (RT-PCR). We found that mechanical stretching of dermal fibroblasts induced a significant 1.6-fold increase in CD44v7 mRNA expression at six hours (p<0.01), which returned to a baseline expression by twelve hours (Figure 4a). CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours increased 3.3 ± 1.3 times relative to control fibroblasts (p<0.05) (Figure 4b,c). Osteopontin alone did not induce CD44v7 mRNA (data not shown). These results were reproducible in three repeat experiments. In contrast, mechanical stretching did not have a significant effect on fibroblastic osteopontin mRNA expression.


Gottron's papules exhibit dermal accumulation of CD44 variant 7 (CD44v7) and its binding partner osteopontin: a unique molecular signature.

Kim JS, Bashir MM, Werth VP - J. Invest. Dermatol. (2012)

Mechanical stretching induces CD44v7 in cultured human dermal fibroblastsConfluent cultured human dermal fibroblasts grown on flexible membranes were subjected to tonic mechanical stretching. All cells were plated simultaneously and stretched for the final 6 or 12 h of incubation, after which RNA was harvested simultaneously. a. CD44v7 mRNA levels were assayed by qRT-PCR, normalized to PPIA mRNA levels, and then expressed relative to unstretched controls. Every data point from a representative experiment is displayed; wide and short horizontal bars represent mean and SEM, respectively. P=0.0017 by ANOVA; (**, P<0.01; n.s., not significant); b. CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours were assayed by western blot and expressed relative to unstretched controls; c. Mean CD44v7 protein detected in one representative experiment; horizontal bar represents SEM. P=0.0321 by ANOVA (*, p<0.05).
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Figure 4: Mechanical stretching induces CD44v7 in cultured human dermal fibroblastsConfluent cultured human dermal fibroblasts grown on flexible membranes were subjected to tonic mechanical stretching. All cells were plated simultaneously and stretched for the final 6 or 12 h of incubation, after which RNA was harvested simultaneously. a. CD44v7 mRNA levels were assayed by qRT-PCR, normalized to PPIA mRNA levels, and then expressed relative to unstretched controls. Every data point from a representative experiment is displayed; wide and short horizontal bars represent mean and SEM, respectively. P=0.0017 by ANOVA; (**, P<0.01; n.s., not significant); b. CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours were assayed by western blot and expressed relative to unstretched controls; c. Mean CD44v7 protein detected in one representative experiment; horizontal bar represents SEM. P=0.0321 by ANOVA (*, p<0.05).
Mentions: Because mechanical stretching has been reported to induce expression of CD44v7 and v8 in porcine ocular trabecular meshwork cells (Keller et al., 2007), we hypothesized that the increased CD44v7 seen in the IP skin of DM patients and healthy volunteers is induced by chronic stretching of the skin over joints. Thus, we stretched primary dermal fibroblasts on cell culture membrane inserts and then determined CD44v7 mRNA expression by quantitative real-time PCR (RT-PCR). We found that mechanical stretching of dermal fibroblasts induced a significant 1.6-fold increase in CD44v7 mRNA expression at six hours (p<0.01), which returned to a baseline expression by twelve hours (Figure 4a). CD44v7 protein in the supernatant of stretched fibroblasts at 6 hours increased 3.3 ± 1.3 times relative to control fibroblasts (p<0.05) (Figure 4b,c). Osteopontin alone did not induce CD44v7 mRNA (data not shown). These results were reproducible in three repeat experiments. In contrast, mechanical stretching did not have a significant effect on fibroblastic osteopontin mRNA expression.

Bottom Line: Mechanically stretching cultured fibroblasts for 6 hours induced CD44v7 mRNA and protein, whereas IFN-γ treatment induced OPN mRNA and protein.OPN alone did not induce CD44v7, but stretching dermal fibroblasts in the presence of OPN increased human acute monocytic leukemia cell line (THP-1) monocyte binding, which is blunted by anti-CD44v7 blocking antibody.C4S, CD44v7, and OPN are three molecules uniquely present in Gottron's papules that contribute to inflammation individually and in association with one another.

View Article: PubMed Central - PubMed

Affiliation: New York University School of Medicine, New York, New York, USA.

ABSTRACT
The accumulated mucin in non-Gottron's dermatomyositis (DM) lesions is primarily chondroitin-4-sulfate (C4S), which is immunomodulatory in vitro. Gottron's papules are a particularly resistant manifestation of DM that often persist after other lesions have resolved with therapy. We examined non-Gottron's DM lesions and Gottron's papule skin biopsies for C4S, CD44 variant 7 (CD44v7), a chondroitin sulfate-binding isoform causally implicated in autoimmunity, and osteopontin (OPN), a CD44v7 ligand implicated in chronic inflammation. Gottron's papule dermis contained more C4S and CD44v7 than non-Gottron's lesions. Normal skin showed less CD44v7 over joints relative to Gottron's lesions. All DM dermis had increased OPN compared with healthy skin. Mechanically stretching cultured fibroblasts for 6 hours induced CD44v7 mRNA and protein, whereas IFN-γ treatment induced OPN mRNA and protein. OPN alone did not induce CD44v7, but stretching dermal fibroblasts in the presence of OPN increased human acute monocytic leukemia cell line (THP-1) monocyte binding, which is blunted by anti-CD44v7 blocking antibody. C4S, CD44v7, and OPN are three molecules uniquely present in Gottron's papules that contribute to inflammation individually and in association with one another. We propose that stretch-induced CD44v7 over joints, in concert with dysregulated OPN levels in the skin of DM patients, increases local inflammatory cell recruitment and contributes to the pathogenesis and resistance of Gottron's papules.

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Related in: MedlinePlus