The effect of amniotic membrane extract on the expression of iNOS mRNA and generation of NO in HaCaT cell by ultraviolet B irradiation.
OBJECTIVE
Amniotic membrane (AM) is the innermost fetal membrane, which contains several proteinase inhibitors and expresses several growth factors. Nitric oxide (NO) has been implicated in the pathogenesis of various inflammatory diseases including sunburn and ultraviolet induced erythema. The expression of inducible nitric oxide synthase (iNOS) is up regulated by UVB irradiation and inhibited by TGF-beta and EGF-beta. We evaluated the effect of AM extract on the expression of iNOS mRNA by UV irradiation in HaCaT cell (immortalized human keratinocyte cell line).


METHODS
HaCaT cells were irradiated UVB 30 mJ/cm2 and AM extract was added. The iNOS mRNA was isolated by RT-PCR and NO production was assessed by spectrophotometric method based on Griess reaction.


RESULTS
The expression of iNOS mRNA was induced by UVB irradiation in HaCaT cell and the expression of iNOS mRNA was higher at 48 h than that at 24 h. AM extract down regulated the induction of iNOS mRNA in HaCaT cell by UVB irradiation. NO generation was increased by UVB irradiation, but down regulated by AM extract treatment in HaCaT cells.


CONCLUSIONS
These results assured that the expression of iNOS mRNA and generation of NO are up regulated by UVB irradiation and showed that AM extract down regulated the induction of iNOS mRNA and decreased generation of NO by UVB irradiation.
Chang
D S
DS
Department of Dermatology, Yongsan Hospital, Chung Ang University School of Medicine, Seoul, Korea.
Seo
S J
SJ
Hong
C K
CK
D000650:Amnion; D002470:Cell Survival / Q000528:radiation effects; D002478:Cells, Cultured / Q000378:metabolism / Q000528:radiation effects; D017931:DNA Primers; D005786:Gene Expression Regulation / Q000528:radiation effects; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000528:radiation effects*; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014466:Ultraviolet Rays
D017931:DNA Primers; D012333:RNA, Messenger; D009569:Nitric Oxide; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II