J Korean Med Sci.  2005 Aug;20(4):649-654. 10.3346/jkms.2005.20.4.649.

Expression and Modulation of LL-37 in Normal Human Keratinocytes, HaCaT cells, and Inflammatory Skin Diseases

• Kim JE
• Kim BJ
• Jeong MS
• Seo SJ
• Kim MN
• Hong CK
• Ro BI

• KMID: 1712748
• DOI: http://doi.org/10.3346/jkms.2005.20.4.649

Abstract

Defensins and cathelicidins (LL-37) are major antimicrobial peptides (AMPs) of the innate immune system of the human skin. In normal non-inflamed skin these peptides are negligible, but their expression can be markedly increased in inflammatory skin disease such as psoriasis. We designed this study to identify the expressions of LL-37 in normal human keratinocyte (NHK) and HaCaT cells after exposure to stimulants and to investigate difference of LL-37 expression accompanied with cell differentiation status, and come to understand difference of susceptibility to infection in atopic dermatitis and psoriasis. Expressions of LL-37 in NHKs and HaCaT cells were evaluated by using RT-PCR, Western blotting, and immunohistochemical (IHC) staining at 6, 12, and 24 hr post stimulation after exposure to Ultraviolet B irradiation and lipopolysaccharide. And expression of LL-37 in skin biopsy specimens from patients with atopic dermatitis and psoriasis was determined by immunohistochemical analysis. In time-sequential analyses of LL-37 expression revealed that LL-37 was expressed in NHKs, but not in HaCaT cells. IHC analysis confirmed the presence of abundant LL-37 in the epidermis of psoriasis. Therefore we deduced that expression of LL-37 is affected by UV irradiation, bacterial infection, and status of cell differentiation.

Keyword

CAP18 lipopolysaccharide-binding protein; LL-37; Keratinocytes; HaCaT Cell; Psoriasis; Dermatitis; Atopic

MeSH Terms

Antimicrobial Cationic Peptides/analysis/*genetics
Blotting, Western
Cell Line
Cells, Cultured
Comparative Study
Defensins/analysis/genetics
Dose-Response Relationship, Drug
Gene Expression/drug effects/radiation effects
Humans
Immunohistochemistry
Keratinocytes/cytology/*metabolism
Lipopolysaccharides/pharmacology
Male
RNA, Messenger/genetics/metabolism
Research Support, Non-U.S. Gov't
Reverse Transcriptase Polymerase Chain Reaction
Skin/cytology/metabolism
Skin Diseases/*genetics/metabolism/pathology

Figure

• 

  Fig. 1 RT-PCR for LL-37 mRNA from normal human keratinocytes (NHK) at 6 hr, 12 hr, and 24 hr after stimulation. CON, control; UVB 20, UVB 20 mJ/cm2; LPS 25, LPS (2.5 µg/mL); LPS 50, LPS (5.0 µg/mL). *Statistically significant compared to control group (p<0.001).

• 

  Fig. 2 Expression of LL-37 protein in NHKs by Western blotting study using polyclonal antibody to LL-37 at 6 hr, 12 hr, and 24 hr after stimulation. CON, control; UVB 20, UVB 20 mJ/cm2; LPS 25, LPS (2.5 µg/mL); LPS 50, LPS (5.0 µg/mL).

• 

  Fig. 3 Immunohistochemical study using polyclonal antibody to LL-37 in NHKs at 12 hr after stimulation. CON, control; UVB 20, UVB 20 mJ/cm2; LPS 25, LPS (2.5 µg/mL); LPS 50, LPS (5.0 µg/mL) (×200).

• 

  Fig. 4 Immunostaining for LL-37 in frozen skin sections from patients with psoriasis and atopic dermatitis. The arrow indicate immune reactivity to LL-37 (×200).

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