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MicroRNA 181b regulates decorin production by dermal fibroblasts and may be a potential therapy for hypertrophic scar.

Kwan P, Ding J, Tredget EE - PLoS ONE (2015)

Bottom Line: Lower decorin levels were found in hypertrophic scar as compared to normal skin, and in deep as compared to superficial dermis.By blocking miR-181b with an antagomiR, it was possible to increase decorin protein expression in dermal fibroblasts.This suggests miR-181b is involved in the differential expression of decorin in skin and wound healing.

View Article: PubMed Central - PubMed

Affiliation: Division of Plastic Surgery, Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada; Wound Healing Research Group, Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
Hypertrophic scarring is a frequent fibroproliferative complication following deep dermal burns leading to impaired function and lifelong disfigurement. Decorin reduces fibrosis and induces regeneration in many tissues, and is significantly downregulated in hypertrophic scar and normal deep dermal fibroblasts. It was hypothesized that microRNAs in these fibroblasts downregulate decorin and blocking them would increase decorin and may prevent hypertrophic scarring. Lower decorin levels were found in hypertrophic scar as compared to normal skin, and in deep as compared to superficial dermis. A decorin 3' un-translated region reporter assay demonstrated microRNA decreased decorin in deep dermal fibroblasts, and microRNA screening predicted miR- 24, 181b, 421, 526b, or 543 as candidates. After finding increased levels of mir-181b in deep dermal fibroblasts, it was demonstrated that TGF-β1 stimulation decreased miR-24 but increased miR-181b and that hypertrophic scar and deep dermis contained increased levels of miR-181b. By blocking miR-181b with an antagomiR, it was possible to increase decorin protein expression in dermal fibroblasts. This suggests miR-181b is involved in the differential expression of decorin in skin and wound healing. Furthermore, blocking miR-181b reversed TGF-β1 induced decorin downregulation and myofibroblast differentiation in hypertrophic scar fibroblasts, suggesting a potential therapy for hypertrophic scar.

No MeSH data available.


Related in: MedlinePlus

The effect of antagomiR-181b on TGF-β1 stimulated NS and HSc fibroblasts.(a) antagomiR-181b reversed DCN downregulation in HSc fibroblasts. Cells were stimulated by TGF-β1 at indicated concentrations and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS, and DCN protein was measured using ELISA on the supernatants (mean ± SEM, n = 3, * P < 0.02, ** P < 0.006). (b) antagomiR-181b reversed myofibroblast differentiation in HSc fibroblasts. Cells were stimulated by TGF-β1 10 ng/mL and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS then permeabilized and stained for α-smooth muscle actin and 10 000 cells per sample measured by flow cytometry (mean ± SEM, n = 3, *** P <0.03).
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pone.0123054.g006: The effect of antagomiR-181b on TGF-β1 stimulated NS and HSc fibroblasts.(a) antagomiR-181b reversed DCN downregulation in HSc fibroblasts. Cells were stimulated by TGF-β1 at indicated concentrations and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS, and DCN protein was measured using ELISA on the supernatants (mean ± SEM, n = 3, * P < 0.02, ** P < 0.006). (b) antagomiR-181b reversed myofibroblast differentiation in HSc fibroblasts. Cells were stimulated by TGF-β1 10 ng/mL and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS then permeabilized and stained for α-smooth muscle actin and 10 000 cells per sample measured by flow cytometry (mean ± SEM, n = 3, *** P <0.03).

Mentions: Based on prior results, blocking miR-181b might treat HSc, so this strategy was examined in matched NS and HSc fibroblasts treated with TGF-β1. As shown in Fig 6A, TGF-β1 stimulation significantly decreased DCN in both NS (P < 0.02) and HSc (P < 0.02) fibroblasts, and antagomiR-181b treatment reversed the decrease in DCN induced by TGF-β1 in HSc fibroblasts, returning DCN to baseline (P < 0.02). Again, DCN protein levels are expressed as fold changes to allow comparison despite inter-individual variation. As shown in Fig 6B, TGF-β1 stimulation significantly increased myofibroblast differentiation (mean 7.99 fold increase compared to baseline, P < 0.03), and antagomiR-181b treatment reversed this effect, significantly decreasing the number of myofibroblasts (mean 3.01 fold increase compared to baseline, P = 0.01).


