Role of autophagy in apoptosis induction by methylene chloride extracts of Mori cortex in NCI-H460 human lung carcinoma cells.

The root of Mori cortex has traditionally been used in Korea for the treatment of cutaneous inflammation, pulmonary asthma, and congestion for thousands of years. The present study was designed to validate the anticancer effects of methylene chloride extracts of the M. cortex root (MEMC) in NCI-H460 human lung carcinoma cells. Exposure to MEMC was found to result in growth inhibition by the induction of caspase­dependent apoptosis in NCI-H460 cells, which correlated with upregulated expression of death receptor (DR)4, DR5 and FasL, downregulation of anti-apoptotic Bcl-2 and Bcl-xL expression, cleavage of Bid, and loss of mitochondrial membrane potential. In addition, autophagosomes, a characteristic finding of autophagy, and markers of autophagy, conversion of microtubule-associated protein light chain-3 (LC3)-I to LC3-II and increased beclin-1 accumulation, were observed in MEMC-treated NCI-H460 cells. Inhibition of autophagy by 3-methyladenine or LC3B small interfering (siRNA) resulted in enhanced apoptotic cell death, suggesting that MEMC-induced autophagy functions as a suppressor of apoptosis. MEMC-induced autophagy was also blocked by N-acetyl-cysteine (NAC) and catalase, indicating that H2O2 can regulate autophagy. Our data demonstrate that MEMC triggers both ROS-mediated autophagy and caspase-dependent apoptosis, and that autophagy plays a protective role against apoptotic cell death.

Induction of G2/M arrest and apoptosis by water extract of Strychni Semen in human gastric carcinoma AGS cells.

The seed of Strychnos nux-vomica (Loganiaceae) has been used in traditional Oriental medicine as a folk remedy for the treatment of cancer. However, the mechanism responsible for the anticancer effects of Strychni Semen is not clearly understood. The study tested whether and how the water extract of Strychni Semen (ESS) treatment would affect the growth of AGS human gastric carcinoma cells. ESS was found to inhibit the growth of AGS cells in a concentration-dependent manner. Cell cycle analysis showed G2/M phase arrest and apoptosis in AGS cells following ESS treatment. ESS-mediated G2/M arrest was found to be associated with up-regulation of cyclin A, Cdc2, tumor suppressor p53 and cyclin dependent kinase (Cdk) inhibitor p21(WAF1/CIP1), whereas the expressions of other G2/M regulatory proteins, including cyclin B1 and Cdk2, were down-regulated compared with the control. The induction of apoptotic cell death by ESS was associated with down-regulation of anti-apoptotic Bcl-2 and up-regulation of pro-apoptotic Bax expression. Further results indicate that caspase-3, caspase-8 and caspase-9 are all activated by ESS, together with cleavage of downstream caspase-3 target proteins. Taken together, the results of this study suggest the involvement of multiple signaling pathways targeted by ESS in mediating G2/M cell cycle arrest and apoptosis in AGS cells, and warrant further investigation.

Effect of extracts from safflower seeds on osteoblast differentiation and intracellular calcium ion concentration in MC3T3-E1 cells.

Although safflower seeds have long been used in Korea as herbal medicines, very little research has been published on the effects of safflower seed on bone formation or bone density. The study reported here therefore examined bone nodule formation, calcium uptake, alkaline phosphatase activity, and intracellular concentration of calcium ion [Ca(2+)](i) in murine osteoblastic cells of the MC3T3-E1 line that were cultured on modified Eagle's minimal essential medium alone (controls) or with addition of 0.1% crude extract of safflower seed (experimental group I) or 0.1% aqueous fraction of safflower seed (experimental group II). Fluorescence spectrometry measurement of ([Ca(2+)](i)) showed significantly accelerated rates of osteoblast differentiation in experimental group I (3 microL of crude extract in 8 x 10(4) cells) and experimental group II (2 microL of aqueous fraction in 8 x 10(4) cells) compared to the control group.

Phosphorylation of p53, induction of Bax and activation of caspases during beta-lapachone-mediated apoptosis in human prostate epithelial cells.

The DNA topoisomerase I inhibitor beta-lapachone, the product of a tree from South America, is known to exhibit various biological properties, the mechanisms of which are poorly understood. We investigated the effects of beta-lapachone on the growth of human prostate epithelial cells. Upon treatment with beta-lapachone, a concentration-dependent inhibition of cell viability was observed and cells developed many of the hallmark features of apoptosis, including condensation of chromatin and DNA fragmentation. The apoptotic effects of beta-lapachone were associated with marked induction of p53 phosphorylation and Bax protein without altering the expression of p53 and Bcl-2 protein. In addition, the proteolytic cleavage of specific target proteins such as poly(ADP-ribose) polymerase, beta-catenin and Rad51, which are hallmarks of apoptosis, were observed, and Western blotting demonstrated that processing/activation of caspases release cytochrome c from the mitochondria into the cytosol and accompany the generation of beta-lapachone-mediating apoptotic cell death. However, beta-lapachone did not affect the levels of c-IAP family proteins. The present results suggest that apoptotic signals evoked by beta-lapachone in human prostate epithelial cells may converge caspases activation through up-regulation of phosphorylation of p53 and Bax rather than down-regulation of c-IAPs family.