Cytotoxic activity, induction of apoptosis and p53 gene expression of Petrosia cf nigricans Methan fraction to HeLa cell line

Sponge has been known as marine invertebrate containing bioactive compounds. These compounds have been explored as antitumor agents. This research was aimed to investigate the cytotoxic activity, induction of apoptosis and p53 gene expression of Petrosia cf. nigricans extract and its fractions to HeLa cell line. Pre activity assay was conducted using Brine Shrimp Lethality Test (BSLT) and the cytotoxic assay was performed using MTT ([3 (4.5 dimethylthiazol 2yl) 2.5 diphenyltetrazolium bromide]) assay The induction of apoptosis assay was performed by double staining and DNA fragmentation using gel electrophoresis. PCR analysis was performed using p53 primer to detect the expression of p53 gene. The BSLT showed that P. cf. nigricans crude extract was considered very active with LC sub(50) value of 23.4 mu g/mL. Cytotoxicity assay indicated that methanol fraction showed a good cytotoxic activity to HeLa cell with LC sub(50) value of 11. 9 mu g/mL. The result of apoptosis assay indicated that the extract could induce the apoptosis. PCR analysis showed positive expression of p53 gene.Original Abstract: Spons merupakan hewan invertebrata laut yang kaya akan kandungan senyawa bioaktif. Senyawa bioaktif dari spons banyak dieksplorasi sebagai bahan obat antitumor. Penelitian ini bertujuan untuk mengetahui aktivitas sitotoksik, induksi apoptosis dan ekspresi gen p53 fraksi metanol spons Petrosia cf. nigricans terhadap sel tumor HeLa. Uji aktivitas pendahuluan dan sitotoksik masing masing dilakukan dengan metode Brine Shrimp Lethality Test (BSLT) dan MTT ([3 (4.5 dimethylthiazol 2yl) 2.5 diphenyltetrazolium bromide]) assay. Induksi apoptosis dilakukan dengan metode pengecatan menggunakan akridin oranye dan etidium bromida. Analisis PCR menggunakan primer p53 dilakukan untuk mengetahui ekspresi gen p53. Hasil uji BSLT memperlihatkan bahwa ekstrak kasar P. cf nigricans memiliki aktivitas tahap awal yang sangat baik dengan LC sub(50) = 23,4 mu g/mL. Hasil uji MTT menunjukkan bahwa fraksi metanol memiliki aktivitas sitotoksik terhadap sel HeLa dengan nilai LC sub(50) sebesar 11,9 mu g/mL. Berdasarkan uji induksi apoptosis metode pengecatan dapat dinyatakan bahwa fraksi metanol mampu menginduksi peristiwa apoptosis pada sel HeLa. Fraksi metanol juga mampu meningkatkan ekspresi gen p53.