MicroRNA 181b regulates decorin production by dermal fibroblasts and may be a potential therapy for hypertrophic scar.

Kwan P, Ding J, Tredget EE - PLoS ONE (2015)

The effect of antagomiR-181b on TGF-β1 stimulated NS and HSc fibroblasts.(a) antagomiR-181b reversed DCN downregulation in HSc fibroblasts. Cells were stimulated by TGF-β1 at indicated concentrations and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS, and DCN protein was measured using ELISA on the supernatants (mean ± SEM, n = 3, * P < 0.02, ** P < 0.006). (b) antagomiR-181b reversed myofibroblast differentiation in HSc fibroblasts. Cells were stimulated by TGF-β1 10 ng/mL and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS then permeabilized and stained for α-smooth muscle actin and 10 000 cells per sample measured by flow cytometry (mean ± SEM, n = 3, *** P <0.03).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4383602&req=5

pone.0123054.g006: The effect of antagomiR-181b on TGF-β1 stimulated NS and HSc fibroblasts.(a) antagomiR-181b reversed DCN downregulation in HSc fibroblasts. Cells were stimulated by TGF-β1 at indicated concentrations and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS, and DCN protein was measured using ELISA on the supernatants (mean ± SEM, n = 3, * P < 0.02, ** P < 0.006). (b) antagomiR-181b reversed myofibroblast differentiation in HSc fibroblasts. Cells were stimulated by TGF-β1 10 ng/mL and transfected with antagomiR-control or antagomiR-181b for 48 hours in DMEM + 2% FBS then permeabilized and stained for α-smooth muscle actin and 10 000 cells per sample measured by flow cytometry (mean ± SEM, n = 3, *** P <0.03).
Mentions: Based on prior results, blocking miR-181b might treat HSc, so this strategy was examined in matched NS and HSc fibroblasts treated with TGF-β1. As shown in Fig 6A, TGF-β1 stimulation significantly decreased DCN in both NS (P < 0.02) and HSc (P < 0.02) fibroblasts, and antagomiR-181b treatment reversed the decrease in DCN induced by TGF-β1 in HSc fibroblasts, returning DCN to baseline (P < 0.02). Again, DCN protein levels are expressed as fold changes to allow comparison despite inter-individual variation. As shown in Fig 6B, TGF-β1 stimulation significantly increased myofibroblast differentiation (mean 7.99 fold increase compared to baseline, P < 0.03), and antagomiR-181b treatment reversed this effect, significantly decreasing the number of myofibroblasts (mean 3.01 fold increase compared to baseline, P = 0.01).

Bottom Line: Lower decorin levels were found in hypertrophic scar as compared to normal skin, and in deep as compared to superficial dermis.By blocking miR-181b with an antagomiR, it was possible to increase decorin protein expression in dermal fibroblasts.This suggests miR-181b is involved in the differential expression of decorin in skin and wound healing.

View Article: PubMed Central - PubMed

Affiliation: Division of Plastic Surgery, Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada; Wound Healing Research Group, Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
Hypertrophic scarring is a frequent fibroproliferative complication following deep dermal burns leading to impaired function and lifelong disfigurement. Decorin reduces fibrosis and induces regeneration in many tissues, and is significantly downregulated in hypertrophic scar and normal deep dermal fibroblasts. It was hypothesized that microRNAs in these fibroblasts downregulate decorin and blocking them would increase decorin and may prevent hypertrophic scarring. Lower decorin levels were found in hypertrophic scar as compared to normal skin, and in deep as compared to superficial dermis. A decorin 3' un-translated region reporter assay demonstrated microRNA decreased decorin in deep dermal fibroblasts, and microRNA screening predicted miR- 24, 181b, 421, 526b, or 543 as candidates. After finding increased levels of mir-181b in deep dermal fibroblasts, it was demonstrated that TGF-β1 stimulation decreased miR-24 but increased miR-181b and that hypertrophic scar and deep dermis contained increased levels of miR-181b. By blocking miR-181b with an antagomiR, it was possible to increase decorin protein expression in dermal fibroblasts. This suggests miR-181b is involved in the differential expression of decorin in skin and wound healing. Furthermore, blocking miR-181b reversed TGF-β1 induced decorin downregulation and myofibroblast differentiation in hypertrophic scar fibroblasts, suggesting a potential therapy for hypertrophic scar.

No MeSH data available.


Related in: MedlinePlus