Anti-Photoaging Effects of Standardized Cirsium setidens (Dunn) Nakai on UVB-Irradiated Hs68 Fibroblasts and Hairless Mice via Modulation of the MAPK/AP-1 Signaling Pathway

다국어 초록 (Multilingual Abstract)

Repetitive exposure to ultraviolet (UV) radiation causes skin damage, leading to inflammation, wrinkles, and aging. Many natural products are being highlighted as anti-aging agents due to their accessibility and minor side effects. Here, we explored the photoprotective effects of a standardized hot water extract from the aerial parts of Cirsium setidens (Dunn) Nakai (CS). In Hs68 cells, CS treatment exhibited cytoprotective effect and reduced inflammation. CS also decreased matrix metalloproteinase (MMP)-1 and -3 expression by inactivation the mitogen-activated protein kinase (MAPK)/activator protein 1 (AP-1) signaling pathway and recovered pro-collagen type I level by activating Smad pathway. CS reduced reactive oxygen species (ROS) production and enhanced antioxidant defense by activating heme oxygenase-1 (HO-1)/nuclear factor erythroid-2-related factor 2 (Nrf2) signaling. In HR-1 mice, oral administration of CS reduced wrinkle formation and epidermal/dorsal thickness, while also restoring hydration. CS inhibited collagen degradation by downregulating MMP-1, -3, and -9 expression via inactivation of the MAPK/AP-1 and stimulation of the transforming growth factor beta (TGF-β)/Smad signaling pathways. Taken together, our data revealed that CS may prevent photoaging by regulating MMPs and collagen synthesis through the MAPK/AP-1 and Smad pathways, respectively.

목차 (Table of Contents)

• Abstract Ⅴ
• 1. Introduction 1
• 2. Material and Methods 3
• 2.1. Materials and reagents 3
• 2.2. CS preparation 4
• 2.3. Determination of qualitative target compound using HPLC 5
• 2.4. Cell culture, UVB exposure, and CS treatment 5
• 2.5. Cell viability assay 5
• 2.6. Quantitative real-time RT-PCR (qRT-PCR) analysis 6
• 2.7. Western blot analysis 7
• 2.8. Measurement of pro-collagen type I production 8
• 2.9. Measurement of intracellular ROS production 8
• 2.10. UVB-irradiated skin photoaging animal experiments 8
• 2.11 Analyze of hepatoxicity and renal toxicity of CS 9
• 2.12. Skin wrinkle formation analysis 9
• 2.13. Histological analysis 10
• 2.14. Assessment of dorsal skin thickness and hydration level 10
• 2.15. Statistical analysis 10
• 3. Results 11
• 3.1. CS standardization through pectolinarin content by HPLC analysis 11
• 3.2. CS attenuates UVB-irradiated photodamage and skin inflammation in Hs68 fibroblasts 12
• 3.3. CS reduces MMPs expression through MAPK/AP-1 signaling pathway and recovers collagen synthesis in UVB-exposed Hs68 fibroblasts 13
• 3.4. CS modulates ROS production via Nrf2/HO-1 signaling pathway in UVB-exposed Hs68 fibroblasts 15
• 3.5. CS does not induce hepatotoxicity and nephrotoxicity in UVB-irradiated HR-1 mice 16
• 3.6. CS alleviates wrinkle formation in the dorsal skin of UVB-irradiated HR-1 mice 17
• 3.7. CS ameliorates skin thickening and dehydration in the dorsal skin of UVB-irradiated HR-1 mice 19
• 3.8. CS prevents collagen degradation in the dorsal skin of UVB-irradiated HR-1 mice 20
• 3.9. CS suppresses MAPK/AP-1 activation and stimulates TGF-β/Smad signaling pathway in the dorsal skin of UVB-irradiated HR-1 mice 23
• 4. Discussion 25
• 5. Conclusion 28
• 6. References 29
• Table & Figure Contents
• Table 1. Information of materials and reagents
• Table 2. Primer sequences for qRT-PCR
• Table 3. Information of antibodies for Western blot analysis
• Figure 1. UVB-irradiated skin photoaging animal model
• Figure 2. High-performance liquid chromatography chromatograms
• Figure 3. Cytoprotective and anti-inflammatory effects of CS in UVB-induced Hs68 cells
• Figure 4. Effects of CS on mRNA expression of MMP-1 and MMP-3, activation of MAPK/AP-1 and Smad 2/3, and production of pro-collagen type I in UVB-induced Hs68 cells
• Figure 5. Effects of CS on ROS production and the Nrf2/HO-1 signaling pathway in UVB-induced Hs68 cells
• Figure 6. Effects of CS on GOT, GPT, and BUN production in HR-1 mice
• Figure 7. Effects of CS on wrinkle formation in UVB-irradiated HR-1 mice
• Figure 8. Inhibitory effects of CS on skin thickening and dehydration in UVB-irradiated HR-1 mice
• Figure 9. Effects of CS on collagen degradation
• Figure 10. Effects of CS on MAPK/AP-1 and TGF-β/Smad signaling pathways in UVB–irradiated HR-1 mice