Tung-Hu Tsai

Oct-4 Expression Maintained Cancer Stem-Like Properties in Lung Cancer-Derived CD133-Positive Cells

CD133 (prominin-1), a 5-transmembrane glycoprotein, has recently been considered to be an important marker that represents the subset population of cancer stem-like cells. Herein we report the isolation of CD133-positive cells (LC-CD133+) and CD133-negative cells (LC-CD133−) from tissue samples of ten patients with non-small cell lung cancer (LC) and five LC cell lines. LC-CD133+ displayed higher Oct-4 expressions with the ability to self-renew and may represent a reservoir with proliferative potential for generating lung cancer cells. Furthermore, LC-CD133+, unlike LC-CD133−, highly co-expressed the multiple drug-resistant marker ABCG2 and showed significant resistance to chemotherapy agents (i.e., cisplatin, etoposide, doxorubicin, and paclitaxel) and radiotherapy. The treatment of Oct-4 siRNA with lentiviral vector can specifically block the capability of LC-CD133+ to form spheres and can further facilitate LC-CD133+ to differentiate into LC-CD133−. In addition, knock-down of Oct-4 expression in LC-CD133+ can significantly inhibit the abilities of tumor invasion and colony formation, and increase apoptotic activities of caspase 3 and poly (ADP-ribose) polymerase (PARP). Finally, in vitro and in vivo studies further confirm that the treatment effect of chemoradiotherapy for LC-CD133+ can be improved by the treatment of Oct-4 siRNA. In conclusion, we demonstrated that Oct-4 expression plays a crucial role in maintaining the self-renewing, cancer stem-like, and chemoradioresistant properties of LC-CD133+. Future research is warranted regarding the up-regulated expression of Oct-4 in LC-CD133+ and malignant lung cancer.

Curcumin and its nano-formulation: The kinetics of tissue distribution and blood–brain barrier penetration

Curcumin has considerable neuro-protective and anti-cancer properties but is rapidly eliminated from the body. By optimizing the HPLC method for analysis of curcumin, this study evaluates how the ability of curcumin to penetrate organs and different regions of the brain is affected by nanoparticulation to increase curcumin circulation time in the body. Curcumin-loaded PLGA nanoparticles (C-NPs) were prepared by the high-pressure emulsification-solvent evaporation method. The mean particle size and entrapment efficiency were 163nm and 46.9%, respectively. The release profile of C-NPs was an initial burst effect followed by sustained diffusion. In distribution studies, curcumin could be detected in the evaluated organs, including liver, heart, spleen, lung, kidney and brain. C-NPs were found mainly in the spleen, followed by the lung. Formulation significantly raised the curcumin concentration in these organs with increases in the AUC, t(1/2) and MRT of curcumin, though this was not apparent in the heart. Curcumin and C-NPs could cross the blood-brain barrier (BBB) to enter brain tissue, where it was concentrated chiefly in the hippocampus. Nanoparticulation significantly prolonged retention time of curcumin in the cerebral cortex (increased by 96%) and hippocampus (increased by 83%). These findings provide further understanding for the possible therapeutic effects of curcumin and C-NPs in further pre-clinical and clinical research.

Periodic maternal separation of neonatal rats produces region- and gender-specific effects on biogenic amine content in postmortem adult brain

Early environment exerts profound effects on mammalian behavioral and neural development. The aim of this study was to describe changes in adult neurochemistry in the rat following repeated neonatal maternal separation (RMS) during the preweaning period, a procedure known to induce enduring behavioral effects. Following RMS, rats show an attenuated locomotor response to novelty, to D‐amphetamine, and attenuated behavioral responses for conditioned incentives as adults. These behavioral effects are broadly opposite in direction to those found following postweaning isolation rearing. Isolation rearing‐induced behavioral changes are associated with profound changes in central monoamine function. Following RMS, adult rats had increased tissue levels of dopamine in both dorsal and ventral striatum. The turnover of dopamine, as determined by the ratio of DOPAC to dopamine, was decreased in the mPFC of RMS subjects. Serotonin levels were reduced in dorsal hippocampus of RMS rats of both sexes and in the mPFC of male RMS rats. Noradrenaline levels were increased in the dorsal hippocampus in female, but not in male, RMS rats. These data provide evidence that, in addition to the adult behavioral consequences, RMS leads to profound, region‐, and gender‐specific changes in brain monoamine content. The developmental specificity of these results is discussed with respect to their possible role in altered behavioral development and psychopathology. Synapse 40:1–10, 2001.

Effect of green tea extract on obese women: a randomized, double-blind, placebo-controlled clinical trial.

