Asperyellone prevents HDF cells from UVB irradiation damages: An elaborated study

Abstract

The current study explores the photo‐protective effect of asperyellone (AY) (a fungal secondary metabolite), assessed under in vitro condition using human dermal fibroblast cell line. AY was isolated from Aspergillus sp. during the resting phase and purified. The initial cytocompatibility assessment on concentrations of AY and the duration of exposure of UVB irradiations were studied respectively. N‐acetyl‐L‐cysteine (NAC) was used as positive control. Cells were then pretreated with optimized concentration of AY (2.0 μM) and NAC (1 mM) for 1 hour and then UVB irradiated (30 mJ/cm 2) for the period of 10 minutes. Results revealed that reactive oxygen species generated upon UVB irradiation found scavenged by the AY pretreatment at a significant level. Furthermore, an appreciable reduction in apoptotic cell count and DNA damages support the scavenging effect of AY. Assessments on the expression of enzymatic and nonenzymatic antioxidants evidently prove the protective role of AY. The reduced expression levels of inflammatory markers (TNF‐α and COX‐2), collagen degraders (MMP 2 and MMP 9), apoptotic protein expressions (Bax and Bcl‐2), and cell‐cycle arrest analyses substantiate the photo‐protective effect of AY similar to NAC (positive control). Thus, the observations made in the current study indicate the possible role of AY as a photo‐protective agent.

Asperyellone pretreatment protects HaCaT cells from UVB irradiation induced oxidative damages: Assessment under in vitro and in vivo conditions and at molecular level

“…After the observations on apoptosis in HaCaT cells upon UVB irradiation and the reduction in apoptotic cells upon pretreatment with AY, further experiments were conducted to assess the changes in cellular integrity by the comet assay. The results were analyzed as described by Iswarya et al Figure B showed that in neat control and AY pretreatment followed by UVB‐irradiated cells, DNA was not fragmented and showed intact nucleoids, whereas, in UVB‐irradiated cells, increased numbers of tail DNA were observed, which implies that DNA damage was initiated by UVB exposure. Figure C shows the changes in the level of DNA damages like % tail DNA, tail length, tail moment, and Olive tail moment upon UVB treatment and the effect of AY pretreatment, and the results are expressed as % DNA damage.…”

“…Solarmeter, model 6.2 (Vector Technologies, Hyderabad, India), was used for UVB intensity measurement. Before the assessment of the UVB protective role of AY and to fix the concentration of AY, HaCaT cell viability with respect to AY at varied concentration (0.1‐100 µM) was evaluated using 3‐(4,5‐Dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2H‐tetrazolium bromide (MTT) according to the procedure described . The stock solution of 1 mg/mL of AY was prepared using DMSO and used for the study.…”

“…The cells were then pretreated with an optimized concentration of AY for 1 hour and then subjected to UVB irradiation (25 mJ/cm 2 ) for 10 minutes with a minimal amount of phosphate buffered saline. After UVB radiation, the cells were then stabilized with fresh medium (containing fetal bovine serum) for a period of 1 hour and then subjected to the determination of intracellular reactive oxygen species (ROS) levels (measured as fluorescence intensity in the presence of H2DCFDA dye at 1 μg mL −1 ), mitochondrial membrane potential (using Rhodamine 123 dye at 10 μg mL −1 ), cellular integrity (alkaline single cell gel electrophoresis (comet assay), analysis of apoptotic proteins (immunohistochemical analyses), and expression of various antioxidant enzyme levels in all the experimental groups according to the previously described procedures . Furthermore, the expression level of various inflammatory and apoptotic markers (both in vitro and in vivo) were determined using a procedure described earlier …”

“…22 Our previous study on the Asperyellone (AY) pigment and the UVB protection efficacy in HDF implied a significant protective effect of AY. 23 Because the epidermal layer of the skin is dominated by keratinocytes cells, which immediately cascade the response after excess UVB exposure, in vitro studies using keratinocytes may provide better results on their protective role. Hence, the present study has been conducted using HaCaT, and the protective role has been assessed under in vitro and in vivo studies.…”