Christopher H.K. Cheng

Lignans and isoflavonoids in plasma and prostatic fluid in men: samples from Portugal, Hong Kong, and the United Kingdom.

Chinese men have lower incidences of prostate cancer compared to men from Europe and North America. Asians consume large quantities of soya, a rich source of isoflavanoids phyto‐oestrogens and have high plasma and urinary levels of these compounds. The mammalian lignans, enterolactone and enterodiol, are another group of weak plant oestrogens and are derived from seeds, cereals and grains. Vegetarians have high plasma and urinary concentrations of lignans.

Efficient targeted gene disruption in Xenopus embryos using engineered transcription activator-like effector nucleases (TALENs).

Transcription activator-like effector nucleases (TALENs) are an approach for directed gene disruption and have been proved to be effective in various animal models. Here, we report that TALENs can induce somatic mutations in Xenopus embryos with reliably high efficiency and that such mutations are heritable through germ-line transmission. We modified the Golden Gate method for TALEN assembly to make the product suitable for RNA transcription and microinjection into Xenopus embryos. Eight pairs of TALENs were constructed to target eight Xenopus genes, and all resulted in indel mutations with high efficiencies of up to 95.7% at the targeted loci. Furthermore, mutations induced by TALENs were highly efficiently passed through the germ line to F1 frogs. Together with simple and reliable PCR-based approaches for detecting TALEN-induced mutations, our results indicate that TALENs are an effective tool for targeted gene editing/knockout in Xenopus.

Akt and Autophagy Cooperate to Promote Survival of Drug-Resistant Glioma

Combined inhibition of PI3K, mTOR, and autophagy promotes glioma cell death. Blocking All Escape Routes Many cancers, including glioma, are associated with increased signaling through the phosphatidylinositol 3-kinase to Akt to mammalian target of rapamycin (PI3K-Akt-mTOR) pathway, which promotes cell growth, proliferation, and survival. This suggests that pharmacological inhibition of key kinases in this pathway could provide an approach to antineoplastic therapy. Disappointingly, however, inhibitors of PI3K, Akt, or mTOR typically block cancer cell growth rather than eliciting the death of malignant cells, limiting their utility as antineoplastic agents. Noting that autophagy, a process of autodigestion that can enable cells to endure periods of stress and nutrient deprivation, could provide a survival mechanism under conditions of decreased PI3K-Akt-mTOR signaling, Fan et al. explored the effects of various combinations of kinase and autophagy inhibitors on glioma cell survival. Inhibition of mTOR complex 1 (mTORC1) with rapamycin induced autophagy; however, cells survived the combination of rapamycin with inhibitors of autophagy by activating Akt signaling. In contrast, the combined inhibition of mTORC1, PI3K, and autophagy, or that of mTORC1, mTORC2, and autophagy, triggered apoptosis—the process of programmed cell death. The authors elicited cell death with combinations of drugs that are either now in use in patients or in clinical trials, raising the hope that this approach could be readily translatable to human therapy. Although the phosphatidylinositol 3-kinase to Akt to mammalian target of rapamycin (PI3K-Akt-mTOR) pathway promotes survival signaling, inhibitors of PI3K and mTOR induce minimal cell death in PTEN (phosphatase and tensin homolog deleted from chromosome 10) mutant glioma. Here, we show that the dual PI3K-mTOR inhibitor PI-103 induces autophagy in a form of glioma that is resistant to therapy. Inhibitors of autophagosome maturation cooperated with PI-103 to induce apoptosis through the mitochondrial pathway, indicating that the cellular self-digestion process of autophagy acted as a survival signal in this setting. Not all inhibitors of mTOR synergized with inhibitors of autophagy. Rapamycin delivered alone induced autophagy, yet cells survived inhibition of autophagosome maturation because of rapamycin-mediated activation of Akt. In contrast, adenosine 5′-triphosphate–competitive inhibitors of mTOR stimulated autophagy more potently than did rapamycin, with inhibition of mTOR complexes 1 and 2 contributing independently to induction of autophagy. We show that combined inhibition of PI3K and mTOR, which activates autophagy without activating Akt, cooperated with inhibition of autophagy to cause glioma cells to undergo apoptosis. Moreover, the PI3K-mTOR inhibitor NVP-BEZ235, which is in clinical use, synergized with the lysosomotropic inhibitor of autophagy, chloroquine, another agent in clinical use, to induce apoptosis in glioma xenografts in vivo, providing a therapeutic approach potentially translatable to humans.

Inhibition of xanthine oxidase by some Chinese medicinal plants used to treat gout.