AIMS To examine the effect of green tea extract (GTE) on obese women and to explore the relationship between GTE and obesity-related hormone peptides. METHODS A randomized, double-blind, placebo-controlled clinical trial was conducted from July 2006 to June 2007 in Taipei Hospital, Taiwan. Seventy-eight of 100 obese women aged between 16 and 60 years with BMI>27 kg/m(2) and who had not received any other weight control maneuvers within the last 3 months completed this study. The subjects were randomly divided into Groups A and B. Group A (n=41) received GTE while Group B (n=37) took cellulose as a placebo, one capsule (400mg) three times each day for 12 weeks. The body weight (BW), body mass index (BMI) and waist circumflex (WC) were measured at the beginning of the study and after 12 weeks of treatment with GTE. The data were compared and expressed as % reduction. RESULTS There was only a 0.3% reduction in BW (0.15 kg) after 12 weeks of treatment with GTE. There was no statistical difference in % reduction in BW, BMI and WC between the GTE and placebo groups. Within group comparison revealed that the GTE group had significant reduction in LDL-cholesterol and triglyceride, and marked increase in the level of HDL-cholesterol, adiponectin and ghrelin. On the other hand, the placebo group showed significant reduction in triglyceride only, and a marked increase in the level of ghrelin alone. CONCLUSIONS This study showed no statistical difference in % reduction in BW, BMI and WC between the GTE and placebo groups after 12 weeks of treatment. The intake of GTE (491 mg catechins containing 302 mg EGCG) for 12 weeks is considered safe as shown by the results.

Effects of lipophilic emulsifiers on the oral administration of lovastatin from nanostructured lipid carriers: Physicochemical characterization and pharmacokinetics

Nanostructured lipid carriers (NLCs) made from mixtures of Precirol and squalene were prepared to investigate whether the bioavailability of lovastatin can be improved by oral delivery. The size, zeta potential, drug-loading capacity, and release properties of the NLCs were compared with those of lipid nanoparticles containing pure Precirol (solid lipid nanoparticles, SLNs) and squalene (lipid emulsions, LEs). Stable nanoparticles with a mean size range of 180-290 nm and zeta potential range of -3 to -35 mV were developed. More than 70% lovastatin was entrapped in the NLCs and LEs, which was significantly higher compared to the SLNs. The in vitro release kinetics demonstrated that lovastatin release could be reduced by up to 60% with lipid nanoparticles containing Myverol as the lipophilic emulsifier, which showed a decreasing order of NLCs>LEs>SLNs. Drug release was further decreased by soybean phosphatidylcholine (SPC) incorporation, with NLCs and SLNs showing the slowest delivery. The oral lovastatin bioavailability was enhanced from 4% to 24% and 13% when the drug was administered from NLCs containing Myverol and SPC, respectively. The in vivo real-time bioluminescence imaging indicated superior stability of the Myverol system over the SPC system in the gastric environment.

Down-modulation of Bcl-XL, release of cytochrome c and sequential activation of caspases during honokiol-induced apoptosis in human squamous lung cancer CH27 cells

Honokiol is a phenolic compound purified from Magnolia officinalis, which induced the apoptotic cell death in several types of human cancer cells. In the present study, the molecular mechanism of honokiol-mediated apoptotic process was examined in human squamous lung cancer CH27 cells. Here, we found that honokiol-induced apoptotic cell death was accompanied by upregulation of Bad and downregulation of Bcl-XL, while honokiol had no effect on the levels of Bcl-2, Bcl-XS, Bag-1, Bax and Bak proteins. Moreover, honokiol treatment caused the release of mitochondrial cytochrome c to cytosol and sequential activation of caspases. Proteolytic activation of caspase-3 and cleavage of PARP, an in vivo substrate for caspase-3, were observed in honokiol-treated CH27 cells. Furthermore, treatment with caspase inhibitors z-DEVD-fmk and z-VAD-fmk markedly blocked honokiol-induced apoptosis. These results demonstrated that modulation of Bcl-XL and Bad proteins, release of mitochondrial cytochrome c and activation of caspase-3, participated in honokiol-triggered apoptotic process in human squamous lung cancer CH27 cells.

Neuroprotection by Imipramine against lipopolysaccharide-induced apoptosis in hippocampus-derived neural stem cells mediated by activation of BDNF and the MAPK pathway

