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Effects of cold atmospheric plasma (CAP) on ß-defensins, inflammatory cytokines, and apoptosis-related molecules in keratinocytes in vitro and in vivo.

Arndt S, Landthaler M, Zimmermann JL, Unger P, Wacker E, Shimizu T, Li YF, Morfill GE, Bosserhoff AK, Karrer S - PLoS ONE (2015)

Bottom Line: In summary, our results suggest that a 2 min application of CAP induces gene expression of key regulators important for inflammation and wound healing without causing proliferation, migration or cell death in keratinocytes.The induction of ß-defensins in keratinocytes describes an absolutely new plasma strategy.Activation of antimicrobial peptides supports the well-known antibacterial effect of CAP treatment, whereas the mechanism of ß-defensin activation by CAP is not investigated so far.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology, University Regensburg, D-93042 Regensburg, Germany.

ABSTRACT
Cold atmospheric plasma (CAP) has been gaining increasing interest as a new approach for the treatment of skin diseases or wounds. Although this approach has demonstrated promising antibacterial activity, its exact mechanism of action remains unclear. This study explored in vitro and in vivo whether CAP influences gene expression and molecular mechanisms in keratinocytes. Our results revealed that a 2 min CAP treatment using the MicroPlaSter ß in analogy to the performed clinical studies for wound treatment induces expression of IL-8, TGF-ß1, and TGF-ß2. In vitro and in vivo assays indicated that keratinocyte proliferation, migration, and apoptotic mechanisms were not affected by the CAP treatment under the applied conditions. Further, we observed that antimicrobial peptides of the ß-defensin family are upregulated after CAP treatment. In summary, our results suggest that a 2 min application of CAP induces gene expression of key regulators important for inflammation and wound healing without causing proliferation, migration or cell death in keratinocytes. The induction of ß-defensins in keratinocytes describes an absolutely new plasma strategy. Activation of antimicrobial peptides supports the well-known antibacterial effect of CAP treatment, whereas the mechanism of ß-defensin activation by CAP is not investigated so far.

No MeSH data available.


Related in: MedlinePlus

Human and murine ß-defensin expression after the CAP treatment.(a, b, c) mRNA expression of human ß-defensin-1 (HBD-1), ß-defensin-2 (HBD-2), and-3 (HBD-3) was analyzed 6 h, 24 h, 48 h, and 72 h in keratinocytes after the CAP treatment for 2 min. (d, e) ELISAs present the relative level of human BD-1 (HBD-1) and human BD-2 (HBD-2) secretion by keratinocytes after the CAP treatment for 2 min compared to control (ctr.). (f) mRNA expression of murine ß-defensins, MBD-1, MBD-2, and MBD-3 in epidermal skin tissue after 5x CAP treatment for 2 min compared to placebo control. *p < 0.05. (g) Immunohistochemistry against murine BD-2 (MBD-2) in murine tissue after 5x CAP treatment versus placebo control. Arrowheads exemplarily indicate MBD-2 positive keratinocytes, whereas a more intensive staining was observed in the CAP treated skin.
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pone.0120041.g004: Human and murine ß-defensin expression after the CAP treatment.(a, b, c) mRNA expression of human ß-defensin-1 (HBD-1), ß-defensin-2 (HBD-2), and-3 (HBD-3) was analyzed 6 h, 24 h, 48 h, and 72 h in keratinocytes after the CAP treatment for 2 min. (d, e) ELISAs present the relative level of human BD-1 (HBD-1) and human BD-2 (HBD-2) secretion by keratinocytes after the CAP treatment for 2 min compared to control (ctr.). (f) mRNA expression of murine ß-defensins, MBD-1, MBD-2, and MBD-3 in epidermal skin tissue after 5x CAP treatment for 2 min compared to placebo control. *p < 0.05. (g) Immunohistochemistry against murine BD-2 (MBD-2) in murine tissue after 5x CAP treatment versus placebo control. Arrowheads exemplarily indicate MBD-2 positive keratinocytes, whereas a more intensive staining was observed in the CAP treated skin.

Mentions: We analyzed the expression of human ß-defensin, HBD-1, HBD-2, and HBD-3, and observed that HBD-2 is significantly induced 6 h and 24 h after the CAP treatment for 2 min (Fig. 4b), whereas HBD-1 and HBD-3 mRNA expression was induced but not significantly by the CAP treatment (Fig. 4a, 4c). A significant induction of HBD-2 was also obtained on protein level 24 h after CAP treatment (Fig. 4e). The protein amount of HBD-1 was not significantly modified after CAP treatment for 2 min (Fig. 4d) and the sensitivity of the BD-3 ELISA was not sufficient to detect HBD-3 protein in the supernatant of both CAP and control treated keratinocytes (data no shown). These results let us speculate that under used treatment conditions HBD-2 is the most important human ß-defensin affected by the CAP treatment. Furthermore, also murine ß-defensins were analyzed in the epidermal skin after 5x CAP treatment for 2 min on mRNA level (Fig. 4f). MBD-2 and MBD-3 were significantly induced after the CAP treatment, whereas MBD-1 was not significantly induced. Immunohistochemistry confirmed an increased expression of murine MBD-2 in epidermal cells after the CAP treatment (Fig. 4g, arrowhead).


