BACKGROUNDAntimicrobial peptides, including cathelicidin LL-37, human beta defensin (HBD)-2, and HBD-3, are important elements of the innate immune response and involved in modulation of the adaptive immunity, and they also play an important role in cutaneous defense against Mycobacterium tuberculosis.

METHODSThe fresh skin tissues and paraffin-embedded biopsy samples from three cutaneous tuberculosis, two tuberculids, and ten healthy individuals were collected. The expressions of LL-37, HBD-2, and HBD-3 mRNA in the lesions of three cutaneous tuberculosis and two tuberculids were detected by quantitative real-time polymerase chain reaction; the protein expressions were detected by immunohistochemistry and Western blotting methods.

RESULTSThe expressions of LL-37 mRNA and protein in the lesions of cutaneous tuberculosis and tuberculids were similar to that of normal skin. The expression of HBD-2 mRNA had an increasing trend in the lesions of cutaneous tuberculosis and tuberculids compared with that of normal skin; however, the expression of HBD-2 protein in the lesions of cutaneous tuberculosis had a decreasing trend compared with that of normal skin, and the expression of HBD-2 protein in the lesions of tuberculids was similar to that of normal skin. The expressions of HBD-3 mRNA and protein in lesions of cutaneous tuberculosis and tuberculids were similar to that of normal skin.

CONCLUSIONSOur study indicated that the expression of HBD-2 and HBD-3 mRNA and protein in lesions of cutaneous tuberculosis may be not consistent with that of tuberculids. However, an inherent limitation of the present study was that the sample size was small, and the roles and regulation mechanisms of LL-37, HBD-2, and HBD-3 in cutaneous tuberculosis and tuberculids need to be further investigated.

Adult ; Aged ; Antimicrobial Cationic Peptides ; genetics ; Female ; Humans ; Male ; Middle Aged ; RNA, Messenger ; analysis ; Tuberculosis, Cutaneous ; metabolism ; beta-Defensins ; genetics

To screen the candidate genes associated with Musca domestica antibacterial peptides using DNA microarray technique, the hybrid probes were designed from the conservative domains of the encoded area of the insect antibacterial peptide genes in GenBank with biology software Designer 2.0, and were synthesized by a chemical process, with the assistance of the automated Gen III Microarray Spotter, those oligo probes were printed on a special ready-made glass, and a cDNA microarray was constructed. The total RNA was extracted from the fat body of Musca domestic third-instar larve induced after 24 hours by Escherichia coli and Staphylococcus aureus, the strands of cDNA were labled with fluoresceine Cy3 using the method of reverse transcription PCR, after prehybridization, hybridization and washing procedure, the results of hybridization were scanned using computer system, and the data were analyzed using the software of MIDAS, fifteen valid hybridization signals were detected through two times of hybridization and scanning (the positive samples as a control were excluded). DNA microarray technique can be successfully applied screen the candidate genes associated with Musca domestica antibacterial peptides, and further provide significant evidence to discover its antibacterial peptide new genes.
Animals ; Antimicrobial Cationic Peptides ; genetics ; Base Sequence ; Gene Expression Profiling ; Genes, Insect ; Houseflies ; genetics ; growth & development ; Larva ; genetics ; growth & development ; Molecular Sequence Data ; Oligonucleotide Array Sequence Analysis ; methods ; Oligonucleotide Probes ; chemistry

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.
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