Tesis

Efek Alfa Mangostin Terhadap Proliferasi dan Aktivasi Sel Stelata Hati yang Diinduksi Asetaldehid Melalui Jalur TGF-β dan ERK 1/2 = The Effect of Alfa Mangosteen to the Activation and Proliferation of Acetaldehyde-Induced Hepatic Stellate Cell through TGF-β and ERK 1/2 Signalling Pathways .

Latar belakang : Fibrosis hati ditandai dengan penimbunan berlebihan matriks ekstraseluler pada cedera hati kronik. HSC memegang peranan sentral dalam proses fibrosis hati. HSC yang teraktivasi merupakan sumber miofibroblas yang berkontribusi terhadap fibrogenesis. Asetaldehid memiliki efek langsung terhadap HSC karena meningkatkan sintesis TGF-β, sitokin profibrogenik utama yang berperan dalam transformasi HSC menjadi aktif. Asetaldehid juga mengaktivasi PKC dan menghasilkan ROS yang selanjutnya mengaktifkan transduksi sinyal ERK1/2. Saat ini belum ada terapi standar fibrosis hati. Alfa mangostin diketahui memiliki aktivitas antiproliferatif dan antioksidan secara in vivo. Penelitian ini menggunakan alfa mangostin untuk mengetahui aktivitasnya pada jalur TGF-β dan ERK1/2 dengan sorafenib sebagai kontrol positif. Metode : Penelitian menggunakan sel HSC LX-2. Sel dibagi dalam 6 kelompok yaitu kelompok normal, asetaldehid, asetaldehid + sorafenib10μM, asetaldehid + alfa mangostin10μM, asetaldehid + alfa mangostin20μM, dan alfa mangostin10μM. Sel dipanen setelah induksi obat selama 24 jam. Proliferasi sel dihitung menggunakan tryphan blue exclusion method. Ekspresi Ki-67, TGF-β, dan TGFβR diukur dengan qRT-PCR. Ekspresi α-SMA dan pERK menggunakan WesternBlot. Kadar TGF-β medium diukur menggunakan ELISA. Kadar ROS intraseluler dengan spektrofotometri. Hasil penelitian : Asetaldehid meningkatkan proliferasi sel dan ekspresi marker fibrogenik pada HSC. Pemberian sorafenib dan alfa mangostin menurunkan viabilitas sel, ekspresi Ki-67 dan pERK. Penurunan tersebut juga diikuti dengan menurunnya ekspresi TGF-β, TGF-βR, and α-SMA, dan penurunan kadar TGF-β dalam medium dan ROS intraseluler. Pada kelompok yang hanya diberikan alfa mangostin, terdapat penurunan viabilitas sel namun penurunan ekspresi biomarker belum terlihat jelas dibandingkan kelompok normal. Kesimpulan : Alfa mangostin menghambat proliferasi dan aktivasi pada HSC yang diinduksi asetaldehid pada jalur TGF-β dan ERK1/2.
Kata kunci : asetaldehid, alfa mangostin, hepatic stellate cell, TGF-β, ERK1/2


Background: Liver fibrosis is characterized by excessive accumulation of extracellular matrix in chronic liver injury. HSC plays a central role in the process of liver fibrosis. Activated HSC is considered as a source of miofibroblasts that contribute to fibrogenesis. Acetaldehyde has a direct effect on HSC because it increases the synthesis of TGF-β, the major profibrogenic cytokine that plays a role in HSC transformation into active. Acetaldehyde also activates PKC and produces ROS which further activates signal transduction pathway ERK1/2. There is currently no standard therapy of liver fibrosis. Alpha mangostin is known to have antiproliferative and antioxidant activity in vivo. In this research, we use alpha mangostin to know its activity on TGF-β and ERK1/2 pathways with sorafenib as positive control. Method: This study using HSC LX-2 cell line. Cells were divided into 6 groups: normal group, acetaldehyde group, acetaldehyde + sorafenib10μM group, acetaldehyde + alfa mangostin 10μM group, acetaldehyde + alfa mangostin 20μM group, and alfa mangostin 10μM group. Cells were harvested after 24 hours of drug induction. Cell proliferation was counted using tryphan blue exclusion method. Expression of Ki-67, TGF-β, and TGF-βR were measured by qRT-PCR. Expression of α-SMA and PERK were evaluated using Western-Blot. TGF-β level in culture medium was measured by ELISA. Intracellular ROS level was analyzed using spectrophotometry. Result: Acetaldehyde increased proliferation and expression of fibrogenic markers on HSC. Sorafenib and alpha mangostin treatments resulted in a reduced cell viability, decreased Ki-67 and pERK 1/2 expressions. These findings were followed with decreased expressions of TGF-β, TGF-βR, and α-SMA, decrease concentration of TGF-β in medium culture and intracelluler ROS. In the group given only alpha mangostin, there was a decrease in cell viability but decreased biomarker expression was not clearly visible compared to the normal group. Conclusion: Alpha mangostin inhibits proliferation and activation acetaldehydeinduced HSC through TGF-β and ERK 1/2 pathways.
Keywords: acetaldehyde, alpha mangostin, hepatic stellate cell, TGF-β, ERK1/2