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Justicidin A‐induced autophagy inhibits apoptosis of human hepatocellular carcinoma Hep 3B cells (644.7)

Justicidin A (JA) is a new and pure arylnaphthalide lignan isolated from Justicia procumbens, which has been used as an herbal remedy in Taiwan and China for fever, cancer, and pain. Our previous report shows that JA induced apoptosis of human hepatocellular carcinoma (HCC) Hep 3B cells, leading to the suppression of tumor cell growth in NOD‐SCID mice. Here we reveal that treatment of JA not only increased the expression of autophagic marker microtubule‐associated protein 1 light chain‐II (LC3‐II) by immunoblotting, but also augmented the formation of acidic vesicular organelles by flow cytometry. Administration of autophagy inhibitor bafilomycin A1 (BAF) increased protein expression of LC3‐II and lysosomal marker LAMP2a and decreased colocalization of LC3 and LAMP2a puncta by confocal microscopy, confirming that JA not only induced autophagy but also completed the autophagic flux. JA‐induced autophagy was observed via activation of Raf/MEK/ERK pathway, and was not associated with inhibition of class I PI3K/Akt/mTOR or induction of class III PI3K/Beclin 1 pathway. The novelty of JA is that pre‐treatment of cells with BAF elevated the percentages of cells at sub‐G1 phase and raised the expression of cleaved caspase 3. All together, our data suggest the use of autophagy inhibitor should be taken into consideration to enhance the chemotherapeutic potential of JA for treatment of human HCC.