Project description:Antrodia cinnamomea (Ac), a traditional medicine and an endemic fungus in Taiwan, has been used in cancer research. Recent research has revealed decreased cell proliferation after treatment of Ac on tumor. In this study, we profiled the 2 hours and 4 hours genome-wide miRNA and mRNA transcriptome by next-generation sequencing techniques to report the early apoptotic effect on Ac fruiting body extract treated human hepatocarcinoma cells, SK-HEP-1, instead of prolonged treatment. Results showed that miRNAs were globally downregulated during the first 2-4 hours in, and solely in, AcFBE-treated SK cells. The inhibition of miRNAs imposed no discrimination against any particular miRNA species, but oncogenic miR-21, miR-191 and two oncogenic clusters miR-17-92 and miR-106b-25 were among the most significantly inhibited miRNAs. In addition to miRNA expression, mRNA transcriptome data indicated the association of apoptosis mechanism with AcFBE treatment. Western blotting indicated a decrease in key proteins Drosha and Dicer required for miRNA biogenesis, and an increase of XRN2 involved in miRNA degradation. Our results suggest that miRNAs appeared to be the prime targets of Ac in disrupting multiple miRNA regulatory pathways and global disruption of miRNA transcriptome resulting in activation of extrinsic and intrinsic (mitochondrial) pathways. Human liver SK-Hep-1 cells with or without Antrodia cinnamomea treatment at 2 hours and 4 hours were sequenced by SOLiD 3 and SOLiD 5500xl to obtain miRNA profiles; mRNA profiles also were profiled by SOLID 3. Mouse liver BNL CL.2 cells with or without Antrodia cinnamomea treatment at 2 hours and 4 hours were sequenced by SOLiD 3 to obtain miRNA profiles.
Project description:Antrodia Camphorata is well known in Taiwan as a folk medicinal fungus with anticancer and anti-inflammatory effects. In this study, we used a human acute myelogenous leukemia cell line, KG-1, as the experimental model and constructed a high-throughput gene screen integrated platform by using a combination of herbal identification, UV-VIS spectrophotometry, high performance liquid chromatography (HPLC), inductively coupled plasma mass spectrometry (ICPMS), and DNA microarray technology. Based on this science-based platform, we developed a practical quantitative method and also established an organic/inorganic chemical fingerprint for the herbal materials. In the mean while, the global gene expression patterns of the KG-1 cells treated with various A. camphorata mycelia extracts (water and methanol extracts) were compared. Furthermore, several possible biological responses and relevant marker gene would be identified by our high-throughput integrated platform. Keywords: KG-1 treated with AC extracts for 4 days, cDNA microarray, biomarker selection.
Project description:Antrodia cinnamomea (Ac), a traditional medicine and an endemic fungus in Taiwan, has been used in cancer research. Recent research has revealed decreased cell proliferation after treatment of Ac on tumor. In this study, we profiled the 2 hours and 4 hours genome-wide miRNA and mRNA transcriptome by next-generation sequencing techniques to report the early apoptotic effect on Ac fruiting body extract treated human hepatocarcinoma cells, SK-HEP-1, instead of prolonged treatment. Results showed that miRNAs were globally downregulated during the first 2-4 hours in, and solely in, AcFBE-treated SK cells. The inhibition of miRNAs imposed no discrimination against any particular miRNA species, but oncogenic miR-21, miR-191 and two oncogenic clusters miR-17-92 and miR-106b-25 were among the most significantly inhibited miRNAs. In addition to miRNA expression, mRNA transcriptome data indicated the association of apoptosis mechanism with AcFBE treatment. Western blotting indicated a decrease in key proteins Drosha and Dicer required for miRNA biogenesis, and an increase of XRN2 involved in miRNA degradation. Our results suggest that miRNAs appeared to be the prime targets of Ac in disrupting multiple miRNA regulatory pathways and global disruption of miRNA transcriptome resulting in activation of extrinsic and intrinsic (mitochondrial) pathways.
Project description:Antrodia Camphorata is well known in Taiwan as a folk medicinal fungus with anticancer and anti-inflammatory effects. In this study, we used a human acute myelogenous leukemia cell line, KG-1, as the experimental model and constructed a high-throughput gene screen integrated platform by using a combination of herbal identification, UV-VIS spectrophotometry, high performance liquid chromatography (HPLC), inductively coupled plasma mass spectrometry (ICPMS), and DNA microarray technology. Based on this science-based platform, we developed a practical quantitative method and also established an organic/inorganic chemical fingerprint for the herbal materials. In the mean while, the global gene expression patterns of the KG-1 cells treated with various A. camphorata mycelia extracts (water and methanol extracts) were compared. Furthermore, several possible biological responses and relevant marker gene would be identified by our high-throughput integrated platform. Keywords: KG-1 treated with AC extracts for 4 days, cDNA microarray, biomarker selection. We used optimal loop design to calculate the gene expression profile. In this design, we used labeled RNA from each sample to labeled three slides for a total of nine arrays. Therefore, the data computations were consisted of 18 measurements (6 samples with 3 replicates) for each of 7334 genes. Besides, duplicated water-treatment samples were used as internal control