Aktivitas Anti-Photoaging Senyawa Analog Kurkumin Tetrahidroheksagamavunon-5 (THHGV-5) Pada Sel Human Dermal Fibroblast Yang Diinduksi UV-B

AISYAH NUR DAMAYANTI, drh. Retno Murwanti, M.P., Ph.D.

2024 | Skripsi | FARMASI

Paparan sinar UV-B jangka panjang akan mengakibatkan photoaging karena UV-B dapat meningkatkan akumulasi ROS dan kerusakan DNA sehingga dibutuhkan agen antioksidan untuk memproteksi kulit dari kerusakan DNA pada sel kulit. Senyawa tetrahidiroheksagamavunon-5 (THHGV-5) merupakan senyawa analog kurkumin yang masih tergolong baru yang disintesis dari senyawa HGV-5. Senyawa ini terbukti memiliki efek antioksidan yang poten dari hasil pengujian dengan DPPH dan reduksi ion feri sehingga diprediksi dapat memberikan proteksi terhadap photoaging yang diakibatkan oleh paparan sinar UVB. Penelitian ini dilakukan untuk menganalisis potensi antiphotoaging senyawa THHGV-5 berdasarkan studi in silico tertarget reseptor kolagenase serta pengaruhnya terhadap akumulasi ROS intraseluler dan jumlah sel senescence pada sel human dermal fibroblast (HDF) yang diinduksi UV-B.  
Analisis molecular docking dilakukan untuk memahami potensi antiphotoaging THHGV-5 melalui afinitas terhadap reseptor kolagenase secara in silico dengan luaran berupa % inhibisi prediktif menggunakan vitamin E sebagai kontrol positif. Senyawa THHGV-5 diuji sitotoksisitasnya secara in vitro menggunakan MTT assay untuk menentukan konsentrasi yang tidak toksik pada sel HDF berdasarkan viabilitas sel. Konsentrasi yang memberikan viabilitas terbaik digunakan untuk analisis ROS intraseluler dan sel senescence melalui Senescence-Associated ?-Galactosidase Assay (SA-?-Gal), analisis data dilakukan menggunakan GraphPad Prism 9 melalui uji statistik Shapiro-Wilk, Levene, dan one-way ANOVA.
Hasil molecular docking menunjukkan bahwa senyawa THHGV-5 mampu berikatan dengan reseptor kolagenase (MMPs) dengan lebih stabil dan poten dibanding vitamin E dengan nilai persen inhibisi prediktif THHGV-5 terhadap fibroblast collagenase sebesar 56,58%. Potensi inhibisi terlihat dari nilai skor docking terkorelasi % inhibisi serta interaksi dengan residu kunci HIS218 dan HIS 228. Selanjutnya untuk hasil penelitian in vitro menunjukkan bahwa senyawa THHGV-5 dalam rentang konsentrasi 6,25 – 100 uM tidak menunjukkan efek toksik terhadap sel HDF. Seri konsentrasi 6,25; 12,5; 25; 50; 100 uM senyawa THHGV-5 dapat mempertahankan viabilitas sel berturut-turut sebesar 98,65±0,34%; 104,7±0,35%; 104,0±1,11%; 103,6±1,58%; dan 102,3±0,23%. Dengan dosis UV-B 1400 mJ/cm2. Konsentrasi 25 uM senyawa THHGV-5 merupakan konsentrasi paling optimal dalam memproteksi terjadinya photoaging akibat sinar UV-B karena dapat mencegah akumulasi ROS intraseluler paling besar dan menurunkan jumlah sel senescence paling banyak secara berturut-turut yaitu 4,37?n 36,60%.

Long-term exposure to UV-B light will result in photoaging because UV-B can increase ROS accumulation and DNA damage so that antioxidant agents are needed to protect the skin from DNA damage in skin cells. The tetrahydrohexagamavunon-5 (THHGV-5) compound is a relatively new curcumin analog compound synthesized from the HGV-5 compound. This compound is proven to have a potent antioxidant effect from the test results with DPPH and ferric ion reduction so that it is predicted to provide protection against photoaging caused by UVB exposure. This study was conducted to analyze the antiphotoaging potential of THHGV-5 compound based on in silico study targeted collagenase receptor and its effect on intracellular ROS accumulation and the number of senescence cells in UV-B-induced human dermal fibroblasts (HDF) cells.  
Molecular docking analysis was carried out to understand the antiphotoaging potential of THHGV-5 through affinity to collagenase receptors in silico with the output in the form of predictive % inhibition using vitamin E as a positive control. THHGV-5 compound was tested for cytotoxicity in vitro using MTT assay to determine the concentration that is not toxic to HDF cells based on cell viability. The concentration that gave the best viability was used for intracellular ROS analysis and cell senescence through Senescence-Associated ?-Galactosidase Assay (SA-?-Gal), data analysis was performed using GraphPad Prism 9 through Shapiro-Wilk, Levene, and one-way ANOVA statistical tests.
The molecular docking results show that THHGV-5 compound is able to bind to collagenase receptors (MMPs) more stably and potently than vitamin E with a predictive inhibition percent value of 56.58%. Potential inhibition is seen from the correlated docking score value of % inhibition and interaction with key residues HIS218 and HIS 228. The results showed that THHGV-5 compounds in the concentration range of 6.25 - 100 uM showed no toxic effects on HDF cells. The concentration series of 6.25; 12.5; 25; 50; 100 uM THHGV-5 compound can maintain cell viability of 98.65±0.34%; 104.7±0.35%; 104.0±1.11%; 103.6±1.58%; and 102.3±0.23%, respectively. With UV-B dose of 1400 mJ/cm2. The concentration of 25 uM THHGV-5 compound is the most optimal concentration in protecting UV-B-induced photoaging because it can prevent the accumulation of intracellular ROS the most and reduce the number of senescence cells the most, namely 4.37% and 36.60%, respectively.

Kata Kunci : photoaging, kurkumin, THHGV-5, sel HDF, UV-B