The enzyme xanthine oxidase catalyses the oxidation of hypoxanthine to xanthine and then to uric acid, which plays a crucial role in gout. A total of 122 traditional Chinese medicinal plants, selected according to the clinical efficacy and prescription frequency for the treatment of gout and other hyperuricemia-related disorders, have been evaluated for the enzyme inhibitory activity. Among the 122 methanol extracts derived from these species, 69 were shown to be inhibitory at 100 microg/ml, with 29 having greater than 50% inhibition. As to the equal amount of water extracts, 40 were disclosed to be active at 100 microg/ml, with 13 possessing more than 50% inhibition. At 50 microg/ml, 58 methanol and 24 water extracts exhibited inhibitory activity, with 15 of the former and two of the latter showing greater than 50% inhibition. The most active was the methanol extract of the twig of Cinnamomum cassia (Lauraceae) (IC(50), 18 microg/ml), which was followed immediately by those of the flower of Chrysanthemum indicum (Asteraceae) (IC(50), 22 microg/ml) and the leaves of Lycopus europaeus (Lamiatae) (IC(50), 26 microg/ml). Among the water extracts, the strongest inhibition of the enzyme was observed with that of the rhizome of Polygonum cuspidatum (Polygonaceae) (IC(50), 38 microg/ml). The IC(50) value of allopurinol used as a positive control was 1.06 microg/ml. The study demonstrated that the effects for these medicinal plants used for the gout treatment were based, at least in part, on the xanthine oxidase inhibitory action.

Bufalin induces autophagy-mediated cell death in human colon cancer cells through reactive oxygen species generation and JNK activation

Colorectal cancer is the second most common cause of cancer death in the world and about half of the patients with colorectal cancer require adjuvant therapy after surgical resection. Therefore, the eradication of cancer cells via chemotherapy constitutes a viable approach to treating patients with colorectal cancer. In this study, the effects of bufalin isolated from a traditional Chinese medicine were evaluated and characterized in HT-29 and Caco-2 human colon cancer cells. Contrary to its well-documented apoptosis-promoting activity in other cancer cells, bufalin did not cause caspase-dependent cell death in colon cancer cells, as indicated by the absence of significant early apoptosis as well as poly(ADP-ribose) polymerase and caspase-3 cleavage. Instead, bufalin activated an autophagy pathway, as characterized by the accumulation of LC3-II and the stimulation of autophagic flux. The induction of autophagy by bufalin was linked to the generation of reactive oxygen species (ROS). ROS activated autophagy via the c-Jun NH(2)-terminal kinase (JNK). JNK activation increased expression of ATG5 and Beclin-1. ROS antioxidants (N-acetylcysteine and vitamin C), the JNK-specific inhibitor SP600125, and JNK2 siRNA attenuated bufalin-induced autophagy. Our findings unveil a novel mechanism of drug action by bufalin in colon cancer cells and open up the possibility of treating colorectal cancer through a ROS-dependent autophagy pathway.

Identification and functional characterization of two alternatively spliced growth hormone secretagogue receptor transcripts from the pituitary of black seabream Acanthopagrus schlegeli

Two cDNA transcripts, namely sbGHSR-1a and sbGHSR-1b, for growth hormone secretagogue receptor (GHSR), were identified from the seabream pituitary. When translated, the sbGHSR-1a encodes for a protein of 385 amino acids (aa) with seven putative transmembrane domains and the sbGHSR-1b contains 295 aa with five putative transmembrane domains. Tissue distribution studies indicated that the two receptors are mainly expressed in the central nervous system of the fish. The sbGHSR-1a transcript has the highest expression level in the pituitary. The sbGHSR-1b transcript, on the other hand, has the highest expression level in the telencephalon. Genomic Southern analysis indicated that there is a single gene for GHSR in the seabream genome. Comparison of the cDNA sequences of sbGHSR1a and sbGHSR1b with the seabream genomic sequence indicated that the presence of the two receptor transcripts is a result of alternative splicing of the single GHSR gene. The two receptor cDNAs were expressed in cultured eukaryotic cells for functional analyses. A variety of structurally diverse growth hormone secretogogues (GHS), including the peptide GHS (GHRP-6 and ghrelin), the benzolactam GHS (L692,585) and the spiropiperidine GHS (L163,255), were able to trigger an elevation of intracellular Ca(2+) ion concentration in HEK293 cells expressing sbGHSR-1a, but not in cells expressing sbGHSR-1b. Microphysiometry revealed that an increase in extracellular acidification rate (EAR) could be detected in CHO cells expressing the sbGHSR-1a receptor when stimulated with GHRP-6. On the contrary, CHO cells expressing the sbGHSR-1b receptor registered no detectable EAR changes. However, when sbGHSR-1b was co-expressed with sbGHSR-1a in HEK293 cells, the signal transduction capacity of sbGHSR-1a was attenuated. This is the first report on the identification of a GHSR-1b transcript from species other than mammals and the demonstration that receptor interaction might provide a possible explanation for the existence and biological significance of the sbGHSR-1b transcript.

