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Abstract Ethno-pharmacological relevance Numerous studies have demonstrated the potent anticancer activity of various Chinese herbs. Actinidia chinensis Planch root (acRoots), a traditional Chinese medicine, functions as an antitumor and detoxifying agent and plays a role in diuresis and hemostasis. Treatment with acRoots confers strong inhibition of tumor growth in various forms of cancer. Here, we evaluated the anticancer activity and molecular mechanisms of Actinidia chinensis Planch root extract (acRoots) on hepatocellular carcinoma (HCC). Materials and methods Our previous study used mRNA chip analyses to identify the genes regulated by acRoots. Further analyses of the altered genes identified a key regulator of genes in response to acRoots. Here, the effects of acRoots on HCC cell proliferation, migration, invasion, and apoptosis were evaluated by cell counting, Transwell and apoptosis assays. In addition, the in vivo anti-HCC effects of acRoots were investigated using an HCC animal model. The expression of a key regulator of genes in response to acRoots was analyzed using quantitative polymerase chain reaction and western blotting. Results Treatment with acRoots (10 mg/mL) had no cytotoxicity in L02 cells and had a positive effect on L02 cell viability; however, it significantly inhibited HCC cell proliferation. Treatment with acRoots downregulated DLX2 gene expression in HCC cells, and high DLX2 expression was associated with advanced stage and poor prognosis in patients with HCC. Treatment with acRoots inhibited proliferation, invasion and migration, clonality, and the epithelial-to-mesenchymal transition, and promoted the apoptosis of HCC cells by downregulating DLX2 expression. HCC cells with higher DLX2 expression were more sensitive to acRoots. Conclusions acRoots inhibited the malignant biological behavior of HCC cells via regulation of the epithelial-mesenchymal transition (EMT) by DLX2. Graphical abstract fx1 [ABSTRACT FROM AUTHOR]

Abstract: Reference genes are used to normalize target genes for relative quantification in gene expression studies. However, different experimental conditions may affect the expression of reference genes, which could lead to erroneous quantitative results. In this study, we performed real-time polymerase chain to investigate the expression of eight reference genes (rpoN, rpoD, dbhA, phaF, 16S rRNA, gst, lexA, and atkA) in Pseudomonas putida mt-2 during degradation of p-xylene. According to their expression stability, geNorm software analysis revealed that rpoN, rpoD, 16S rRNA, and atkA were suitable reference genes with highly stable expression, whereas phaF and dbhA were not suitable due to unstable expression. When normalized either to phaF or dbhA, xylA and xylE expression were significantly different compared to the expression levels normalized with the normalization factor (NF4) obtained from the four most stable reference genes (rpoN, -rpoD, -16S rRNA, and -atkA). The use of unstably expressing reference genes resulted in an over- or underestimation of target gene expression, a delay in maximal gene expression, and an increase in gene expression in the absence of inducer. While experimental results indicated that the relative maximum expression of xylA and xylE occurred at different times, unstable reference genes indicated that the maximum expression occurred at the same time. Our study indicates that a valid set of reference genes covering a broad expression range is recommended to accurately normalize and quantify the relative expression levels of the target gene(s) transcripts in many microbial processes. [Copyright &y& Elsevier]