Ann Dermatol.  2010 Aug;22(3):290-299. 10.5021/ad.2010.22.3.290.

Epidermal Hyperplasia and Elevated HB-EGF are More Prominent in Retinoid Dermatitis Compared with Irritant Contact Dermatitis Induced by Benzalkonium Chloride

• Lee JE
• Chang JY
• Lee SE
• Kim MY
• Lee JS
• Lee MG
• Kim SC

• KMID: 2265362
• DOI: http://doi.org/10.5021/ad.2010.22.3.290

Abstract

BACKGROUND
'Retinoid dermatitis' is a retinoid-induced irritant contact dermatitis (ICD). The mechanism of retinoid dermatitis may be different from that of other ICDs. However, it remains uncertain how topical retinoid induce ICD.
OBJECTIVE
We compared several aspects of contact dermatitis induced by topical retinol and benzalkonium chloride (BKC) on hairless mice skin.
METHODS
2% retinol or 2.5% BKC was applied to hairless mice and transepidermal water loss (TEWL), ear thickness, histologic and immunohistochemical findings were compared. We also compared mRNA expression of inflammatory cytokines, epidermal differential markers, cyclooxygenases (COXs) and heparin binding epidermal growth factor like growth factor (HB-EGF).
RESULTS
Topical application of 2% retinol and 2.5% BKC increased TEWL and ear thickness in similar intensity. Epidermal hyperplasia was more prominent in retinol treated skin. Proliferating cell nuclear antigen, involucrin and loricrin expression were higher in retinol-treated skin than in BKC-treated skin. Filaggrin, however, was more expressed in BKC-treated skin. The mRNA expression of IL-8, TNF-alpha, COX-2, involucrin, loricrin and filaggrin were increased in both retinol- and BKC-treated skin in similar intensity. HB-EGF was more significantly increased in retinol-treated skin.
CONCLUSION
Elevated HB-EGF and epidermal hyperplasia are more prominent features of retinoid dermatitis than in BKC-induced ICD.

Keyword

Benzalkonium chloride; HB-EGF; Irritant contact dermatitis; Retinoid

MeSH Terms

Animals
Benzalkonium Compounds
Cytokines
Dermatitis
Dermatitis, Contact
Ear
Epidermal Growth Factor
Heparin
Hyperplasia
Intercellular Signaling Peptides and Proteins
Interleukin-8
Intermediate Filament Proteins
Membrane Proteins
Mice
Mice, Hairless
Proliferating Cell Nuclear Antigen
Prostaglandin-Endoperoxide Synthases
Protein Precursors
RNA, Messenger
Skin
Tumor Necrosis Factor-alpha
Vitamin A
Water Loss, Insensible
Benzalkonium Compounds
Cytokines
Epidermal Growth Factor
Heparin
Intercellular Signaling Peptides and Proteins
Interleukin-8
Intermediate Filament Proteins
Membrane Proteins
Proliferating Cell Nuclear Antigen
Prostaglandin-Endoperoxide Synthases
Protein Precursors
RNA, Messenger
Tumor Necrosis Factor-alpha
Vitamin A
Water Loss, Insensible

Figure

• 

  Fig. 1 Transepidermal water loss (TEWL) measured from the skin topically treated with retinol is significantly elevated than that measured in skin treated with ethanol. TEWL measured from the skin topically treated with benzalkonium chloride (BKC) was also significantly elevated, compared to that measured in skin treated with acetone. However, there was no significant difference between TEWL measured in retinol- and BKC-applied skin. Data are presented as mean±SEM, *p<0.01. Statistical significance was determined using Student's t test, empty box: ethanol-treated skin, shaded box: retinol-treated skin, striped box: acetone-treated skin, filled box: BKC-treated skin.

• 

  Fig. 2 Ear thickness is significantly increased in both retinoland benzalkonium chloride (BKC)-treated ears compared to ethanol- and acetone-treated ears, however, there is no significant difference between retinol- and BKC-treated ear. Data are presented as mean±SEM, *p<0.01. Statistical significance was determined using Student's t test, empty box: ethanol-treated skin, shaded box: retinol-treated skin, striped box: acetone-treated skin, filled box: BKC-treated skin.

• 

  Fig. 3 Both retinol and benzalkonium chloride (BKC)-induced epidermal hyperplasia and inflammation. H&E-stained sections of ethanol-applied mouse ear (A), retinol-treated (B), acetone-treated ear (C), BKC-treated ear (D). Both retinol and BKC treatments result in epidermal hyperplasia, increased intercellular edema and dermal inflammatory cell infiltration (B and D). Epidermal hyperplasia and separation of the stratum corneum are more pronounced in retinol-applied ear skin compared to BKC-applied ear skin (H&E stain, ×400).

