The effect of amniotic membrane extract on the expression of iNOS mRNA and generation of NO in HaCaT cell by ultraviolet B irradiation

Background/Purpose: 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/cm(2) 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. Conclusion: 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.