Structural and functional multiplicity of the kisspeptin/GPR54 system in goldfish (Carassius auratus)

To ascertain the neuroendocrine function of the kisspeptin/GPR54 system in non-mammalian species, full-length cDNAs encoding for Kiss1 and Kiss2 as well as their putative cognate receptors GPR54a and GPR54b, were isolated from goldfish (Carassius auratus). The deduced protein sequences between Kiss1 and Kiss2 in goldfish share very low similarity, but their putative mature peptides (kisspeptin-10) are relatively conserved. RT-PCR analysis demonstrated that the goldfish kiss1 gene (gfkiss1) is highly expressed in the optic tectum-thalamus, intestine, kidney, and testis, while the goldfish kiss2 gene (gfkiss2) is mainly detected in the hypothalamus, telencephalon, optic tectum thalamus, adipose tissue, kidney, heart, and gonads. The two receptor genes (gfgpr54a and gfgpr54b) are highly expressed in the brain regions including telencephalon, optic tectum thalamus, and hypothalamus. Both mature goldfish kisspeptin-10 peptides (gfKiss1-10 and gfKiss2-10) are biologically active as they could functionally interact with the two goldfish receptors expressed in cultured eukaryotic cells to trigger the downstream signaling pathways with different potencies. The actions of gfKiss1-10 and gfKiss2-10 on LH secretion were further investigated in vitro and in vivo. Intraperitoneal administration of gfKiss1-10 to sexually mature female goldfish could increase the serum LH levels. However, this peptide does not significantly influence LH release from goldfish pituitary cells in primary culture, indicating that the peptide does not exert its actions at the pituitary level. On the other hand, gfKiss2-10 appears to be a much less potent peptide as it exhibits no significant in vivo bioactivity and is also inactive on the primary pituitary cells.

Inhibition of xanthine oxidase by liquiritigenin and isoliquiritigenin isolated from Sinofranchetia chinensis.

Abstract. The methanol extract of the stem of Sinofranchetia chinensis inhibited the activity of xanthine oxidase in vitro. Bioassay-guided purification led to the isolation of liquiritigenin and isoliquiritigenin as the main xanthine oxidase inhibitors. This inhibition of enzyme activity was found to be dose dependent, with an IC50 value of approximately 49.3 μM for liquiritigenin and 55.8 μM for isoliquiritigenin. Lineweaver-Burk transformation of the inhibition data indicated that the inhibition was of a mixed type for both liquiritigenin and isoliquiritigenin. For liquiritigenin, the Ki and KI were determined to be 14.0 μM and 151.6 μM, respectively. For isoliquiritigenin, the Ki and KI were determined to be 17.4 μM and 81.9 μM, respectively. These results suggest that these natural products could be used to treat conditions where the inhibition of xanthine oxidase is warranted.

Oncoproteomics: current trends and future perspectives

Oncoproteomics is the application of proteomics technologies in oncology. Functional proteomics is a promising technique for the rational identification of biomarkers and novel therapeutic targets for cancers. Recent progress in proteomics has opened new avenues for tumor-associated biomarker discovery. With the advent of new and improved proteomics technologies, such as the development of quantitative proteomic methods, high-resolution, -speed and -sensitivity mass spectrometry and protein arrays, as well as advanced bioinformatics for data handling and interpretation, it is now possible to discover biomarkers that can reliably and accurately predict outcomes during cancer management and treatment. However, there are several difficulties in the study of proteins/peptides that are not inherent in the study of nucleic acids. New challenges arise in large-scale proteomic profiling when dealing with complex biological mixtures. Nevertheless, oncoproteomics offers great promise for unveiling the complex molecular events of tumorigenesis, as well as those that control clinically important tumor behaviors, such as metastasis, invasion and resistance to therapy. In this review, the development and advancement of oncoproteomics technologies for cancer research in recent years are expounded.

Cultural Dimensions, Gender, and the Nature of Self-concept: A Fourteen-country Study

Ratings of the importance of and satisfaction with 20 areas of the self were obtained from 3604 first or second year social science undergraduates from 14 countries (15 cultures). Factor analysis at the culture by gender level supported four factors for both sets of ratings. The resulting factor scores were analyzed for mean differences according to the cultural dimension of Individualism-Collectivism by Gender and by correlations with other cultural dimenions and economic indicators. It was found that participants from the 10 collectivist cultures placed greater salience for their self-concepts on “family values” than did those from the individualist cultures. However, this cultural difference was not found for “social relationships”. The expected gender differences, with females valuing “family values” and “social relationships” more highly, were found only for the individualist countries. The findings indicate that there may be a strong cultural level interaction effect between gender and Individualism-Collectivism on the nature of self-conceptions, and that the “family” and “social” aspects of self-concept in collectivist countries need to be considered separately.