Depression is accompanied by the activation of the inflammatory-response system, and increased production of proinflammatory cytokines may play a role in the pathophysiology of depressive disorders. Imipramine (IM), a tricyclic antidepressant drug, has recently been shown to promote neurogenesis and improve the survival rate of neurons in the hippocampus. However, whether IM elicits a neuroprotective or anti-inflammatory effect, or promotes the differentiation of neural stem cells (NSCs) remains to be elucidated. In this study, we cultured NSCs derived from the hippocampal tissues of adult rats as an in vitro model to evaluate the NSCs drug-modulation effects of IM. Our results showed that 3 microM IM treatment significantly increased the survival rate of NSCs, and up-regulated the mRNA and protein expression of brain-derived neurotrophic factor (BDNF) and Bcl-2 in Day-7 IM-treated NSCs. Similar to BDNF-treated effect, incubation of NSCs with 3 microM IM increased Bcl-2 protein levels and further prevented lipopolysaccharide (LPS)-induced apoptosis through the activation of the mitogen-activated protein kinase (MAPK)/extracellular-regulated kinase (ERK) pathway. Inhibition of BDNF expression with small interfering RNA (siRNA), or blocking the MAPK pathway with U0126 further significantly decreased Bcl-2 protein levels and abrogated the neuroprotective effects of IM against LPS-induced apoptosis in NSCs. In addition, the percentages of serotonin and MAP-2-positive neuronal cells in the Day 7 culture of IM-treated NSCs were significantly increased. By using microdialysis with high performance liquid chromatography-electrochemical detection, the functional release of serotonin in the process of serotoninergic differentiation of IM-treated NSCs was concomitantly increasing and mediated by the activation of the BDNF/MAPK/ERK pathway/Bcl-2 cascades. In sum, the study results indicate that IM can increase the neuroprotective effects, suppress the LPS-induced inflammatory process, and promote serotoninergic differentiation in NSCs via the modulation of the BDNF/MAPK/ERK pathway/Bcl-2 cascades.

Drug-drug interactions of silymarin on the perspective of pharmacokinetics.

Silymarin, which is extracted from the milk thistle (Silybum marianum), has been used for centuries for treating hepatic disorders and its hepatoprotective effects have been known for hundreds of years. Silymarin is a mixture of polyphenoic flavonoids, which include silibinin (silybin A and silybin B), isosilyin A and B, silychristin A and B, silydianin and other phenol compounds. The pharmacokinetics of silibinin shows fast absorption and elimination. Silymarin undergoes phase I and phase II metabolism, especially phase II conjugation reactions, it undergoes multiple conjugation reactions, and is primarily excreted into bile and urine. Silymarin has a good safety profile, but little is known regarding its potential for drug interaction. Silymarin has limited effect on the pharmacokinetics of several drugs in vivo; despite silymarin decreasing the activity of cytochrome P-450 (CYPs) enzymes, UDP-glucuronosyltransferase (UGT) enzyme, and reducing P-glycoprotein (P-gp) transport. Health-care practitioners should caution patients against co-administration of silymarin and pharmaceutical drugs.

Hydroxyl radical in living systems and its separation methods.

It has recently been shown that hydroxyl radicals are generated under physiological and pathological conditions and that they seem to be closely linked to various models of pathology putatively implying oxidative stress. It is now recognized that the hydroxyl radical is well-regulated to help maintain homeostasis on the cellular level in normal, healthy tissues. Conversely, it is also known that virtually every disease state involves free radicals, particularly the most reactive hydroxyl radical. However, when hydroxyl radicals are generated in excess or the cellular antioxidant defense is deficient, they can stimulate free radical chain reactions by interacting with proteins, lipids, and nucleic acids causing cellular damage and even diseases. Therefore, a confident analytical approach is needed to ascertain the importance of hydroxyl radicals in biological systems. In this paper, we provide information on hydroxyl radical trapping and detection methods, including liquid chromatography with electrochemical detection and mass spectrometry, gas chromatography with mass spectrometry, capillary electrophoresis, electron spin resonance and chemiluminescence. In addition, the relationships between diseases and the hydroxyl radical in living systems, as well as novel separation methods for the hydroxyl radical are discussed in this paper.

Determination of melamine in rat plasma, liver, kidney, spleen, bladder and brain by liquid chromatography–tandem mass spectrometry

In this study, we describe a method for the analysis of melamine in rat plasma, liver, kidney, spleen, bladder, and brain using trichloroacetic acid precipitation with mixed-mode cation-exchange solid-phase extraction and hydrophilic interaction chromatography coupled to tandem mass spectrometry detection. Method validation was investigated completely, including linearity, precision, accuracy, matrix effect, extraction recovery, and carryover for the determination of melamine. The method exhibited a good linear range covering 20-500 ng/mL, and the overall precision ranged from 1.6 to 16.3%, with the accuracy varying from -7.9 to 15.1%. The mean matrix effects of melamine in rat plasma, liver, kidney, spleen, bladder, and brain ranged from 66.2+/-6.7 to 95.5+/-13.2%, and the mean recoveries for melamine varied from 79.8+/-8.2 to 113.0+/-9.6%. Rat kidney showed the highest level among the organs (192.5% of the plasma melamine level), and the average concentration of melamine in the brain was only 7.5% of the plasma melamine concentration. This work has pointed out that even with the application of two popular preparation procedures (acid precipitation and solid-phase extraction) of melamine, the matrix effect in analyzing biological samples still exists in certain kinds of matrices.