Dehydroandrographolide enhances innate immunity of intestinal tract through up-regulation the expression of hBD-2.

BACKGROUND: Dehydroandrographolide (DA) is one of major active components in the well-known oriental herbal medicine Andrographis paniculata (Burm.f) Nees which belongs to the Acanthaceae family. DA is used for the treatment of infections in China. However, DA has not been found to significantly inhibit bacterial and viral growth directly. The current study investigates the effect of DA on the expression of human ß -defensin-2 (hBD-2) in human intestinal epithelial cells and the possible signaling pathways. METHODS: Human intestinal epithelial HCT-116 cells were incubated with 1-100 µM DA for 2-24 h. RT-PCR and Western blot were used to assess the expression of hBD-2. The specific inhibitors were used and the levels of phosphorylation of signaling molecules were detected for dissecting the signaling pathways leading to the induction of hBD-2. RESULTS: MTT assay showed there was no obvious cytotoxicity for HCT-116 cells by 1-100 µM DA treatment. RT-PCR and Western blot assays showed that DA (1-100 µM) could up-regulate the expression of hBD-2, and the effect lasted longer than 24 h. By using SB203580 and SB202190 (inhibitors of p38), the enhancement of hBD-2 expression were significantly attenuated. However, inhibitor of ERK and inhibitor of JNK could not block the effect of DA. Furthermore, Western blot found activation of p38 but not ERK and JNK in DA-treated HCT-116 cells. CONCLUSION: The results suggested that DA enhanced innate immunity of intestinal tract by up-regulating the expression of hBD-2 through the p38 MAPK pathways.

Apoptosis induced by ß,ß-dimethylacrylshikonin is associated with Bcl-2 and NF-κB in human breast carcinoma MCF-7 cells.

ß,ß-dimethylacrylshikonin (DA) is a natural naphthoquinone derivative compound of Lithospermum erythrorhizon with various biological activities. The present study aimed to investigate the inhibitory effects and underlying mechanisms of DA in human breast carcinoma MCF-7 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that DA inhibited the proliferation of MCF-7 cells in a dose- and time-dependent manner. The half maximal inhibitory concentration of DA with regard to the proliferation of MCF-7 cells was 0.050±0.016 mM. The characteristics of cell apoptosis, including cell shrinkage, nuclear pyknosis and chromatin condensation, were all observed in DA-treated cells. DA decreased the expression levels of Bcl-2 and increased the expression of Bax and caspase-3 compared with those in the control. DA inhibited the activity of the nuclear factor (NF)-κB pathway, by downregulating the expression of the p65 subunit, and inhibited the Iκb phosphorylation. DA inhibits the proliferation of MCF-7 cells in vitro by inducing apoptosis through the downregulation of Bcl-2, upregulation of Bax and partial inactivation of the NF-κB pathway.

Andrographolide exerted its antimicrobial effects by upregulation of human ß-defensin-2 induced through p38 MAPK and NF-κB pathway in human lung epithelial cells.

Andrographis paniculata (Burm. f) Nees is a traditional herbal medicine for the treatment of infection and inflammation in China. Andrographolide (andro) is one of the major components. Human ß-defensin-2 (hBD-2) is an inducible antimicrobial peptide that plays an important role in innate immunity. The present study aimed to investigate the effect of andro on upregulation of hBD-2 and the key signaling pathways involved in andro-induced hBD-2 expression. Real-time reverse transcription - PCR and Western blot assays showed that andro (1.0-10 µmol/L) can upregulate the expression of hBD-2 in a dose-dependent manner. Further studies suggested that hBD-2 mRNA and protein expression in responsive to andro were attenuated by pretreatment with SB203580 (an inhibitor of p38 mitogen-activated protein kinase (p38 MAPK)), MG-132 (an inhibitor of nuclear factor κB (NF-κB)), and an NF-κB activator inhibitor, but not by an inhibitor of ERK (PD98059) or by an inhibitor of JNK(SP600125). Moreover, we found that a second p38 MAPK inhibitor (SB202190) significantly blocked andro-mediated hBD-2 induction in SPC-A-1 lung epithelial cells. Finally, the p-c-Jun transcription factor activity assay also showed that AP-1 activity was induced by andro compared with the untreated group. We conclude that andro may exert its antimicrobial effects by upregulating the expression of hBD-2 through the p38 MAPK and NF-κB pathway.

Inhibitory effects of ß,ß-dimethylacrylshikonin on hepatocellular carcinoma in vitro and in vivo.

ß,ß-Dimethylacrylshikonin is one of the most abundant naphthoquinones in the root extracts of Lithospermum erythrorhizon Sieb. et Zucc. (Boraginaceae), which have been reported to have antitumor effects. This study evaluated the antiproliferative activity of ß,ß-dimethylacrylshikonin on human hepatocellular carcinoma (HCC) cells both in vitro and in vivo. In vitro, the MTT assay showed that ß,ß-dimethylacrylshikonin inhibited the proliferation of SMMC-7721 cells in both dose- and time-dependent manners with its 50% inhibitory concentration (IC(50) ) at 48 h being 15.01 ± 0.76 µg/mL. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) and Hoechst staining detected the characteristics of cell apoptosis in ß,ß-dimethylacrylshikonin-treated cells and the apoptotic rates of treated groups were increased in a dose-dependent manner. Flow cytometric analysis revealed that ß,ß-dimethylacrylshikonin could block the cell cycle arrest at G2 phase. Furthermore, ß,ß-dimethylacrylshikonin down-regulated the mRNA and protein expression of Bcl-2 but up-regulated that of Bax. The cleaved caspase-3 protein was also detected in treated cells. The experiment in vivo showed that ß,ß-dimethylacrylshikonin significantly suppressed the growth of H(22) transplantable hepatoma, and induced the activation of caspase-3 determined by immunohistochemistry. The results indicate that ß,ß-dimethylacrylshikonin has significant antitumor effects on hepatocellular carcinoma both in vitro and in vivo.