The Protective Effect of Gallic Acid on UVB Induced Photodamage in Human Immortalized Keratinocytes by Regulating the TLR4/NF-κB Pathway

This paper was to investigate the protective effect of gallic acid on UVB (ultraviolet B) induced photodamage in human immortalized keratinocytes by regulating the TLR4 (Toll-like receptor 4)/NF-κB (nuclear factor kappa B) pathway. HaCaT cells were grouped into blank group (untreated), UVB group (exposed only to UVB), gallic acid in different concentrations (5, 10 and 20 μg/mL) groups, and gallic acid (20 μg/mL)+lipopolysaccharide (10 μg/mL) groups. Except for the blank group, all other cells were exposed to UVB after corresponding drug treatment. MTT method was used to detect cell viability. Kit method was used to detect the activity of SOD (superoxide dismutase) and the levels of MDA (malondialdehyde), TNF-α (tumor necrosis factor-α), and IL-6 (interleukin-6), respectively. TUNEL staining method was used to detect cell apoptosis. DCFH-DA probe and chromogenic substrate were used to detect ROS (reactive oxygen species), caspase (cysteinyl aspartate specific proteinase)-3, and caspase-9 activities, respectively. Immunoblotting method was used to detect the protein expression of TLR4, NF-κB, iNOS (inducible nitric oxide synthase), and COX-2 (cyclooxygenase-2). The mice were divided into blank, UVB, 50 mg/kg gallic acid and gallic acid+lipopolysaccharide groups, and UVB induced photodamage in the skin of mice in the UVB, 50 mg/kg gallic acid and gallic acid+lipopolysaccharide groups, which were treated with 50 mg/kg gallic acid. Compared with the blank group, the HaCaT cell survival rate and SOD activity in the UVB group were lower (P<0.05), and the ROS fluorescence intensity, MDA, inflammatory factor (TNF-α and IL-6) levels, apoptosis index (TUNEL positive cell rate, caspase-3 and caspase-9 activities), TLR4, p-NF-κB, iNOS and COX-2 protein levels were higher (P<0.05). Compared with the UVB group, the HaCaT cell survival rate and SOD activity were higher in the gallic acid different concentration groups (P<0.05), and the ROS fluorescence intensity, MDA, inflammatory factor levels, apoptosis index, TLR4, p-NF-κB, iNOS and COX-2 protein levels were lower (P<0.05). Compared with the 20 μg/mL gallic acid group, the HaCaT cell survival rate and SOD activity in the gallic acid+lipopolysaccharide group were lower (P<0.05), and the ROS fluorescence intensity, MDA, inflammatory factor levels, apoptosis index, TLR4, p-NF-κB, iNOS and COX-2 protein levels were higher (P<0.05). The skin condition of mice in the 50 mg/kg gallic acid group was better than that of the UVB group; the skin of mice in the gallic acid+lipopolysaccharide group was more severely damaged than that of the 50 mg/kg gallic acid group. Gallic acid alleviates UVB induced photodamage in human immortalized keratinocytes by inhibiting the TLR4/NF-κB pathway

CSTR: 32200.14.cjcb.2025.02.0004