Effects of cold atmospheric plasma (CAP) on ß-defensins, inflammatory cytokines, and apoptosis-related molecules in keratinocytes in vitro and in vivo.

Arndt S, Landthaler M, Zimmermann JL, Unger P, Wacker E, Shimizu T, Li YF, Morfill GE, Bosserhoff AK, Karrer S - PLoS ONE (2015)

Human and murine ß-defensin expression after the CAP treatment.(a, b, c) mRNA expression of human ß-defensin-1 (HBD-1), ß-defensin-2 (HBD-2), and-3 (HBD-3) was analyzed 6 h, 24 h, 48 h, and 72 h in keratinocytes after the CAP treatment for 2 min. (d, e) ELISAs present the relative level of human BD-1 (HBD-1) and human BD-2 (HBD-2) secretion by keratinocytes after the CAP treatment for 2 min compared to control (ctr.). (f) mRNA expression of murine ß-defensins, MBD-1, MBD-2, and MBD-3 in epidermal skin tissue after 5x CAP treatment for 2 min compared to placebo control. *p < 0.05. (g) Immunohistochemistry against murine BD-2 (MBD-2) in murine tissue after 5x CAP treatment versus placebo control. Arrowheads exemplarily indicate MBD-2 positive keratinocytes, whereas a more intensive staining was observed in the CAP treated skin.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4359157&req=5

pone.0120041.g004: Human and murine ß-defensin expression after the CAP treatment.(a, b, c) mRNA expression of human ß-defensin-1 (HBD-1), ß-defensin-2 (HBD-2), and-3 (HBD-3) was analyzed 6 h, 24 h, 48 h, and 72 h in keratinocytes after the CAP treatment for 2 min. (d, e) ELISAs present the relative level of human BD-1 (HBD-1) and human BD-2 (HBD-2) secretion by keratinocytes after the CAP treatment for 2 min compared to control (ctr.). (f) mRNA expression of murine ß-defensins, MBD-1, MBD-2, and MBD-3 in epidermal skin tissue after 5x CAP treatment for 2 min compared to placebo control. *p < 0.05. (g) Immunohistochemistry against murine BD-2 (MBD-2) in murine tissue after 5x CAP treatment versus placebo control. Arrowheads exemplarily indicate MBD-2 positive keratinocytes, whereas a more intensive staining was observed in the CAP treated skin.
Mentions: We analyzed the expression of human ß-defensin, HBD-1, HBD-2, and HBD-3, and observed that HBD-2 is significantly induced 6 h and 24 h after the CAP treatment for 2 min (Fig. 4b), whereas HBD-1 and HBD-3 mRNA expression was induced but not significantly by the CAP treatment (Fig. 4a, 4c). A significant induction of HBD-2 was also obtained on protein level 24 h after CAP treatment (Fig. 4e). The protein amount of HBD-1 was not significantly modified after CAP treatment for 2 min (Fig. 4d) and the sensitivity of the BD-3 ELISA was not sufficient to detect HBD-3 protein in the supernatant of both CAP and control treated keratinocytes (data no shown). These results let us speculate that under used treatment conditions HBD-2 is the most important human ß-defensin affected by the CAP treatment. Furthermore, also murine ß-defensins were analyzed in the epidermal skin after 5x CAP treatment for 2 min on mRNA level (Fig. 4f). MBD-2 and MBD-3 were significantly induced after the CAP treatment, whereas MBD-1 was not significantly induced. Immunohistochemistry confirmed an increased expression of murine MBD-2 in epidermal cells after the CAP treatment (Fig. 4g, arrowhead).

Bottom Line: In summary, our results suggest that a 2 min application of CAP induces gene expression of key regulators important for inflammation and wound healing without causing proliferation, migration or cell death in keratinocytes.The induction of ß-defensins in keratinocytes describes an absolutely new plasma strategy.Activation of antimicrobial peptides supports the well-known antibacterial effect of CAP treatment, whereas the mechanism of ß-defensin activation by CAP is not investigated so far.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology, University Regensburg, D-93042 Regensburg, Germany.

ABSTRACT
Cold atmospheric plasma (CAP) has been gaining increasing interest as a new approach for the treatment of skin diseases or wounds. Although this approach has demonstrated promising antibacterial activity, its exact mechanism of action remains unclear. This study explored in vitro and in vivo whether CAP influences gene expression and molecular mechanisms in keratinocytes. Our results revealed that a 2 min CAP treatment using the MicroPlaSter ß in analogy to the performed clinical studies for wound treatment induces expression of IL-8, TGF-ß1, and TGF-ß2. In vitro and in vivo assays indicated that keratinocyte proliferation, migration, and apoptotic mechanisms were not affected by the CAP treatment under the applied conditions. Further, we observed that antimicrobial peptides of the ß-defensin family are upregulated after CAP treatment. In summary, our results suggest that a 2 min application of CAP induces gene expression of key regulators important for inflammation and wound healing without causing proliferation, migration or cell death in keratinocytes. The induction of ß-defensins in keratinocytes describes an absolutely new plasma strategy. Activation of antimicrobial peptides supports the well-known antibacterial effect of CAP treatment, whereas the mechanism of ß-defensin activation by CAP is not investigated so far.

No MeSH data available.


Related in: MedlinePlus