• 

  Fig. 4 Epidermal hyperplasia was also measured quantitatively by the number of keratinocyte layers in the epidermis. The number of keratinocyte layers is increased in both retinol- and benzalkonium chloride (BKC)-treated ears, compared to ethanoland acetone-treated ears. The number of keratinocyte layers is significantly increased in retinol-treated skin, compared to BKC-treated skin. Data are presented as mean±SEM, *p<0.01. Statistical significance was determined using Student's t test, empty box: ethanol-treated skin, shaded box: retinol-treated skin, striped box: acetone-treated skin, filled box: BKC-treated skin.

• 

  Fig. 5 Immunohistochemical stain - proliferation cell nuclear antigen (PCNA). (A) Normal epidermis, (B) Retinol, (C) Benzalkonium chloride (BKC). PCNA expressions in retinol- or BKC-applied skin are increased than those in normal or vehicle-applied skin. PCNA expression is more pronounced in retinol-applied skin than BKC-applied skin (×400).

• 

  Fig. 6 Immunohistochemical stain - involucrin, loricrin, filaggrin. (A, D, G) Normal epidermis, (B) Involucrin in retinol, (C) Involucrin in benzalkonium chloride (BKC), (E) Loricrin in retinol, (F) Loricrin in BKC, (H) Filaggrin in retinol, (I) Filaggrin in BKC. Involucrin expressions in retinol- or BKC-treated skin are increased. Involucrin expression is slightly pronounced in retinol-treated skin, compared to BKC-treated skin. Loricrin expressions in retinol- or BKC-treated skin are increased. Loricrin expression is markedly pronounced in retinol-treated skin compared to BKC-treated skin Filaggrin expressions in retinol- or BKC-treated skin are increased. Filaggrin expression is slightly pronounced in BKC-treated skin compared to retinol-treated skin (×400).

• 

  Fig. 7 (A) Agarose gel electrophoresis of each cDNA amplified with reverse transcriptase-polymerase chain reaction (RT-PCR) (IL-8, TNF-α, COX-1, COX-2 and HB-EGF). (B) COX-1 levels by RT-PCR between 4 groups. No significant difference in COX-1 level between the 4 groups was observed (p=0.997, ANOVA). (C) IL-8 levels by RT-PCR between the 4 groups. IL-8 in retinol- or benzalkonium chloride (BKC)-applied skin is significantly increased compared to ethanol- or acetone-treated skin, respectively (p=0.012, p=0.006). However, no significant difference in IL-8 level between retinol- and BKC-treated groups was observed (p=0.157). (D) TNF-α levels by RT-PCR between the 4 groups. TNF-α in retinol- or BKC-treated skin is significantly increased compared to ethanol- or acetone-treated skin, respectively (p=0.001, p=0.046). No significant difference in TNF-α level between retinol- and BKC-treated groups was observed (p=0.460). (E) COX-2 levels by RT-PCR between the 4 groups. Unlike COX-1 results, COX-2 in retinol- or BKC-applied skin is significantly increased compared to ethanol- or acetone-treated skin, respectively (p<0.0001, p=0.005). No significant difference in COX-2 level between retinol- and BKC-treated groups was observed (p=0.221). (F) HB-EGF levels by RT-PCR between the four groups. HB-EGF in retinol- or BKC-treated skin is significantly increased compared to ethanol- or acetone-treated skin, respectively (p=0.01, p=0.0017). Moreover, HB-EGF in retinol-applied skin was significantly increased compared to BKC-treated skin (p=0.029). Data are presented as mean±SEM, *p<0.05, †p<0.01. Statistical significance was determined using Student's t-test, empty box: ethanol-treated skin, shaded box: retinol-treated skin, striped box: acetone-treated skin, filled box: BKC-treated skin.

• 

  Fig. 8 (A) Agarose gel electrophoresis of each cDNA amplified with reverse transcriptase-polymerase chain reaction (RT-PCR) (involucrin, loricrin, and filaggrin). (B) Involucrin in retinol- or benzalkonium chloride (BKC)-treated skin is significantly increased compared to ethanol- or acetone-treated skin respectively (p=0.017, p=0.020). However, no significant difference in involucrin level between retinol- and BKC-treated groups was observed. (C) Loricrin in retinol- or BKC-treated skin is significantly increased compared to ethanolor acetone-treated skin, respectively (p=0.025, p=0.0007). However, no significant difference in loricrin level between retinol- and BKC-treated groups was observed. (D) Filaggrin in retinol- or BKC-treated skin is significantly increased compared to ethanol- or acetone-treated skin, respectively (p=0.001, p=0.001). However, no significant difference in filaggrin level between retinol- and BKC-treated groups was observed. Data are presented as mean±SEM, *p<0.05, †p<0.01. Statistical significance was determined using Student's t test, empty box: ethanol-applied skin, shaded box: retinol-applied skin, striped box: acetone-applied skin, filled box: BKC-applied skin.

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